Nour Cindy Patinote
email: cindy.patinote@umontpellier.fr

Bou Karroum

Georges Moarbess

Natalina Cirnat

Issam Kassab

Pierre-Antoine Bonnet

Carine Deleuze-Masquéfa

Nour Bou Karroum

Pierre- Antoine Bonnet

Agonist and antagonist ligands of toll-like receptors 7 and 8: Ingenious tools for therapeutic purposes

diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

I. Introduction

The innate immunity is composed of several families of pattern recognition receptors (PPRs). The latter receptors serve to identify pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs). The PPRs act as the primary defense against pathogenic entities and control the activation and progression of the adaptive immunity by activating the production not only of proinflammatory cytokines, chemokines and interferons, but also B and T cells. Among the PPRs, the Toll-like Receptors (TLRs) are of special interest. Their discovery more than 30 years ago has improved knowledge in the regulation of innate immunity, inflammation and cytokines induction.

TLRs are divided into two groups depending on their subcellular localization, which largely correlate with the type of molecular patterns they are able to recognize. Among endosomal TLRs, TLR7 and TLR8 have a high degree of sequence homology and function similarity. Both receptors recognize viral single-stranded RNAs, as well as synthetic tricyclic derivatives belonging to the imidazo [4,5-c]quinoline series such as resiquimod [START_REF] Chouchou | Deleuze-Masquéfa, Imidazo[1,2-a]quinoxalines Derivatives Grafted with Amino Acids: Synthesis and Evaluation on A375 Melanoma Cells[END_REF], a TLRs 7/8 agonist (Figure 1). Many novel TLRs 7/8 agonists from different chemical series have been developed these last years. The immunostimulating activity of these agonists allows them to intervene in the therapy of several diseases and to be valuable vaccine adjuvant candidates. In contrast, chronic immune activation by a sustained TLR activation is a hallmark of several infectious, autoimmune and malignant diseases. Therefore, the development of TLRs 7/8 antagonists may also play an important role in the therapy of these diseases. The evidence of the therapeutic potential of TLRs 7/8 ligands has prompted many pharmaceutical companies and research institutes to design, synthetize and optimize TLRs modulators. Our research team has attached great importance to the immunomodulatory potential of heterocycles known for their antitumoral and antiparasitic activities. [START_REF] Chouchou | Deleuze-Masquéfa, Imidazo[1,2-a]quinoxalines Derivatives Grafted with Amino Acids: Synthesis and Evaluation on A375 Melanoma Cells[END_REF][START_REF] Hajj | EAPB0503: An Imiquimod analog with potent in vitro activity against cutaneous leishmaniasis caused by Leishmania major and Leishmania tropica[END_REF][START_REF] Zghaib | New imidazoquinoxaline derivatives: Synthesis, biological evaluation on melanoma, effect on tubulin polymerization and structure-activity relationships[END_REF][START_REF] Khier | Metabolism and Pharmacokinetics of EAPB0203 and EAPB0503, Two Imidazoquinoxaline Compounds Previously Shown to Have Antitumoral Activity on Melanoma and T-Lymphomas[END_REF][START_REF] Khier | Pharmacology of EAPB0203, a novel imidazo[1,2a]quinoxaline derivative with anti-tumoral activity on melanoma[END_REF][START_REF] Deleuze-Masquefa | New Imidazo[1,2-A]quinoxalines and Derivates Thereof for the Treatment of Cancer[END_REF][7][START_REF] Saliba | EAPB0503, a novel imidazoquinoxaline derivative, inhibits growth and induces apoptosis in chronic myeloid leukemia cells[END_REF] The structural analogies between our synthetized imidazo [1,5-a]quinoxalines, pyrazolo [1,5-a]quinoxalines, imidazo[1,2-a]pyrazines and imiquimod, set the future lines of our research both in chemistry and biology, allowing us to focus on the TLRs 7/8 immunomodulatory effects of bi-and tri-cyclic derivatives. [START_REF] Bou Karroum | Novel and Selective TLR7 Antagonists among the Imidazo[1,2-a]pyrazines, Imidazo[1,5-a]quinoxalines, and Pyrazolo[1,5-a]quinoxalines Series[END_REF][START_REF] Patinote | Imidazo[1,2-a]pyrazine, Imidazo[1,5-a]quinoxaline and Pyrazolo[1,5a]quinoxaline derivatives as IKK1 and IKK2 inhibitors[END_REF][START_REF] Moarbess | New IKK inhibitors: Synthesis of new imidazo[1,2a]quinoxaline derivatives using microwave assistance and biological evaluation as IKK inhibitors[END_REF] In this review, we gather more than 340 publications until January 2020 relating to TLR7 and TLR8 modulators. By the extent of these data, we show in this review that the modulation of TLR activity is not easy despite many SAR studies around various recognized chemical cores. Not only minimal structures but also macromolecules have been considered and prepared by researchers. In the first chapter, we briefly present data concerning the expression, the structure and the signaling pathways of TLR7 and TLR8. This information will help to better understand the importance of targeting these receptors and the interest of preparing compounds able to modulate the inherent biological responses. We then focus in a second and a third chapter on the implication of TLR7 and TLR8 agonists and antagonists, respectively, in therapeutic treatments, and describe the natural and synthetic ligands focusing mainly on small organic compounds. In a final chapter, we have exposed all the clinical cases using agonist and antagonist ligands clearly referring to receptors.

II. Presentation of the TLR7 and TLR8 receptors

Localization and expression

TLRs are expressed on immune cells such as macrophages, dendritic cells, B and T cells, neutrophils, natural killer cells (NK), monocytes, eosinophils and non-immune cells such as fibroblasts, keratinocytes, the epithelial cells of the intestinal, respiratory and urogenital tracts. [START_REF] Iwasaki | Toll-like receptor control of the adaptive immune responses[END_REF][START_REF] Hornung | Quantitative expression of toll-like receptor 1-10 mRNA in cellular subsets of human peripheral blood mononuclear cells and sensitivity to CpG oligodeoxynucleotides[END_REF] TLR1, TLR2, TLR4, TLR5, TLR6 and TLR10, identified in mice and humans, are expressed on the cellular plasma membrane while TLR3, TLR7, TLR8 and TLR9 are expressed on the endosome membrane in mice and humans. [START_REF] Brubaker | Innate immune pattern recognition: a cell biological perspective[END_REF][START_REF] Leifer | Molecular mechanisms of regulation of Toll-like receptor signaling[END_REF] TLR11, TLR12 and TLR13 are endosomal receptors specific for mice. [START_REF] Ewald | Nucleic acid sensing Toll-like receptors in autoimmunity[END_REF][START_REF] Yarovinsky | TLR11 activation of dendritic cells by a protozoan profilinlike protein[END_REF][START_REF] Koblansky | Recognition of profilin by Toll-like receptor 12 is critical for host resistance to Toxoplasma gondii[END_REF][START_REF] Wang | Structure of the mouse Toll-like receptor 13 ectodomain in complex with a conserved sequence from bacterial 23S ribosomal RNA[END_REF] TLR7 is predominantly expressed in plasmacytoid dendritic cells (pDC) and B cells. [START_REF] Hornung | Quantitative expression of toll-like receptor 1-10 mRNA in cellular subsets of human peripheral blood mononuclear cells and sensitivity to CpG oligodeoxynucleotides[END_REF][START_REF] Edwards | Toll-like receptor expression in murine DC subsets: lack of TLR7 expression by CD8 alpha+ DC correlates with unresponsiveness to imidazoquinolines[END_REF][START_REF] Barr | TLR-mediated stimulation of APC: Distinct cytokine responses of B cells and dendritic cells[END_REF] Low levels of TLR7 have also been observed in non-immune cells such as hepatocytes, epithelial cells and keratinocytes. [START_REF] Tengroth | Functional effects of Toll-like receptor (TLR)3, 7, 9, RIG-I and MDA-5 stimulation in nasal epithelial cells[END_REF][START_REF] Lee | Activation of anti-hepatitis C virus responses via Toll-like receptor 7[END_REF][START_REF] Schaefer | Toll-like receptor (TLR) expression and TLR-mediated cytokine/chemokine production by human uterine epithelial cells[END_REF][START_REF] Li | Roles of TLR7 in Activation of NF-κB Signaling of Keratinocytes by Imiquimod[END_REF] Contrary to TLR7, TLR8 is more strongly expressed in myeloid dendritic cells, monocytes and to a lesser extent in pDC. [START_REF] Hornung | Quantitative expression of toll-like receptor 1-10 mRNA in cellular subsets of human peripheral blood mononuclear cells and sensitivity to CpG oligodeoxynucleotides[END_REF][START_REF] Crozat | Crosstalk between components of the innate immune system: promoting anti-microbial defenses and avoiding immunopathologies[END_REF] Under certain circumstances, TLR7 expression is inducible in cells expressing a basal, undetectable level of TLR7, including immune cells such as macrophages, myeloid dendritic cells and non-immune cells such as hepatocytes and keratinocytes. [START_REF] Lester | Toll-like receptor expression and responsiveness are increased in viraemic HIV-1 infection[END_REF][START_REF] Roelofs | The expression of toll-like receptors 3 and 7 in rheumatoid arthritis synovium is increased and costimulation of toll-like receptors 3, 4, and 7/8 results in synergistic cytokine production by dendritic cells[END_REF][START_REF] Yin | TLR7-expressing cells comprise an interfollicular epidermal stem cell population in murine epidermis[END_REF] For example, TLR7 mRNA expression is induced by the stimulation of IFNγ in macrophage level. Similarly, viral infections such as hepatitis C, the human immunodeficiency virus (HIV) and the influenza A virus lead to upregulation of TLR7 expression in hepatocytes, circulating immune cells and primary macrophages respectively. [START_REF] Lester | Toll-like receptor expression and responsiveness are increased in viraemic HIV-1 infection[END_REF][START_REF] Miettinen | IFNs activate toll-like receptor gene expression in viral infections[END_REF] Under these circumstances, it seems that the cytokines induced by the viral infection are responsible for triggering the induction of TLR7 expression. In fact, infection of primary macrophages by the influenza virus leads to upregulation of TLR7 mRNA in a type I IFN dependent manner. [START_REF] Miettinen | IFNs activate toll-like receptor gene expression in viral infections[END_REF] In addition, the increased expression of TLRs 7 and 8 is observed in cancer and autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus (SLE). [START_REF] Roelofs | The expression of toll-like receptors 3 and 7 in rheumatoid arthritis synovium is increased and costimulation of toll-like receptors 3, 4, and 7/8 results in synergistic cytokine production by dendritic cells[END_REF][START_REF] Sheyhidin | Overexpression of TLR3, TLR4, TLR7 and TLR9 in esophageal squamous cell carcinoma[END_REF][START_REF] Komatsuda | Up-regulated expression of Toll-like receptors mRNAs in peripheral blood mononuclear cells from patients with systemic lupus erythematosus[END_REF]

Composition and signaling pathways

Each TLR is composed of an extracellular domain with leucine-rich repeat motifs (LRRs) that are responsible for the recognition of pathogen elements PAMPs and/or DAMPs, a transmembrane domain, and a cytoplasmic domain comprising a Toll / IL-1 receptor (TIR) that is homolog to the IL1 receptor and is consequently responsible for signal transduction. TLRs interact with their PAMPs or DAMPs as a homo or heterodimer along with a co-receptor or an accessory molecule. [START_REF] Kawai | Signaling to NF-kappaB by Toll-like receptors[END_REF][START_REF] Botos | The structural biology of Toll-like receptors[END_REF] Both TLRs 7 and 8 receptor types have a high degree of sequence homology and function similarity. [START_REF] Heil | The Toll-like receptor 7 (TLR7)-specific stimulus loxoribine uncovers a strong relationship within the TLR7, 8 and 9 subfamily[END_REF][START_REF] Heil | Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8[END_REF][START_REF] Jurk | Human TLR7 or TLR8 independently confer responsiveness to the antiviral compound R-848[END_REF][START_REF] Shibata | Guanosine and its modified derivatives are endogenous ligands for TLR7[END_REF] Basically, TLR signaling is divided into two pathways: the MyD88-dependent pathway and the TRIFdependent pathway. [START_REF] Kawai | Signaling to NF-kappaB by Toll-like receptors[END_REF] Activation of TRIF/TRAF3 pathway leads to the recruitment of IKKε/TBK1, the phosphorylation of IRF3 and the expression of IFNβ. As this way is not involved in the signaling pathway of TLR7/8, we will not give more details. In fact, the signaling pathways of TLR7 and TLR8 are dependent on MyD88 protein (Figure 2).

MyD88 is composed of a TIR domain and a death domain. Following the activation of TLRs 7/8, MyD88 interacts, through its death domain, with the death domains of members of the IRAK (IL-1 receptorassociated kinase) protein kinase family, including IRAK1, IRAK2, IRAK4 and IRAK-M. [START_REF] Akira | Toll-like receptor signalling[END_REF][START_REF] West | Recognition and signaling by toll-like receptors[END_REF] IRAK4 phosphorylates and activates IRAK1. After phosphorylation of IRAK4 and IRAK1, these two kinases dissociate from MyD88 and interact with TRAF6 with E3 ubiquitin ligase activity. TRAF6, together with the ubiquitin E3: UBC13 and UEV1A conjugation enzyme, favor the lysine-related polyubiquitination of TRAF6 itself and the TAK1 protein kinase complex. [START_REF] Adhikari | Ubiquitin-mediated activation of TAK1 and IKK[END_REF] The latter is a member of the MAPKKK family and forms a complex with the regulatory subunits TAB1, TAB2 and TAB3 that will interact with the TRAF6-generated polyubiquitin chains to allow activation of TAK1. [START_REF] Adhikari | Ubiquitin-mediated activation of TAK1 and IKK[END_REF] TAK1 then activates two signaling pathways : NF-κβ and MAPK pathways. The IKK complex is composed of two catalytic subunits: IKKα and IKKβ and a regulatory subunit NEMO (or IKKγ). TAK1 binds to the IKK complex via ubiquitin chains, allowing it to phosphorylate and activate IKKβ. The IKK complex phosphorylates the inhibitory protein of NF-κβ: Iκβα, which will undergo degradation in the cytoplasm, thus allowing NF-κβ to translocate to the nucleus to induce the expression of proinflammatory genes. In addition, TAK1 also activates members of the MAPKs family such as ERK1/2, p38 and JNK, which mediate the activation of the AP-1 transcription factor, responsible for the expression of proinflammatory cytokines and IFNβ (Figure 2). [START_REF] Wang | TAK1 is a ubiquitin-dependent kinase of MKK and IKK[END_REF][START_REF] Kawasaki | Toll-like receptor signaling pathways[END_REF] Moreover, Ito et al. showed in 2002 that TLR7 is expressed in myeloid and plasmacytoid dendritic cells. They studied the production of IL12 and IFNα by dendritic cells during TLR7 agonist stimulation. They found that the cytokine induction pattern differs between myeloid dendritic cells (mDCs) and pDCs. pDCs produce IFNα while mDCs produce IL12. [START_REF] Ito | Interferon-alpha and interleukin-12 are induced differentially by Toll-like receptor 7 ligands in human blood dendritic cell subsets[END_REF] Given the large amounts of IFNα produced by pDCs expressing TLRs 7 and 9, much work has been published in the literature to elucidate the signaling pathway leading to activation and secretion of IFNα especially by dendritic cells. TLR7, TLR8 and TLR9 induce antiviral responses by the production of IFNα as well as pro-inflammatory cytokines. These three receptors use the MyD88 adapter protein to initiate the signaling pathways. The IRF7 transcription factor (Interferon regulatory factor 7) is responsible for the expression and production of IFNα. MyD88 interacts directly with IRF7 at the endosome. [START_REF] Kawai | Interferon-alpha induction through Toll-like receptors involves a direct interaction of IRF7 with MyD88 and TRAF6[END_REF] IRF7 also interacts with TRAF6, another adapter molecule that operates downstream of MyD88, and after receptor activation (TLRs 7, 8 or 9), IRF7 is activated in a MyD88 and TRAF6 dependent manner. Splenic pDCs from IRF7-deficient mice show a significant decrease in IFNα induction following viral infection or exposure to synthetic TLR7 or 9 ligands. [START_REF] Honda | IRFs: master regulators of signalling by Toll-like receptors and cytosolic patternrecognition receptors[END_REF] On the other hand, this induction is normal in IRF1, IRF3 or IRF5-deficient pDCs. This shows that induction of IFNα in pDCs requires IRF7. [START_REF] Honda | IRFs: master regulators of signalling by Toll-like receptors and cytosolic patternrecognition receptors[END_REF] In addition, MyD88 mutation studies have shown that this protein interacts with IRF7 via its death domain. This death domain also interacts with the serine / threonine kinase family (IRAK), which will transduce the signal between MyD88 and TRAF6, indicating that IRAKs are involved in the signaling of IRF7. [START_REF] Uematsu | Interleukin-1 receptor-associated kinase-1 plays an essential role for Toll-like receptor (TLR)7-and TLR9-mediated interferon-{alpha} induction[END_REF] pDCs from IRAK1 or IRAK4deficient mice are unable to produce IFNα upon activation of TLRs 7, 8 or 9. [START_REF] Honda | IRFs: master regulators of signalling by Toll-like receptors and cytosolic patternrecognition receptors[END_REF] In addition, one study has shown that IKKα is also essential for activation of IRF7 [START_REF] Hoshino | IkappaB kinase-alpha is critical for interferon-alpha production induced by Toll-like receptors 7 and 9[END_REF], indicating that activation of IRF7 requires a cascade of IRAK4-IRAK1-IKKα protein kinases. Studies have also shown that TRAF3 plays an important role in this IRF7-dependent signaling. [START_REF] Honda | IRFs: master regulators of signalling by Toll-like receptors and cytosolic patternrecognition receptors[END_REF] In addition to IRF7, IRF5 also interacts with MyD88 and TRAF6. Unlike IRF7, which binds to the MyD88 death domain, IRF5 interacts with the middle region (known as the intermediate domain) and part of the MyD88 TIR domain. [START_REF] Takaoka | Integral role of IRF-5 in the gene induction programme activated by Toll-like receptors[END_REF] Activation of the MyD88-dependent signaling pathway by TLR7 or TLR9 ligands leads to translocation of IRF5 to the nucleus where it will activate the expression of pro-inflammatory cytokines. [START_REF] Schoenemeyer | The interferon regulatory factor, IRF5, is a central mediator of toll-like receptor 7 signaling[END_REF] In 2005, Schoenemeyer et al. have shown that stimulation of TLR7 and TLR8 by resiquimod induces the activation of IRF5 as well as IRF7, and they also found that IRF5 is a central mediator in TLRs 7/8 signaling pathway. IRF5 contributes to the induction of IFN type I in human cells, and in addition, is important not only for IFNα induction but also for IFNβ induction. [START_REF] Schoenemeyer | The interferon regulatory factor, IRF5, is a central mediator of toll-like receptor 7 signaling[END_REF] In 2009, a study showed that mDCs, and not pDCs or macrophages, are capable of inducing a large amount of IFNβ after bacterial degradation in phagolysosomes and such response requires the intervention of TLR7, MyD88 and IRF1. [START_REF] Mancuso | Bacterial recognition by TLR7 in the lysosomes of conventional dendritic cells[END_REF] Consequently, those signaling pathways lead to the activation of transcription factors NF-κβ and AP1, which regulate the expression of inflammatory cytokines, and IFN inducible genes. Briefly, despite the phylogenetic and structural similarities between TLR7 and TLR8, these TLRs differ functionally in their cytokine profiles induced by human PBMCs as well as in PBMC-isolated cell populations. TLR7 is functionally associated with the production of IFNα. TLR8 is involved in the production of pro-inflammatory cytokines such as TNFα and IL12.

