Aicha Talha

Cécile Favreau

Maxence Bourgoin

Guillaume Robert

Patrick Auberger

Lahcen El Ammari

Mohamed Saadi

Rachid Benhida

Anthony R Martin
email: anthony.martin@umontpellier.fr

Khalid Bougrin
email: kbougrin@yahoo.fr

Lahcen El

Ultrasound-assisted one-pot three-component synthesis of new isoxazolines bearing sulfonamides and their evaluation against hematological malignancies

Keywords: Sulfonamides-Isoxazolines, One-pot synthesis, Radical processes, Ultrasound, Hematological Malignancies

à la 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.

Introduction

Nitrogen-based heterocyclic chemistry is an important and unique class of organic molecules with immense utility in medicinal chemistry [1,[START_REF] Draye | Ultrasound for Drug Synthesis: A Green Approach[END_REF]. A major goal of current medicinal research is to identify new, safer and more effective treatments for a wider range of neoplastic conditions, with the hope of reducing morbidity and mortality. This led to the development of targeted therapies for cancer [3], including differentiation therapies [4]. Nevertheless, the quest for new anti-cancer drugs to combat leukemic cells is still an ongoing field of research [START_REF] Coombs | Acute promyelocytic leukemia: where did we start, where are we now, and the future[END_REF]6]. In line with this consideration, and in continuation of our recent research in identifying new and original routes to bioactive anticancer agents [7], we synthesized a series of new isoxazolinelinked sulfonamide structures and evaluated their biological activity against CML and APL.

Isoxazoline derivatives are a very important class of N,O-containing heterocycles that received significant attention in modern organic chemistry. They are widely present in different biologically active molecules such as anticancer [8], antimicrobial [START_REF] Kotian | Potassium iodate (KIO3) as a novel reagent for the synthesis of isoxazolines: evaluation of antimicrobial activity of the products[END_REF], antibacterial [START_REF] Kudryavtseva | Synthesis and Antibacterial Activity of New Acridone Derivatives Containing an Isoxazoline Fragment[END_REF], antiinflammatory and analgesics [11]. Additionally, sulfonamides are recurrent motifs in therapeutic agents exhibiting a broad spectrum of biological activities, such as antibacterial [START_REF] Özbek | Synthesis, characterization and antimicrobial activity of new aliphatic sulfonamide[END_REF], antitumor [13], diuretic [START_REF] Husain | Synthesis and in vivo diuretic activity of some new benzothiazole sulfonamides containing quinoxaline ring system[END_REF] and hypoglycemic [START_REF] Ahmadi | Synthesis and Evaluation of the Hypoglycemic and Hypolipidemic Activity of Sulfonamide-benzothiazole Derivatives of Benzylidene-2, 4-thiazolidnedione[END_REF] (Figure 1). Isoxazolines are frequently synthesized by 1,3-dipolar cycloadditions between an alkene and a nitrile oxide, in the presence of oxidants such as CrO2 [START_REF] Roy | Enhanced rate of intramolecular nitrile oxide cycloaddition and rapid synthesis of isoxazoles and isoxazolines[END_REF], MnO2 [START_REF] Hashimoto | Synthesis of isoxazoles en route to semi-aromatized polyketides: dehydrogenation of benzonitrile oxide-para-quinone acetal cycloadducts[END_REF], MgO(Ac)2 [START_REF] Nishiwaki | Onestep synthesis of differently bis-functionalized isoxazoles by cycloaddition of carbamoylnitrile oxide with β-keto esters[END_REF],

Ce III /NaI/I2 [START_REF] Bartoli | Microwave-Assisted Cerium (III)-Promoted Cyclization of Propargyl Amides to Polysubstituted Oxazole DerivativesEur[END_REF], N-chlorosuccinimide [START_REF] Pérez | Synthesis of 3, 5-disubstituted isoxazoles and isoxazolines in deep eutectic solvents[END_REF] and hypervalent iodine reagents [START_REF] Zheng | Synthesis of oxazoles from enamides via phenyliodine diacetate-mediated intramolecular oxidative cyclization[END_REF]. However, all these methods involve the use of either powerful oxidants, toxic solvents or drastic reaction conditions. Thus, we developed a new synthetic method for isooxazoline preparation using an inexpensive, ecological and water-soluble oxidant, namely the trichloroisocyanuric acid (TCCA) [26]. It is coupled with ultrasounds as an efficient activation method as recently disclosed for the synthesis of this type of heterocycle [START_REF] Pacheco | Facile sonochemical synthesis of novel pyrazolyne derivates at ambient conditions[END_REF]. TCCA was first used as a disinfectant and industrial deodorant, also used in swimming pools. In organic chemistry, TCCA proved efficient in chlorination reactions under mild conditions, as well as oxidation of ethers, thioethers, aldehydes, acetals and alcohols [START_REF] Gaspa | Metal-free direct oxidation of aldehydes to esters using TCCA[END_REF]. TCCA was recently used in the heterocyclic synthesis of 2-substituted oxazolines, imidazolines, thiazolines and coumarin, at room temperature [29].

More recently Zhang et al., [START_REF] Zhang | Trichloroisocyanuric acid promoted cascade cyclization/trifluoromethylation of allylic oximes: Synthesis of trifluoromethylated isoxazolines[END_REF] and Bhatt et al., [START_REF] Bhatt | Trichloroisocyanuric acid-mediated synthesis of 1, 5-fused 1, 2, 4-triazoles from N-heteroaryl benzamidines via intramolecular oxidative N-N bond formation[END_REF] reported a novel method for the synthesis of isoxazoles/isoxazolines based on an intramolecular oxidative cyclization using a large excess of TCCA in MeCN or EtOH and long reaction time (ca. 10 h). In addition, Aghapour et al., [START_REF] Aghapour | Tandem and Selective Conversion of Tetrahydropyranyl and Silyl Ethers to Oximes Catalyzed with Trichloroisocyanuric Acid[END_REF] successfully used TCCA as a catalyst, in combination with hydroxylamine hydrochloride, for the preparation of oximes starting from tetrahydropyran (THP) ethers under solvent-free conditions at 110°C. To our knowledge, there are no reports in the literature on the preparation of isoxazolines in the presence of TCCA under ultrasonic irradiation in water. Inspired by the methods described above for the synthesis of isoxazolines and isoxazoles, we disclose herein the one-pot two-step synthesis of novel isoxazoline substituted sulfonamides (3a-r) promoted by TCCA at room temperature. It directly starts from aldehydes (1a-f) and operates in aqueous medium under ultrasonic irradiation (Scheme 1). The important features of this new method are its ease, straightforwardness and environmentally friendly protocol.

