MD, MSc Steeve Doizi
email: steeve.doizi@aphp.fr

MD Guido Kamphuis
email: g.m.kamphuis@amc.uva.nl

MD Guido Giusti

J L Palmero
email: jlpalmero@hospital-ribera.com

MD Jake M Patterson
email: jake.patterson@nhs.net

MD Silvia Proietti
email: proiettisil@gmail.com

MD Michael Straub
email: michael.straub@tum.de

Jean De La Rosette
email: j.j.delarosette@amc.uva.nl

Jose Luis

Jean De

MD Olivier Traxer
email: olivier.traxer@aphp.fr

Prof Olivier Traxer

Keywords: cystoscopy, double-J stent, ureteral stent, stents, device removal

removal outcomes, cost analysis and microbiology will help to delineate the possible place of Isiris™ in the current practice.

INTRODUCTION

Since their introduction in 1978, double-J stents have become a fundamental tool in the urological armamentarium and their placement is one of the most common procedures performed in modern urology [START_REF] Finney | Experience with new double J ureteral catheter stent[END_REF]. Although these stents are inserted for various indications, the most common is probably after an ureterorenoscopic procedure to prevent the incidence of postoperative ureteric obstruction and renal colic secondary to ureteral edema. Ureteric stenting has also been advocated to facilitate the passage of stone fragments secondary to the passive ureteric dilation [START_REF] Auge | Ureteral stents and their use in endourology[END_REF][START_REF] Harmon | Ureteroscopy: current practice and long-term complications[END_REF][START_REF] Júnior | Ureteroscopic stone removal in the distal ureter. Why change?[END_REF][START_REF] Knudsen | Stenting after ureteroscopy: pros and cons[END_REF][START_REF] Rapoport | Ureteral access sheath use and stenting in ureteroscopy: effect on unplanned emergency room visits and cost[END_REF]. Although this common practice is still debated, it has been reported in large series that approximately 80% of urologists place a ureteric stent after uncomplicated ureteroscopy for stone disease [START_REF] Nabi | Outcomes of stenting after uncomplicated ureteroscopy: systematic review and meta-analysis[END_REF][START_REF] Song | Meta-analysis of postoperatively stenting or not in patients underwent ureteroscopic lithotripsy[END_REF][START_REF] Haleblian | Ureteral stenting and urinary stone management: a systematic review[END_REF][START_REF] Tang | Placement of ureteral stent after uncomplicated ureteroscopy: do we really need it?[END_REF][START_REF] Netto Nr | Routine ureteral stenting after ureteroscopy for ureteral lithiasis: is it really necessary?[END_REF][START_REF] Fx | Routine stenting after ureteroscopy: think again[END_REF][START_REF] Gerber | Use of stents after ureteroscopic stone removal[END_REF][START_REF] De La Rosette | The clinical research office of the endourological society ureteroscopy global study: indications, complications, and outcomes in 11,885 patients[END_REF][START_REF] Auge | Practice patterns of ureteral stenting after routine ureteroscopic stone surgery: a survey of practicing urologists[END_REF]. Once the double-J stent has been placed, this has to be removed after a while. Currently, there are two options common in practice to remove these stents: to use the extraction string suture with which the current stents may be packaged, or a cystoscopic removal using a flexible or rigid cystoscope with a grasper. For the latter procedure, this requires a dedicated place with video equipment as well as cystoscopes and graspers. These instruments are fragile and have to be disinfected after each procedure, which may limit the number of procedures due to their availability [START_REF] Mcgill | Durability of flexible cystoscopes in the outpatient setting[END_REF][START_REF] Söylemez | A cheap minimally painful and widely usable alternative for retrieving ureteral stents[END_REF][START_REF] Canales | An independent analysis of flexible cystoscope repairs and cost[END_REF][START_REF] Fuselier | Liquid sterilization versus high level disinfection in the urologic office[END_REF].

For these reasons, a single use flexible cystoscope with integrated grasper dedicated to double-J stent removal has been developed.

The objective of the current study was to evaluate the first clinical performance of the Isiris™ flexible cystoscope for double-J stent removal.

MATERIAL AND METHODS

Study design and participants

A prospective cohort study was conducted in six tertiary reference centers in Europe (France, Germany, Italy, Spain, the Netherlands, and United-Kingdom) in September 2015.

All consecutive patients included were 18 years or older and had to undergo a double-J stent removal. The following preoperative data were prospectively collected: gender, indication for double-J stent placement, stent characteristics (length, diameter, type and manufacturer), duration of stent implantation and type of anesthesia performed for stent removal. All patients gave informed consent to undergo a cystoscopic removal of stent with Isiris™, and the principles of the Declaration of Helsinki were followed.

Isiris™

Isiris™ (Porgès-Coloplast) is a single use digital flexible cystoscope, CE approved, with an integrated grasper designed for double-J stent removal (Figures 1, 2 and3). Isiris™ is designed for ureteric stent removal only and not to perform regular diagnostic cystoscopy.

This flexible cystoscope has a 16-Fr outer diameter from the tip to the main part, no working channel for insertion of instruments, minimum of 80˚ deflection in upward and 90° in downward directions and a length of 39 centimeters (Figures 1 and4). The handle includes an irrigation connector, a deflection lever for upward and downward directions and a grasper activation button (Figure 1). The digital camera is made of a complementary metal oxide semiconductor (CMOS) sensor located at the tip of the endoscope and provides 0° direct view with 85° field of vision. The scope is connected via a cable to a reusable dedicated LCD portable monitor (Figure 3). The dimensions of display on monitor are 8.5 inches for a resolution of 800x600 pixels. A USB port is integrated in the monitor allowing connecting a USB drive to record the case if needed. The Isiris™ cystoscope cannot be sterilized.

Procedures

According to local protocol, preoperative urine analysis and culture were performed and appropriate prophylactic antibiotics were given. Procedures were performed as office-based cystoscopy in the outpatient clinic if no other intervention was planned. Patients were placed in dorsolithotomy position. The procedure was performed without anesthesia, local anesthesia (intraurethral lidocaine gel instillation) or general anesthesia if patient had to undergo an ureterorenoscopic procedure. All procedures were conducted by experienced endourologists. Each procedure was performed as a regular flexible cystoscopy, starting with disinfection of the genitalia with antisepsis according to local protocol followed by Isiris™ cystoscope insertion. Then, the stent was removed using the integrated grasper by activating the button located on the handle.

Evaluation criteria

The main outcome was the evaluation of success for stent removal.

The secondary outcomes were the performance characteristics of the instrument as experienced by the surgeon according to the following criteria evaluated during the procedure:

-Image quality displayed on the monitor with native resolution: rated according to a Likert scale of "bad" to "very good".

-Active deflection: upward and downward deflections were rated according to a Likert scale of "bad" to "very good" by the surgeon during the cystocopy.

-Maneuverability: surgeons rated the maneuverability on a Likert scale of "bad" to "very good".

-Grasper activation: participants rated the ability to use the grasper by activating the button using a Likert scale of "very difficult" to "very easy".

-Grasper functionality: participants rated the ability of grasper to catch the stent using a Likert scale of "bad" to "very good".