Therapeutic purposes a. Antiviral and antibacterial actions

In response to viral infections, the host activates its innate immune system whose response is characterized by IFNs, inflammatory cytokines and chemokines production. This response to viral infection is mediated through several TLR signaling pathways including TLR3, TLRs 7/8/9, TLR4 and TLR2. The contribution of each TLR is different depending on the virus, cell type and infection model examined. TLR7 and 8 are able to detect GU-rich and AU-rich ssRNA sequences of RNA viruses. [START_REF] Heil | Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8[END_REF][START_REF] Diebold | Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA[END_REF] They are known to serve as endosomal PRRs for a number of ssRNA viruses as influenza, HIV-1, vesicular stomatitis virus, Sendai virus, flaviviruses, and coronaviruses. [START_REF] Heil | Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8[END_REF][START_REF] Melchjorsen | Activation of Innate Defense against a Paramyxovirus Is Mediated by RIG-I and TLR7 and TLR8 in a Cell-Type-Specific Manner[END_REF][START_REF] Brown | HIV-1 Activates Macrophages Independent of Toll-Like Receptors[END_REF][START_REF] Beignon | Endocytosis of HIV-1 activates plasmacytoid dendritic cells via Tolllike receptor-viral RNA interactions[END_REF][START_REF] Triantafilou | Human cardiac inflammatory responses triggered by Coxsackie B viruses are mainly Tolllike receptor (TLR) 8-dependent[END_REF][START_REF] Wang | Flavivirus Activation of Plasmacytoid Dendritic Cells Delineates Key Elements of TLR7 Signaling beyond Endosomal Recognition[END_REF][START_REF] Town | Toll-like receptor 7 mitigates lethal West Nile encephalitis via interleukin 23-dependent immune cell infiltration and homing[END_REF][START_REF] Patel | Novel drugs targeting Toll-like receptors for antiviral therapy[END_REF][START_REF] Cervantes-Barragan | Control of coronavirus infection through plasmacytoid dendritic-cell-derived type I interferon[END_REF] Moreover, TLR7 signaling is well described in human and mice for type I IFN production by pDCs in response to ssRNA viral stimuli [START_REF] Diebold | Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA[END_REF], and TLR8 signaling mainly results in NF-κB activation and subsequent inflammatory cytokine expression. [START_REF] Liu | IPC: Professional Type 1 Interferon-Producing Cells and Plasmacytoid Dendritic Cell Precursors[END_REF] Shortly after TLR7 and TLR8 description, it was shown that both receptors respond to potent antiviral imidazoquinoline compounds. These low molecular weight synthetic compounds have been shown to mediate the TLR7 and TLR8 inflammatory responses. [START_REF] Gorden | Synthetic TLR agonists reveal functional differences between human TLR7 and TLR8[END_REF] TLRs 7/8 play key roles in antiviral and antibacterial responses.

b. Adjuvant strategies

Vaccination imprints the immune system with the experience of pathogen exposure resulting in the production of humoral immune response by Ag-specific memory B cells. The efficacy of the resulting immunization strongly varies with the ingredients of the vaccine. As the vaccine employed in the early stages of vaccinology were live attenuated or killed organisms, adjuvants were not systematically required. Currently employed adjuvants in human vaccines are aluminum salts, oil in water emulsions (e.g. MF59) and TLR9 adjuvants which have limitations and require multiple administrations to induce protection and drive Th2-over Th1-polarized immunity. Moreover, vaccinology moves toward the use of inactivated antigens which lack the immunological information needed to enhance an adaptive immunity response. Hence, the development of novel adjuvants and delivery systems to replace the ones actually used has become crucial to improve vaccine-induced protective immunity. [START_REF] Reed | Key roles of adjuvants in modern vaccines[END_REF] TLR7 and TLR8 together mediate recognition of purinerich ssRNA to elicit an immune response to pathogens recognized in the endosome. TLRs 7 and 8 are implicated in the recognition of naturally derived uridine-rich ssRNAs of influenza and HIV. [START_REF] Heil | Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8[END_REF] In addition TLR7 and TLR8 recognize bacterial RNA. [START_REF] Mancuso | Bacterial recognition by TLR7 in the lysosomes of conventional dendritic cells[END_REF][START_REF] Ugolini | Recognition of microbial viability via TLR8 drives T FH cell differentiation and vaccine responses[END_REF] Furthermore, TLR7 and 8 are expressed in human plasmacytoid DCs (pDCs), in T and B cells, monocytes and macrophages. [START_REF] Cervantes | TLR8: the forgotten relative revindicated[END_REF][START_REF] Vasilakos | The use of Toll-like receptor 7/8 agonists as vaccine adjuvants[END_REF] Naive human B cells express low levels of TLR7 and, whereas activated and memory human B cells also express a broader range of TLRs including TLR7. B cell intrinsic TLR7 signaling may play a role in B cell responses during chronic infections which could be used to activate memory B cells and boost humoral immune responses during immunization. [START_REF] Castiblanco | Co-Stimulation of BCR and Toll-Like Receptor 7 Increases Somatic Hypermutation, Memory B Cell Formation, and Secondary Antibody Response to Protein Antigen[END_REF] Synthetic small molecules agonists, which activate TLRs 7 and 8 and could be used as adjuvants, have been identified and presented herein. However, the challenge in the development of these molecules remains their formulation.

c. Cancer targeting

Specific and potent immunostimulatory molecules targeting TLRs 7/8 lead to stimulate innate and adaptive immunity, product cytokines and activate cytotoxic cells. TLRs agonists could be promising elements for novel therapies. Whereas many studies suggest that the activation of TLRs may have potential applications in oncotherapy, others have proved that TLRs signaling is also implicated in tumorigenesis. Even if fighting cancers by TLRs targeting is always under investigation, human clinical trials often failed. Many patients are immunosuppressed due to the traditional anticancer therapies. By this way, it is difficult to induce a positive immune response able to help organism to combat tumors. Understanding the role of TLRs in all cancer cells types remains the main obstacle for clinical therapeutic application. Combination of modulators and traditional approaches (chemotherapy or radiotherapy) could be the key for a successful anticancer therapy. [START_REF] Chi | Anti-tumor Activity of Toll-Like Receptor 7 Agonists[END_REF][START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF][START_REF] Liu | The Role of Toll-Like Receptors in Oncotherapy[END_REF]

d. Allergy and asthma treatments

The primary step of type I allergic diseases (such as allergic rhinitis, asthma, drug-induced anaphylaxis) is usually a strong pro-inflammatory Th2 and IgE-mediated responses against antigens. It can be counterbalanced by the induction of TLR signaling inducing a cytokine response and activating antigen presenting cells, leading to a Th1 response. Agonists that activate TLRs, particularly TLR7, TLR8 and TLR9 would be expected to induce a strong Th1 response and have been a focus for novel therapeutics to treat asthma and allergies. [71,[START_REF] Matesic | Modulating toll-like receptor 7 and 9 responses as therapy for allergy and autoimmunity[END_REF] Among allergic diseases, asthma is a complex and chronic inflammatory disease of the airways characterized by airway hyper-responsiveness, eosinophilic infiltration, reversible airflow obstruction, airway remodeling, mucus hypersecretion, and goblet cell hyperplasia. Asthma affects more than 350 million people worldwide. Allergy and atopy are the main causes of asthma. Genetic and environmental triggers modulating the activation and regulation of the immune system are also implicated in the pathophysiology of asthma. Bronchial inflammation, smooth muscle spasm, and mucus production in allergic asthma are triggered by IL-4, IL-5, and IL-13, which are released by Th2 cells. TLR7 ligands play an important role in reduction of airway inflammation, promoting Th1 responses in immune cells, reversing airway hyper-responsiveness, and preventing airway remodeling (including smooth muscle proliferation and goblet cell hyperplasia). Acute and chronic asthma attacks can be managed by precisely identifying the regulators of these pathogenic pathways and targeting their molecular mediators. [START_REF] Athari | Targeting cell signaling in allergic asthma, Signal Transduct[END_REF][START_REF] Dong | Holding the Inflammatory System in Check: TLRs and Their Targeted Therapy in Asthma[END_REF] 2. Natural ligands Given the structural similarity of the firstly described agonists of TLRs 7/8 with nucleic acids, Heil et al. studied the effect of single-stranded ribonucleic acids (ssRNAs) on the activation of antigen-presenting cells via TLR7 and TLR8. They showed that guanosine (G) and uridine (U)-rich oligonucleotides induce the production of TNFα by human PBMCs and that ssRNAs are the natural ligands of TLR7 and TLR8 (Figure 3). [START_REF] Heil | Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8[END_REF] The same year, Diebold et al. have shown that the ssRNA of influenza virus is a TLR7 agonist, and that TLR7 has a preference for a particular RNA motif that is uridine. [START_REF] Gorden | Synthetic TLR agonists reveal functional differences between human TLR7 and TLR8[END_REF] In addition, studies have shown that adenosine-and uridine-rich oligoribonucleotides (ORNs) are capable of activating TLR8, without any effect on TLR7, while guanosine-rich ORNs activate TLR7 and TLR8-dependent signaling. [START_REF] Diebold | Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA[END_REF] In quest of TLR7 and TLR8 agonists, a range of chemical modifications was done to natural ORN ligands in order to increase their selectivity for TLR7 and/or TLR8 and stability against nucleases. [START_REF] Diebold | Nucleic acid agonists for Tolllike receptor 7 are defined by the presence of uridine ribonucleotides[END_REF][START_REF] Forsbach | Identification of RNA sequence motifs stimulating sequencespecific TLR8-dependent immune responses[END_REF][START_REF] Lan | Stabilized immune modulatory RNA compounds as agonists of Tolllike receptors 7 and 8[END_REF][START_REF] Lan | Toll-like receptor 7 selective synthetic oligoribonucleotide agonists: synthesis and structure-activity relationship studies[END_REF] In 2015, Tanji et al. showed for the first time, that TLR8 recognizes the ssRNA degradation products at two different sites: the first site recognizes uridine while the second recognizes short ORNs such as UG and UUG. Uridine alone is not able to activate TLR8 in the absence of the ORNs at the second site. The ORN binding to the second site increases the affinity of uridine at the first site and therefore leads to activation of TLR8. [START_REF] Tanji | Toll-like receptor 8 senses degradation products of single-stranded RNA[END_REF] Based on the above results, Shibata et al. showed that TLR7 responds to guanosine analogs: guanosine, 2'-deoxyguanosine (dG), 8-hydroxydeoxyguanosine (8-OHdG), in the presence of oligoribonucleotides (Figure 4). [START_REF] Shibata | Guanosine and its modified derivatives are endogenous ligands for TLR7[END_REF] Guanosine alone or ssRNA alone (polyU) are not capable of activating TLR7, however a combination of both leads to receptor activation. Similarly, ssRNA enhances the affinity of guanosine, dG, 8-OHG, and 8-OHdG to TLR7. Considering that these nucleotides are metabolites present in the physiological or pathological state, they can be considered as endogenous ligands of TLR7. These results were confirmed by Zhang et al by showing in 2016 that TLR7 also recognizes the degraded form of ssRNA : guanosine binds to the first site while the oligonucleotide (polyU) binds to the second site, and the combination of these two products activates the dimerization of TLR7 in a synergistic way. [START_REF] Zhang | Structural Analysis Reveals that Toll-like Receptor 7 Is a Dual Receptor for Guanosine and Single-Stranded RNA[END_REF] Figure 4 : Guanosine derivatives Furthermore, many articles were published showing the role of small-interfering RNA (siRNAs) in stimulating the immune system. After siRNAs recognition by TLR7 and TLR8, cytokines are released. [START_REF] Hornung | Sequence-specific potent induction of IFN-alpha by short interfering RNA in plasmacytoid dendritic cells through TLR7[END_REF][START_REF] Judge | Sequence-dependent stimulation of the mammalian innate immune response by synthetic siRNA[END_REF] In 2012, Fabbri et al. show that tumor-secreted miRNA-21 and miRNA-29 can function as ligands by binding to murine TLR7 and human TLR8 receptors in immune cells, triggering a TLR-mediated pro-metastatic inflammatory response that ultimately may lead to tumor growth and metastasis. [START_REF] Fabbri | MicroRNAs bind to Toll-like receptors to induce prometastatic inflammatory response[END_REF] 3. Synthetic ligands Many synthetic ligands as potent agonists of TLRs 7/8 have been designed worldwide. They have been classified in this review by increasing number of (hetero)cycles a. Pyrimidine and purine base derivatives a.1. Pyrimidine derivatives and smaller fragment First described by Skulnick et al. [START_REF] Skulnick | Pyrimidinones.1.2-Amino-5-halo-6-aryl-4(3JEf)-pyrimidinones[END_REF], Bropirimine (2, also known as ABPP for 2-amino-5-bromo-6phenylpyrimidin-4-ol, Figure 5) is an orally active immunostimulant acting as IFN inducer in several species, immunomodulator of macrophages, natural killer cells and cytokines. [START_REF] Stringfellow | Interferon induction by 5-halo-6-phenyl pyrimidinones[END_REF][START_REF] Lotzová | 5-halo-6-phenyl pyrimidinones: new molecules with cancer therapeutic potential and interferon-inducing capacity are strong inducers of murine natural killer cells[END_REF][START_REF] Lotzová | Stimulation of natural killer cells in two randombred strains of athymic rats by interferon-inducing pyrimidinone[END_REF] Bropirimine is described as antiviral and antineoplastic agent. [START_REF] Sidwell | Antiviral and immunomodulating inhibitors of experimentally-induced Punta Toro virus infections[END_REF] It was tested as agent against melanoma [START_REF] Li | Mechanism of antitumor action of pyrimidinones in the treatment of B16 melanoma and P388 leukemia[END_REF], bladder tumor [START_REF] Tei | In vitro antitumor activity of bropirimine against urinary bladder tumor cells[END_REF][START_REF] Witjes | Treatment of carcinoma in situ of the urinary bladder with bropirimine[END_REF][START_REF] Sarosdy | A review of clinical studies of bropirimine immunotherapy of carcinoma in situ of the bladder and upper urinary tract[END_REF], prostate cancer [START_REF] Sarosdy | Oral bropirimine immunotherapy of rodent prostate cancer[END_REF] and renal cell carcinoma [START_REF] Fujioka | Antitumor effects of oral administration of an interferon-inducing pyrimidinone, Bropirimine, on murine renal-cell carcinoma[END_REF]. In an attempt to design a TLR8 agonist containing benzimidazole core as vaccine adjuvant, Beesu et al. found that the smallest possible fragment of the benzimidazole core allowing retention of TLR8-agonistic activity corresponds to 1-pentyl-4-phenyl-1H-imidazol-2-amine (4, Figure 5). [START_REF] Beesu | Identification of a Human Toll-Like Receptor (TLR) 8-Specific Agonist and a Functional Pan-TLR Inhibitor in 2-Aminoimidazoles[END_REF] The crystal structure of this compound bound to the TLR8 ectodomain displayed binding interactions that are common to other TLR8 agonists. This compound showed marked attenuated pro-inflammatory properties in ex vivo human blood models. a. 2. Adenine derivatives 8-hydroxyadenine derivatives as TLRs agonists have attracted significant interest. In 2002, Hirota et al. synthesized a wide variety of compounds based on the 8-hydroxyadenine scaffold (Figure 6), to test their potential to secrete IFN-α for the treatment of HCV. [START_REF] Hirota | Discovery of 8-hydroxyadenines as a novel type of interferon inducer[END_REF] SAR studies showed that the presence of a benzyl group at the N9 position as well as a C8 hydroxyl group in compound 5 is essential for IFN induction. The introduction of a R substituent such as alkyl, alkylthio, alkylamino and alkoxy groups to the 8-hydroxyadenine moiety resulted in a remarkable increase in activity. The 2-alkylthio (6 to 8), 2-butylamino (9) and 2-butoxy [START_REF] Patinote | Imidazo[1,2-a]pyrazine, Imidazo[1,5-a]quinoxaline and Pyrazolo[1,5a]quinoxaline derivatives as IKK1 and IKK2 inhibitors[END_REF] analogs showed important activities after oral administration in mice. IFN-inducing activity of compound 11 is 10 times higher than that of imiquimod after oral administration in mice. Furthermore, compound 11 has a better oral tolerance compared to imiquimod (Figure 20) in mice. These results indicate that compound 11 has a better efficacy and safety profile compared to imiquimod. Based on these data, Kurimoto et al. synthesized a series of 8-hydroxyadenines having various alkoxy or C2 alkylthio groups and evaluated them for their ability to induce IFN both in vitro and in vivo. [START_REF] Kurimoto | Synthesis and evaluation of 2-substituted 8-hydroxyadenines as potent interferon inducers with improved oral bioavailabilities[END_REF] They found that SM-360320 (12, also known as CL087, Figure 7) is a potent inducer of IFN and has good oral bioavailability. SM-360320 is a TLR7 agonist and is able to activate immune responses against the HCV, as shown by Lee et al. [START_REF] Lee | Activation of anti-hepatitis C virus responses via Toll-like receptor 7[END_REF] It induces IFNα in human peripheral blood mononuclear cells with an EC50 value of 0.14 µM. [START_REF] Jin | Synthesis and immunostimulatory activity of 8-substituted amino 9-benzyladenines as potent Toll-like receptor 7 agonists[END_REF] At 10 µM, SM-360320 was shown to reduce HCV levels by 60% in 24 hours in the human hepatocyte cell line Huh7, which carries a HCV replicon. [START_REF] Lee | Activation of anti-hepatitis C virus responses via Toll-like receptor 7[END_REF] It induces IFN in mice with a minimal effective dose of approximately 0.03 mg/kg, making it approximately 100-fold more potent than imiquimod. [START_REF] Kurimoto | Synthesis and evaluation of 2-substituted 8-hydroxyadenines as potent interferon inducers with improved oral bioavailabilities[END_REF] The mechanism of action is still studied, notably by Russo et al., which showed a higher concentration of this TLR7 agonist in the MIIC of pDC, the acidic compartment protonating and charging positively the compound. [START_REF] Russo | Small molecule Toll-like receptor 7 agonists localize to the MHC class II loading compartment of human plasmacytoid dendritic cells[END_REF] SM-360320 is not only a potential treatment for hepatitis C [START_REF] Thomas | Investigating Toll-Like Receptor Agonists for Potential To Treat Hepatitis C Virus Infection[END_REF] but also for colon cancer [START_REF] Dharmapuri | An oral TLR7 agonist is a potent adjuvant of DNA vaccination in transgenic mouse tumor models[END_REF]. SM-276001 is a selective and potent TLR7 agonist (13, Figure 7), displaying a minimum effective concentration (MEC) of 3 nM, which is comparable to resiquimod (Figure 1). SM-276001 demonstrated potent at a dose of 0.1 mg/kg by oral administration in mice, a dose dependent manner IFN-inducing activity in monkeys and did not cause emesis in ferrets even at a dose of 30 mg/kg. Isobe et al. also showed that the maximum plasma concentration of SM-276001 was 1019 ng/mL (ca. 3.1 µM), which was approximately 1000fold higher than the MEC value. [START_REF] Isobe | Synthesis and biological evaluation of novel 9-substituted-8-hydroxyadenine derivatives as potent interferon inducers[END_REF] Oral administration of SM-276001 leads to the induction of IFNα, TNFα and IL-12p40, the activation of T and B lymphocytes, NK and NKT cells and a reduction in tumor burden in the Balb/c syngeneic Renca and CT26 models. In an ovarian model cancer that spontaneously metastasizes, SM-276001 is able to reduce pulmonary metastasis and a spreading in lymph nodes when dosed either intratracheally or orally. [START_REF] Koga-Yamakawa | Intratracheal and oral administration of SM-276001: A selective TLR7 agonist, leads to antitumor efficacy in primary and metastatic models of cancer[END_REF] Telormedix, a clinical-stage biopharmaceutical company, proposes diverse prodrugs or formulations of potent TLRs 7/8 agonists. The first generation was represented by TMX-101, which is a liquid formulation of imiquimod originally developed as alternative for BCG to produce a local immune response leading to antitumor activity but circumventing the use of live attenuated mycobacteria preventing side effects. Prepared for intravesical instillation of imiquimod and developed by UroGen Pharma (formerly TheraCoat), TMX-101 (Vesimune™) was already tested for pharmacokinetics and pharmacodynamics [START_REF] Arentsen | Pharmacokinetics and toxicity of intravesical TMX-101: a preclinical study in pigs[END_REF] and was the subject of clinical trials. [START_REF] Donin | A phase 2 study of TMX-101, intravesical imiquimod, for the treatment of carcinoma in situ bladder cancer[END_REF][START_REF] Falke | Results of a phase 1 dose escalation study of intravesical TMX-101 in patients with nonmuscle invasive bladder cancer[END_REF][START_REF] Arends | Pharmacokinetic, Pharmacodynamic, and Activity Evaluation of TMX-101 in a Multicenter Phase 1 Study in Patients With Papillary Non-Muscle-Invasive Bladder Cancer[END_REF] TMX-202 (14, Figure 8) is a second-generation SM-360320 prodrug, which conjugates the TLR7 ligand to a C12 phospholipid via a benzoic acid functional group. [START_REF] Falke | A placebo-controlled efficacy study of the intravesical immunomodulators TMX-101 and TMX-202 in an orthotopic bladder cancer rat model[END_REF] Discovered through a collaboration with the University of California San Diego, TMX-202 has already successfully completed many in vivo studies. TMX-202 is safe and efficacious without causing excessive adverse effects and could be an alternative to imiquimod for the treatment of proliferative skin disorders. [START_REF] Crain | Inhibition of keratinocyte proliferation by phospholipid-conjugates of a TLR7 ligand in a Myc-induced hyperplastic actinic keratosis model in the absence of systemic side effects[END_REF] It is in preclinical development for the topical treatment of skin cancers, bladder cancers, and other indications. TMX-302 and TMX-306 (15 and 16 respectively, Figure 8) are PEGylated SM-360320 purine-like compounds characterized by TLR7 partial agonist activity. The clinical application of TMX-302 in lung disorders should be examined with caution because of its direct pro-inflammatory effects. TMX-306 seems to be comparatively more effective and safer, deserving further investigations in drug development particularly for silicosis. This third generation is focused on developing candidates for treating autoimmune diseases. [START_REF] Ferreira | Potential of PEGylated Toll-Like Receptor 7 Ligands for Controlling Inflammation and Functional Changes in Mouse Models of Asthma and Silicosis[END_REF] CL264, a 9-benzyl-8-hydroxyadenine derivative, containing a glycine moiety on the benzyl group and similar to SZU-101, induces NF-κB activation and IFNα secretion in TLR7 expressing cells (18, Figure 9). CL264 is a TLR7 specific agonist, which does not activate TLR8 even at high concentrations. [START_REF] Hirota | Discovery of 8-hydroxyadenines as a novel type of interferon inducer[END_REF] Another base analogue commercialized by Invivogen™ is CL307 (19, Figure 10) which is prepared by linking a spermine to CL264. CL307 is a very potent TLR7 agonist inducing NF-κB activation even at low concentrations (10 ng/mL).