Scheme 1. One-pot synthesis of sulfonamides 4-substituted-isoxazolines catalyzed by TCCA in ethanol/water under sonication.

Experimental

Material and methods

All organic solvents were purchased from commercial sources and used as received or dried using standard procedures; all chemicals were purchased from Aldrich, Merck and used without further purification. Analytical thin layer-chromatographies (TLC) have been performed on pre-coated silica gel plates (Kieselgel 60 F254, E. Merck, Germany), and chromatograms were visualized by UV-light irradiation (254 and 360 nm), then by staining with ninhydrin or H2SO4/EtOH. Purifications by column chromatography have been performed using silica gel, 100-200 mesh (Merck, Germany). NMR spectroscopies were recorded in dry deuterated solvent (DMSO or chloroform) a Bruker AC 200, or on a Bruker AC 400 spectrometer at 200 MHz or 400 MHz for 1 H NMR and 50 MHz or 101 MHz for 13 C NMR; δ is expressed in ppm related to TMS (0 ppm) as internal standard. Splitting patterns are designated as follow: s (singlet), d (doublet), t (triplet), m (multiplet), br (broad). Coupling constants (J values) are listed in Hertz (Hz). Mass spectra (ESI-MS) were recorded on a Bruker Daltonics Esquire 3000+, and the samples were diluted in methanol. Melting points were measured on a Wagner and Munz Köfler Bench System. The purity of compounds was further verified by HPLC analysis was performed on a Jasco LC-Net II /ADC apparatus using phenomenex columns (conditions: unless otherwise stated: 1.0 mL/min, gradient 75% A / 25% B (1 min.) then increasing to 0% A / 100% B in 6 min. and a plateau of 3 min. before returning to 75% A / 25% B in 1 min. A is water and B is CH3CN, both containing 1‰ HCOOH). Purity of all compounds was found to be ≥95% as determined by HPLC using UV detection at 254 nm. The ultrasound-assisted reactions were carried out in an "'Elmasonic S 30 / S 30 H UltraSonic Bath Cleaner", with an effective ultrasound power of 80 W at a frequency of 47 kHz. This ultrasonic cleaner has internal dimensions W / D / H (mm) of 240 / 137 / 100 with a liquid retention capacity of 2.75 L. The reactions were carried out in a flask with a capacity of 25 mL in suspension in the center of the cleaning bath, 5 cm below the surface of the liquid. The reaction temperatures were controlled by adding or removing water from the ultrasonic bath.

Synthesis

General procedure for the synthesis of allyl-sulfonamides (2a-l):

The primary amine (1 equiv) and K2CO3 (2 equiv) were dissolved in EtOH/H2O (v/v, 1:1) (20 ml) and alkyl/phenyl sulfonyl chloride (1.1 equiv) was added dropwise. After completion of the reaction (as monitored via TLC), allyl bromide (1.1 equiv) was added dropwise with constant stirring or sonication until complete consumption of the starting material (6-10 h under stirring or 15-20 min under sonication). The salts were then removed by filtration, and filtrate was concentrated under reduced pressure. The crude material was purified on silica gel column chromatography (cyclohexane/ethyl acetate: 9/1 to 6/4), to afford the desired pure products 2al.

N-Allyl-N-(4-bromo-phenyl)-benzenesulfonamide (2a):

Brown solid; MP 63-65 °C, TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.67; 1 H NMR (300 MHz, DMSO-d6) δ 7.73 -7.64 (m, 1H), 7.58 (d,J = 4.3 Hz,4H),7.50 (d,J = 8.8 Hz,2H), 7.00 (d,J = 8.8 Hz,2H),1H),2H),4.19 (dd,J = 6.0,1.2 Hz,2H). 13 C NMR (75 MHz, DMSO-d6) δ 133.8, 133.1, 132.3 (2C), 130.8 (2C), 129.8 (2C), 127.7 (2C), 119.6, 52.9; MS (EI): m/z = 351.99 [M + H] + .

N-Allyl-N-phenyl-methanesulfonamide (2b):