-Estimation of procedure duration with Isiris™ compared to the usual double-J stent removal with flexible cystoscope: participants rated the duration between shorter, similar or longer than the usual one.

-Overall satisfaction: participants rated their overall satisfaction using a Likert scale of "bad" to "very good".

The following data were also collected: need of assistant and his role.

Per operative complications and technical failures were collected. No data were captured on postoperative complications.

Statistical analysis

Qualitative variables were described as numbers and percentage. Quantitative variables were described as median [interquartile range] values.

RESULTS

Patient characteristics

A total of 83 procedures were performed with Isiris™. The study included 45 men (54%) and 38 women (46%). 82% of procedures were performed in the endoscopy room while the others were in the operating room as further urological intervention was planned. Double-J stent diameters were 7-Fr, 6-Fr and 4.8-Fr in 60%, 36% and 4% respectively. Length of stent was 24 cm, 26 cm, 28 cm in 31.3%, 14.5% and 30.1% respectively. The median duration of stent implantation was 28 days [14; 60]. Indications for stent placement were drainage after a ureterorenoscopy procedure for stone disease in 73.5% of cases, ureterorenoscopy procedure for upper urinary tract tumor in 7.3%, ureteral stenosis, renal transplant and other indications in 6% for each (Table 1).

Primary outcome

In five patients, stent removal was not possible. Four patients had an incrusted double-J stent and in one patient the stent migrated into the ureter. In the four cases with encrusted double-J stent, stent removal with conventional flexible cystoscope and grasper was attempted and did not succeed. Finally, the five patients had to be scheduled for an ureterorenoscopy procedure to remove the stent.

In the other 78 patients, all double-J stents were removed successfully (Table 2).

No complication occurred during the procedures and no technical failure has been experienced.

Secondary outcomes

Image quality was rated as "very good" in 72.3%, "good" in 22.9% and "fair" in 4.8% of cases.

Deflection was rated as "very good" in 78.3%, "good" in 20.5% and "fair" in 1.2% of cases.

Maneuverability was rated as "very good" in 72.3%, "good" in 25.3% and "fair" in 2.4% of cases.

Grasper activation button was rated as "very easy" to use in 69.9%, "easy" in 27.7% and "fair" in 2.4%.

Grasper functionality was rated as "very good" in 73.5%, "good" in 22.9%, "fair" in 2.4% and "poor" in 1.2% of cases.

Duration of procedure compared to conventional cystoscopic double-J removal was considered "shorter" in 37.4%, "similar" in 57.8% and "longer" in 4.8% of cases.

Overall satisfaction was "very good" in 68.7%, "good" in 22.9%, "acceptable" in 7.2% and "poor" in 1.2% of cases (Table 2).

An assistant was present during the procedure in 51.8% of cases. His role consisted of connecting the irrigation and scope to the monitor in 36.4%, preparing the endoscopic room in 24.2% or because the hospital regulations required an assistant in 39.4%.

DISCUSSION

This study evaluated the first single use digital flexible cystoscope integrating a grasper dedicated to double-J stent removal. We showed this device obtained a 94% success rate for ureteric stent withdrawal and very good results in almost 70% of cases according to the Likert scale rating for evaluation of image quality, deflection, maneuverability and grasper.

The five double-J stents that could not be successfully removed by the Isiris™ were also unsuccessfully engaged by conventional cystoscopy.

As this is the first disposable device dedicated to double-J stent removal and no comparison with similar instruments can be done, the release of Isiris™ addresses some questions. First of all, the question arises if this instrument is functional for its purpose and safe to use. In this first evaluation we confirmed that double-J stents can be removed safely with very good results regarding of image quality, deflection, maneuverability and instrumentation of the integrated grasper. The next question is related to costs. Since this endoscope is single use and does not require a dedicated place with equipment to remove the stents, cleaning and storage, the only direct costs are the one of the Isiris™ cystoscope. Besides this, the set up with the Isiris™ might change the need for an assistant in the room. However, this has to be balanced with the cost of a double-J stent removal using a flexible cystoscope and grasping forceps. Many studies focused on the cost of ureteroscopy but no evaluation on the direct and indirect costs of cystoscopic double-J stent removal is available. The only study partly addressing this question is the one of Netto and colleagues where they assessed the costeffectiveness of routine ureteral stenting after ureteroscopic stone removal. They found that the hospital charge per patient for stent use after uncomplicated ureteroscopy for stone removal was $2,445.31 with and $3,727.82 without the string left in place. However, they specified that stent removal was performed in the operating room, which may add an extra charge compared to the office cystoscopic removal setting [START_REF] Netto Nr | Routine ureteral stenting after ureteroscopy for ureteral lithiasis: is it really necessary?[END_REF]. Another issue is the flexible cystoscopes durability, implying repair costs. Although the durability of flexible ureteroscopes has often been reported, only few studies assessed this issue on flexible cystoscopes. In 2013, McGill et al prospectively evaluated the durability and repair costs of six flexible cystoscopes over a study period of 14 months on an outpatient setting. They excluded of their analysis the costs associated with cystoscope purchase. They found a total of five failures occurring in four cystoscopes, with a mean of 412.8 procedures per cystoscope. Damages occurred after 70, 194, and 236 uses for three cystoscopes and one cystoscope had 2 failures after 168 and 255 uses. This meant a cost of cystoscope maintenance of $5.41 per procedure during the study period. They also underlined that cystoscopes damages occurred earlier in cystoscopes with a higher percentage of uses secondary to stent removals, biopsies, and fulgurations [START_REF] Mcgill | Durability of flexible cystoscopes in the outpatient setting[END_REF]. In 2011, Söylemez et al reported a purchase cost of $18.342 for a flexible cystoscope and $2.057 for a 3-Fr flexible grasper. The maintenance interval for each instrument was 10 times with a repair cost of $1.487 and $491 for the flexible cystoscope and forceps respectively [START_REF] Söylemez | A cheap minimally painful and widely usable alternative for retrieving ureteral stents[END_REF]. In a retrospective study, Canales et al showed that flexible cystoscopes required repair every 2 to 3 years over a four year study period. However, they did not report the number of uses before repair and the mechanism of damage [START_REF] Canales | An independent analysis of flexible cystoscope repairs and cost[END_REF]. And finally, Fuselier and Mason found that 7 cystoscopes were damaged over a two year study period including approximately 2,000 procedures.

Similarly, no details on number of procedures performed before repair and cause of failure were reported and it was unclear whether the endoscopes were used or new before the onset of the study [START_REF] Fuselier | Liquid sterilization versus high level disinfection in the urologic office[END_REF]. None of the studies included costs related to cleaning instruments as well as costs related to handling them.

The other issue addressed is the risk of urinary tract infection after flexible cystoscopy.

Flexible cystoscopes mostly undergo high level disinfection (HLD) rather than sterilization between patients [START_REF] Noronha | A 21st century nosocomial issue with endoscopes[END_REF]. Thus, the question of pathogenic cross contamination between patients is addressed given that there is now an alternative with this single use cystoscope.