AdiFectin™ (20, also known as CL347, Figure 10), a derivative obtained by conjugation of CL307 with a bis(phytanyl)phosphonate group, is a weaker TLR7 agonist than CL307, but in contrast to CL307, is able to efficiently complex nucleic acids resulting in a strong IFN response. The compound is able to form positively charged liposomes, which can encapsulate DNA (or RNA) thanks to the lipid moiety. Repeated in vivo studies showed that pDNA/AdiFectin™ complexes display robust anti-tumor activity. Tumor growth was markedly reduced resulting in a 50% survival rate. Notably, mice that achieved long-term clearance of tumor following AdiFectin™ treatment were protected from subsequent tumor rechallenge suggesting the generation of a tumor-specific memory immune response. Adilipoline™ (21, also known as CL413, Figure 11), a derivative obtained by linearly linking a hydroxyadenine derivative with the terminal acid function of Pam2CSK4 (a TLR2 agonist), is a good ligand for TLRs 2/7. In vivo tumor studies have demonstrated that CL413 is a potent antitumor agent. Intratumoral injection of CL413 in established B16 tumors resulted in tumor regression. However, in contrast to CL347, no protection after tumor rechallenge was observed. CL531 [START_REF] Tengroth | Functional effects of Toll-like receptor (TLR)3, 7, 9, RIG-I and MDA-5 stimulation in nasal epithelial cells[END_REF] corresponds to the conjugation of a hydroxyadenine derivative to the lateral chain of the second lysine of Pam2CSK4, resulting in a dual TLRs 2/7 agonist (Figure 11). CL572 [START_REF] Lee | Activation of anti-hepatitis C virus responses via Toll-like receptor 7[END_REF] contains a monoacyl-ethyl-cystein group grafted to a hydroxyadenine via a glutamic acid derivative and is also described by Invivogen™ as a dual TLRs 2/7 agonist (Figure 11). Among the 8-oxoadenine family, Biggadike et al. prepared derivatives bearing saturated oxygen or nitrogen heterocycles as N-9 substituting group. [START_REF] Biggadike | Discovery of 6-Amino-2-{[(1S)-1-methylbutyl]oxy}-9-[5-(1-piperidinyl)pentyl]-7,9-dihydro-8H-purin-8-one (GSK2245035), a Highly Potent and Selective Intranasal Toll-Like Receptor 7 Agonist for the Treatment of Asthma[END_REF] To treat allergic asthma, GSK2245035 (24) is selected as intranasal candidate (Figure 12). It resulted in vitro in suppression of Th2 cytokine response to allergens and in vivo to local upregulation of TLR7-mediated cytokine IP-10. Clinical evaluations on patients confirm the pharmacological interest of this compound. 13) is a TLR7-selective agonist that leads to the induction of IFNα/γ, IP-10, TNFα, IL-1Ra and IL-12p70, and after intravenously injections to a reduction in tumor burden in syngeneic models of renal cancer (Renca), metastatic osteosarcoma (LM8) and colorectal cancer (CT26). Moreover, the efficacy of DSR-29133 was significantly improved when administered in combination with low-dose fractionated radiotherapy. Long-term surviving mice originally treated with DSR-29133 and radiotherapy were protected by a tumor-specific memory immune response which could prevent tumor growth. [START_REF] Dovedi | Intravenous administration of the selective toll-like receptor 7 agonist DSR-29133 leads to anti-tumor efficacy in murine solid tumor models which can be potentiated by combination with fractionated radiotherapy[END_REF] a.3. Guanine derivatives In 2003, Lee et al. proved that guanosine analogs such as loxoribine (27, also known as RWJ 21757) and isatoribine (28, also known as Thiaoxoguanosine, TOG, 7T80G, ANA-225, ANA-245, ICN-10146, ICN25, Immunosine, Immusine, KN 25, N10146, N12143, N14316, NARI-10146) activate immune cells exclusively via TLR7 by a pathway that requires endosomal maturation (Figure 14). [START_REF] Lee | Molecular basis for the immunostimulatory activity of guanine nucleoside analogs: activation of Toll-like receptor 7[END_REF] Previous work showed in vitro NFκβ activity as immunomodulator agents but it could not be linked to in vivo observations. [START_REF] Bonnet | Modulation of leukocyte genetic expression by novel purine nucleoside analogs. A new approach to antitumor and antiviral agents[END_REF] Loxoribine (27, Figure 14) displays interesting anti-tumor activities. Significant inhibition of B16 melanoma lung tumor metastasis was observed in mice receiving a single injection of 2 mg loxoribine at day 3 of tumor growth. The greatest inhibition (96%) corresponds to four injections of loxoribine on alternate days starting the day before tumor injection. The combination of loxoribine and IL-2 significantly greater inhibition of metastasis. As guanosine analogs display adjuvant activity in B cell systems, loxoribine was evaluated as adjuvant in a tumor protection model. Mice immunized with both irradiated tumor cells and loxoribine developed a significantly lower number of lung tumors when challenged by live B16 tumor cells, whereas mice injected with either vaccine or loxoribine alone were not protected. [START_REF] Pope | 7-Allyl-8-oxoguanosine (loxoribine) inhibits the metastasis of B16 melanoma cells and has adjuvant activity in mice immunized with a B16 tumor vaccine[END_REF] Loxoribine was also tested in the treatment of chronic lymphocytic leukemia. [START_REF] Spaner | A phase I/II trial of TLR-7 agonist immunotherapy in chronic lymphocytic leukemia[END_REF][START_REF] Ntoufa | Distinct Innate Immunity Pathways to Activation and Tolerance in Subgroups of Chronic Lymphocytic Leukemia with Distinct Immunoglobulin Receptors[END_REF][START_REF] Spaner | Toll-like receptor agonists in the treatment of chronic lymphocytic leukemia[END_REF] By this way, loxoribine may be useful in tumor therapy as an immunomodulator or as an adjuvant. Isatoribine (28, Figure 14), which synthesis was described by Nagahara et al. [START_REF] Nagahara | Thiazolo[4,5-d]pyrimidine nucleosides. The synthesis of certain 3-β-D-ribofuranosylthiazolo[4,5d]pyrimidines as potential immunotherapeutic agents[END_REF], enhances IFNα levels through its interaction with TLR7 and stimulate of the patient's own immune system. With a potent toxicity profile in multiple species, this compound displays antiviral activity in a variety of murine systems (semliki forest, san angelo, banzi, rat-corona, EMC viruses). [START_REF] Smee | Broad-spectrum activity of 8-chloro-7deazaguanosine against RNA virus infections in mice and rats[END_REF][START_REF] Smee | Broad-spectrum in vivo antiviral activity of 7-thia-8-oxoguanosine, a novel immunopotentiating agent[END_REF][START_REF] Smee | Roles of interferon and natural killer cells in the antiviral activity of 7-thia-8-oxoguanosine against Semliki Forest virus infections in mice[END_REF][START_REF] Michael | Alkylpurines as immunopotentiating agents. Synthesis and antiviral activity of certain alkylguanines[END_REF] Moderate to absent antiviral effects were observed against HSV-2, VSV infections or influenza B pneumonia. Isatoribine was also evaluated on chronic hepatitis C infection. [START_REF] Horsmans | Isatoribine, an agonist of TLR7, reduces plasma virus concentration in chronic hepatitis C infection[END_REF] On February 2007, this drug was no longer listed on Anadys Pharmaceuticals, Inc. development pipeline. c]pyridine compounds by incorporating substituents which were previously determined to be optimal agonists (N1-benzyl and C2-butyl, Figure 15) in the imidazoquinoline series. These compounds were studied for their potency to induce the IFNα production. [START_REF] Yoo | Structure-activity relationships in Toll-like receptor 7 agonistic 1H-imidazo[4,5-c]pyridines[END_REF] Compound 29 showed TLR7 agonistic activity (EC50 = 1.57 μM) with negligible activity on TLR8. Direct aryl-aryl bonds at the R1 position abolish all activities, but the TLR7 agonist effect is restored with analogues having a benzyl or phenethyl group. For example, the specific TLR7 agonist compound 30 (EC50 = 0.26 μM) displayed high IFNα induction in human PBMCs. Since some of these compounds are potent inducers of type I IFN, with an attenuated pro-inflammatory profiles, they could be potent adjuvants without local or systemic inflammation reactions. [START_REF] Yoo | Determinants of activity at human Toll-like receptors 7 and 8: quantitative structure-activity relationship (QSAR) of diverse heterocyclic scaffolds[END_REF] In quest of new vaccine adjuvants, Beesu et al. synthesized a wide variety of heterocyclic compounds that showed good selectivity for TLR8, such as compound 31 (Figure 16) belonging to the benzimidazol-2amine series (EC50 (TLR8) = 1.13 μM). [START_REF] Beesu | Human Toll-like receptor 8-selective agonistic activities in 1-alkyl-1H-benzimidazol-2-amines[END_REF][START_REF] Beesu | Structure-Based Design of Human TLR8-Specific Agonists with Augmented Potency and Adjuvanticity[END_REF][START_REF] Beesu | Human Toll-like Receptor (TLR) 8-Specific Agonistic Activity in Substituted Pyrimidine-2,4-diamines[END_REF] 17) displays a remarquable TLR8 agonist activity with an EC50 value of 9 nM. [START_REF] Beesu | Human Toll-like receptor 8-selective agonistic activities in 1-alkyl-1H-benzimidazol-2-amines[END_REF][START_REF] Beesu | Structure-Based Design of Human TLR8-Specific Agonists with Augmented Potency and Adjuvanticity[END_REF][START_REF] Beesu | Human Toll-like Receptor (TLR) 8-Specific Agonistic Activity in Substituted Pyrimidine-2,4-diamines[END_REF] Selgantolimod (31, also known as GS-9688, Figure 17), a TLR8 agonist, induces cellular immune mediators, such as IL-12 and IL-8, as well as antiviral cytokines TNF-α and IFN-γ in vitro in human PBMCs. Selgantolimod activates NK and mucosal associated invariant T cells, stimulates cluster of differentiation (CD)-8+ T-cell proliferation, while lowering programmed cell death protein 1 expression by HBV-specific CD8+-T cells in vitro. Selgantolimod-induced cytokines reduce HBV DNA, RNA, and antigen levels in HBVinfected primary human hepatocytes. The agonist is actually under investigation to treat HIV infection and hepatitis B virus. [START_REF] Geleziunas | Modulators of toll-like receptors for the treatment of hiv, US20170071944A1[END_REF]