Orange solid; MP 67-69°C. TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.58; 1 H NMR (400 MHz, Chloroform-d) δ 7. 45 -7.31 (m, 5H), 5.96 -5.72 (m, 1H), 5.24 -5.12 (m, 2H), 4.34 -4.27 (m, 2H), 2.94 (s, 3H). 13 C NMR (101 MHz,132.8,129.4 (2C),128.5 (2C),128.1,119.1,53.7,[START_REF] Gaspa | Trichloroisocyanuric Acid: a versatile and efficient chlorinating and oxidizing reagent[END_REF] 3H),2H),1H),2H),4.21 (d,J = 6.3 Hz,2H). 13 C NMR (101 MHz, Chloroform-d) δ 138. 9, 138.3, 132.7, 132.7, 128.9 (2C), 128.8 (4C), 127.9, 127.6 (2C), 118.9, 53.6 (d,J = 9.0 Hz,2H),1H),2H), (m, 2H), 3.82 (s, 3H), 2.92 (s, 3H). 13 C NMR (101 MHz, Chloroform-d) δ 159. 2, 133.1, 131.7, 130.1(2C), 119.1, 114.5 (2C), 55.4, 53.9, 37.9; MS (EI): m/z = 242.08 [M + H] + . 5.90 -5.72 (m, 1H), 5.23 -5.01 (m, 2H), 4.31 -4.18 (m, 2H), 3.80 (s, 3H), 3.04 (q, J = 7.4 Hz, 2H), 1.39 (t, J = 7.4 Hz, 3H). 13 C NMR (101 MHz, Chloroform-d) δ 159. 1, 133.5, 131.6, 130.3 (2C), 118.7, 114.5 (2C), 55.4, 54.4, 45.4, 8 83 -7.79 (m, 2H), 7.57 -7.54 (m, 1H), 7.53 -7.49 (m, 2H), 5.67 -5.56 (m, 1H), 5.21 -5.07 (m, 2H), 3.81 (d, J = 6.4 Hz, 2H), 3.09 (dd, J = 8.4, 6.8 Hz, 2H), 1.53 (dd, J = 15.0, 7.5 Hz, 2H), 0.85 (t, J = 7.4 Hz, 3H). 13 C NMR (101 MHz,133.1,132.4,129.1 (2C),127.1 (2C),118.6,50.5,49.1,21.4,11.1;MS (EI) 2.2.1.7. N-allyl-N-(4-methoxyphenyl) benzenesulfonamide (2g): Gray Solid, MP 91-94°C, TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.54; 1 H NMR (200 MHz,5H),6.92 (d,J = 9.1 Hz,2H),6.78 (d,J = 9.1 Hz,2H),1H),2H), 4.14 (d,J = 6.3 Hz,2H),3.78 (s,3H). 13 C NMR (50 MHz, Chloroform-d) δ 158. 9, 138.5, 132.8, 132.5, 131.4, 130.2 (2C), 128.7 (2C), 127.6 (2C), 118.7, 114. (2C), 55.4, 53.9; MS (EI): -benzenesulfonamide (2h): Green Solid, MP 65-67°C, TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.6; 1 H NMR (400 MHz, Chloroform-d) δ 7. 3H),2H),7.28 (d,J = 8.7 Hz,2H),6.98 (d,J = 8.7 Hz,2H),1H),2H),4.18 (d,J = 6.3 Hz,2H). 13 C NMR (101 MHz, Chloroform-d) δ 137. 9, 137.4, 133.7, 132.9, 132.3, 130.1 (2C), 129.1 (2C), 128.9 (2C), 127.6 (2C), 119.3, 53.5 (d,J = 8.8 Hz,2H),1H),2H),4.29 (d,J = 6.4 Hz, 2H), 3.05 (q, J = 7.4 Hz, 2H), 1.38 (t, J = 7.4 Hz, 3H). 13 C NMR (101 MHz,133.5,132.9,129.9 (2C),129.4 (2C),119.2,54.1,45.8,8.1

N-Allyl-N-(4-methoxy

: m/z = 240.1 [M + H] + .
m/z = 304,09 [M + H] + .
; MS (EI): m/z = 260.04 [M + H] + .

N-Allyl-N-p-tolyl-ethanesulfonamide (2k):

Beige Solid, MP 66-69°C, TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.52; 1 H NMR (400 MHz, Chloroform-d) δ 7.25 (d, J = 8.4 Hz, 2H), 7.19 (d,J = 8.3 Hz,2H),1H),2H),4.30 (d,J = 6.3 Hz, 2H), 3.06 (q, J = 7.4 Hz, 2H), 2.37 (s, 3H), 1.40 (t, J = 7.4 Hz, 3H). 13 C NMR (101 MHz, Chloroform-d) δ 137.8, 136.6, 133.5, 130.8 (2C), 130.2 (2C), 118.6, 49.9, 45.6, 21.2, 7; MS (EI): m/z = 240.10 [M + H] + . (d,J = 8.3 Hz,2H),1H),2H),4.30 (d,J = 6.3 Hz,2H),2.37 (s,3H). 13 C NMR (101 MHz, Chloroform-d) δ 140.7, 133.48, 131.1, 130.8 (2C), 130.2 (2C), 118.6, 49.9, 45.6

N-Allyl-N-(4-chloro-phenyl)-methanesulfonamide (2l)

: Green Solid, MP 97-99°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.38; 1 H NMR (400 MHz, Chloroform-d) δ 7.25 (d, J = 8.3 Hz, 2H), 7.19
; MS (EI): m/z = 246,03 [M + H] + .

General procedure for the one-pot synthesis of compounds (3a-r):

To a solution of aldehyde (1 equiv) and hydroxylamine hydrochloride (1.5 equiv) in EtOH/H2O (v/v, 1:1) (20 mL) TCCA (0.5 equiv) and allyl-sulfonamides 2a-l (1 equiv) were added successively and the mixture was sonicated according to the time reported in Table 3. Reactions were then extracted with CH2Cl2 (20 mL x 2). The organic phase was washed with water (20 mL) and saturated brine solution (20 mL), dried over MgSO4 and concentrated in vacuum to give the crude product, which was purified by flash silica gel chromatography (cyclohexane/ EtOAc: 9/1 to 7/3) to afford the pure adducts 3a-r. 4-Chloro-phenyl)-N-[3-(4-chloro-phenyl)-isoxazolin-5-ylmethyl] benzenesulfonamide (3a): White Solid, Mp 195-197°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.14; 1 H NMR (400 MHz, Chloroform-d) δ 7.62 (t, J = 7.4 Hz, 3H), 7.60 -7.55 (m, 2H), 7.50 (t, J = 7.7 Hz, 2H), 7.41 (d,J = 8.6 Hz,2H),2H), 7.00 (d,J = 8.7 Hz,2H),1H), 3.82 (dd, J = 13.9, 7.0 Hz, 1H), 3.72 (dd, J = 13.9, 4.9 Hz, 1H), 3.52 (dd, J = 16.9, 6.6 Hz, 1H), 3.40 (dd, J = 16.9, 10.5 Hz, 1H). 13 C NMR (101 MHz, Chloroform-d) δ 155.6, 137.8, 137.3, 136.3, 134.3, 133.2, 129.9 (2C), 129.4 (2C), 129.1 (4C), 128.1 (2C), 127.6 (2C), 127.6, 78.9, 53.2, 37.8; MS (EI): m/z = 460.40 [M + H] + ; HPLC analysis (luna column, λ = 254 nm, purity 100 %, t R = 4.55min). 6, 154.8, 134.4, 133.2, 131.5, 130.8, 130.1 (2C), 129.2, 128.5, 125.8, 114.9 (2C), 79.1, 55.5, 53.6, 37.7, 37.5; MS (EI): m/z = 429.04 [M + H] + ; HPLC analysis (luna column, λ = 254 nm, purity 100 %, t R = 4.15min)