In the field of endourology, Chang et al first reported in 2013 15 patients developing ertapenem-resistant Enterobacter cloacae urinary tract infections due to a contaminated ureteroscope [START_REF] Chang | Outbreak of ertapenemresistant Enterobacter cloacae urinary tract infections due to a contaminated ureteroscope[END_REF]. However, we have to emphasize that the risk of urinary tract infection after flexible cystoscopy is low. In two large prospective studies, Herr et al reported that 1.9% of patients undergoing a diagnostic flexible cystoscopy developed a febrile urinary tract infection within 30 days after the procedure [START_REF] Herr | Should antibiotics be given prior to outpatient cystoscopy? A plea to urologists to practice antibiotic stewardship[END_REF][START_REF] Herr | The risk of urinary tract infection after flexible cystoscopy in patients with bladder tumor who did not receive prophylactic antibiotics[END_REF]. In another large study, the authors found a 9% incidence of bacteriuria after cystoscopy and only 1.2% developed a symptomatic urinary tract infection [START_REF] Johnson | Oral ciprofloxacin or trimethoprim reduces bacteriuria after flexible cystoscopy[END_REF]. Other studies reported an incidence of bacteriuria up to 7.8% and symptomatic urinary tract infection up to 3% [START_REF] Clark | Urinary infection following out-patient flexible cystoscopy[END_REF][START_REF] Lugagne | Infectious risks of outpatient cystoscopy in men with sterile urine[END_REF][START_REF] Almallah | Urinary tract infection and patient satisfaction after flexible cystoscopy and urodynamic evaluation[END_REF][START_REF] Wilson | Is antibiotic prophylaxis required for flexible cystoscopy? A truncated randomized double-blind controlled trial[END_REF][START_REF] García-Perdomo | Efficacy of antibiotic prophylaxis in patients undergoing cystoscopy: a randomized clinical trial[END_REF][START_REF] Doizi | First clinical evaluation of a new single-use flexible ureteroscope (LithoVue™): a European prospective multicentric feasibility study[END_REF][31][32][33][34][35][36][37][38][39].

And finally, what is the environmental impact of the single use devices? Further studies should be conducted comparing this issue between single use endoscopes with reusable scopes requiring disinfection with highly toxic detergents.

The present study has several limitations. First, we did not compare the clinical performance of Isiris™ to a control group including current practice to remove ureteric stents i.e. flexible cystoscope with grasping forceps. This may impact the strength of our results. However the current study should be considered as a preliminary evaluation of this new device. All the concerns discussed earlier highlight the need of further studies to define the place of Isiris™ in the current practice. A comparison with conventional flexible cystoscopic stent removal is needed where the clinical performance, direct and indirect costs with cost-effectiveness analysis are assessed. However, such studies comparing the costs of care display great variation across countries due to differences in practices methods of calculating costs and billing. Furthermore, studies comparing microbiological evaluation and the risk of urinary tract infection between this new device and traditional stent removal with flexible cystoscope are needed.

This new concept of double-J stent removal and single use endoscopes such as flexible ureteroscopes may open a new era in the field of endourology [START_REF] Doizi | First clinical evaluation of a new single-use flexible ureteroscope (LithoVue™): a European prospective multicentric feasibility study[END_REF]. However, further evaluations are required to delineate the position of these instruments in the current practice.

CONCLUSIONS

This preliminary study evaluated the Isiris™, the first single use digital flexible cystoscope integrating a grasper dedicated to double-J stent removal. The evaluation of instrument characteristics of image quality, active deflection, maneuverability and grasper functionality by the surgeons on Likert scales was good. There were no complications or technical failures.

In five cases the double-J stent required additional ureteroscopic surgical removal due to stent encrustation or stent migration into the ureter.

The position of single use endoscopes in day to day practice will depend on functional outcome in larger series, local practice and local cost analysis and concerns about possible infections.

RESEARCH INVOLVING HUMAN PARTICIPANTS AND/OR ANIMALS

Ethical approval: "All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards." Informed consent: "Informed consent was obtained from all individual participants included in the study."

LEGENDS

Characteristics

First

  clinical evaluation of a new single-use flexible cystoscope dedicated to double-J stent removal (Isiris™): a European prospective multicenter study Steeve Doizi, Guido Kamphuis, Guido Giusti, Jl Palmero, Jm Patterson, Silvia Proietti, Michael Straub, Jean de La Rosette, Olivier Traxer To cite this version: Steeve Doizi, Guido Kamphuis, Guido Giusti, Jl Palmero, Jm Patterson, et al.. First clinical evaluation of a new single-use flexible cystoscope dedicated to double-J stent removal (Isiris™): a European prospective multicenter study. World Journal of Urology, 2017, 35 (8), pp.1269 -1275. 10.1007/s00345-016-1986-0. hal-01840055

Figure 1 .

 1 Figure 1. Isiris™ flexible cystoscope. Plain arrow: grasper activation button. Square dotted arrow: irrigation connector. Round dotted arrow: deflection lever.

Figure 2 .

 2 Figure 2. Activation of grasper. Maximum length of protruded grasper is 18 mm. Minimum grasper opening distance is 4.5 mm.

Figure 3 .

 3 Figure 3. Isiris™ connected via its cable to the reusable dedicated LCD portable monitor. A: Activation of the grasper. B: Grasper catching a double-J stent.

Figure 4 .

 4 Figure 4. Downward (left) and upward (right) deflections.

Table 1 .

 1 Demographic data.

Table 2 .

 2 Primary and secondary outcomes.

Data are presented as median with interquartile rangeTable 1 .

 1 Demographic data.

	Number of procedures	83
	Gender, n (%)
	Male/Female	45/38 (54/46)
	Double-J stent characteristics, n (%)
	Diameter (Fr)
	4.8	3	(4)
	6		(36)
	7		(60)
	Length (cm)
	16	1	(1.2)
	18	2	(2.4)
	22	3	(3.6)
	24	(31.3)
	26	(14.5)
	28	(30.1)
	Other	(16.9)
	Duration of stent implantation, median days	[14 ; 60]
	Anesthesia, n (%)
	None	(14.5)
	Local		(71)
	General	(14.5)
	Double-J stent removal indication, n (%)
	Stone	(73.5)
	Upper Urinary Tract Tumor	6	(7.3)
	Ureteral stenosis	5	(6)
	Renal transplantation	5	(6)
	Ureteral reimplantation	1	(1.2)
	Unspecified	5	(6)

ACKNOWLEDGMENTS

The authors would like to thank Porgès-Coloplast for providing Isiris™ endoscopes in each centre in order to evaluate its performance.