.2. Saturated derivatives

Pteridinone-based compounds were identified as potent and selective TLR7 agonists, leading to the discovery of GS-9620 (34, also known as vesatolimod, Figure 18), a drug with multiple potent applications. [START_REF] Roethle | Identification and Optimization of Pteridinone Toll-like Receptor 7 (TLR7) Agonists for the Oral Treatment of Viral Hepatitis[END_REF][START_REF] Turnas | PRECLINICAL CHARACTERIZATION OF GS-9620, A POTENT AND SELECTIVE ORAL TLR7 AGONIST[END_REF] This orally bioavailable small molecule has 30-fold selectivity over TLR8 (EC50 (TLR7) = 291 nM, EC50 (TLR8) = 9 µM). GS-9620 selectively induces IFN-α, cytokines and chemokines. TLR7 activation by GS-9620 is likely associated with compound-induced conformational changes. Activation of NF-κβ and Akt pathways in primary plasmacytoid dendritic cells occurs as immediate downstream cellular responses to GS-9620 stimulation. GS-9620 possesses high absorption properties in the gastrointestinal tract but has moderate clearance during first-pass hepatic metabolism. These properties minimize systemic exposure of GS-9620 following oral administration while targeting exposure to, and stimulation of, innate immune cells within the gastrointestinal tract and/or liver. [START_REF] Fosdick | Pharmacokinetic and Pharmacodynamic Properties of GS-9620, a Novel Toll-Like Receptor 7 Agonist, Demonstrate Interferon-Stimulated Gene Induction without Detectable Serum Interferon at Low Oral Doses[END_REF][START_REF] Niu | Toll-like receptor 7 agonist GS-9620 induces prolonged inhibition of HBV via a type I interferon-dependent mechanism[END_REF][START_REF] Rebbapragada | Molecular Determinants of GS-9620-Dependent TLR7 Activation[END_REF] Furthermore, GS-9620 could activate TLR7 signaling in immune cells to induce clearance of virus infected cells. The minimum effective concentrations for IFN-α induction were similar in pDCs and in PBMCs from HCV-positive donors. It has been shown to stimulate a TLR7 immune response in different animal models of chronic hepatitis B infection (mice, woodchuck, nonhuman primates, …). [START_REF] Fosdick | Pharmacokinetic and Pharmacodynamic Properties of GS-9620, a Novel Toll-Like Receptor 7 Agonist, Demonstrate Interferon-Stimulated Gene Induction without Detectable Serum Interferon at Low Oral Doses[END_REF][START_REF] Menne | Sustained efficacy and seroconversion with the Toll-like receptor 7 agonist GS-9620 in the Woodchuck model of chronic hepatitis B[END_REF] GS-9620 was administered to HBV infected three chimpanzees for 8 weeks with an interval of 1 week. [START_REF] Lanford | an oral agonist of Toll-like receptor-7, induces prolonged suppression of hepatitis B virus in chronically infected chimpanzees[END_REF] Consequently, serum concentrations of HBV surface antigen and HBV antigen, and the number of HBV antigen-positive hepatocytes, were decreased as hepatocyte apoptosis increased. In a phase 1 clinical trial to evaluate the safety and tolerability of GS-9620, treatment of GS-9620 results in dose dependent increases in select cytokines, chemokines, and ISGs beginning at 2mg and is safe in a single dose up to 12 mg. Increases in percentages of immunocytes, like T cells, B cells and NK cells, expressing CD69 were also noted in subjects receiving GS-9620 treatment. [START_REF] Gane | The oral toll-like receptor-7 agonist GS-9620 in patients with chronic hepatitis B virus infection[END_REF][START_REF] Lopatin | Safety, pharmacokinetics and pharmacodynamics of GS-9620, an oral Toll-like receptor 7 agonist[END_REF][148] A dose-dependent pharmacodynamic induction of interferonstimulated genes (ISGs) was demonstrated without a significant systemic induction of IFNα expression or related symptoms. No significant hepatitis B surface antigen decline was observed. [START_REF] Janssen | Safety, efficacy and pharmacodynamics of vesatolimod (GS-9620) in virally suppressed patients with chronic hepatitis B[END_REF][START_REF] Boni | TLR7 Agonist Increases Responses of Hepatitis B Virus-Specific T Cells and Natural Killer Cells in Patients With Chronic Hepatitis B Treated With Nucleos(T)Ide Analogues[END_REF] Moreover, GS-9620 is able in vitro to inhibit HIV-1 infection in human peripheral blood mononuclear cells. [START_REF] Bam | TLR7 Agonist GS-9620 Is a Potent Inhibitor of Acute HIV-1 Infection in Human Peripheral Blood Mononuclear Cells[END_REF] It was also demonstrated that GS-9620 induced HIV RNA expression ex vivo in PBMC isolated from patients on suppressive antiretroviral therapy. [START_REF]TLR7 Agonist GS-9620 Activates HIV-1 in PBMCs From HIV-Infected Patients on cART | CROI Conference[END_REF][START_REF] Sloan | Targeting HIV Reservoir in Infected CD4 T Cells by Dual-Affinity Re-targeting Molecules (DARTs) that Bind HIV Envelope and Recruit Cytotoxic T Cells[END_REF][START_REF] Tsai | Toll-Like Receptor 7 Agonist GS-9620 Induces HIV Expression and HIV-Specific Immunity in Cells from HIV-Infected Individuals on Suppressive Antiretroviral Therapy[END_REF] Although GS-9620 was inactive against HIV in purified CD4+ T cells and macrophages, HIV replication was potently inhibited by conditioned medium derived from GS-9620-treated pDC cultures when added to CD4+ T cells prior to infection. GS-9620-treated PBMCs primarily showed increased production of IFN-α, and cotreatment with IFN-α-blocking antibodies reversed the HIV-1-inhibitory effect of GS-9620. Supplementary studies demonstrated that GS-9620 inhibited a post-entry event in HIV replication at a step coincident with or prior to reverse transcription. GS-9620 induced phagocytic cell maturation and improved effector-mediated killing of HIV-infected CD4 T cells by the HIV envelope-specific broadly neutralizing antibody PGT121. All data show that the compound enhanced antibody-mediated clearance of HIV-infected cells by improving immune effector functions targeting specifically those cells. GS-9620 is under clinical evaluation for reducing latent reservoirs in virally suppressed HIV-infected patients. The viral reservoir in latently infected cells is responsible for viral rebound in the vast majority of HIV-1-infected individuals who stop taking antiretroviral therapy. The key challenge to cure for HIV-1 infection is represented by the more extensively studied therapeutic strategies known as "Shock and kill". These strategies are aimed toward the reactivation of the latent reservoir using a latency-reversal agent (LRA) with the subsequent killing of the reactivated cell either by the cytotoxic arm of the immune system, including NK and CD8 T cells, or by viral cytopathic mechanisms. [START_REF] Macedo | Targeting Cellular and Tissue HIV Reservoirs With Toll-Like Receptor Agonists[END_REF][START_REF] Sarabia | HIV-1 Latency and Latency Reversal: Does Subtype Matter?[END_REF] Borducchi et al. show that administration of the V3 glycan-dependent bNAb PGT121 together with GS-9620 during ART delayed viral rebound following discontinuation of ART in simian-human immunodeficiency virus (SHIV)-SF162P3-infected rhesus monkeys in which ART was initiated during early acute infection. [START_REF] Borducchi | Antibody and TLR7 agonist delay viral rebound in SHIV-infected monkeys[END_REF] More contrasted results were obtained by Del Prete et al. [START_REF] Del Prete | TLR7 agonist administration to SIV-infected macaques receiving early initiated cART does not induce plasma viremia[END_REF] GS-986, another structurally-unpublished TLR7 agonist, was also evaluated to reduce latent reservoir in SIV-infected rhesus monkeys. [START_REF] Borducchi | Ad26/MVA therapeutic vaccination with TLR7 stimulation in SIV-infected rhesus monkeys[END_REF] 18), as demonstrated by the induction of interferon-γ induced protein (IP-10). [START_REF] Mcgowan | Design and synthesis of tetrahydropyridopyrimidine based Toll-Like Receptor (TLR) 7/8 dual agonists[END_REF] The pharmacokinetic profile of 52a1 in mice confirmed the low systemic exposure. In addition, 52a1 showed low plasma protein binding (30% bound in mice plasma), no inhibition of major CYP450 isozymes (CYP450 > 10 μM: 3A4, 2C8, 2C9, 2D6, 1A2, 2C19) and lacked off-target activity across a multi-receptors panel (> 10 μM against histamine, dopamine, and serotonin subtypes). SC1 (36, also known as Pluripotin, Figure 18), a TLR7 agonist, is an inhibitor of extracellular signalregulated kinase 1 (ERK1, MAPK3) and RasGAP with Kd values of 98 nM and 212 nM respectively. It maintains embryonic stem cell self-renewal in the absence of feeder cells and exogenous factors. Long-term SC1expanded murine embryonic stem cells can be differentiated into cells of the three primary germ layers in vitro and also can generate chimeric mice and contribute to the germ line in vivo. [START_REF] Chen | Self-renewal of embryonic stem cells by a small molecule[END_REF] Intravenously administered SC1 mediates systemic release of type I IFN, but not of pro-inflammatory cytokines such as TNFα and IL6, and results in activation of circulating immune cells. Intratumoral CD8+ T cells and CD11b+ are significantly increased, pDCs are strongly activated and macrophages are M1 phenotype polarized, whereas myeloid-derived suppressor cells are decreased. SC1 prevents spontaneous lung metastasis, retards tumor growth, results in tumor rejection, prolongs survival and, most importantly, protects from tumor rechallenge, as demonstrated by Vascotto et al. in melanoma and colorectal cancer tumor models. SC1 treatment of mice inhibits the growth of established syngeneic tumors and results in significantly prolonged survival. Tumorfree mice treated with SC1 are protected from subsequent tumor rechallenge. [START_REF] Vascotto | Intravenous delivery of the toll-like receptor 7 agonist SC1 confers tumor control by inducing a CD8+ T cell response[END_REF][START_REF] Vormehr | A non-functional neoepitope specific CD8+ T-cell response induced by tumor derived antigen exposure in vivo[END_REF] SC1 is also efficient in preventing lung metastases in a pulmonary metastatic Renca model. [START_REF] Hamm | Cancer immunotherapeutic potential of novel small molecule TLR7 and TLR8 agonists[END_REF] Mice bearing the NK cellsensitive lymphoma RMA-S are cured by repeated s. c. SC1 administrations as efficiently as with resiquimod. No relevant toxicities were observed. Mechanistically, SC1 reverses NK cell anergy and restores NK cellmediated tumor cell killing in an IFN-dependent manner. [START_REF] Wiedemann | A novel TLR7 agonist reverses NK cell anergy and cures RMA-S lymphoma-bearing mice[END_REF] SC-2 (unpublished structure) is a dual-specific TLRs 7/8 agonist which displays a more pronounced pro-inflammatory cytokines release than SC-1. Additional experiments showed a significant up-regulation of activation marker CD69 on NK cells and increased cytolytic activity of peripheral blood cells. [START_REF] Hamm | Cancer immunotherapeutic potential of novel small molecule TLR7 and TLR8 agonists[END_REF] b.3. [6+7] fused systems VTX-2337 (37, also known as Motolimod, Figure 19) is a selective and potent benzazepine small molecule agonist of TLR 8 with EC50 of 100 nM and displays 50-fold selectivity over TLR7. By coordinated activation of TLR8 and NLRP3, VTX-2337 activates NK cells and augments ADCC. [START_REF] Lu | VTX-2337 is a novel TLR8 agonist that activates NK cells and augments ADCC[END_REF][START_REF] Dietsch | Coordinated Activation of Toll-Like Receptor8 (TLR8) and NLRP3 by the TLR8 Agonist, VTX-2337, Ignites Tumoricidal Natural Killer Cell Activity[END_REF] When treated with VTX-2337, TLR8 stimulates TNF-α and IL-12 production at lower concentrations in human PBMCs. It also induces TNF-α and IL-12 secretion from monocytes and myeloid DCs through the NF-κB pathway. IFNγ secretion was observed when NK cells were treated with VTX-2337, which can enhance the lytic capability and antibody-dependent cell-mediated cytotoxicity of NK cells. VTX-2337 also improves the efficacy of pegylated liposomal doxorubicin in treatment of ovarian cancer in a mouse model with humanized immune system that has been reconstituted with human CD34+ cells. This compound is evaluated for treatment of a variety of cancers, including head and neck cancer, colorectal, pancreatic, melanoma, breast, renal cell carcinoma, non small-cell lung carcinoma, and other solid neoplasms. Evaluated as a stand-alone drug for treatment of lymphoma, VTX-2337 was essentially used in combination with other drugs. [START_REF] Lu | VTX-2337 is a novel TLR8 agonist that activates NK cells and augments ADCC[END_REF][START_REF] Monk | Integrative Development of a TLR8 Agonist for Ovarian Cancer Chemoimmunotherapy[END_REF] Thanks to its potential immunostimulating and anticancer activities, VTX-2337 is currently under clinical development as an immunotherapy for multiple oncology indications such as ovarian cancer and squamous cell carcinoma of the head and neck. [START_REF] Chow | Phase Ib Trial of the Toll-like Receptor 8 Agonist, Motolimod (VTX-2337), Combined with Cetuximab in Patients with Recurrent or Metastatic SCCHN[END_REF] VTX-1463 is an intranasal formulation of VTX-2337 under phase I clinical trial for allergic rhinitis. [START_REF] Hershberg | Methods for the treatment of allergic diseases[END_REF][START_REF] Royer | VTX-1463, a novel TLR-8 agonist, attenuates nasal congestion after ragweed challenge in sensitized beagle dogs[END_REF] Figure 19 : VTX series and analogues Among the VTX family (Figure 19) [START_REF] Howbert | Methods of Synthesis of Benzazepine Derivatives[END_REF], VTX-294 (also known as VTX-763) was defined as an ultra-potent TLR8 agonist that activates newborn and adult leukocytes and is a candidate vaccine adjuvant in both early life and adulthood. VTX-294 was 100x more active on TLR8-than TLR7-transfected HEK cells (EC50 (TLR8) = 50 nM vs. EC50 (TLR7) = 5700 nM) defining by this way its selectivity for TLR8. VTX-294-induced TNF and IL-1β production were comparable in newborn cord and adult peripheral blood and was more potent than MPLA, R848 or CL075. Combination of VTX-294 and MPLA induced greater blood TNF and IL-1β responses than combination of R-848 and MPLA. VTX-294 also potently induced expression of cytokines and co-stimulatory molecules HLA-DR and CD86 in human newborn MoDCs. [START_REF] Dowling | The Ultra-Potent and Selective TLR8 Agonist VTX-294 Activates Human Newborn and Adult Leukocytes[END_REF] VTX-763, another TLR8 agonist, is in preclinical study to treat autoimmune inflammation. VTX-463, a dual TLRs 7/8 agonist, is in preclinical study to treat allergies.

TL8-506, a benzoazepine analogue to motolimod (38, Figure 19), is also a TLR8 ligand more potent than resiquimod and 3M-002. TL8-506 is 500x more active on HEL-Blue hTLR8 than TLR7-transfected HEK293 (HEK-Blue™ hTLR7) cells (EC50 = 30 nM vs 15 µM). TL8-506 is 50x and 25x more potent in inducing NF-κB activation in TLR8-transfeceted HEK293 cells than resiquimod and 3M-002 (EC50 = 30 nM vs 1500 nM and 800 nM respectively). [START_REF] Yamamoto | STING agonists activate latently infected cells and enhance SIV-specific responses ex vivo in naturally SIV controlled cynomolgus macaques[END_REF][START_REF] Tang | Toll-Like Receptor 8 Agonist Strengthens the Protective Efficacy of ESAT-6 Immunization to Mycobacterium tuberculosis Infection[END_REF] c. (Hetero)Tricyclic compounds c.1. Imidazo[4,5-c]quinoline series Among the imidazo[4,5-c]quinolines series, imiquimod and its derivative resiquimod (Figure 20) were the first synthetic molecules described in literature as TLR8 and/or TLR7 agonists. [START_REF] Jurk | Human TLR7 or TLR8 independently confer responsiveness to the antiviral compound R-848[END_REF][START_REF] Hemmi | Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway[END_REF] Various derivatives have been then designed. It is commercialized as Aldara® or Zyclara®, respectively 5% and 3.75% imiquimod-containing creams. It was first described for its antiviral activity against certain viruses such as herpes simplex II (HSV-II) [START_REF] Harrison | Modification of immunological responses and clinical disease during topical R-837 treatment of genital HSV-2 infection[END_REF], sendai virus [START_REF] Megyeri | Stimulation of interferon and cytokine gene expression by imiquimod and stimulation by Sendai virus utilize similar signal transduction pathways[END_REF] and papilloma virus [START_REF] Hengge | Topical immunomodulators for the treatment of external genital warts, cutaneous warts and molluscum contagiosum[END_REF]. This molecule does not directly inhibit viral replication, it is an immunomodulator whose viral activity is related to the inducing power of IFNα and other cytokines. It was approved in 1997 by the Food and Drug Administration (FDA) for the treatment of external genital and perianal warts (condylomata acuminata). In 2004, its use was extended to the treatment of actinic keratosis and superficial basal cell carcinoma. [START_REF] Miller | Imiquimod applied topically: a novel immune response modifier and new class of drug[END_REF][START_REF] Tyring | Imiquimod applied topically: A novel immune response modifier[END_REF] In 2002, Hemmi et al. have shown that imiquimod and its derivative resiquimod (Figure 20) exert their antiviral and antitumor properties via TLR7 (EC50 = 10.7 µM). [START_REF] Hemmi | Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway[END_REF] They showed that TLR7-deficient mice were not able to produce pro-inflammatory cytokines following stimulation with imiquimod and resiquimod. Once bound to its TLR7 receptor, imiquimod triggers a Myd88-dependent signaling pathway that drives activation of NF-κB. Once activated, NF-κB migrates into the nucleus and induces the transcription of proinflammatory cytokines: TNFα, IL-1, IL-6, IL-8 and IL-12 as well as IFNα. The consequence of this effect is a marked stimulation of the immune response against infected and tumor cells. [START_REF] Hanna | Imiquimod in dermatology: an overview[END_REF] The effect of imiquimod was also demonstrated by skin recruitment and activation of pDCs. [START_REF] Urosevic | Disease-independent skin recruitment and activation of plasmacytoid predendritic cells following imiquimod treatment[END_REF] In addition, topically applied imiquimod induces the maturation of epidermal Langerhans cells in vivo and stimulates the migration of these antigen-presenting cells to the lymph nodes where they induce a specific T-cell response. [START_REF] Suzuki | Imiquimod, a topical immune response modifier, induces migration of Langerhans cells[END_REF] In addition, imiquimod can also enhance adaptive immunity by indirectly stimulating the production of IFNγ, a cytokine involved in a Th1 response. [START_REF] Rogge | Selective expression of an interleukin-12 receptor component by human T helper 1 cells[END_REF][START_REF] Weber | Induction of pro-inflammatory cytokine production in thymocytes by the immune response modifiers Imiquimod and Gardiquimod TM[END_REF] At high concentrations, imiquimod induces apoptosis of tumor cells via the activation of Bcl-2 proteins and the caspase family. [START_REF] Schön | The antitumoral mode of action of imiquimod and other imidazoquinolines[END_REF] Besides FDA approved indications, topical imiquimod has been used in the treatment of several infectious skin disorders (non-anagenital cutaneous warts [START_REF] Muzio | Treatment of non-genital warts with topical imiquimod 5% cream[END_REF], Molluscum Contagiosum [START_REF] Van Der Wouden | Interventions for cutaneous molluscum contagiosum[END_REF], cutaneous leishmaniasis [START_REF] Tuon | Treatment of New World cutaneous leishmaniasis--a systematic review with a meta-analysis[END_REF], etc.), neoplastic (fungoid mycosis [START_REF] Suchin | Treatment of stage IA cutaneous T-Cell lymphoma with topical application of the immune response modifier imiquimod[END_REF], vascular tumors [START_REF] Sanchez-Carpintero | Clinical and histopathologic observations of the action of imiquimod in an epithelioid hemangioendothelioma and Paget's mammary disease[END_REF], cutaneous metastases [START_REF] Steinmann | Topical imiquimod treatment of a cutaneous melanoma metastasis[END_REF], etc.) and other disorders such as ring granuloma [START_REF] Kuwahara | Granuloma annulare resolved with topical application of imiquimod[END_REF], primary follicular mucinosis [START_REF] Alonso De Celada | Treatment of primary follicular mucinosis with imiquimod 5% cream[END_REF]. Moreover, topical imiquimod might be considered as adjuvant for vaccination. [START_REF] Suchin | Treatment of stage IA cutaneous T-Cell lymphoma with topical application of the immune response modifier imiquimod[END_REF][START_REF] Fehres | Topical rather than intradermal application of the TLR7 ligand imiquimod leads to human dermal dendritic cell maturation and CD8+ T-cell cross-priming[END_REF] Imiquimod was also tested as potent agent against prostate [START_REF] Han | In vitro and in vivo growth inhibition of prostate cancer by the small molecule imiquimod[END_REF], bladder [START_REF] Donin | A phase 2 study of TMX-101, intravesical imiquimod, for the treatment of carcinoma in situ bladder cancer[END_REF][START_REF] Hayashi | Intravesical Toll-like receptor 7 agonist R-837: optimization of its formulation in an orthotopic mouse model of bladder cancer[END_REF][START_REF] Camargo | Intravesical Immunomodulatory Imiquimod Enhances Bacillus Calmette-Guérin Downregulation of Nonmuscle-invasive Bladder Cancer[END_REF], breast [START_REF] Adams | Topical TLR7 agonist imiquimod can induce immune-mediated rejection of skin metastases in patients with breast cancer[END_REF][START_REF] Adams | Clinical Trial Evidence of the Antitumor Activity of Topical Imiquimod for Breast Cancer Skin Metastases[END_REF][START_REF] Rozenblit | Transcriptomic profiles conducive to immunemediated tumor rejection in human breast cancer skin metastases treated with Imiquimod[END_REF][START_REF] Kohrt | Breast Cancer Treatment with Imiquimod: Applying an Old Lotion to a New Disease[END_REF] cancers and squamous carcinoma [START_REF] Bhatta | Therapeutic effect of Imiquimod enhanced ALA-PDT on cutaneous squamous cell carcinoma[END_REF][START_REF] Ahn | Toll-like receptor 7 agonist, Imiquimod, inhibits oral squamous carcinoma cells through apoptosis and necrosis[END_REF].

Resiquimod (1, also known as R-848, Figure 20) is an imidazo [4,5-c]quinoline compound stimulating the immune system by activating TLR7 and TLR8 (EC50 (TLR7) = 1.5 ± 0.3 µM, EC50 (TLR8) = 4.5 ± 3.2 µM). It was developed by 3M pharmaceuticals laboratories in the early 1980s in attempts to identify nucleoside-like structures inhibiting infection with HSV-2. [START_REF] Bernstein | Daily or weekly therapy with resiquimod (R-848) reduces genital recurrences in herpes simplex virus-infected guinea pigs during and after treatment[END_REF] Although Imiquimod and its derivative resiquimod belong to the same chemical family, resiquimod can activate both TLR7 and TLR8, whereas imiquimod is only TLR7 agonist. [START_REF] Jurk | Human TLR7 or TLR8 independently confer responsiveness to the antiviral compound R-848[END_REF][START_REF] Hemmi | Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway[END_REF] These differences in TLR binding presumably explain the important cytokine induction subsequent TLRs activation. The cytokine profile observed following exposure of human PBMCs to resiquimod is similar to that of imiquimod, nevertheless the level of expression of pro-inflammatory cytokines (TNFα, IL-1β and IL-6) as well as IFNα is approximately 100 times higher when using the 1 μg.mL -1 of resiquimod compared to 1.2 μg.mL -1 of imiquimod. [START_REF] Tomai | Immunomodulating and antiviral activities of the imidazoquinoline S-28463[END_REF][START_REF] Testerman | Cytokine induction by the immunomodulators imiquimod and S-27609[END_REF] In response to resiquimod, dendritic cells secrete IL-6, IL-12, TNFα and IFNα [START_REF] Ahonen | Dendritic cell maturation and subsequent enhanced T-cell stimulation induced with the novel synthetic immune response modifier R-848[END_REF], monocytes secrete IL-1, IL-6, IL-8, IL-12, TNFα and IFNα [START_REF] Wagner | Modulation of TH1 and TH2 cytokine production with the immune response modifiers, R-848 and imiquimod[END_REF].