2.2.2.1.N-(

2.2.2.2.N-[3-(3,4-Dichloro-phenyl)-isoxazolin-5-ylmethyl]-N-(4-methoxy-phenyl

2.2.2.3.N-(4-Methoxy-phenyl)-N-[3-(3-nitro-phenyl)-isoxazolin-5-ylmethyl]-benzene

sulfonamide (3c): White Solid, MP 145-147°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.5; 1 H NMR (300 MHz, DMSO-d6) δ 8.37 -8.26 (m, 2H), 8.09 -8.02 (m, 1H), 7.78 -7.64 (m, 2H), 7.56 (d,J = 4.4 Hz,4H),4H),1H),3.84 (dd,J = 14.2,6.8 Hz,1H),3.67 (dd,J = 15.9,11.0 Hz,1H),3.54 (dd,J = 20.5,9.8 Hz,1H),3.37 (dd,J = 12.8,5.6 Hz, 1H). 13 C NMR (75 MHz, DMSO-d6) δ 159. 2, 156.1, 148.5, 138.2, 133.6, 133.2, 131.8, 131.3, 131.1, 130.7(2C), 129.7 (2C), 127.8 (2C), 125.1, 121.3, 114.6 (2C), 79.8, 55.7, 54.1, 37.5; MS (EI): m/z = 467.12 [M + H] + ; IR (KBr, cm -1 ): 1134 (S=O),1162 (S=O), 732-646 (5HAr-Adjacent), 1506 (N=O), 1420 (Ph-O-C), 3312-2700 (C=CAr); HPLC analysis (luna column, λ = 254 nm, purity 95 %, t R = 4.11min).

2.2.2.4.N-(4-Bromo-phenyl)-N-[3-(3-nitro-phenyl)-isoxazolin-5-ylmethyl]-benzene

sulfonamide (3d): White Solid, MP 167-169°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.57; 1 H NMR (300 MHz, DMSO-d6) δ 8. 36 -8.26 (m, 2H), 8.06 (d, J = 7.9 Hz, 1H), 7.79 -7.67 (m, 2H), 7.58 (d, J = 1.5 Hz, 2H), 7.56 (s, 2H), 7.53 (d, J = 8.7 Hz, 2H), 7.00 (d, J = 8.7 Hz, 2H), 4.86 -4.68 (m, 1H), 3.88 (dd, J = 14.5, 6.7 Hz, 1H), 3.76 (dd, J = 14.5, 4.5 Hz, 1H), 3.58 (dd, J = 17.3, 10.8 Hz, 1H), 3.34 (dd, J = 13.6, 10.5 Hz, 1H). 13 C NMR (75 MHz, DMSO-d6) δ 156.1, 148. 5, 138.9, 137.6, 133.9, 133.2, 132.4 (2C), 131.3, 131.2 (2C), 131.1, 129.8 (2C), 127.7 (2C), 125.1, 121.6, 121.3, 80.1, 53.5, 37.4; MS (ESI +): m/z = 516.01 [M + H] + ; HPLC analysis (luna column, λ = 254 nm, purity 95 %, t R = 4.33min).

N-(4-Methoxy-phenyl)-N-(3-phenyl-isoxazolin-5-ylmethyl)-benzenesulfonamide (3e):

White Solid, MP 127-129°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.52; 1 H NMR (300 MHz, DMSO-d6) δ 7.68 (m, 1H), 7.57 (d, J = 4.4 Hz, 4H), 6.91 (d, J = 9.1 Hz, 2H), 6.83 (d, J = 9.1 Hz, 2H), 5.66 (ddt, J = 17.2, 10.2, 6.0 Hz, 1H), 5.14 -4.94 (m, 2H), 4.14 (d, J = 6.1 Hz, 2H), 3.71 (s, 3H). 13 C NMR (75 MHz, DMSO-d6) δ 158. 9, 138.3, 133.5, 133.5, 131.5, 130.2 (2C),129.7 (2C), 127.7 (2C), 119.1, 114.4 (2C), 55.7, 53.5; MS (EI): m/z = 422.13 [M + H] + ; HPLC analysis (luna column, λ = 254 nm, purity 96 %, t R = 4.34 min). = 8.5 Hz, 2H), 4.98 -4.84 (m, 1H), 3.85 (dd, J = 13.9, 6.8 Hz, 1H), 3.76 (dd, J = 13.9, 4.9 Hz, 1H), 3.59 (dd, J = 16.9, 6.7 Hz, 1H), 3.48 (dd, J = 16.9, 10.6 Hz, 1H). 13 C NMR (101 MHz, Chloroform-d) δ 154. 9, 148.5, 137.8, 137.3, 134.4, 133.3, 132.3, 130.9, 129.9 (2C), 129.9, 129.5 (2C), 129.1 (2C), 127.6 (2C), 124.7, 121.6, 79.5, 53.2, 37.5; MS (EI): m/z = 471.91 [M + H] + ; HPLC analysis (luna column, λ = 254 nm, purity 100 %, t R = 4.30 min).