SUPPORT/FINANCIAL DISCLOSURES


MD Fanny Mochel
email: fanny.mochel@upmc.fr

Triheptanoin for the Treatment of Brain Energy Deficit: A 14-Year Experience

Keywords: Krebs cycle, astrocytes, GLUT1, Huntington disease, magnetic resonance spectroscopy

Triheptanoin is an odd-chain triglyceride with anaplerotic properties-that is, replenishing the pool of metabolic intermediates in the Krebs cycle. Unlike even-chain fatty acids metabolized to acetyl-CoA only, triheptanoin can indeed provide both acetyl-CoA and propionyl-CoA, two key carbon sources for the Krebs cycle. Triheptanoin was initially used in patients with long-chain fatty acid oxidation disorders. The first demonstration of the possible benefit of triheptanoin for brain energy deficit came from a patient with pyruvate carboxylase deficiency, a severe metabolic disease that affects anaplerosis in the brain. In an open-label study, triheptanoin was then shown to decrease nonepileptic paroxysmal manifestations by 90% in patients with glucose transporter 1 deficiency syndrome, a disease that affects glucose transport into the brain. 31 P magnetic resonance spectroscopy studies also indicated that triheptanoin was able to correct bioenergetics in the brain of patients with Huntington disease, a neurodegenerative disease associated with brain energy deficit. Altogether, these studies indicate that triheptanoin can be a treatment for brain energy deficit related to altered anaplerosis and/or glucose metabo-lism.

BACKGROUND ON THE USE OF TRIHEPTANOIN IN HUMANS Triheptanoin

Triheptanoin is a medium-chain triglyceride of three 7-carbon fatty acids with a molecular formula of C 24 H 44 O 6 . Triheptanoin is metabolized rapidly in the gut to form glycerol and heptanoate (Fig. 1). Glycerol is converted to pyruvate, and heptanoate can be further metabolized in the liver to C4-ketone bodies (3hydroxybutyrate and acetoacetate) and C5-ketone bodies (3-hydroxypentanoate and 3-ketopentanoate) [START_REF] Kinman | Parenteral and enteral metabolism of anaplerotic triheptanoin in normal rats[END_REF][START_REF] Deng | Interrelations between C4 ketogenesis, C5 ketogenesis, and anaplerosis in the perfused rat liver[END_REF] (Fig. 1). Unlike even-chain fatty acids metabolized to acetyl-CoA only, heptanoate can provide both acetyl-CoA and propionyl-CoA, two key carbon sources for the Krebs cycle (Fig. 1). In the brain, heptanoate and/or C5-ketone bodies are metabolized primarily through the glial Krebs cycle [START_REF] Marin-Valencia | Heptanoate as a neural fuel: energetic and neurotransmitter precursors in normal and glucose transporter I-deficient (G1D) brain[END_REF][START_REF] Hadera | Triheptanoin partially restores levels of tricarboxylic acid cycle intermediates in the mouse pilocarpine model of epilepsy[END_REF] (Fig. 2). Triheptanoin has been used clinically for over a decade in more than 200 subjects in human studies of a variety of different diseases, including about 70 children [START_REF] Roe | Treatment of cardiomyopathy and rhabdomyolysis in long-chain fat oxidation disorders using an anaplerotic odd-chain triglyceride[END_REF][START_REF] Roe | Anaplerotic diet therapy in inherited metabolic disease: therapeutic potential[END_REF][START_REF] Mochel | Dietary anaplerotic therapy improves peripheral tissue energy metabolism in patients with Huntington's disease[END_REF][START_REF] Pascual | Triheptanoin for glucose transporter type I deficiency (G1D): modulation of human ictogenesis, cerebral metabolic rate, and cognitive indices by a food supplement[END_REF][START_REF] Mochel | Triheptanoin dramatically reduces paroxysmal motor disorder in patients with GLUT1 deficiency[END_REF], and many of them received over 5 years of treatment. These data support the safety of triheptanoin in humans, usually when administered at a dose between 1 and 2.5 g/kg body weight depending on age. Triheptanoin is split into glycerol and three molecules of heptanoate in the gut. In the liver, glycerol is converted to pyruvate, and heptanoate can be further metabolized to C4-ketone bodies (3-hydroxybutyrate and acetoacetate) and C5ketone bodies (3-hydroxypentanoate and 3-ketopentanoate). In the liver, or after export to the muscle, the kidney, or the brain, C5ketone bodies are further metabolized to acetyl-CoA and propionyl-CoA, unlike even-chain fatty acids metabolized to acetyl-CoA only.

In the brain, a direct uptake of heptanoate is also possible by passive diffusion (see Fig. 2). Fig. 2. Pyruvate carboxylase deficiency, GLUT1 syndrome deficiency, and treatment with triheptanoin. Illustration of astrocyteneuron metabolic coupling, highlighting metabolic specificities from each cell type-for instance, predominant expression of pyruvate carboxylase and glutamine synthetase in astrocytes versus predominant expression of glutaminase in neurons. Heptanoate and/or C5 ketones are metabolized primarily through the glial Krebs cycle [START_REF] Marin-Valencia | Heptanoate as a neural fuel: energetic and neurotransmitter precursors in normal and glucose transporter I-deficient (G1D) brain[END_REF][START_REF] Hadera | Triheptanoin partially restores levels of tricarboxylic acid cycle intermediates in the mouse pilocarpine model of epilepsy[END_REF]. Therefore, impaired astrocytic anaplerosis due to reduced oxaloacetate resulting from pyruvate carboxylase deficiency, or impaired astrocytic glucose metabolism resulting from GLUT1 deficiency, can be compensated by the provision of Krebs cycle intermediates from heptanoate and/ or C5-ketone bodies.GLUT, glucose transporter; MCT, monocarboxylate transporter; glucose-6-P, glucose-6-phosphate; pyr, pyruvate; lac, lactate; C5KB, C5-ketone bodies; OAA, oxaloacetate; KC, Krebs cycle. into energy. The standard of care is to restrict dietary fat, avoid fasting, suppress lipolysis, and use even-carbon medium-chain triglyceride and carnitine supplementation. However, patients under these therapeutic measures are still exposed to symptoms reflecting insufficient provision of energy, in particular cardiomyopathy, rhabdomyolysis, and muscle weakness. Based on the hypothesis that this energy deficiency is caused by the reduced availability of Krebs cycle intermediates required for optimal oxidation of acetyl-CoA, triheptanoin was first tested in 3 patients harboring a deficiency in very-long-chain acyl-CoA dehydrogenase and led to the rapid clinical improvement of their cardiac and muscle functions [START_REF] Roe | Treatment of cardiomyopathy and rhabdomyolysis in long-chain fat oxidation disorders using an anaplerotic odd-chain triglyceride[END_REF]. A similar therapeutic benefit was later reported in 7 patients exhibiting carnitine palmitoyltransferase II deficiency [START_REF] Roe | Carnitine palmitoyltransferase II deficiency: successful anaplerotic diet therapy[END_REF]. More recently, a retrospective analysis of 24 patients with long-chain fatty acid oxidation disorders treated for up to 12.5 years showed a significant decrease in mean hospitalization days per year and hypoglycemia event rate per year [START_REF] Vockley | Long-term major clinical outcomes in patients with long chain fatty acid oxidation disorders before and after transition to triheptanoin treatment-a retrospective chart review[END_REF]. A therapeutic effect of triheptanoin has also been reported in the management of acute cardiomyopathy associated with long-chain fatty acid oxidation disorders in 10 children, including 8 infants [START_REF] Vockley | Triheptanoin treatment in patients with pediatric cardiomyopathy associated with long chain-fatty acid oxidation disorders[END_REF].