Even if the development of topical resiquimod for the treatment of genital herpes in humans has been discontinued due to inconsistent results in clinical trials, many studies have shown the interest of resiquimod in the treatment of cutaneous tumors. In 2001, resiquimod was considered as potential anti-HSV-2 treatment but due to a lack of efficacy in clinical trials, its development in this indication was stopped. [START_REF] Fife | Effect of resiquimod 0.01% gel on lesion healing and viral shedding when applied to genital herpes lesions[END_REF][START_REF] Wu | Resiquimod: a new immune response modifier with potential as a vaccine adjuvant for Th1 immune responses[END_REF] Nevertheless, this compound proved to be effective in the treatment of actinic keratosis. At skin level, the target cells of resiquimod are mainly monocytes and dendritic cells. PDCs express TLR7 and TLR9, while MDCs express TLRs 2, 3, 4, 5 and 8, therefore resiquimod by stimulating both TLR 7 and TLR 8 can induce a strong cytokine response including type-I IFN and pro-inflammatory cytokines, which may explain the antitumor effect of resiquimod in skin cancer. [START_REF] Meyer | Resiquimod, a topical drug for viral skin lesions and skin cancer[END_REF] In basal cell carcinomas (BCC) tumor cells, imiquimod showed direct and independent of death receptor binding pro-apoptotic activity. This activity was also observed with resiquimod, however, to a much smaller extent compared with imiquimod. [START_REF] Schön | Tumor-selective induction of apoptosis and the small-molecule immune response modifier imiquimod[END_REF] Resiquimod can indirectly provoke an apoptosis induction in tumor cells by inducing the expression of TRAIL (TNF-related apoptosisinducing ligand) on dendritic cells and thus increasing their cytotoxic potency. [START_REF] Stary | Tumoricidal activity of TLR7/8-activated inflammatory dendritic cells[END_REF][START_REF] Kalb | TRAIL(+) human plasmacytoid dendritic cells kill tumor cells in vitro: mechanisms of imiquimod-and IFN-α-mediated antitumor reactivity[END_REF] In 2015, Rook et al. have shown that topical resiquimod can also improve effector T cell functions in cutaneous T-cell lymphoma and consequently induce regression of the disease. [START_REF] Rook | Topical resiquimod can induce disease regression and enhance T-cell effector functions in cutaneous T-cell lymphoma[END_REF] Furthermore, resiquimod has proven to be an effective adjuvant against several diseases as cancer given its ability to induce local activation of immune cells, stimulate the production of pro-inflammatory cytokines and improve the presentation of antigen by dendritic cells leading to the activation of cellular immune responses like CD8 + cytotoxic T lymphocytes. [START_REF] Wu | Resiquimod: a new immune response modifier with potential as a vaccine adjuvant for Th1 immune responses[END_REF][START_REF] Tomai | TLR-7 and -8 agonists as vaccine adjuvants[END_REF][START_REF] Sabado | Resiquimod as an immunologic adjuvant for NY-ESO-1 protein vaccination in patients with high-risk melanoma[END_REF] Resiquimod is also tested as potent agent in glioma [START_REF] Grauer | TLR Ligands in the Local Treatment of Established Intracerebral Murine Gliomas[END_REF], acute myeloid leukemia [START_REF] Smits | The Toll-like receptor 7/8 agonist resiquimod greatly increases the immunostimulatory capacity of human acute myeloid leukemia cells[END_REF][START_REF] Ignatz-Hoover | The role of TLR8 signaling in acute myeloid leukemia differentiation[END_REF] and breast cancer [START_REF] Yin | Toll-like receptor 7/8 agonist, R848, exhibits antitumoral effects in a breast cancer model[END_REF] c]quinoline compounds as well as structure-activity relationship studies that influence IFN induction. [START_REF] Gerster | Synthesis and structure-activity-relationships of 1H-imidazo[4,5c]quinolines that induce interferon production[END_REF] The presence of an alkyl or hydroxyalkyl chain at R1 position and no substitution at R2 show good IFN induction in human PBMCs (Figure 21). However, the activity is not improved compared to the unsubstituted compound at R1. When longer alkyl chains (heptyl or more) or bulkier alkyl substituents (i.e., tert-butyl) are considered at R1, the activity is abolished. The activity is also lost when a phenyl group is directly linked to the R1 position. The substitution by an alkyl group at the R2 position improves the IFN induction for small chains, butyl bringing the better activity (MEC = 0.01 µg/mL or 0.05 µg/mL for butyl [START_REF] West | Recognition and signaling by toll-like receptors[END_REF] or pentyl (41) respectively). Phenoxymethyl and benzyl groups at R2 also increase activity. The amine in C4 position is essential for the activity. All other modifications on this position rule out the IFN production. Overall, a simple substitution on the benzene ring appears to be detrimental since most of the synthesized compounds are not efficient in inducing IFN. However, C7-substituted compounds preserve the activity of the unsubstituted scaffold. 44)) series (Figure 23). [START_REF] Gorden | Synthetic TLR agonists reveal functional differences between human TLR7 and TLR8[END_REF] Compounds 3M-001 and 3M-002 are selective agonists for TLR7 and TLR8 respectively.

Compound 3M-001 (also known as 852A, 3M-852A, PF-04878691, PF-4878691, S-32865, Figure 23) was used in preclinical studies for ovarian, cervix and breast cancers. [START_REF] Geller | Toll-Like Receptor-7 (TLR-7) Agonist Administered Subcutaneously in a Prolonged Dosing Schedule in Heavily Pretreated Recurrent Breast, Ovarian, and Cervix Cancers[END_REF] It has undergone clinical trials for melanoma (phase II) [START_REF] Dummer | An exploratory study of systemic administration of the toll-like receptor-7 agonist 852A in patients with refractory metastatic melanoma[END_REF] and lymphocytic leukemia (phase I/II). [START_REF] Spaner | A phase I/II trial of TLR-7 agonist immunotherapy in chronic lymphocytic leukemia[END_REF] Compound 3M-002 (also known as CL075, Figure 23) has low water solubility and is a highly potent immunostimulant that can be systemically toxic. A polymersome nanocarrier encapsulating CL075 (CL075-PS) mimicked the immunomodulatory effects of live BCG vaccine and improved innate and adaptive neonatal immune responses. [START_REF] Dowling | Toll-like receptor 8 agonist nanoparticles mimic immunomodulating effects of the live BCG vaccine and enhance neonatal innate and adaptive immune responses[END_REF] The in vitro immunostimulatory activities of CL075-PS on human newborn and adult monocyte-derived dendritic cells (MoDCs) were benchmarked against conventional adjuvants and human vaccines, including the live attenuated BCG vaccine, which elicits moderate TH1 immunity in neonates and is safe and effective at birth. Compared with BCG, CL075-PS induced greater production of IL-12p70, a cytokine that enhances TH1-polarized immune responses and promotes cytotoxic T-cell proliferation and survival. Considering all their results, Dowing et al. suggest a strong potential for CL075-PS to serve as a dual antigen/adjuvant vaccine delivery system for human neonatal vaccines.

Compound 3M-003 is an agonist of both TLR7 and TLR8 (Figure 23). In the same structural family, 3M-011 (46, Figure 23) is described as a potent TLRs 7/8 agonist. It stimulates type I IFN and other cytokines such as TNF-α, IL-12, and IFN-γ from rat peripheral blood mononuclear cells, induces IL-12 and COX-2 expression in mDC from HIV+ and HIV-individuals, and inhibits H3N2 influenza viral replication in the nasal cavity. It potentiates NK cytotoxicity, and shows antitumor effects in scid/B6 mice and scid/NOD mice. That is why, 3M-011 is evaluated not only in pancreatic, gastro-intestinal and colon cancers [START_REF] Schölch | Radiotherapy combined with TLR7/8 activation induces strong immune responses against gastrointestinal tumors[END_REF][START_REF] Huber | Combined treatment with TLR7/8 agonists and carbon ion radiation induces local and abscopal immune responses in gastrointestinal tumor models[END_REF], but also in bladder and renal cancers [START_REF] Schwartz | Antitumor Effects of an Imidazoquinoline in Renal Cell Carcinoma[END_REF][START_REF] Eric | Antitumor Effects of Imidazoquinolines in Urothelial Cell Carcinoma of the Bladder[END_REF][START_REF] Kauffman | Toll-Like Receptor 7 Agonist Therapy with Imidazoquinoline Enhances Cancer Cell Death and Increases Lymphocytic Infiltration and Proinflammatory Cytokine Production in Established Tumors of a Renal Cell Carcinoma Mouse Model[END_REF].

3M-052 (47, also known as MEDI9197 or Telratolimod, Figure 23) is an injectable TLRs 7/8 agonist corresponding to a lipid modified imidazoquinoline able to form a tissue depot with gradual, sustained release, allowing local TLR triggering activity without systemic cytokine release to finally modify the tumoral microenvironment. 3M-052 displays potential immunostimulating and antitumor activities [START_REF] Smirnov | Vaccine adjuvant activity of 3M-052: An imidazoquinoline designed for local activity without systemic cytokine induction[END_REF][START_REF] Smirnov | Intratumoral immunotherapy with the TLR7/8 agonist 3M-052[END_REF] Compound 3M-052 was used as potent agent in melanoma studies. [START_REF] Singh | Effective innate and adaptive anti-melanoma immunity through localized TLR-7/8 activation[END_REF][START_REF] Singh | Induction of systemic anti-melanoma immunity through intratumoral TLR-7/8 activation[END_REF] Even if 3M-052 is retained at the sight of injection limiting systemic exposure upon intratumoral administration, it also generated systemic antitumor immunity. The agonist is able to suppress not only both injected and distant uninjected wild-type poorly immunogenic B16-F10 melanomas in mice, but also in the more immunogenic version B16.OVA melanoma. Treated tumors showed increased level of CCL2 chemokines and infiltration of M1 phenotypeshifted macrophages, which could kill tumor cells directly through production of nitric oxide and CCL2. 3M-052 therapy potentiated checkpoint blockade therapy (with anti-CTLA-4 or anti-PD-L1 antibodies), that was also confirmed by Mullins et al. [START_REF] Mullins | Intratumoral immunotherapy with TLR7/8 agonist MEDI9197 modulates the tumor microenvironment leading to enhanced activity when combined with other immunotherapies[END_REF] Zhao et al. showed that combining 3M-052 with CpG ODN increases cytotoxic T cell activity and Th1 cytokine production while downregulating the activity of immunosuppressing myeloid derived suppressor cells. This combination allows tumor suppression in CT26 colon cancer bearing mice and establishment of long-term immunity. [START_REF] Zhao | Combination therapy targeting toll like receptors 7, 8 and 9 eliminates large established tumors[END_REF] In 2017, Hoeven et al. described 3M-052 formulation development studies. [START_REF] Van Hoeven | A Formulated TLR7/8 Agonist is a Flexible, Highly Potent and Effective Adjuvant for Pandemic Influenza Vaccines[END_REF] The hydrophobicity of 3M-052 allows its incorporation in lipid-based formulations (anionic, cationic, PEGylated or neutral liposomes or oil-in-water emulsion), which were able to protect both mice and ferrets from infections caused by lethal H5N1 homologous virus. Furthermore, they demonstrated the ability of 3M-052 adjuvant formulations to broaden responses to H5N1 hemagglutinin-based antigens. Gardiquimod (46, Figure 24) is an agonist of TLR7 but not of human TLR8, although it activates both porcine TLR7 and TLR8. [START_REF] Zhu | Porcine TLR8 and TLR7 are both activated by a selective TLR7 ligand, imiquimod[END_REF] It enhances the immunosuppressive activity of T regulatory cells at 0.5 to 2 µg/mL. [START_REF] Forward | Signaling through TLR7 enhances the immunosuppressive activity of murine CD4+CD25+ T regulatory cells[END_REF] Gardiquimod is effective as a mucosal adjuvant for Norwalk virus-like particles and inhibits infection of macrophages and T cells by HIV-1. [START_REF] Velasquez | An Intranasally Delivered Toll-Like Receptor 7 Agonist Elicits Robust Systemic and Mucosal Responses to Norwalk Virus-Like Particles[END_REF][START_REF] Buitendijk | Gardiquimod: A Toll-Like Receptor-7 Agonist That Inhibits HIV Type 1 Infection of Human Macrophages and Activated T Cells[END_REF] It inhibits proliferation and migration while inducing apoptosis in pancreatic cancer cells in vitro. [START_REF] Zou | Activation of Tolllike receptor 7 inhibits the proliferation and migration, and induces the apoptosis of pancreatic cancer cells[END_REF] Guardiquimod was also evaluated on melanoma. [START_REF] Ma | The TLR7 agonists imiquimod and gardiquimod improve DC-based immunotherapy for melanoma in mice[END_REF] It should be noted that gardiquimod is a more active TLR7 agonist than imiquimod. Based on these data, Shukla et al. hypothesized that the C2 ethylaminomethylene substituent of the gardiquimod brings sufficient polarity to prevent its efficient transcutaneous penetration and stimulation of plasmacytoid dendritic cells. Hydrophobic analogues of gardiquimod were later synthesized and their TLR7 and TLR8 agonist activities were studied using HEK293 cells expressing either TLR7 or TLR8. [START_REF] Shukla | Structure-activity relationships in human toll-like receptor 7-active imidazoquinoline analogues[END_REF] Compounds with modifications of the secondary amine at the C2 position are not convenient to keep the agonist activity. Replacing the imidazole ring with a triazole ring leads to a total loss of activity. The 4-amino group is absolutely necessary to obtain maximum activity. In a series of synthesized lipophilic compounds carrying a benzyl group in the N1 position and a C2 alkyl chain show a link between the length of the alkyl chain and the TLR7 agonist activity. The optimal activity was observed for compound 47 carrying a C2 butyl chain (Figure 24) with a minimum effective concentration at which IFN induction was observed of 0.5 µg/mL as suggested by Gerster et al. [START_REF] Gerster | Synthesis and structure-activity-relationships of 1H-imidazo[4,5c]quinolines that induce interferon production[END_REF] The rearrangement of the substituents in N1 and C2 of gardiquimod and of compound 47 leads to the identification of the highly active compound 48, agonist TLR7 with an EC50 of 8.6 nM (Figure 24). These compounds are selective for TLR7 without any agonistic activity for TLR8. 25). [START_REF] Shi | Discovery of Imidazoquinolines with Toll-Like Receptor 7/8 Independent Cytokine Induction[END_REF] They showed that the elimination of the C2 ethoxymethyl chain of resiquimod results in a significant decrease in the TLR7 agonist activity and a complete loss of the TLR8 agonist activity. The activity is restored and even improved, by the addition of a butyl chain in C2 position (EC50 TLR7 = 0.1±0.02 µM, EC50 TLR8 = 0.1± 0.003 µM). The unsubstituted in C2 position and carrying a methoxycarbonyl group in C7 position compound lacked TLRs 7/8 agonist activity, however, it was able to induce a high production of pro-inflammatory cytokines (TNFα, IL12, IL1β). This suggests the existence of an alternative mechanism of action by which imidazoquinolines can stimulate the production of cytokines. The addition of the alkyl (butyl) chain in C2 position restored TLR7 and TLR8 agonist activity. All this indicates that C7 substitution may be important for cytokine production, whereas TLRs 7/8 agonist activity requires C2 substitution. Based on these data, Schiaffo et al. synthesized imidazo [4,5-c]quinolines by introducing substituents in R2 position (Figure 26). The results show that the TLRs 7/8 agonist activity is correlated with the length of the alkyl chain in C2 position, with maximum TLR 7 activity for derivatives with butyl and TLR8 activity with pentyl. A similar SAR study also identified that the production of IL1β, IL12 and IFNγ depends both on the length of the C2 alkyl chain and on the N1 substitution. The compounds also stimulate the production of IFNα and IL-10, but with less pronounced structural correlations. [START_REF] Schiaffo | Structure-activity relationship analysis of imidazoquinolines with Toll-like receptors 7 and 8 selectivity and enhanced cytokine induction[END_REF] Larson et al. proposed imidazo[4,5-c]quinoline derivatives substituted in N1 position as TLR7 and TLR8 agonists (Figure 27). While the selectivity of the compounds for TLR8 can be obtained by the introduction of an ethyl, propyl or butylamino group on N1 position [START_REF] Larson | Design and Synthesis of N1-Modified Imidazoquinoline Agonists for Selective Activation of Toll-like Receptors 7 and 8[END_REF], analysis of the SAR indicates that TLR7 activity is less sensitive to N1 modification. However, a high TLR7 affinity can be obtained by extending the aminoalkyl chain length to pentyl and p-methyl benzyl. Compared to selective TLR7 agonists and mixed TLRs 7/8 agonists, the TLR8 agonist reported better cytokine induction with higher levels of IL1β, IL12 and IFNγ. In the perspective of developing new vaccine adjuvants, Kokatla et al. synthesized thiazolo[4,5c]quinolines (Figure 28), analogous to CL075 (Figure 23). [START_REF] Kokatla | Tolllike receptor-8 agonistic activities in C2, C4, and C8 modified thiazolo[4,5-c]quinolines[END_REF] Synthesized alkylthiazoloquinolines with methyl, ethyl, propyl (CL075) and butyl groups in the R1 position have comparable TLR8 agonist activities. The activity moderately decreases with the pentyl-bearing compound at R1 position and is completely lost for the compounds with a longer alkyl chain. Virtually all changes in the R2 position led to loss of agonist activity. The alkylation of the amine at position R3 is not tolerated. The compound 55 with a butyl in R1 was also endowed with TLR7 agonist activity. Immunization in rabbits with an antigen adjuvated with compound 55 showed significant improvements in antigen-specific antibody titers. Given the pledge of TLR8 agonists in the production of pro-inflammatory cytokines including TNFα and IL12 and activation of a Th1 response in neonatal antigen presenting cells, researchers focused on the synthesis of selective TLR8 agonists for use as vaccine adjuvants, in particular to improve the immune responses in newborns. [START_REF] Cervantes-Barragan | Control of coronavirus infection through plasmacytoid dendritic-cell-derived type I interferon[END_REF] showed TLR7 agonist activities with EC50 of 0.55, 0.73 and 0.19 μM respectively, as well as TLR8 agonist activities with respective EC50 of 0.18, 3.94 and 0.056 μM (Figure 30). Imidazo[1,2-a]quinoxaline [START_REF] Liu | IPC: Professional Type 1 Interferon-Producing Cells and Plasmacytoid Dendritic Cell Precursors[END_REF] showed a selectivity towards TLR8 with an EC50 of 7.99 μM. This activity was confirmed by a study of the secretion of cytokines by human PBMCs. Compounds 58, 59 and 60 showed an ability to produce IFNα, whereas compound 61, a TLR8 agonist, was not able to produce it. In addition, these compounds have been able to produce pro-inflammatory cytokines (TNFα, IL6, IL1β, ...), type II interferon (IFNγ) as well as chemokines. Docking studies were performed with the most active compound 60 and the ectodomain of human TLR8.