2.2.2.6.N-(4-Chloro-phenyl)-N-[3-(3-nitro-phenyl)-isoxazolin-5-ylmethyl

N-(4-Chloro-phenyl)-N-(3-phenyl-isoxazolin-5-ylmethyl)-benzenesulfonamide (3g):

White Solid, Mp 148-150°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.34; 1 H NMR (400 MHz, Chloroform-d) δ 7.70 (dd, J = 6.5, 3.1 Hz, 2H), 7.64 -7.58 (m, 3H), 7.51 -7.41 (m, 5H), 6.97 (d, J = 9.0 Hz, 2H), 6.84 (d,J = 9.0 Hz,2H),1H),3.76 (dd,J = 20.5,7.4 Hz, 1H), 3.69 (dd, J = 13.5, 4.9 Hz, 1H), 3.55 (dd, J = 16.9, 6.5 Hz, 1H), 3.42 (dd, J = 16.9, 10.4 Hz, 1H). 13 C NMR (101 MHz, Chloroform-d) δ 159.3, 156.5, 137.8, 132.9, 131.6, 130.2, 129.9 (2C)

N-[3-(3-Nitro-phenyl)-isoxazolin-5-ylmethyl]-N-p-tolyl-methanesulfonamide (3q):

White Solid, Mp 177-179°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.

N-((3-(3-Nitro-phenyl)-isoxazolin-5-yl)methyl)-N-propyl-benzenesulfonamide (3r):

White Solid, Mp 102-104°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0. Hz, 1H), 3.13 (dd, J = 14.8, 7.1 Hz, 1H), 1.61 (q, J = 7.4 Hz, 3H), 0.88 (t, J = 7.4 Hz, 3H). 13 C NMR (101 MHz,148.5,139.2,132.8,132.3,131.1,129.8,129.2 (2C),127.1 (2C),124.6,121.6,80.9,51.9,50.9,37.6,21.8,11.1;MS (EI): m/z = 403.12 [M + H] + ; IR (KBr, cm -1 ): 1336 (S=O), 1188 (S=O), 1046 (Propyl),1500 (N=O), 3312-2700 (C=CAr) and

(C-HAr) elongation 818-670 (3HAr-Adjacent); HPLC analysis (luna column, λ = 254 nm, purity 99 %, t R = 4.12min).

General procedure for the radical trapping experiment

To a solution of p-chlorobenzaldehyde 1a (1.0 equiv, 70 mg) and hydroxylamine hydrochloride

(1.5 equiv, 52. 1 mg) in EtOH/H2O (v/v, 1:1) (10 mL) TCCA (0.5 equiv, 58.1 mg), TEMPO (0.5 equiv, 78.1 mg) and alkene 2h (1 equiv, 153.5 mg) were added successively and the mixture was sonicated for 20 min using ultrasonic bath (47 kHz) at 25°C (the temperature of the reaction has been maintained by the addition of crushed ice). After completion of the reaction (TLC), the reaction mixture was concentrated under pressure to remove ethanol and then extracted with CH2Cl2 (20 mL). The organic phase was washed with water (20 mL) and saturated brine solution (20 mL), dried over MgSO4 and concentrated in vacuum. The resulting residue was then purified by silica gel column chromatography (cyclohexane/ EtOAc: 9/1 to 7/3) to afford the pure 3a (≈ 125 mg), with inseparable mixture of trapped-TEMPO products.

Crystallography

One crystal suitable for X-ray diffraction analysis was coated with dry perfluoropolyether, mounted on glass fiber, and fixed in a cold nitrogen stream to the goniometer head. Data collection was performed on a Bruker D8 VENTURE Super DUO diffractometer, using graphite monochromatized Mo radiation (Mo Kα radiation, l = 0.71073 Å). The data were reduced (SAINT) and corrected for absorption effects by the multiscan method (SADABS).

The corresponding crystallographic data were deposited with the Cambridge Crystallographic Data Centre as supplementary publications. CCDC 2040890 3k. The data can be obtained free of charge via: https://www.ccdc.cam.ac.uk/structures/.

Results and discussion

Effect of sonication on one-pot, tandem synthesis of allyl sulfonamides

Firstly, we synthesize different allyl-sulfonamides 2a-l directly from phenyl/alkyl sulfonyl chloride and a primary amine in presence of potassium carbonate, and subsequently allyl bromide is added; this process was performed in a one-pot under ultrasound activation [START_REF] Aghapour | Tandem and Selective Conversion of Tetrahydropyranyl and Silyl Ethers to Oximes Catalyzed with Trichloroisocyanuric Acid[END_REF].

The results presented in Table 1 clearly show the effect of ultrasound on the acceleration of the reaction rate (15 -27 min) compared with conventional agitation (6 -13 h). Of note, both methods afforded the expected allyl-sulfonamides (2 a-l) in comparable yields.

Effect of sonication and different oxidant on one-pot synthesis of sulfonamides 4substituted-isoxazolines

Next, we examined the reactivity of p-chlorobenzaldehyde 1a (1.0 equiv1) with hydroxylamine hydrochloride (1.5 equiv), and alkene 2h (1.0 equiv) as the benchmark reaction for the 1,3dipolar cycloaddition under oxidative condition leading to sulfonamides 4-substitutedisoxazolines (Table 2). For this purpose, different oxidizing agents were screened, with or without base, under conventional stirring or sonication, at room temperature (25°C).

Table 2. Optimization of the reaction conditions for the 'one-pot' synthesis of sulfonamideisoxazolines under sonication. a aryl aldehyde/ HONH 2 .HCl (1/1.5 mmol), in EtOH/H 2 O (v/v, 1:1) at 25°C.

b Pure isolated yield under stirring and sonication (ultrasonic bath, 47 kHz).

c Reaction conditions with 0.5 equiv of TCCA.

d Reaction performed with 0.3 equiv of TCCA.

First, we proceeded to the optimization of the one-pot two steps conversion of the aldehyde 1a

to the corresponding nitrile oxide, using different oxidants (1.0 equiv.) with or without base, and its cycloaddition with the model N-allyl compound 2h (Table 2, entries 2-7). We first confirmed that the presence of an oxidant is mandatory (entry 1). Therefore, we subsequently assessed different oxidants (N-chlorosuccinimide, chloranil and chloramine-T), in the presence/absence of a base. It appeared that the reaction performs poorly when the base is omitted. Low yields of isolated product 3a were obtained under such conditions (10-36%, Table 2, entries 2, 4, 6). However, once the base is added the reactions proceed more smoothly and the isolated yields were improved. For instance, a 40% yield increase was observed with p-Chloranil in the presence of K2CO3 (Table 2, entries 4 and 5). Similarly, a 50% yield increase was observed for N-chlorosuccinimide in the presence of Et3N (Table 2, entries 2 and 3). Of note, the combination of ceric ammonium nitrate (CAN) as the oxidant, and K2CO3 [START_REF] Aghapour | Tandem and Selective Conversion of Tetrahydropyranyl and Silyl Ethers to Oximes Catalyzed with Trichloroisocyanuric Acid[END_REF] afforded the expected product 3a in good yield (75%, entry 8).