PYRUVATE CARBOXYLASE DEFICIENCY:

A SHIFT TOWARD BRAIN ENERGY METABOLISM The first demonstration of the possible benefit of triheptanoin for brain energy deficit came from a patient with pyruvate carboxylase (PC) deficiency [START_REF] Mochel | Pyruvate carboxylase deficiency: clinical and biochemical response to anaplerotic diet therapy[END_REF]. PC converts pyruvate to oxaloacetate and therefore plays an important role in refilling the pool of catalytic intermediates of the Krebs cycle in peripheral tissues and in the brain. Briefly, a 6-day-old infant presented with the most severe phenotype of PC deficiency that associates hepatic failure, severe lactic acidosis, ketoacidosis, and hyperammonemia with elevated citrullinemia. Enteral administration with a formula containing triheptanoin (about 35% of total caloric intake) resulted within 24 hours in decreased lactate and lactate/pyruvate ratio, indicating improved ratio of cytosolic NADH/NAD. Triheptanoin also led to decreased plasma ammonia and citrulline, likely reflecting the increased availability of oxaloacetate, and therefore aspartate through the malateaspartate shuttle [START_REF] Mochel | Pyruvate carboxylase deficiency: clinical and biochemical response to anaplerotic diet therapy[END_REF]. These immediate metabolic changes were associated with the complete restoration of hepatic function in less than 48 hours. Over the following months of treatment, increased levels of C5-ketone bodies, glutamine, and g-aminobutyric acid (GABA) were measured in the cerebrospinal fluid (CSF), associated with improved myelin synthesis on sequential brain magnetic resonance imaging [START_REF] Mochel | Pyruvate carboxylase deficiency: clinical and biochemical response to anaplerotic diet therapy[END_REF].

Further to the immediate correction of peripheral energy deficiency, reflected by the prompt restoration of hepatic function, the subsequent neurometabolic response in this PC-deficient infant is of particular interest considering the exclusive glial localization of PC in the brain (Fig. 2). Besides the well-known metabolic dichotomy between neurons and glial cells-that is, neurons relying highly on oxidative metabolism and astrocytes on aerobic glycolysis [START_REF] Zhang | An RNA-sequencing transcriptome and splicing database of glia, neurons, and vascular cells of the cerebral cortex[END_REF][START_REF] Volkenhoff | Glial glycolysis is essential for neuronal survival in drosophila[END_REF]brain energetic organization relies indeed on the narrow metabolic coupling between these different cell types. The glutamate-glutamine(-GABA) shuttle between neurons and astrocytes first illustrated such metabolic interactions more than 30 years ago with the selective expression of glutamine synthetase by astrocytes that is essential for the recycling of glutamate released from neurons in glutamatergic neurotransmission [START_REF] Shank | Pyruvate carboxylase: an astrocyte-specific enzyme implicated in the replenishment of amino acid neurotransmitter pools[END_REF] (Fig. 2). More recently, a tricellular compartmentation of brain energy metabolism between neurons, astrocytes, and oligodendrocytes has been proposed-although still the subject of controversy [START_REF] Hertz | The astrocyte-neuron lactate shuttle: a challenge of a challenge[END_REF][START_REF] Dienel | Lack of appropriate stoichiometry: strong evidence against an energetically important astrocyte-neuron lactate shuttle in brain[END_REF]-via a lactate shuttle involving astrocytes at the synaptic level [START_REF] Supplie | Respiration-deficient astrocytes survive as glycolytic cells in vivo[END_REF] (Fig. 2) and oligodendrocytes at the axonal level [START_REF] Funfschilling | Glycolytic oligodendrocytes maintain myelin and long-term axonal integrity[END_REF]. Hence, the PC case study suggested that the astrocytic anaplerotic defect secondary to PC deficiency could be bypassed by C4and C5-ketone bodies derived from triheptanoin, or heptanoate itself as shown with octanoate [START_REF] Ebert | Energy contribution of octanoate to intact rat brain metabolism measured by 13C nuclear magnetic resonance spectroscopy[END_REF]. It also paved the way to research on neurometabolic diseases where the cerebral anaplerosis is deficient and/or glucose metabolism is impaired.

GLUCOSE TRANSPORTER 1 DEFICIENCY SYNDROME: A DISEASE MODEL FOR GLUCOSE AND ANAPLEROSIS DEFICIENCY

Glucose transporter 1 (GLUT1) deficiency syndrome (GLUT1-DS), characterized in 1991 by De Vivo et al., is caused by impaired glucose transport across the bloodbrain barrier and into astrocytes (Fig. 2) [START_REF] Vivo | Defective glucose transport across the blood-brain barrier as a cause of persistent hypoglycorrhachia, seizures, and developmental delay[END_REF]. GLUT1-DS is usually associated with a mutation in the solute carrier family 2, member 1 (SLC2A1) gene, encoding the GLUT1 protein responsible for transporting glucose across the blood-brain barrier. However, genetic analysis can fail at identifying variants despite a very suggestive clinical and biochemical phenotype [START_REF] Verrotti | Glut1 deficiency: when to suspect and how to diagnose?[END_REF]. Most commonly, GLUT1-DS is inherited in an autosomal dominant manner, but the vast majority of heterozygous SLC2A1 mutations occur de novo. The laboratory hallmark of GLUT1-DS is low CSF glucose concentration-usually < 2.2 mmol/l-associated with low CSF lactate-usually < 1 mmol/l [START_REF] Klepper | GLUT1 deficiency syndrome in clinical practice[END_REF]. The low CSF lactate in most patients with GLUT1-DS [START_REF] Leen | Cerebrospinal fluid analysis in the workup of GLUT1 deficiency syndrome: a systematic review[END_REF] is thought to reflect the glucose shortage in the brain [START_REF] Marin-Valencia | Glut1 deficiency (G1D): epilepsy and metabolic dysfunction in a mouse model of the most common human phenotype[END_REF] (Fig. 2). Decreased 3-O-methyl-D-glucose uptake assay is a functional measure of glucose transport across the red blood cell membrane that can very useful for the diagnosis of GLUT1-DS [START_REF] Yang | Glut1 deficiency syndrome and erythrocyte glucose uptake assay[END_REF], although rarely available on a clinical basis as it implies the use of radioactivity and precautions in blood sample storage and preanalytical steps [START_REF] Klepper | GLUT1 deficiency syndrome in clinical practice[END_REF]. Recently, we evaluated a novel diagnostic test on red blood cells using flow cytometry, readily available in clinical practice, in 30 patients with GLUT1-DS with predominant movement disorders, 18 patients with movement disorders due to other genetic defects, and 346 healthy controls [START_REF] Gras | A simple blood test expedites the diagnosis of GLUT1 deficiency syndrome[END_REF]. Our diagnostic rate was 78%, including patients with normal lumbar puncture or genetic analysis, and no false positive. This simple and rapid test opens exciting perspectives for the screening of GLUT1-DS in any patient, child or adult, presenting with cognitive impairment, epilepsy, ataxia and/or dystonia, or paroxysmal movement disorder.