. Thymidine (dT) derivatives

Poly(dT) is a thymidine 17-mer homopolymer phosphorothioate ODN commercialized by Invivogen™ and described as a modulator of human TLRs 7/8 (Figure 32). Poly(dT) alone has no significant effect on TLRs 7/8. Nevertheless, in combination with imidazoquinolines such as resiquimod or CL075, poly(dT) increases TLR8 mediated signaling and abolishes TLR7 mediated signaling. [START_REF] Jurk | Modulating responsiveness of human TLR7 and 8 to small molecule ligands with T-rich phosphorothiate oligodeoxynucleotides[END_REF][START_REF] Gorden | Oligodeoxynucleotides differentially modulate activation of TLR7 and TLR8 by imidazoquinolines[END_REF] Moreover, co-incubation of poly(dT) and an imidazoquinoline induces NF-κβ activation in HEK293 cells transfected with murine TLR8and primary TLR8-expressing mousse cells. [START_REF] Gorden | Cutting edge: activation of murine TLR8 by a combination of imidazoquinoline immune response modifiers and polyT oligodeoxynucleotides[END_REF] Figure 32 : Thymidine nucleoside d.2. RNA-based agonists MCT465, developed by MultiCell Technologies Inc., is a high molecular weight synthetic double-stranded RNA (dsRNA) designed to activate the innate immune system through TLRs 3/7/8 signaling. with immune enhancing properties. In preclinical studies, MCT-465 successfully reduced pulmonary influenza virus levels 1,000-fold in animal models. It also demonstrated effectiveness in reducing virus levels of HIV and HCV in animal models. MCT465 is currently ongoing preclinical trials for liver cancer (hepatocellular carcinoma), breast cancer (triple-negative breast cancer TNBC), and others.[259] According to the company, MCT-465 could be indicated as cancer adjuvant therapy alone or combined with currently available or novel therapies. MCT-465 can also be combined with MCT-475, a patent pending antigen-presenting immunoglobulin therapeutic, for the treatment of various cancers such as breast and metastatic colorectal carcinoma.

CV8102 is a ssRNA-based TLRs 7/8 and retinoic acid-inducible gene I (RIG-I) agonist containing (noncoding and uncapped) several polyU-repeats complexed with a polymeric carrier formed by arginine-rich disulfide-crosslinked cationic peptides. It was developed by CureVac company as adjuvant to enhance immunogenicity of poorly immunogenic antigens. Upon intratumoral injection, CV8102 stimulates a Th1 response and activates a systemic cytotoxic T-lymphocyte-mediated immune response against the tumor cells when simultaneously exposed to tumor-associated antigens. CV8102 is actually under clinical investigations for oncological indications. [START_REF] Doener | RNA-based adjuvant CV8102 enhances the immunogenicity of a licensed rabies vaccine in a first-in-human trial[END_REF][START_REF] Heil | Species-Specific Recognition of Single-Stranded RNA via Toll-like Receptor 7 and 8[END_REF][START_REF] Heidenreich | A novel RNA-based adjuvant combines strong immunostimulatory capacities with a favorable safety profile[END_REF]

e. Structurally-unpublished potent agonists

The researches and developments of agonist ligands are clearly booming. Certain referenced products display interesting activities without detailed chemical structure. Indications on their belonging to chemical families are sometimes given (Table 1).

RO7020531

TLRs 7/8 Cure chronic HBV infection [START_REF] Gane | RO7020531, a novel prodrug of a toll-like receptor 7 agonist, is safe, well tolerated and activates TLR signaling in healthy volunteers[END_REF] AZD8848 (DSP-3025) -TLR7

Long-term remission in allergic disorders such as bronchial asthma and allergic rhinitis [START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF] DSP-0509 TLR7 Potent immunostimulatory and antineoplastic activities [START_REF] Ota | Abstract 4726: Novel intravenous injectable TLR7 agonist, DSP-0509, synergistically enhanced antitumor immune responses in combination with anti-PD-1 antibody[END_REF][START_REF]DSP 0509 -AdisInsight[END_REF]

BNT411 -TLR7

Combination with chemotherapy and checkpoint inhibitors in preclinical development [START_REF] Pipeline | [END_REF] JNJ-4964 (AL-034 or TQ-A3334) -TLR7 Clinical development for the treatment of chronic hepatitis B (CHB) [START_REF] Hu | Orally Administered TQ-A3334 to Evaluate the Safety,Tolerability,Pharmacokinetics and Pharmacodynamics Single and Multiple Ascending Doses in Healthy Chinese Subjects[END_REF][START_REF] Herschke | PS-076-Antiviral activity of JNJ-4964 (AL-034/TQ-A3334), a selective toll-like receptor 7 agonist, in AAV/HBV mice after oral administration for 12 weeks[END_REF][START_REF] De Creus | P1-014: Preclinical in-vivo characterization of an oral TLR7 agonist, AL-034 (or TQ-A3334)[END_REF][START_REF] Li | P1-015: Oral administration of JNJ-4964 (AL-034/TQ-A3334), a selective toll-like receptor 7 agonist, in AAV/HBV mice for 12 weeks, resulted in potent and sustained anti-HBV activity with HBsAg seroconversion and detectable HBsAg-specific T cells and B cells[END_REF] NKTR-262 TLRs 7/8 Induction of antigen-specific immunity and tumor regression effects [START_REF] Adams | Toll-like receptor agonists in cancer therapy[END_REF][START_REF] Kivimäe | Harnessing the innate and adaptive immune system to eradicate treated and distant untreated solid tumors[END_REF] IV. Antagonist ligands

Beside their role as defensive tools of the immune system, over-activation or dysregulation of the TLR signaling are causing a panel of diseases. TLRs 7 and 8 are today well known for their implication in the development of autoimmune diseases, cancers and in the progression of the acquired immunodeficiency syndrome (AIDS). Countering uncontrolled TLR-mediated signaling has been a major subject of research in the last decades by designing TLR agonist ligands and to a much lesser extent TLR antagonist ligands. TLR antagonists are regulators that inhibit or reduce activation of TLR-mediated cytokine cascades and check over-reactive uncontrolled adaptive immune responses. TLR antagonists are generally modified agonists that bind TLRs but fail to induce the signal transduction. [START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF][START_REF] Patra | Recent progress in the development of Toll-like receptor (TLR) antagonists[END_REF] Recently highlighting the existence of antagonist pockets on TLRs 7/8 will also allow specific structure-based design using molecular modeling tools to produce new potent structures. [START_REF] Bou Karroum | Novel and Selective TLR7 Antagonists among the Imidazo[1,2-a]pyrazines, Imidazo[1,5-a]quinoxalines, and Pyrazolo[1,5-a]quinoxalines Series[END_REF][START_REF] Zhang | Small-molecule inhibition of TLR8 through stabilization of its resting state[END_REF] They represent novel therapies to treat or prevent diseases cited herein. [START_REF] Severa | IFN-β Therapy Regulates TLR7-Mediated Response in Plasmacytoid Dendritic Cells of Multiple Sclerosis Patients Influencing an Anti-Inflammatory Status[END_REF] 1. Therapeutic purposes a. Implications in autoimmune diseases Scientific literature show that in autoimmune diseases, such as lupus and psoriasis, TLRs 7 and 8 recognize endogenous immune complexes (IC) containing nucleic acids, and induce pro-inflammatory cytokines contributing to the progression of the disease. [START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF][START_REF] Lai | Natural Modulators of Endosomal Toll-Like Receptor-Mediated Psoriatic Skin Inflammation[END_REF] TLRs 7 and 8 activation by ICs leads to IL12, IL6, TNFα, IL1β and IFNα expression and is associated with the presence of anti-DNA and anti-RNA autoantibodies in patients with systemic lupus erythematosus (SLE). [START_REF] Marshak-Rothstein | Toll-like receptors in systemic autoimmune disease[END_REF][START_REF] Finke | Endogenous type I interferon inducers in autoimmune diseases[END_REF] These autoantibodies cause organ damage by binding directly to host tissues or forming IC that deposit in vascular tissue, thereby inducing destructive inflammation. [START_REF] Tsokos | Systemic lupus erythematosus[END_REF] Several studies using TLRdeficient mice have investigated the role of TLRs 7, 8 and 9 in the development of SLE in mice prone to lupus. The deletion of TLR7 in mice prone to lupus led to failure in antibody production, decreased lymphocyte activation, decreased serum IgG levels and attenuation of the disease. [START_REF] Christensen | Toll-like receptor 7 and TLR9 dictate autoantibody specificity and have opposing inflammatory and regulatory roles in a murine model of lupus[END_REF] The role of TLR8 in autoimmune diseases has not been extensively studied, perhaps because of a lack of response to TLR8 ligands in murine models. There is a difference in the response immune profile after TLR 7 or TLR8 stimulation. [START_REF] Jurk | Human TLR7 or TLR8 independently confer responsiveness to the antiviral compound R-848[END_REF] TLR7 responses are characterized by a high production of type I IFNs such as IFNα, which appears to play a major role in autoimmune diseases, while TLR8 stimulates very low levels of IFNα but induces a strong production of other pro-inflammatory cytokines. [START_REF] Gorden | Synthetic TLR agonists reveal functional differences between human TLR7 and TLR8[END_REF][START_REF] Jurk | Modulating responsiveness of human TLR7 and 8 to small molecule ligands with T-rich phosphorothiate oligodeoxynucleotides[END_REF][START_REF] Forsbach | Characterization of conserved viral leader RNA sequences that stimulate innate immunity through TLRs[END_REF] However, TNFα produced during TLR8 activation may exert pro-inflammatory effects in autoimmune diseases. [START_REF] Vielhauer | Functions of TNF and its receptors in renal disease: distinct roles in inflammatory tissue injury and immune regulation[END_REF][START_REF] Bandt | Lessons for lupus from tumour necrosis factor blockade[END_REF] TLR8 being stimulated by ribonucleoproteins [START_REF] Vollmer | Immune stimulation mediated by autoantigen binding sites within small nuclear RNAs involves Toll-like receptors 7 and 8[END_REF] and TLR8 mRNA over-regulated in patients with Sjögren's syndrome [START_REF] Gottenberg | Activation of IFN pathways and plasmacytoid dendritic cell recruitment in target organs of primary Sjögren's syndrome[END_REF] indicates a possible role of TLR8 in autoimmune processes. In psoriasis, a chronic autoimmune inflammatory disease of the skin, extracellular RNA forms complexes with the cationic antimicrobial peptide LL37. These complexes are highly protected against degradation of RNAs and have access to the endosomal compartments of pDCs and mDCs. The RNA-LL37 complexes induce the activation of TLR7 in PDCs and trigger IFNα secretion. RNA-LL37 complexes also trigger direct activation of mDCs to secrete TNFα and IL6 and differentiate into mature dendritic cells. This maturation of MDCs is triggered by endosomal TLR8 and is enhanced by the concomitant activation of pDCs to produce IFNα. These data identify RNA-LL37 complexes as endogenous triggers of TLR7 and TLR8 in human dendritic cells thus leading to the development of psoriasis. [START_REF] Ganguly | Self-RNA-antimicrobial peptide complexes activate human dendritic cells through TLR7 and TLR8[END_REF] b. Implications in cancers

In contrast to the therapeutic benefits of TLRs 7/8 agonists on immune cells, several studies have shown that TLRs 7/8 stimulation promote tumor progression. In 2010, Cherfils-Vicini et al. have shown that TLR7 and TLR8 are highly expressed on primary tumor cells of patients with non-small cell lung cancer (NSCLC). Additionally to the chemoresistance in patients with NSCLC, TLR7 agonist induce an important protumurogenic effect in vitro. [START_REF] Cherfils-Vicini | Triggering of TLR7 and TLR8 expressed by human lung cancer cells induces cell survival and chemoresistance[END_REF] These effects were also verified in immunodeficient (NOD/SCID) and immunocompetent (C57BL/6) murine models, using subcutaneously grafted lung carcinoma cells. [START_REF] Chatterjee | TLR7 promotes tumor progression, chemotherapy resistance, and poor clinical outcomes in non-small cell lung cancer[END_REF] In both models, the repeated administration of TLR7 agonists, such as CL264, loxoribine or imiquimod resulted in increased tumor volume. This protumorigenic effect could be mediated by direct stimulation of tumor cells expressing TLR7 or by increased recruitment and differentiation of immunosuppressive cells into the tumor microenvironment. Furthermore, research teams find similar results in various tumor models. Stimulation of TLR7 in a pancreatic cancer model expressing TLR7 induces tumor growth acceleration and reduction in antitumor molecules expression. [START_REF] Ochi | Toll-like receptor 7 regulates pancreatic carcinogenesis in mice and humans[END_REF][START_REF] Vaz | Intervention on toll-like receptors in pancreatic cancer[END_REF] Similarly, TLR7 stimulation in hepatocellular carcinoma model expressing TLR7 boost malignant cells proliferation. [START_REF] Mohamed | Effect of toll-like receptor 7 and 9 targeted therapy to prevent the development of hepatocellular carcinoma[END_REF] c. TLR7 implication in AIDS Chronic immune activation is a leading cause of progressive immunodeficiency of human immunodeficiency virus type 1 (HIV-1) infection. HIV ssRNA is a potent immune activator that triggers TLR7mediated signaling. The HIV virus itself directly contributes to the activation of the immune system and its dysfunction by stimulating TLR7. [START_REF] Baenziger | Triggering TLR7 in mice induces immune activation and lymphoid system disruption, resembling HIV-mediated pathology[END_REF] Recent studies have shown that activation of TLR7 in human CD4+ T cells triggers an anergic state that may contribute to CD4+ T cell hyporeactivity after infection with HIV-1 and may also increase the spread of this virus. This unexpected role of TLR7, whose activation facilitates infection of CD4+ T cells by HIV-1, was blocked using a TLR7 inhibitor. The presence of this inhibitor has led to an attenuation of the infection. The authors concluded that HIV-1 use the anergic state induced by TLR7 activation to support its own spread. [START_REF] Lederman | A surprising role for TLR7[END_REF] The inhibition of TLRs 7 and 8 activation by synthetic compounds is therefore a promising research area for potential treatment of autoimmune diseases, cancers and AIDS. TLRs 7/8 antagonists bind to the receptor and inhibit its activation, thereby suppressing the inherent immune response. Till 2020, antagonist molecules are compounds developed generally as structural analogues to TLRs 7/8 agonists. As in the case of TLR7 and TLR8 agonists, currently known antagonists can be divided into two groups (oligonucleotides and bi-and tricyclic heterocyclic compounds) that we discus below.

Synthetic ligands a. Imidazole derivatives

In their search for TLR8 agonists containing a benzimidazole moiety, Beesu et al. have prepared an inactive agonist (63, Figure 33) that competitively inhibited TLR8, while inhibited TLRs 2, 3, 4, 5, 7, and 9 by an unknown mechanism. [START_REF] Beesu | Identification of a Human Toll-Like Receptor (TLR) 8-Specific Agonist and a Functional Pan-TLR Inhibitor in 2-Aminoimidazoles[END_REF] The micromolar affinity of this pan-TLR antagonist toward multiple TLRs in various cell lines suggests that it could be further a promising anti-inflammatory agent. A kinase-screening of this compound showed relative specificity for calmodulin kinases. [START_REF] Ugolini | Recognition of microbial viability via TLR8 drives T FH cell differentiation and vaccine responses[END_REF], which proved to be the most active with an IC50 of 67 nM and a strong ability to inhibit the production of pro-inflammatory cytokines (Figure 34). Docking studies reveal that CU-CPT8m is sandwiched between the two monomers of TLR8 and is localized in a hydrophobic pocket at the proteinprotein interface of the two monomers TLR8. This new binding site is close, but distinct from the identified agonist site, whose occupation prevents the activation of TLR8. This implies a new inhibitory mechanism by CU-CPT8m. After these encouraging results, the authors carried out a structural optimization which led to two new TLR8 inhibitors belonging to the quinolines series structurally similar to CU-CPT8m: CU-CPT9a (65, IC50 = 0.5 nM) and CU-CPT9b (66, IC50 = 0.7 nM) (Figure 34). By blocking the newly identified site, these TLR8 inhibitors appear to antagonize the binding of TLR8 agonists such as R848 and uridine, and/or block a conformational change necessary for the receptor activation. In addition, authors explored the effects of CU-CPT8m and CU-CPT9a in human samples from various patients with inflammatory and autoimmune diseases. The study showed that treatment with CU-CPT9a has potent anti-inflammatory effects in the samples of patients with osteoarthritis (OA), rheumatoid arthritis (RA), and adult Still's disease, thus confirming previously published work showing that TLR8 may play a role in inflammatory disorders and autoimmune diseases. Known as antimalarial drugs chloroquine (67, CQ) and hydroxychloroquine (68, HCQ) (Figure 35), have been used to treat autoimmune diseases (arthritis and SLE) after World War II. [START_REF] Fox | Anti-malarial drugs: possible mechanisms of action in autoimmune disease and prospects for drug development[END_REF][START_REF] Ruiz-Irastorza | Clinical efficacy and side effects of antimalarials in systemic lupus erythematosus: a systematic review[END_REF] These weak bases tend to accumulate in the acidic intracellular compartments of endosomes and lysosomes and modulate their pH leading to suppression of autoantigen presentation, blockade of endosomal TLR signaling, and decrease in cytokine production. [START_REF] Kužnik | Mechanism of Endosomal TLR Inhibition by Antimalarial Drugs and Imidazoquinolines[END_REF][START_REF] Lee | The role of antimalarial agents in the treatment of SLE and lupus nephritis[END_REF] Other mechanisms include inhibition of MAPK signaling and phospholipase A2, antiproliferation, photoprotection as well as reduction of matrix metalloproteinase-9 activities. [START_REF] Weber | Inhibition of Mitogen-Activated Protein Kinase Signaling by Chloroquine[END_REF][START_REF] Kim | Hydroxychloroquine potentiates Fas-mediated apoptosis of rheumatoid synoviocytes[END_REF][START_REF] Wozniacka | The influence of antimalarial treatment on IL-1β, IL-6 and TNF-α mRNA expression on UVB-irradiated skin in systemic lupus erythematosus[END_REF][START_REF] Lesiak | Effect of chloroquine phosphate treatment on serum MMP-9 and TIMP-1 levels in patients with systemic lupus erythematosus[END_REF] All these mechanisms of action highlight the anti-inflammatory and immuno-suppressive activity of this antimalarial drugs by acting on endosomal TLR signaling (TLRs 7/8/9). Moreover, CQ was under investigation for diseases associated with uncontrolled acute or chronic inflammation (severe sepsis [START_REF] Yasuda | Chloroquine and inhibition of Toll-like receptor 9 protect from sepsis-induced acute kidney injury[END_REF]), to prevent and treat viral infections (HIV, influenza, and dengue) [START_REF] Paton | Chloroquine for influenza prevention: a randomised, double-blind, placebo controlled trial[END_REF][START_REF] Borges | Chloroquine use improves dengue-related symptoms[END_REF], cardiovascular diseases [START_REF] Cui | Chloroquine pretreatment inhibits toll-like receptor 3 signaling after stroke[END_REF] or cancer by notably modulating autophagy [START_REF] Mohamed | Effect of toll-like receptor 7 and 9 targeted therapy to prevent the development of hepatocellular carcinoma[END_REF]. Long term HCQ administration ameliorate hypertension and aortic endothelial dysfunction in a SLE mouse model. [START_REF] Manuel | Chronic Hydroxychloroquine Improves Endothelial Dysfunction and Protects Kidney in a Mouse Model of Systemic Lupus Erythematosus[END_REF] Figure 35: Worldwide-known antimalarial drugs