We then pursued our investigation using TCCA as the oxidant. While the other oxidants required presence of a base, TCCA could be used alone, without affecting the isolated yields.

In fact, the desired product 3a was obtained in excellent yield, up to 87%, and short reaction time (Table 2, entries 9 and 10). It is noteworthy that the ultrasound assistance speeded up the reaction by 20-fold (12 min vs 4 h) compared with conventional stirring.

Afterwards, we focused on reducing the quantity of TCCA, decreasing from 1.0 equiv to 0.5 equiv promoted the reaction similarly in terms of yield and reaction time (Table 2, entry 11; 89 %). Although, a further decrease in the amount of TCCA (from 0.5 to 0.3 equiv.) induced a reduction in yield (Table 2, entry 12; 65%). After optimizing the experimental conditions, we explored the scope of this new one-pot reaction to synthesize a series of isoxazolines (Table 3). All experiments were performed in short overall reaction time (12-22 min) and isolated yields ranged between 52 and 86% (Table 3). A variety of aldehyde substituents were explored first, by using either electron-donating groups such as methoxy and methyl or electron-withdrawing groups such as chlorine, bromine and nitro. All these substitutions were well tolerated. The corresponding cycloadducts were obtained in good yields (Table 3, entries 1-4, 11-14 and17- 18). In addition, the substrates with an electron donor or acceptor group all reacted well to generate the desired isoxazolines (3a-r) with good to high yields.

High efficiency synthesis of sulfonamides 4-substituted-isoxazolines 3a

The isoxazolines (3a-r), were fully characterized by IR, 1 H, 13 The regioselectivity of the cycloaddition was further ascertained by X-ray diffraction of single crystal of 3k. The single crystals were obtained by slow evaporation from a saturated ethanolic solution (Figure 3). The crystallographic data (see ESI) relative to the structure of 3k has been deposited with the Cambridge Crystallographic Data Centre [START_REF]Crystallographic data for 3k, has been deposited at the Cambridge Crystallographic Data Centre with the deposition numbers[END_REF].

The study of acceleration mechanism under sonication

In this work, the remarkable acceleration and yield improvement observed under US can be explained by the simultaneous coexistence of chemical and mechanical effects of ultrasound [35]; the chemical effects generating free radicals with a high propensity to react due to the implosion of microbubbles and the mechanical effects that enhance heterogeneous solid-liquid or liquid-liquid reactions due to microjets formed during cavitation [START_REF] Hagenson | Comparison of the effects of ultrasound and mechanical agitation on a reacting solid-liquid system[END_REF]. In fact, cavitation accelerates much more easily mass transport between the two phases such as alkene and aldehyde that are poorly soluble at the beginning of the reaction. The 1,3-dipolar cycloaddition reaction can take place in both concerted or radical mechanism [37]. Depending on the reaction conditions used, TCCA can release either an electrophilic chlorine atom (Cl + ) or a chlorine radical (Cl . ) selectively promoting two plausible concerted and /or radical mechanistic approaches [START_REF] Gaspa | Trichloroisocyanuric Acid: a versatile and efficient chlorinating and oxidizing reagent[END_REF].

The use of direct-ultrasonic irradiation, the radical or radical-ion intermediates are generated more possibly in water [39]. For the present hypothesis, a series of control experiments were carried out (Scheme 2). The reaction yields have dramatically changed under the investigated conditions. When a radical scavenger, TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy), was added under sonication, a significant decrease in yield of 3a (≈ 55 %) was observed along with the formation of other non-identified products trapped by TEMPO (Scheme 2, a), which suggests that the reaction under ultrasonication evolves according the two mechanisms, i.e., radical and ionic. This observation was supported with other experiments under O2 or air as the radical initiator, without TEMPO under suitable conditions. The reaction gave the isoxazoline product 3a with excellent yields, 90% and 89% respectively (Scheme 2, b and c). Based on the above-mentioned experimental observations, a plausible mechanism has been proposed as shown in scheme 3. Firstly, the chlorine radical atom (Cl . ) and dichloroisocyanuric acid radical were generated by the homolytic cleavage of N-Cl bond in TCCA assisted by ultrasound [START_REF] Gaspa | Metal-Free Preparation of α-H-Chlorinated Alkylaromatic Hydrocarbons by Sunlight[END_REF]. The amidyl radical N-centered will then react as a proton acceptor during the first step of reaction with hydroxylamine hydrochloride to provide the aldoxime (not isolated) [START_REF] Aghapour | Tandem and Selective Conversion of Tetrahydropyranyl and Silyl Ethers to Oximes Catalyzed with Trichloroisocyanuric Acid[END_REF], which is then converted to hydroximoyl chloride with TCCA [START_REF] Rodrigues | A simple and efficient method for the synthesis of nitrile oxide from aldoxime using trichloroisocyanuric acid[END_REF]42]. The latter was subsequently oxidized to unstable bi-radical nitrile oxide (II) [43] followed by a rapid 1,3cycloaddition reaction with allyl-sulfonamide 2, to deliver the corresponding isoxazoline product (Path A, Scheme 3).

Following similar approach, an electrophilic chlorine atom (Cl + ) and cyanurate anion were generated from TCCA by mechanical effect of ultrasound that promotes an ionic process through the generation of nitrile oxide (III) which reacts with the alkene to give the corresponding isoxazoline by 1,3-cycloaddition reaction (Path B, scheme 3).