Because glucose is the primary source of energy for the brain, this disorder results in a chronic state of cerebral energy deficiency. The phenotype typically comprises psychomotor retardation and permanent motor disorders, associated with paroxysmal manifestations including seizures and nonepileptic paroxysmal episodes [START_REF] Leen | Glucose transporter-1 deficiency syndrome: the expanding clinical and genetic spectrum of a treatable disorder[END_REF]. With age, patients tend to experience less seizures but more frequent paroxysmal movement disorders [START_REF] Gras | GLUT1 deficiency syndrome: an update[END_REF], likely reflecting dynamic changes in brain energy metabolism over years. Ketogenic diets, which provide C4-ketone bodies to the brain and compensate for the lack of glucose, represent the standard of care in GLUT1-DS [START_REF] Klepper | Glut1 deficiency syndrome and novel ketogenic diets[END_REF]. When properly administered, ketogenic diets are well tolerated and are highly effective in controlling epilepsy in GLUT1-DS, but seizures may recur [START_REF] Klepper | Seizure control and acceptance of the ketogenic diet in GLUT1 deficiency syndrome: a 2-to 5-year follow-up of 15 children enrolled prospectively[END_REF]. Even when seizures are controlled, patients may continue to have cognitive and behavioral deficits [START_REF] Klepper | Introduction of a ketogenic diet in young infants[END_REF].

In patients with milder forms of the disease, paroxysmal movement disorders, especially dyskinesia, may be the main or the sole manifestations of the disease and can occur at any age [START_REF] Pons | The spectrum of movement disorders in Glut-1 deficiency[END_REF]. Ketogenic diets seem to be less efficient in nonepileptic paroxysmal manifestations, and sometimes result in more complex movement disorders [START_REF] Leen | Glucose transporter-1 deficiency syndrome: the expanding clinical and genetic spectrum of a treatable disorder[END_REF]. Moreover, many patients, especially adolescents and adults, have difficulty complying with the constraints of these long-term diets and their side effects. Therefore, triheptanoin was tested in GLUT1-DS patients with nonepileptic paroxysmal manifestations for whom a ketogenic diet was not a therapeutic option (ClinicalTrials.gov identifier: NCT02014883). Unlike ketogenic diets, which provide C4-ketone bodies and therefore acetyl-CoA only, triheptanoin provides both acetyl-CoA and propionyl-CoA. An open-label study was divided into three phases of 2 months each (baseline, treatment, and withdrawal), and eight patients with GLUT1-DS, including four children, were enrolled. During the baseline phase, GLUT1-DS patients experienced a mean of 30.8 ( 6 27.7; 10-85) paroxysmal manifestations compared with a mean of 2.8 ( 6 2.9; 0-7) when treated with triheptanoin (P 5 0.028) [START_REF] Mochel | Triheptanoin dramatically reduces paroxysmal motor disorder in patients with GLUT1 deficiency[END_REF]. This represented a 90% symptom reduction. Conversely, during the withdrawal phase, symptoms reappeared with a mean of 24.2 ( 6 21.9; 5-63) paroxysmal manifestations [START_REF] Mochel | Triheptanoin dramatically reduces paroxysmal motor disorder in patients with GLUT1 deficiency[END_REF]. Upon study completion, all patients wished to continue treatment with triheptanoin, and all but one are still treated today, almost 3 years after the study start, with very positive outcomes (personal unpublished data). Functional 31 P magnetic resonance spectroscopy (MRS) was performed to quantify phosphocreatine (PCr) and inorganic phosphate (Pi) within the occipital cortex during (activation) and after (recovery) a visual stimulus-see further details in the next section on Huntington disease. Under treatment with triheptanoin, the ratio of Pi/PCr improved during activation, contrasting with a deterioration of the spectroscopic profile when treatment was withdrawn [START_REF] Mochel | Triheptanoin dramatically reduces paroxysmal motor disorder in patients with GLUT1 deficiency[END_REF].

Despite the absence of a control group, the magnitude of the clinical effect combined with the spectroscopic responses argued against a placebo effect.

Furthermore, another open-label study conducted in epileptic GLUT1-deficient patients showed a reduction of spike waves on electroencephalogram about 90 min after ingesting triheptanoin [START_REF] Pascual | Triheptanoin for glucose transporter type I deficiency (G1D): modulation of human ictogenesis, cerebral metabolic rate, and cognitive indices by a food supplement[END_REF]. In a mouse model of GLUT1-DS, it was shown that triheptanoin can lead to the delivery of heptanoate to the brain, which can then be metabolized into glucose and neurotransmitter intermediates [START_REF] Marin-Valencia | Heptanoate as a neural fuel: energetic and neurotransmitter precursors in normal and glucose transporter I-deficient (G1D) brain[END_REF]. Since GLUT1-DS is primarily related to glucose shortage in the brain, these data indicate that triheptanoin can be a treatment for brain energy deficit related to altered anaplerosis and/or glucose metabolism. Nonetheless, a larger controlled study is needed to establish whether triheptanoin can be an alternative therapeutic approach in GLUT1-DS, especially for patients who object to or cannot comply with the constraints of ketogenic diets.