The quinazoline derivative CpG-52364 [START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF] from Coley Pharmaceutical Group Inc. (Figure 36), a TLRs 7/8/9 antagonist (TLRs 7/8 ratio = 0.8), inhibits disease progression of SLE, rheumatoid arthritis and other autoimmune diseases in animal models. [START_REF] Lipford | Quinazoline derivative useful as toll-like receptor antagonist[END_REF][START_REF] Parkinson | The future of toll-like receptor therapeutics[END_REF][START_REF] Watanabe | Dihydropyrrolo[2,3d]pyrimidines: Selective Toll-Like Receptor 9 Antagonists from Scaffold Morphing Efforts[END_REF] It also increases potency compared to HCQ alone. In combined therapy, it added efficacy by preventing the formation of antiDNA antibodies in SLE-prone mice. CpG-52364 is evaluated in a phase I clinical trial for SLE therapy (NCT00547014). [START_REF] Honda | IRFs: master regulators of signalling by Toll-like receptors and cytosolic patternrecognition receptors[END_REF]TLR7 agonist) and its 3H regioisomer 70 in order to explore the SAR of substituents on the imidazoline ring (Figure 37). Compound 70 revealed to be completely inactive as a TLR7 agonist, having surprisingly weak antagonistic effect. A des-amino precursor of the 3H-regioisomer 71 was more potent as TLR7 antagonist, with an IC50 of 7.5 μM. [START_REF] Shukla | Preliminary evaluation of a 3H imidazoquinoline library as dual TLR7/TLR8 antagonists[END_REF] Figure 37 : Quinoline compounds Among the synthesized compounds [START_REF] Shukla | Preliminary evaluation of a 3H imidazoquinoline library as dual TLR7/TLR8 antagonists[END_REF], the 72 quinolinium bearing p-methoxybenzyl substituents at the N3 and N5 positions was identified as active antagonist of TLR7 and TLR8 with IC50 of 2.79 µM and 4.55 μM, respectively (Figure 38). Compounds 73 and 74 carrying the propargyl group on N3, have also shown a good TLRs 7/8 antagonist activity (10 µM range) and they may prove to be useful supports for additional alkylation in the N5 position with electron-rich substituents as for compound 72. Dimers linked to C2, C4, C8 and N1-aryl positions were also evaluated. [START_REF] Shukla | Toll-like receptor (TLR)-7 and -8 modulatory activities of dimeric imidazoquinolines[END_REF] C4, C8 and N1-aryl linked dimers were TLR7 agonists. Only N1-aryl dimer with a spacer of 12 carbon atoms was a double agonist of TLR7 and TLR8. C2 dimer with a propylene spacer (75) was a maximal antagonist of both TLR7 and TLR8 with IC50 of 3.1 and 3.2 μM respectively (Figure 39). This compound also showed inhibition of pro-inflammatory cytokine production in human PBMCs. Targeting different domains of a TLR could block its activation. This inhibition could also be done indirectly by targeting effectors in relationship with these TLRs. This part will not be explicit due to its remoteness with our main researches.

e.1. Nucleic acids

Lamphier et al. found two TLR7 antagonist small compounds, AT791 (79, Figure 42) and E6446 (80, Figure 42), able to interact with nucleic acid (DNA or RNA) agonists in acidic intracellular compartments. These orally bioavailable compounds can inhibit short-term induction of inflammatory cytokines by DNA. In a mouse MRL/lpr spontaneous model of lupus, E6446 slowed the development of circulating antinuclear antibodies and modestly suppressed anti-dsDNA with no observable impact on proteinuria or mortality. E6446 is effective in preventing hyperinflammation and lethality caused by the parasite Plasmodium berghei in a mouse model of cerebral malaria. [START_REF] Franklin | Therapeutical targeting of nucleic acid-sensing Toll-like receptors prevents experimental cerebral malaria[END_REF] AT791 is evaluated on SLE or autoimmune disease. [START_REF] Lamphier | Novel Small Molecule Inhibitors of TLR7 and TLR9: Mechanism of Action and Efficacy In Vivo[END_REF] [START_REF] Judge | Design of noninflammatory synthetic siRNA mediating potent gene silencing in vivo[END_REF] It has also been reported that chemical modifications of nucleotides in mammalian RNA, including 2'OMe modifications, prevents them from activating dendritic cells. [START_REF] Karikó | Suppression of RNA recognition by Toll-like receptors: the impact of nucleoside modification and the evolutionary origin of RNA[END_REF] In 2007, Robbins et al. tested the ability of 2'OMe RNA to antagonize TLR7 immune stimulation. [START_REF] Robbins | 2'-O-methyl-modified RNAs act as TLR7 antagonists[END_REF] They reported that 2'OMe RNA directly inhibits cytokines induction in murine and human systems by immunostimulatory RNA and by a TLR7 agonist Loxoribine. The inhibitory activity of 2'OMe RNA has also been shown in vivo. These results indicate that 2'OMe RNA acts as a TLR7 antagonist.

Anti-miRNA target TLRs 7/8 binding off viral and bacterial infections. [START_REF] Sarvestani | Sequence-dependent off-target inhibition of TLR7/8 sensing by synthetic microRNA inhibitors[END_REF] b. Oligonucleotides Oligodeoxynucleotides (ODNs) containing unmethylated CpG dinucleotides act as TLR9 agonists and induce Th1 type immune responses. [START_REF] Hemmi | A Toll-like receptor recognizes bacterial DNA[END_REF] On the other hand, the substitution of cytosine in the CpG motif by 2'-O-methyl-cytosine, 2'-O-methyl-5-methyl-cytosine or 5-methyl-deoxycytosine or the substitution of guanosine by 2'-O-methyl-guanosine leads to loss of immunostimulatory activity; these oligonucleotides therefore appear to be TLR7 and TLR9 antagonists. [START_REF] Yu | Modifications incorporated in CpG motifs of oligodeoxynucleotides lead to antagonist activity of toll-like receptors 7 and 9[END_REF] Dynavax Technologies has designed immunoregulatory oligonucleotides (IROs) having 2′-O-methyl or methoxyethyl sugar modifications as inhibitors of TLR7 and/or TLR9. [START_REF] Barrat | Methods of treatment using TLR7 and/or TLR9 inhibitors, US8940310B2[END_REF] Further, Dynavax Technologies has developed IROs with unique inhibitory sequences for TLR7, 8, and 9 that suppress autoimmune and inflammatory diseases. [START_REF] Guiducci | Human toll-like receptor inhibitors and methods of use thereof, US9228184B2[END_REF] The novel approach can be applied to treat a variety of autoimmune diseases, such as SLE or cutaneous lupus erythematosus (CLE), systemic sclerosis, polymyositis, dermatomyositis, RA, and Sjorgren's syndrome.

Based on this work, several recent studies have focused on the synthesis of oligonucleotides using the chemical modifications mentioned above. [START_REF] Kandimalla | Design, synthesis and biological evaluation of novel antagonist compounds of Toll-like receptors 7, 8 and 9[END_REF][START_REF] Zhu | A novel antagonist of Toll-like receptors 7, 8 and 9 suppresses lupus disease-associated parameters in NZBW/F1 mice[END_REF][START_REF] Schmitt | Identification of an optimized 2'-O-methylated trinucleotide RNA motif inhibiting Toll-like receptors 7 and 8[END_REF] Idera Pharmaceuticals has designed unique CpG oligonucleotide sequences containing an immune stimulatory motif with a 7-deaza-dG or arabino-G modification and an immune regulatory motif with 2′-O-methylribonucleotides that act as antagonists of TLRs 7, 8, and 9. [START_REF] Kandimalla | Immune regulatory oligonucleotide (iro) compounds to modulate toll-like receptor based immune response, US20140193396A1[END_REF] IROs have shown promising results in preclinical studies and effectively prevented hyperactivation of these TLRs in response to agonists. They are foreseen as future therapies for TLRs 7, 8, and 9-mediated inflammatory disorders, including various autoimmune and inflammatory diseases and cancers initiated by pathogens.

Chemical modifications and/or internucleotide linkages introduced into the IROs were found to increase inhibition or suppression of TLRs 7, 8, and 9. Their leader, IMO-8400 (also known as Bazlitoran, sequence: CTATCTNNNNTTCTCTNN), is an oligonucleotide antagonist of TLR7, TLR8 and TLR9, currently in clinical development (phase IIa) for the treatment of plaque psoriasis (NCT01899729) and dermatomyositis (NCT02612857). [START_REF] Balak | IMO-8400, a toll-like receptor 7, 8, and 9 antagonist, demonstrates clinical activity in a phase 2a, randomized, placebo-controlled trial in patients with moderate-to-severe plaque psoriasis[END_REF] Treatment with this antagonist regulates IL-23-induced gene expression as demonstrated by Suarez-Farinas et al. [START_REF] Suárez-Fariñas | Suppression of molecular inflammatory pathways by Toll-like receptor 7, 8, and 9 antagonists in a model of IL-23-induced skin inflammation[END_REF] This compound was very effective in preventing inflammation and disease development in both lupus and psoriasis mouse models. [START_REF] Jiang | IMO-8400, a novel TLR7, TLR8 and TLR9 antagonist, inhibits disease development in mouse models of psoriasis (119.8)[END_REF][START_REF] Zhu | IMO-8400, a novel TLR7, TLR8 and TLR9 antagonist, inhibits disease development in lupus-prone NZBW/F1 mice (119.12)[END_REF] Similar to IMO-8400, IMO-9200, a TLRs 7/8/9 antagonist, is evaluated on auto-immune diseases. From a phase I clinical trial, it showed a safe and well-tolerated profile in healthy subjects. IMO-9200 was evaluated as potent treatment against inflammatory bowel disease. [START_REF] Zhu | Sa1757 Targeting Innate Immune Receptors to Treat Inflammatory Bowel Disease: Preclinical Activity of IMO-9200, an Antagonist of TLRS 7, 8, and 9 in Mouse Models of Colitis[END_REF] IMO-3100, dual antagonist of TLR7 and TLR9, is being tested for the treatment of inflammatory and autoimmune disease, such as SLE, rheumatoid arthritis, multiple sclerosis, psoriasis, lupus and colitis. Evaluation of the effect of IMO-3100 on gene expression in human peripheral blood monocytes showed that the drug could significantly reduce the expression of inflammatory genes, such as IL-17A, β-defensin, CXCL1, and keratin 16, inhibited TLR ligand-induced expression of TNF-α and IFNα. [START_REF] Lai | Natural Modulators of Endosomal Toll-Like Receptor-Mediated Psoriatic Skin Inflammation[END_REF][START_REF] Suárez-Fariñas | Suppression of molecular inflammatory pathways by Toll-like receptor 7, 8, and 9 antagonists in a model of IL-23-induced skin inflammation[END_REF][START_REF] Jiang | IMO-3100, an Antagonist of Toll-Like Receptors 7 and 9, Modulates Gene Expression and Regulatory Networks Induced by Ligands (48.25)[END_REF] IMO-4200, a dual agonist of TLR7 and TLR8, is a synthetic RNA-based dual agonist of TLR7 and TLR8, identified as a lead drug candidate for the treatment of hematological malignancies. IMO-4200 stimulates immune responses mediated through TLR7 and TLR8, which are expressed in human dendritic cells, B-cells, monocytes, and macrophages. In preclinical mouse models of cancer, IMO-4200 has shown anticancer activity involving both innate and adaptive immune responses. IMO-4200, when administered in combination with approved cancer therapy drugs, rituximab (an anti-CD20 antibody) or bortezomib (a proteasome inhibitor), showed significantly increased antitumor activities compared to the single-agent effects in several preclinical lymphoma models and increased survival compared to treatment with either agent alone. Notably with rituximab, IMO-4200 is able to increase activation of natural killer cells, indicating an improved immune response and to enhance clearance of circulating tumor cells compared to treatment with agent alone. [START_REF] Wang | Supra-additive effect of IMO-4200, a novel TLR7 and TLR8 dual agonist, with rituximab and cytotoxics in preclinical models of hematologic malignancies[END_REF][START_REF] Wang | IMO-4200, a Novel TLR7 and TLR8 Dual Agonist, Enhances Antitumor Effect of Ofatumumab, Rituximab and Cytotoxics in Preclinical Models of Hematological Malignancies[END_REF] Based antagonist. This compound demonstrated a promising therapeutic effect in SLE through the suppression of both TLR7 and TLR9, potentiated the inhibitory activity and efficiently improved its capacity to reduce TLR7/9-mediated immune signaling in human immune cells. [START_REF] Römmler | Guanine-Modified Inhibitory Oligonucleotides Efficiently Impair TLR7-and TLR9-Mediated Immune Responses of Human Immune Cells[END_REF][START_REF] Römmler | Guanine Modification of Inhibitory Oligonucleotides Potentiates Their Suppressive Function[END_REF] Among their derivatives, Römmler et al. designed diverse antagonists as described below (Table 2). Several oligonucleotides antagonists of TLRs 7 and 9 have been developed such as immunoregulatory sequences (IRS) that are short DNA sequences. The most promising compound is IRS 954 (5'-TGCTCCTGGAGGGGTTGT-3' sequence, also known as DV-1079), which was developed for the treatment of SLE. [START_REF] Barrat | Nucleic acids of mammalian origin can act as endogenous ligands for Toll-like receptors and may promote systemic lupus erythematosus[END_REF] This compound was found to block IFN-α production by human plasmacytoid predendritic cells. IRS-954 treatment showed a reduction of serum levels of nucleic-acid specific autoantibodies and improvement of disease symptoms in lupus-prone mice (decrease of proteinurea, glomerulonephritis and end-organ damage) resulting in an increasing survival. [START_REF] Barrat | Treatment of lupus-prone mice with a dual inhibitor of TLR7 and TLR9 leads to reduction of autoantibody production and amelioration of disease symptoms[END_REF] IRS-954 could also reverse TLR7/9-mediated glucocorticoid resistance of SLE, and thus could be potentially used as corticosteroid-sparing drug. [START_REF] Guiducci | TLR recognition of self nucleic acids hampers glucocorticoid activity in lupus[END_REF] Administration of the drug to HIV-stimulated peripheral blood monocyte cells also led to a decrease in IFNα production, suggesting a potential therapeutic opportunity for treating HIV infection. [START_REF] Pawar | Inhibition of Toll-like receptor-7 (TLR-7) or TLR-7 plus TLR-9 attenuates glomerulonephritis and lung injury in experimental lupus[END_REF] IRS 661 (5'-TGCTTGCAAGCTTGCAAGCA-3' sequence) is a specific TLR7 antagonist evaluated by Pawar et al. on a lupus mouse model. [START_REF] Barrat | Nucleic acids of mammalian origin can act as endogenous ligands for Toll-like receptors and may promote systemic lupus erythematosus[END_REF][START_REF] Pawar | Inhibition of Toll-Like Receptor-7 (TLR-7) or TLR-7 plus TLR-9 Attenuates Glomerulonephritis and Lung Injury in Experimental Lupus[END_REF] Studies on autoimmune diabetes were also conducted. [START_REF] Zhang | Toll like receptor 7 antagonist IRS661 inhibits insulitis and autoimmune diabetes in non-obese diabetic mice (P5227)[END_REF][START_REF] Lee | Toll-like receptor 7 stimulation promotes autoimmune diabetes in the NOD mouse[END_REF] DV-1179, another TLRs 7/9 dual antagonist, entered a phase Ib/IIa study for its safety in both healthy volunteers and patients with active SLE. Although DV-1179 administration was well tolerated, it failed to achieve the endpoints of reducing the IFNα-regulated genes. Such an unfortunate outcome ended the clinical development of this compound ever since. [START_REF]Dynavax Regains Full Rights to Investigational TLR 7/9 Inhibitor DV1179 Following Expiration of Collaboration With GSK[END_REF] V. Clinical trials Herein, we gather all the clinical trials where TLRs 7/8 are mentioned and in which studies relate to the biological activities of the named compound (last update on January 2020, Table 3). [START_REF] Hennessy | Targeting Toll-like receptors: emerging therapeutics?[END_REF][START_REF] Vanpouille-Box | Pharmacological modulation of nucleic acid sensorstherapeutic potential and persisting obstacles[END_REF][START_REF] Anwar | Recent clinical trends in Toll-like receptor targeting therapeutics[END_REF][366][367][368] Among the three FDA or EMA-approved TLR agonists, only one of them targets a TLR which is discussed in this review: the TLR7 agonist imiquimod. Regarding others, BCG is agonist of TLRs 2/3/4 and possibly 9, and monophosphoryl lipid A (MPL) is agonist of TLR 4. Indeed, despite efforts from researchers, few pharmacological modulators reach final stages of clinical studies either as drugs or as adjuvants, due to obstacles that are difficult to overcome. For example, toxicity related to immunomodulating compounds could be modulated by delivering potent therapies locally (intratumorally or intradermally) thus directly targeting pathological sites and/or cell populations. [START_REF] Milling | Delivering safer immunotherapies for cancer[END_REF] Moreover, animal models do not reflect exactly the human immunological pathways, which did not allow appreciating the potentialities of certain compounds. With less genetic variability in animal models, a margin of error is possible to predict safety but active doses in humans. Specific molecular alterations or mutations due to the evolution of pathologies could also prevent potent modulators to mediate the expected therapeutic effect, in addition of host-related characteristics, such as gender, age or gene polymorphic variants. Despite all the barriers mentioned, the number of clinical trials relating to the biological activities of TLRs 7/8 ligands perfectly illustrates the interest of researchers in modulating them for therapeutic purposes.

VI. Conclusion

Toll-like receptors 7/8 demonstrate a key role in the immunology by protecting the host body against invading pathogens or endogenously released hazardous molecules. By improving or blocking their activities, TLRs are capable of modifying the inherent signaling pathways and thus mediating host responses, rendering them ingenious tools to be targeted for therapeutic purposes. A wide spectrum of diseases is associated with TLRs, which directly or indirectly aggravate the pathological conditions. Many pharmaceutical companies and research institutions are thereby developing specific TLR modulators. Some of them have been pre-clinically and clinically evaluated.

TLRs ligands are primarily used as adjuvants to boost the immunity system and to trigger humoral and/or cell-mediated responses. Agonists are also developed as drugs to fight viral infections and to prevent and/or treat cancers. Antagonists are developed for downregulating TLRs overactivity as tools for the treatment of inflammatory and autoimmune diseases. Synthetic TLRs 7/8 ligands are not only small molecules belonging to diverse chemical series, but also macromolecules, such as oligonucleotides, peptides and proteins. Rational drug design of small molecule agonists and antagonists may be helped by the recent knowledge of TLRs three-dimensional structures. The potential of non-traditional approaches such as the use of miRNAs should not be overlooked.

Trials involving TLRs ligands as adjuvants are numerous than those considering them as drugs. Moreover, the imbalance in the number of agonists and antagonists under clinical investigations could be explained by the recent understanding of the implication in pathologies of signaling dysregulation. After an explosion in the number of clinical studies launched, we have witnessed a decrease in recent years. Even if some ligands have performed well in advanced clinical trials, clinical success is generally limited. Researchers still have to overcome some obstacles. Indeed, if animal models allow to appreciate biological activities of the TLRs 7/8 modulators, they do not reflect exactly what happen in human trials due to genetic variability among others. Specific molecular alterations or mutations may also prevent potent modulators to mediate the expected therapeutic effect. Researchers aim to reduce systemic toxicity of immunomodulating compounds by modulating their formulation (liposome, hydrogel, nanoparticle instigations) or way of administration (intratumorally or intradermally). Many studies revealed the importance of combined therapy including TLRs modulators to obtain convenient results. Immunotherapy has been revealing as a good approach to complete traditional clinical management.