Taking into account all these considerations, we suggested that the mechanistic outcome of this heterogeneous reaction might be explained by a switch from a radical mechanism (Path A, scheme 3) to a concerted mechanism (Path B, scheme 3) upon sonication [44]. In both pathways the reaction affords only one regiosiomer, the 4-substituted isooxazoline. This high regioselectivity can be explained by considering the steric interactions that prevent the formation of C5-substituted isoxazolines in favor of the exclusive formation of C4-substituted isoxazolines [START_REF] Freeman | 4-Isoxazolines (2, 3-dihydroisoxazoles)[END_REF]. In fact, the acceleration observed in this reaction is maybe due to the cooperative effect of ultrasound activation and the TCCA catalytic capacity.

Scheme 3. Plausible mechanism of the isoxazoline formation under heterogeneous ultrasound conditions

Biology

All these newly synthesized isooxazoline 3a-r, were evaluated for their anti-cancer activity against K562 cell line (CML) at 1 µM 10 µM and 50 µM (48 h) using XTT and DAPI assays.

This first screening against K562 cells revealed a couple of compounds (3h and 3l) decreases the cell viability between 10 and 50 µM. Thus, we decided to investigate further the biological To confirm that 3h induced a caspase dependent apoptotic cell death, we performed an AV/DAPI labeling in the presence of a pan-caspsae inhibitor (QvD) on HL60 cell line at 24 and with QvD completely restored it. In fact, the co-treatment induced a cell viability comparable with control experiment (untreated cells). This experiment shows us in a formal way that the death induced by compound 3h is a caspases-dependent process at 24 and 48 h, thus confirming the first results obtained by western blot (Figure 4).

Conclusion

In summary, we reported the synthesis, chemical characterization and anticancer activity of a novel functionalized 3,5-disubstituted sulfonamide-isoxazoline series. These molecules were synthesized using a new, versatile and efficient "two-step one-pot" methodology through a 1,3dipolar cycloaddition reaction. This environmentally friendly process was carried out efficiently in an aqueous medium. It is based on the use of inexpensive, environmentally friendly TCCA as an oxidant to generate nitrile oxides in-situ. In addition, the use of a cooperative effect of ultrasound activation and TCCA catalytic capacity allows significant reaction rate acceleration. Among this series of isoxazoline analogues one compound, 3h, impairs the cell viability of leukemia K562 and HL-60 cells. In addition, 3h behaved as an apoptotic cell death inducer. Altogether, these results clearly demonstrated the potential of this class of molecules for the optimization of apoptosis inducer owing to their ease of synthesis the herein report two-step one-pot strategy.

Figure 1 .

 1 Figure 1. Selected examples of therapeutic agents containing an isoxazoline or sulfonamide function [16-19].

  -phenyl)-ethanesulfonamide (2e): Black solid, MP 70-72°C, TLC (cyclohexane/AcOEt, 6/4, v/v) Rf = 0.40; 1 H NMR (400 MHz, Chloroform-d) δ 7.26 (d, J = 9.0 Hz, 2H), 6.89 (d, J = 9.0 Hz, 2H),

  )-methane sulfonamide (3b): White Solid, MP 170-172°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.52 ; 1 H NMR (200 MHz, Chloroform-d) δ 7.70 -7.64 (m, 1H), 7.42 (d, J = 1.4 Hz, 2H), 7.27 -7.17 (m, 3H), 6.86 (d, J = 9.0 Hz, 2H), 4.87 -4.69 (m, 1H), 3.84 (dd, J = 14.1, 6.0 Hz, 1H), 3.75 (s, 3H), 3.72 (dd, J = 7.3, 2.1 Hz, 1H), 3.64 (dd, J = 9.5, 4.6 Hz, 1H), 3.26 ((dd, J = 14.1, 6.0 Hz, 1H), 2.89 (s, 3H). 13 C NMR (50 MHz, Chloroform-d) δ 159.

  ]-benzene sulfonamide (3f): White Solid, Mp 152-154°C, TLC (cyclohexane/AcOEt, 8/2, v/v) Rf = 0.29; 1 H NMR (400 MHz, Chloroform-d) δ 8.49 (s, 1H), 8.30 (d, J = 8.0 Hz, 1H), 8.06 (d, J = 7.7 Hz, 1H), 7.67 -7.56 (m, 4H), 7.51 (t, J = 7.6 Hz, 2H), 7.32 (d, J = 8.5 Hz, 2H), 7.01 (d, J

  ) Time (h) Yield (%) b Time (min) Yield (%)

  C NMR, 2D NMR, MS and their HPLC traces were acquired. For example, the 1 H NMR spectrum of N-[3-(3,4-Dichlorophenyl)-isoxazolin-5-ylmethyl]-N-propyl-benzenesulfonamide 3k showed a multiplet with a chemical shift δ= 5.06 -4.92 ppm for the H-isoxazolinic proton, two doublets appear at δ= 3.10 and 3.39 ppm characteristic of the two CH2-isoxazolinic protons, two other signals in the form of doublets at δ= 3.54 and 3.46 ppm correspond to the two protons of methylene, thus the presence of the signals between 4.92-7.85 ppm attributable to the different aromatic protons. The 13 C NMR spectrum showed characteristic signals at 11.13 ppm (CH2-CH2-CH 3 ), 21.85 ppm (CH2-CH 2 -CH3), 37.70 ppm (CH 2isoxazoline ), 51.96 ppm (CH 2 -CH2-CH3), 50.99 ppm (N-CH 2 -CH), 80.69 ppm (CH isoxazoline ) and 125.87-132.81, 127.13, 128.55, 129.24, 133.16 ppm attributable for aromatic carbons.

Figure 2

 2 Figure 2 Characteristic 1 H, 13 C NMR of 3k.

Figure 3 .

 3 Figure 3. The ORTEP diagram of novel single crystal of product (3k).

Scheme 2 .