TARGETING BRAIN ENERGY DEFICIT IN NEURODEGENERATIVE DISEASES: HUNTINGTON DISEASE

The initial detailed description of Huntington disease (HD) was that of George Huntington, a medical practitioner of Pomeroy, Ohio, in 1872. HD is inherited as an autosomal dominant trait. In individuals with HD, a polymorphic trinucleotide repeat sequence (CAGn), near the 5 0 end of the HTT gene, is expanded beyond the normal repeat range, leading to the translation of an expanded polyglutamine sequence in the huntingtin protein (Htt) [START_REF] Huntington's Disease | A novel gene containing a trinucleotide repeat that is expanded and unstable on Huntington's disease chromosomes[END_REF]. The clinical features of HD usually emerge in adulthood between 30 and 50 years of age, after which illness progresses steadily over a period of 15 to 25 years. Patients with HD may present with motor signs (chorea, bradykinesia, motor impersistence), with behavioral signs (irritability, anxiety, mood disturbance), or with both [START_REF] Paulsen | Detection of Huntington's disease decades before diagnosis: the Predict-HD study[END_REF]. The pathology of HD includes prominent neuronal loss and gliosis in the caudate nucleus and putamen along with regional and more diffuse atrophy. Energy metabolism has been under the scope of HD research for many years because of several observations in both patients and models of the disease. A reduction in ATP production was shown in brain of HD mice, even before the onset of motor deficits [START_REF] Gines | Specific progressive cAMP reduction implicates energy deficit in presymptomatic Huntington's disease knock-in mice[END_REF]. Positron emission tomography studies conducted in HD patients showed that glucose consumption is reduced in the brain, providing strong evidence for hypometabolism, especially in the basal ganglia, even in asymptomatic mutation carriers [START_REF] Antonini | Striatal glucose metabolism and dopamine D2 receptor binding in asymptomatic gene carriers and patients with Huntington's disease[END_REF]. The underlying cause of this early energy deficit in HD brain is currently unknown, but impaired glycolysis [START_REF] Browne | The energetics of Huntington's disease[END_REF], Krebs cycle [START_REF] Tabrizi | Biochemical abnormalities and excitotoxicity in Huntington's disease brain[END_REF], and/or oxidative phosphorylation [START_REF] Milakovic | Mitochondrial respiration and ATP production are significantly impaired in striatal cells expressing mutant huntingtin[END_REF] are likely involved. Mutant Htt was shown to decrease the expression of PGC-1a in the striatum of HD mice and patients, through transcriptional inhibition [START_REF] Cui | Transcriptional repression of PGC-1alpha by mutant huntingtin leads to mitochondrial dysfunction and neurodegeneration[END_REF]. PGC-1a is a transcriptional coactivator that regulates key energetic metabolic pathways, both in the brain and peripheral tissues [START_REF] Lin | Metabolic control through the PGC-1 family of transcription coactivators[END_REF]. The possible role of PGC-1a in HD was initially suspected from the observation of selective striatal lesions in PGC-1a knockout mice. Downregulation of PGC-1a in HD striatum was then shown to affect mitochondrial energy metabolism [START_REF] Lin | Defects in adaptive energy metabolism with CNS-linked hyperactivity in PGC-1alpha null mice[END_REF], possibly by impairing oxidative phosphorylation [START_REF] Cui | Transcriptional repression of PGC-1alpha by mutant huntingtin leads to mitochondrial dysfunction and neurodegeneration[END_REF]. In addition, the inhibition of succinate dehydrogenase, by 3-nitropropionate or malonate, mimicking HD neuropathology in baboons [START_REF] Palfi | Chronic 3-nitropropionic acid treatment in baboons replicates the cognitive and motor deficits of Huntington's disease[END_REF] and mice [START_REF] Brouillet | Partial inhibition of brain succinate deshydrogenase is sufficient to initiate striatal degeneration in rat[END_REF], suggests that a lack of substrates for the Krebs cycle and the respiratory chain is implicated in the energy deficit of HD brain.

Weight loss, reported in patients from the earliest stages of the disease, has been an important clinical observation related to energy deficiency in HD [START_REF] Mochel | Early energy deficit in Huntington disease: identification of a plasma biomarker traceable during disease progression[END_REF][START_REF] Aziz | Weight loss in Huntington disease increases with higher CAG repeat number[END_REF][START_REF] Goodman | The metabolic profile of early Huntington's disease-a combined human and transgenic mouse study[END_REF]. Using a multivariate statistical analysis of plasma components quantified by 1 H MRS, a distinctive signature was identified in HD mutation carriers that presented with hypercatabolism-that is, weight loss despite higher caloric intake, which was not explained by inflammatory processes or hormonal dysfunction. This metabolic signature was characterized by decreased levels of the branched chain amino acids (BCAAs)-valine, leucine, and isoleucine [START_REF] Mochel | Early energy deficit in Huntington disease: identification of a plasma biomarker traceable during disease progression[END_REF]-possibly reflecting their mitochondrial oxidation to provide two key intermediates for the Krebs cycle, acetyl-CoA and succinyl-CoA. As decreased BCAA levels were also detected in the CSF of HD patients [START_REF] Perry | Plasma-aminoacid levels in Huntington's chorea[END_REF], BCAA oxidation may also be activated in the HD brain to compensate for the local energy deficit. Consequently, therapies aiming at providing substrates to the Krebs cycle stood as possible therapeutic approaches in HD.

The evaluation of the benefit of therapies targeting energy metabolism is not trivial in HD as there are no biochemical parameters (unlike PC) or paroxysmal symptoms (unlike GLUT1-DS) directly related to energy deficiency. Therefore, the identification of biomarkers of energy metabolism in HD is critical. Hence, in vivo 31 P MRS was used to study cellular energy homeostasis because it allows the detection of high-energy phosphates, such as ATP and PCr. A pilot open-label study was first conducted in six early-affected HD patients to monitor the tolerability of triheptanoin and evaluate the possible short-term efficacy on muscle metabolism [START_REF] Mochel | Dietary anaplerotic therapy improves peripheral tissue energy metabolism in patients with Huntington's disease[END_REF]. Two patients developed muscle acidosis at baseline while performing a low-intensity exercise at baseline that resolved after 4 days of treatment with triheptanoin.

Measurements of muscle perfusion also suggested an effect of triheptanoin on hyperemic responses to exercise [START_REF] Mochel | Dietary anaplerotic therapy improves peripheral tissue energy metabolism in patients with Huntington's disease[END_REF]. The next step was to translate 31 P MRS methods to the brain to identify dynamic biomarkers of energy deficiency. A visual stimulation paradigm was applied, coupled with 31 P MRS, to measure the concentrations of high-energy phosphates before, during, and after activation of the occipital lobe [START_REF] Sappey-Marinier | Effect of photic stimulation on human visual cortex lactate and phosphates using 1H and 31P magnetic resonance spectroscopy[END_REF]. In two independent populations of healthy controls, an increase of about 10% of Pi/PCr ratio was observed during visual stimulation compared with rest, and a return to rest levels at recovery [START_REF] Mochel | Abnormal response to cortical activation in early stages of Huntington disease[END_REF][START_REF] Adanyeguh | Triheptanoin improves brain energy metabolism in patients with Huntington disease[END_REF] (Fig. 3). This ratio is directly related to ADP levels that regulate mitochondrial oxidative metabolism; Pi/PCr ratio thus provides an index of mitochondrial oxidative regulation, as shown in Fig. 3. 31 P magnetic resonance spectroscopy and brain energy metabolism. A: Representative 31 P MR spectrum from the occipital cortex of a control subject at 3T (pulse-acquire, 240 repetitions, TR 5 2 sec, 8-min total acquisition time). A 2-Hz Lorentzian line broadening was applied. Compared with previous studies [START_REF] Sappey-Marinier | Effect of photic stimulation on human visual cortex lactate and phosphates using 1H and 31P magnetic resonance spectroscopy[END_REF], the use of a surface coil adapted to the region of interest (occipital cortex), the use of a simple pulse-acquire sequence, and very good shimming all contributed to the excellent signal-to-noise ratio and spectral quality in our study. Thereby, the Pi peak was well separated from PME and PDE peaks, and therefore reliably quantified. PME, phosphomonoesters; Pi, inorganic phosphate; PDE, phosphodiesters; PCr, phosphocreatine; ATP, adenosine triphosphate; NADH, nicotinamide adenine dinucleotide. B: Pi/PCr ratio before, during, and after visual stimulation in healthy controls from two independent studies [START_REF] Mochel | Abnormal response to cortical activation in early stages of Huntington disease[END_REF][START_REF] Adanyeguh | Triheptanoin improves brain energy metabolism in patients with Huntington disease[END_REF]. We consistently observed a 10% increase of Pi/PCr ratio during visual stimulation compared with rest and a return to rest levels at recovery. muscle [START_REF] Wiener | Abnormal skeletal muscle bioenergetics during exercise in patients with heart failure: role of reduced muscle blood flow[END_REF]. Contrary to controls, HD patients at an early stage of the disease displayed no change in Pi/PCr ratios during activation and recovery, likely reflecting altered brain bioenergetics [START_REF] Mochel | Early alterations of brain cellular energy homeostasis in Huntington disease models[END_REF][START_REF] Adanyeguh | Triheptanoin improves brain energy metabolism in patients with Huntington disease[END_REF]. This abnormal profile remained stable over 1 month without intervention but was corrected, with increased Pi/PCr ratio during visual stimulation, after 1 month of treatment with triheptanoin [START_REF] Adanyeguh | Triheptanoin improves brain energy metabolism in patients with Huntington disease[END_REF]. This proof-ofconcept study strongly suggested that triheptanoin is able to improve the brain metabolic profile of early-affected HD patients. Further to this short-term effect of triheptanoin on brain energy metabolism, its effect on surrogate markers of HD like caudate atrophy, which have been reported as prominent in a multicentric HD study [START_REF] Tabrizi | Predictors of phenotypic progression and disease onset in premanifest and early-stage Huntington's disease in the TRACK-HD study: analysis of 36-month observational data[END_REF], and/or motor functions shall now be evaluated.