Even if there are similarities in the TLRs 7/8 mechanism of actions, specificities remain to be elucidated to specifically induce a response from a single receptor. Moreover, animal models do not reflect the potent human response to a drug because of the differences in TLRs expression and activity between humans and mice. Continuing researches around TLR biology would allow finding novel therapies to address unmet medical needs, such as cancers, diabetes, rheumatoid arthritis, systemic lupus erythematosus, Alzheimer's disease, and chronic neuropathic pain.

Figure 1 :

 1 Figure 1 : A synthetic tricyclic derivatives belonging to the imidazo[4,5-c]quinoline

Figure 2 :

 2 Figure 2 : Signal transduction of TLRs 7/8 receptors leading to the production pro-inflammatory cytokines III. Agonist ligands TLR signaling pathways activate innate immunity and help in modulating adaptive immunity. Hence, they are essentially useful and developed in immunotherapy. In clinical trials, two third of TLR agonist ligands are first exploited as adjuvants versus one third as potent drugs for various therapies. Herein, the therapeutic purposes of agonist ligands of TLRs 7/8 are discussed, and natural and synthetic ligands are then presented.

Figure 3 :

 3 Figure 3 : Nucleobases and nucleosides

Figure 5 :

 5 Figure 5 : Pyrimidine derivatives Beesu et al. have undergone SAR studies in the 2,4-diamine pyrimidine series, focusing on C5 substituents modulation.[95] Compound 3 (N-4-butyl-6-methyl-5-(3-morpholinopropyl)-pyrimidine-2,4diamine, Figure 5) exhibits important simultaneous TLR7 and TLR8 agonist activities, with remarkable activation of several subsets of dendritic cells (EC50 (TLR7) = 0.046 μM, EC50 (TLR8) = 0.280 μM).

Figure 6 :

 6 Figure 6 : General structure of 9-benzyl-8-hydroxyadenine derivatives

Figure 7 :

 7 Figure 7 : SM derivatives

Figure 8 :

 8 Figure 8 : TMX derivatives

Figure 9 :

 9 Figure 9 : 8-hydroxyadenine derivatives

Figure 10 : 8 -

 108 Figure 10 : 8-hydroxyadenine conjugated derivatives

Figure 11 :

 11 Figure 11 : TLRs 2/7 agonists based on the 8-hydroxyadenine moiety

Figure 12 :

 12 Figure 12 : 8-oxoadenine derivative

Figure 13 :

 13 Figure 13 : DSR derivatives DSR-29133 (26, Figure13) is a TLR7-selective agonist that leads to the induction of IFNα/γ, IP-10, TNFα, IL-1Ra and IL-12p70, and after intravenously injections to a reduction in tumor burden in syngeneic models of renal cancer (Renca), metastatic osteosarcoma (LM8) and colorectal cancer (CT26). Moreover, the efficacy of DSR-29133 was significantly improved when administered in combination with low-dose fractionated radiotherapy. Long-term surviving mice originally treated with DSR-29133 and radiotherapy were protected by a tumor-specific memory immune response which could prevent tumor growth.[START_REF] Dovedi | Intravenous administration of the selective toll-like receptor 7 agonist DSR-29133 leads to anti-tumor efficacy in murine solid tumor models which can be potentiated by combination with fractionated radiotherapy[END_REF]

Figure 14 :

 14 Figure 14 : Guanosine derivatives b. (Hetero)Bicyclic compounds b.1. [5+6] fused systems Yoo et al. synthesized new imidazo[4,5-c]pyridine compounds by incorporating substituents which were previously determined to be optimal agonists (N1-benzyl and C2-butyl, Figure15) in the imidazoquinoline series. These compounds were studied for their potency to induce the IFNα production.[START_REF] Yoo | Structure-activity relationships in Toll-like receptor 7 agonistic 1H-imidazo[4,5-c]pyridines[END_REF] Compound 29 showed TLR7 agonistic activity (EC50 = 1.57 μM) with negligible activity on TLR8. Direct aryl-aryl bonds at the R1 position abolish all activities, but the TLR7 agonist effect is restored with analogues having a benzyl or phenethyl group. For example, the specific TLR7 agonist compound 30 (EC50 = 0.26 μM) displayed high IFNα induction in human PBMCs. Since some of these compounds are potent inducers of type I IFN, with an attenuated pro-inflammatory profiles, they could be potent adjuvants without local or systemic inflammation reactions.[START_REF] Yoo | Determinants of activity at human Toll-like receptors 7 and 8: quantitative structure-activity relationship (QSAR) of diverse heterocyclic scaffolds[END_REF]

Figure 15 :

 15 Figure 15 : Core-structure of the imidazo[4,5-c]pyridine compounds

Figure 16 :

 16 Figure 16 : Benzimidazol-2-amine derivative b.2. [6+6] fused systems b.2.1. Unsaturated derivatives Among all the prepared heterocyclic derivatives, Beesu et al. found that the quinoline derivative 30 (Figure17) displays a remarquable TLR8 agonist activity with an EC50 value of 9 nM.[START_REF] Beesu | Human Toll-like receptor 8-selective agonistic activities in 1-alkyl-1H-benzimidazol-2-amines[END_REF][START_REF] Beesu | Structure-Based Design of Human TLR8-Specific Agonists with Augmented Potency and Adjuvanticity[END_REF][START_REF] Beesu | Human Toll-like Receptor (TLR) 8-Specific Agonistic Activity in Substituted Pyrimidine-2,4-diamines[END_REF]

Figure 17 :

 17 Figure 17 : Pyridine derivatives b.2.2. Saturated derivativesPteridinone-based compounds were identified as potent and selective TLR7 agonists, leading to the discovery of GS-9620 (34, also known as vesatolimod, Figure18), a drug with multiple potent applications.[START_REF] Roethle | Identification and Optimization of Pteridinone Toll-like Receptor 7 (TLR7) Agonists for the Oral Treatment of Viral Hepatitis[END_REF][START_REF] Turnas | PRECLINICAL CHARACTERIZATION OF GS-9620, A POTENT AND SELECTIVE ORAL TLR7 AGONIST[END_REF] This orally bioavailable small molecule has 30-fold selectivity over TLR8 (EC50 (TLR7) = 291 nM, EC50 (TLR8) = 9 µM). GS-9620 selectively induces IFN-α, cytokines and chemokines. TLR7 activation by GS-9620 is likely associated with compound-induced conformational changes. Activation of NF-κβ and Akt pathways in primary plasmacytoid dendritic cells occurs as immediate downstream cellular responses to GS-9620 stimulation. GS-9620 possesses high absorption properties in the gastrointestinal tract but has moderate clearance during first-pass hepatic metabolism. These properties minimize systemic exposure of GS-9620 following oral administration while targeting exposure to, and stimulation of, innate immune cells within the gastrointestinal tract and/or liver.[START_REF] Fosdick | Pharmacokinetic and Pharmacodynamic Properties of GS-9620, a Novel Toll-Like Receptor 7 Agonist, Demonstrate Interferon-Stimulated Gene Induction without Detectable Serum Interferon at Low Oral Doses[END_REF][START_REF] Niu | Toll-like receptor 7 agonist GS-9620 induces prolonged inhibition of HBV via a type I interferon-dependent mechanism[END_REF][START_REF] Rebbapragada | Molecular Determinants of GS-9620-Dependent TLR7 Activation[END_REF]

Figure 18 :

 18 Figure 18 : Annulated pyrimidine derivatives Among a panel of tetrahydropyridopyrimidines, MacGowan et al. identified 52a1 (35) a potent dual TLRs 7/8 agonist (least effective concentrations: 1.5 µM hTLR7, 3.4 µM hTLR8, Figure18), as demonstrated by the induction of interferon-γ induced protein (IP-10).[START_REF] Mcgowan | Design and synthesis of tetrahydropyridopyrimidine based Toll-Like Receptor (TLR) 7/8 dual agonists[END_REF] The pharmacokinetic profile of 52a1 in mice confirmed the low systemic exposure. In addition, 52a1 showed low plasma protein binding (30% bound in mice plasma), no inhibition of major CYP450 isozymes (CYP450 > 10 μM: 3A4, 2C8, 2C9, 2D6, 1A2, 2C19) and lacked off-target activity across a multi-receptors panel (> 10 μM against histamine, dopamine, and serotonin subtypes).

Figure 20 :

 20 Figure 20 : Imidazo[4,5-c]quinoline derivativesImiquimod (39, also known as R-837, Figure20) is a tricyclic nitrogen molecule belonging to the imidazo[4,5-c]quinoline series, developed 3M Pharmaceuticals laboratories (Saint Paul, MN, USA). It is commercialized as Aldara® or Zyclara®, respectively 5% and 3.75% imiquimod-containing creams. It was first described for its antiviral activity against certain viruses such as herpes simplex II (HSV-II)[START_REF] Harrison | Modification of immunological responses and clinical disease during topical R-837 treatment of genital HSV-2 infection[END_REF], sendai virus[START_REF] Megyeri | Stimulation of interferon and cytokine gene expression by imiquimod and stimulation by Sendai virus utilize similar signal transduction pathways[END_REF] and papilloma virus[START_REF] Hengge | Topical immunomodulators for the treatment of external genital warts, cutaneous warts and molluscum contagiosum[END_REF]. This molecule does not directly inhibit viral replication, it is an immunomodulator whose viral activity is related to the inducing power of IFNα and other cytokines. It was approved in 1997 by the Food and Drug Administration (FDA) for the treatment of external genital and perianal warts (condylomata acuminata). In 2004, its use was extended to the treatment of actinic keratosis and superficial basal cell carcinoma.[START_REF] Miller | Imiquimod applied topically: a novel immune response modifier and new class of drug[END_REF][START_REF] Tyring | Imiquimod applied topically: A novel immune response modifier[END_REF]

  studies. c.1.3. Other imidazo[4,5-c]quinoline derivatives In 2005, Gerster et al. of 3M pharmaceuticals laboratory published an article showing the different synthetic strategies of imidazo[4,5-

Figure 21 :

 21 Figure 21 : Imidazo[4,5-c]quinolone derivatives developed by 3M pharmaceuticals

Figure 22 :

 22 Figure 22 : Imidazoquinoline developed by Invivogen™

Figure 23 :

 23 Figure 23 : 3M compounds

Figure 24 :

 24 Figure 24 : Gardiquimod and derivatives In 2012, Shi et al. synthesized an imidazo[4,5-c]quinoline series by replacing hydrogen of the C7 carbon with a methoxycarbonyl group (Figure25).[START_REF] Shi | Discovery of Imidazoquinolines with Toll-Like Receptor 7/8 Independent Cytokine Induction[END_REF] They showed that the elimination of the C2 ethoxymethyl chain of resiquimod results in a significant decrease in the TLR7 agonist activity and a complete loss of the TLR8 agonist activity. The activity is restored and even improved, by the addition of a butyl chain in C2 position (EC50 TLR7 = 0.1±0.02 µM, EC50 TLR8 = 0.1± 0.003 µM). The unsubstituted in C2 position and carrying a methoxycarbonyl group in C7 position compound lacked TLRs 7/8 agonist activity, however, it was able to induce a high production of pro-inflammatory cytokines (TNFα, IL12, IL1β). This suggests the existence of an alternative mechanism of action by which imidazoquinolines can stimulate the production of cytokines. The addition of the alkyl (butyl) chain in C2 position restored TLR7 and TLR8 agonist activity. All this indicates that C7 substitution may be important for cytokine production, whereas TLRs 7/8 agonist activity requires C2 substitution.

Figure 25 :

 25 Figure 25 : Core-structure of the imidazo[4,5-c]quinoline series [248]

Figure 26 :

 26 Figure 26 : Core-structure of the imidazo[4,5-c]quinoline series [249]

Figure 27 :

 27 Figure 27 : Imidazo[4,5-c]quinoline derivatives substituted in N1 position

Figure 28 :

 28 Figure 28 : CL075 analogues

Figure 29 Yoo

 29 Figure 29

Figure 30 :

 30 Figure 30 : Quinoline and quinoxaline derivatives

Figure 31 :

 31 Figure 31 : A TLR7 agonist belonging to Novartis pipeline

Figure 33 :

 33 Figure 33 : Benzimidazole moiety containing agonist b. Bicycles analogs Recently, Zhang et al. identified, for the first time, human TLR8 small antagonist molecules with a novel inhibition mechanism.[304] SAR studies on the pyrazolo[1,5-a]pyrimidine series led to identification of the compound: CU-CPT8m[START_REF] Ugolini | Recognition of microbial viability via TLR8 drives T FH cell differentiation and vaccine responses[END_REF], which proved to be the most active with an IC50 of 67 nM and a strong ability to inhibit the production of pro-inflammatory cytokines (Figure34). Docking studies reveal that CU-CPT8m is sandwiched between the two monomers of TLR8 and is localized in a hydrophobic pocket at the proteinprotein interface of the two monomers TLR8. This new binding site is close, but distinct from the identified agonist site, whose occupation prevents the activation of TLR8. This implies a new inhibitory mechanism by CU-CPT8m.

Figure 34 :

 34 Figure 34 : CU-CPT derivatives

Figure 36 :

 36 Figure 36 : Quinazoline derivative

Figure 38 :

 38 Figure 38 : Quinoline and quinolinium derivatives

Figure 39 :

 39 Figure 39 : Dimer compound

Figure 40 :

 40 Figure 40 : Pyrazolo[1,5-a]quinoxaline derivatives

Figure 41 :

 41 Figure 41 : Artemisinin analogue

Figure 42 :

 42 Figure 42 : Antagonist compounds interacting with nucleic acids e.2.D helixPiao and al. found that 2R9 peptide (PQRFCKLRKIMNT sequence) derived from helix D of the TIR domain of TLR2 specifically targets TIRAP and blocks TLRs 2/4/7/9, but not TLR3 and TNF-α signaling. As confirmed by in vitro studies, cell imaging and co-immunoprecipitation, 2R9 binds TIRAP with nanomolar affinity as a result of a fast association rate coupled with slow dissociation of the complex. 2R9 binds albumin with micromolar affinity. 2R9 diminished systemic cytokine responses elicited in vivo by synthetic TLR2 and TLR7 agonists. It inhibited the activation of macrophages infected with influenza strain A/PR/8/34 (PR8) and significantly improved the survival of PR8-infected mice. Thus, 2R9 represents a TLR-targeting agent that blocks protein interactions downstream of activated TLRs.[START_REF] Piao | A Decoy Peptide that Disrupts TIRAP Recruitment to TLRs Is Protective in a Murine Model of Influenza[END_REF] 3. Macromoleculesa. Modified nucleic acidsIn 2006, Judge et al. have shown that the incorporation of 2'-O-methyl (2'OMe)-uridine or 2'OMeguanosine residues in siRNA or in ssRNA eliminates their immunostimulatory activity both in vitro and in vivo.[START_REF] Judge | Design of noninflammatory synthetic siRNA mediating potent gene silencing in vivo[END_REF] It has also been reported that chemical modifications of nucleotides in mammalian RNA, including 2'OMe modifications, prevents them from activating dendritic cells.[START_REF] Karikó | Suppression of RNA recognition by Toll-like receptors: the impact of nucleoside modification and the evolutionary origin of RNA[END_REF] In 2007, Robbins et al. tested the ability of 2'OMe RNA to antagonize TLR7 immune stimulation.[START_REF] Robbins | 2'-O-methyl-modified RNAs act as TLR7 antagonists[END_REF] They reported that 2'OMe RNA directly inhibits cytokines induction in murine and human systems by immunostimulatory RNA and by a TLR7 agonist Loxoribine. The inhibitory activity of 2'OMe RNA has also been shown in vivo. These results indicate that 2'OMe RNA acts as a TLR7 antagonist.

  on the inhibitory oligodeoxynucleotide 2088 (INH-ODN 2088), INH-ODN-24888 was designed (T*C*C*T*G*G*C*mE*G*G*G*A*A*G*T sequence, * = phosphorothioate-binding) as a guanine-modified inhibitory oligonucleotide TLRs 3/7/9

  *C*T*G*G*C*G*G*G*G*A*A*G*T 3, 7, 9 21595 T*C*C*T*G*G*C*E*G*G*G*A*A*G*T 7, 9 20844 T*C*C*T*G*G*C*G*E*G*G*A*A*G*T 7, 9 24888 T*C*C*T*G*G*C*mE*G*G*G*A*A*G*T 3, 7, 9 24991 C*C*T*G*G*C*mE*G*G*G 3, 7, 9 105870 T*A*A*T*G*G*C*E*G*G*G*A*A*G*T 7 105871 T*A*A*T*G*G*C*mE*G*G*G*A*A*G*T 3, 7 * = phosphorothioate-binding, E = 7-deaza-2'-deoxyguanosine, mE = 7-deaza-2'-O-methyl-guanosine

Table 1 : Potent agonists

 1

	Ligands	Chemical family	Agonist	Biological evaluations	References
	S28690	Imidazoquinoline derivative	TLR7	Immunogeneicity effect on chronic lymphocytic leukemia	[263-266]
	SB 9922	Nucleic acid hybrid	TLR7	Potent vaccine adjuvant used in Mycobacterium tuberculosis combination with BCG vaccine against	[267,268]
	RO6870868
	and RO6864018 (ANA773 or	Tosylate prodrug of isatoribine	TLR7	Treatment for hepatitis C and cancers	[269-271]
	RG7795)

Table 3 : Clinical trials involving TLRs 7/8 agonists and antagonists

 3 : D for drug, A for adjuvant. b: C for completed, T for terminated, O for ongoing, S for suspended.

	Ligands
	(synonyms or explanations given	In combination with	TLRs	Investigators	Pathologies Applications	Type a	Phase	Clinical Trial Identification	Status b
	once)
	Agonists
	852A PF-04878691 S-32865 3M852A		7	3M Pharmaceuticals	Inoperable melanoma (Unresectable metastatic cutaneous melanoma)	D	II	EudraCT2004-003910-41	C
	852A		7	Masonic Cancer Center University of Minnesota, Pfizer	Breast, ovarian, endometrial, and cervical cancer	D	II	NCT00319748	C
					Hepatitis C Virus (Multiple-dose
					escalation study, safety,
	852A		7	Pfizer	tolerability, pharmacokinetics and pharmacodynamics	D	I	NCT00810758	C
					evaluation in healthy
					volunteers)
	852A		7	Pfizer	Inoperable metastatic Melanoma	D	II	NCT00189332	C
	Imiquimod R837	radiotherapy	7	University of Oklahoma, NCI	Melanoma and metastatic cancer	D	I	NCT00453050	O
				NYU Langone
	Imiquimod	cyclophosphamide and radiotherapy	7	Health, New York University School of	Metastatic and recurrent breast cancer	A	I and II	NCT01421017	C
				Medicine, NCI
				New York University
	Imiquimod		7	School of Medicine, NYU	Breast cancer Breast neoplasms	D	II	NCT00899574	C
				Langone Health
	Imiquimod		7	The University of Hong Kong	Influenza viral infection	A	III	NCT02103023	C
	Imiquimod		7	Medical University of Vienna	Human Papillomavirus	D	II	NCT00941811	C

a

Acknowledgements

Authors would thank Laurianne Adin for helping to find additional detailed data during the preparation of this manuscript.

Agonists