 2 Scheme 2. Radical scavenging and control experiments.

  Figure 4-A). This was performed in a DAPI assay by flow cytometry. The percentage of HL-60 DAPI positive cells (dead cells) increased from 18% in the untreated control population (and at the dose of 12.5 µM) to 31, 44 and 67% in cultures treated by 3h for 48 hours with respectively 25, 50 and 100 µM doses. The determined value of EC50 (i.e. the concentration leading to a cell viability rate of 50%) of the compound 3h on HL-60 cell was found at 62 ± 2 µM. In addition, to get insights into the mode of action of this new compound (3h), we had a look at cell death mechanism induced by 3h. HL-60 cells were treated for 48 h with 94 μM of 3h and PARP (poly ADP-ribose polymerase, marker of apoptosis) and caspase 3 (apoptosisrelated cysteine peptidase) cleavage were analyzed by Western blot (Figure 4-B, 3-C).Compound 3h display a strong induction of apoptosis when HL60 cells were treated at the EC50 concentration as suggested by cleavage of PARP and more evidently caspase 3.

Figure 4 .

 4 Figure 4. Upper-left panel (A) HL-60 cells were treated with increasing doses (12.5-100 µM) of 3h and cell viability was evaluated after 48 h with DAPI staining and flow cytometry analysis. Upper-right panel (B) HL-60 cells were treated with 94 μM of 3h (48 h), and expression of cleaved forms of PARP and caspase 3 were analyzed by Western blot. HSP60 was used as loading control. Lower panel (C) Quantification of the cleaved form of PARP and caspase 3, normalized against loading control HSP60.

Figure 5 .

 5 Figure 5. HL-60 cells were treated during 24 h or 48 h with 94 μM of 3h in presence or absence of QvD (Av = Annexin V).

  Conceptualization, Methodology, Writing, Cécile Favreau: biology studies Maxence Bourgoin: biology studies, Guillaume Robert: biology studies, Patrick Auberger: Supervision biology studies, Lahcen EL Ammari: Software, Validation, Mohamed Saadi: Software, Validation, Rachid Benhida: Supervision, review, Anthony R. Martin: Supervision, Writing -review & editing, Khalid Bougrin: Supervision, Project administration, Writing -review & editing.

propyl-benzenesulfonamide (2f): Brown oil

	.1; MS
	(EI): m/z = 256.09 [M + H] + .
	2.2.1.6. N-Allyl-N-, TLC (cyclohexane/AcOEt, 6/4,
	v/v) Rf = 0.40; 1 H NMR (400 MHz, Chloroform-d) δ 7.

Table 1 .

 1 One-pot synthesis of allyl-sulfonamides under sonication.

2a-l Entry a Allyl R 1 R 2 Conventional stirring Sonication Time (h) Yield (%) b Time (min) Yield (%) b

	1	2a	Ph	4-Br C6H4	11	70	27	75
	2	2b	Me	C6H5	9	60	18	66
	3	2c	Ph	C6H5	12	75	25	79
	4	2d	Me 4-MeO C6H4	8	56	16	65
	5	2e	Et 4-MeO C6H4	10	78	17	81
	6	2f	Ph	C3H7	6	66	19	69
	7	2g	Ph 4-MeO C6H4	10.5	75	21	83
	8	2h	Ph	4-Cl C6H4	14	55	26	69
	9	2i	Me	4-Me C6H4	9.5	60	15	69
	10	2j	Et	4-Cl C6H4	8.5	64	21	68
	11	2k	Et	4-Me C6H4	9	72	18	76
	12	2l	Me	4-Cl C6H4	13	77	22	86

a Phenyl/alkyl sulfonyl chloride (1.1 mmol), primary amine (1.0 mmol), K 2 CO 3 (2.0 mmol) in EtOH/H 2 O (20 ml, 1:1) at 25°C, then allyl bromide (1.1 mmol). b Isolated yield after purification.

Table 3 .

 3 Generalization of the one-pot two-steps procedure under sonication.

	Entry a Isoxazoline R 1	R 2	Ar	Time (min) Yield (%) b
	1	3a	Ph	4-Cl C6H4	4-Cl C6H4	17	86
	2	3b	Me 4-MeO C6H4	3,4-di Cl C6H3	18	77
	3	3c	Ph 4-MeO C6H4	3-NO2 C6H4	20	81
	4	3d	Ph	4-Br C6H4	3-NO2 C6H4	22	63
	5	3e	Ph 4-MeO C6H4	C 6H5	16	68
	6	3f	Ph	4-Cl C6H4	3-NO2 C6H4	21	81
	7	3g	Ph	4-Cl C6H4	C 6H5	16	59
	8	3h	Ph	C6H5	3,4-di Cl C6H3	16	75
	9	3i	Ph	C6H5	4 -M e C 6H4	14	85
	10	3j	Ph	C6H5	3,4,5-tri MeO C6H2	12	67
	11	3k	Ph	C3H7	3,4-di Cl C6H3	16	65
	12	3l	Me	C6H5	3,4-di Cl C6H3	14	84
	13	3m	Ph 4-MeO C6H4	3,4-di Cl C6H3	16	70
	14	3n	Et 4-MeO C6H4	3-NO2 C6H4	18	86
	15	3o	Et	4-Cl C6H4	C 6H5	19	58
	16	3p	Me	4-Cl C6H4	C 6H5	18	53
	17	3q	Me	4-Me C6H4	3-NO2 C6H4	19	52
	18	3r	Ph	C3H7	3 -N O 2 C6H4	17	60
	a Reactions carried out with (1 mmol) of aldehyde, (1.5 mmol) NH 2 OH.HCl, (0.5 mmol) TCCA and (1 mmol)
	alkene, in EtOH/H 2 O (10 ml, 1 :1) at 25°C under sonication (ultrasonic bath, 47 kHz).

b Yield of products after purification.

Acknowledgements

This work was financially supported by Erasmus+ program, key action 1 "Mobility of persons for education and training" (2018-2019), CNRST-Morocco, UM5-Rabat, Université Côte d'Azur-Nice (UCA) and UM6P-Benguérir, and are gratefully acknowledged.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Supporting Information

Supplementary data include the experimental procedures for the synthesis of all the compounds, their spectroscopic characterization (NMR, IR, and HPLC), representative spectra and additional biological assays.