PERSPECTIVES

In the era of upcoming gene therapies for orphan diseases, triheptanoin remains a fascinating small molecule because of its peculiar composition with an odd number of carbons and, therefore, its ability to target the Krebs cycle. While initially developed for inborn errors of metabolism causing energy deficiency in peripheral tissues such as long-chain fatty acid oxidation disorders, the neurometabolic response observed in an infant with PC deficiency, although not reproduced so far [START_REF] Breen | Unsuccessful treatment of severe pyruvate carboxylase deficiency with triheptanoin[END_REF], constituted a first step toward the exploration of the therapeutic benefit of triheptanoin for brain energy deficit with impaired Krebs cycle and/or glucose metabolism. Although it still needs to be confirmed in a larger controlled study, the data available in GLUT1-DS strongly suggest that triheptanoin can improve brain energy metabolism with overt clinical benefit. Thanks to the development of dynamic tools to assess brain energy metabolism, it also becomes possible to determine whether other neurometabolic or neurodegenerative diseases, like HD, may benefit from anaplerotic approaches with triheptanoin.

Several promising studies have been conducted in preclinical models of other neurological diseases affecting brain energy homeostasis [START_REF] Willis | Anticonvulsant effects of a triheptanoin diet in two mouse chronic seizure models[END_REF][START_REF] Francis | Dietary triheptanoin rescues oligodendrocyte loss, dysmyelination and motor function in the nur7 mouse model of Canavan disease[END_REF][START_REF] Hadera | Triheptanoin partially restores levels of tricarboxylic acid cycle intermediates in the mouse pilocarpine model of epilepsy[END_REF][START_REF] Park | Anaplerotic triheptanoin diet enhances mitochondrial substrate use to remodel the metabolome and improve lifespan, motor function, and sociability in MeCP2-null mice[END_REF]. In the nur7 mouse model of Canavan disease, a leukodystrophy caused by the deficiency in aspartoacylase that hydrolyzes N-acetylaspartic acid into aspartic acid and acetate, triheptanoin reduced oxidative stress, promoted oligodendrocyte survival, and increased myelin content in the brain [START_REF] Francis | Dietary triheptanoin rescues oligodendrocyte loss, dysmyelination and motor function in the nur7 mouse model of Canavan disease[END_REF]. Of note, the therapeutic benefit was much more prominent in mice treated right after birth compared with older mice [START_REF] Francis | Dietary triheptanoin rescues oligodendrocyte loss, dysmyelination and motor function in the nur7 mouse model of Canavan disease[END_REF]. In male mice hemizygous for Mecp2 knockout, a model for Rett syndrome, triheptanoin increased longevity and improved motor function and social interaction together with reduced mitochondrial pathology [START_REF] Park | Anaplerotic triheptanoin diet enhances mitochondrial substrate use to remodel the metabolome and improve lifespan, motor function, and sociability in MeCP2-null mice[END_REF]. In two mouse models of chronic seizures, triheptanoin exerted anticonvulsant effects [START_REF] Willis | Anticonvulsant effects of a triheptanoin diet in two mouse chronic seizure models[END_REF] associated with increased Krebs cycle intermediates in the brain of the epileptic mice [START_REF] Hadera | Triheptanoin partially restores levels of tricarboxylic acid cycle intermediates in the mouse pilocarpine model of epilepsy[END_REF]. Trials in humans shall now evaluate the clinical benefit of triheptanoin on seizure control in idiopathic epilepsy, as well as metabolic causes of epilepsy such as GLUT1-DS. A short-term stabilization of disease progression was also reported in a pilot open-label study conducted in patients affected by adult polyglucosan body disease due to glycogen brancher enzyme deficiency [START_REF] Roe | Adult polyglucosan body disease (APBD): anaplerotic diet therapy (triheptanoin) and demonstration of defective methylation pathways[END_REF], which must be reproduced in a randomized controlled trial.

QUESTIONS TO BE ADDRESSED

Together with the therapeutic developments of triheptanoin, important questions remain to be addressed. If the energy deficiency associated with the diseases mentioned above is prominent enough, one can expect that improving energy homeostasis will slow, or even halt, disease progression. However, such an intervention should probably be initiated at the early stages of the disease when cell loss is not too far advanced. For genetic conditions, ideally the intervention should be started before the onset of symptoms-that is, at the premanifest stage of the disease-which seems ethically acceptable considering the safety profile of triheptanoin in humans. However, the evaluation of the benefit of such intervention will remain challenging in the many neurological diseases where biochemical or clinical biomarkers of energy deficiency are not available.

Although currently not a major issue for patients, one can hope that future pharmaceutical developments will allow the identification of a more convenient formula for triheptanoin than the current liquid oil. This may be particularly useful for long-term compliance, especially in subjects with minimal or no symptoms who lead a normal socioprofessional life. Nonetheless, it is important to keep in mind that the administration of triheptanoin does require dietary adjustments, especially the replacement of a good portion of the regular fat by triheptanoin and some restrictions in sugar consumption.

Last, but not least, further work on the dose regimen of triheptanoin should be performed. In most studies, triheptanoin has been administered to provide about 35% of total caloric intake, in part based on the traditional dose of regular medium-chain triglyceride used for patients with long-chain fatty acid oxidation disorders. A number of publications also provide data on absorption and metabolism of triheptanoin in rodents when administered intravenously and orally at doses up to 40% of the recommended caloric intake [START_REF] Kinman | Parenteral and enteral metabolism of anaplerotic triheptanoin in normal rats[END_REF][START_REF] Gu | Parenteral and enteral metabolism of anaplerotic triheptanoin in normal rats. II. Effects on lipolysis, glucose production, and liver acyl-CoA profile[END_REF]. The low heptanoate and glycerol blood concentrations during intraduodenal infusion of triheptanoin in rats indicated that the glycerol and heptanoate derived from intestinal hydrolysis was taken up and cleared by the liver [START_REF] Kinman | Parenteral and enteral metabolism of anaplerotic triheptanoin in normal rats[END_REF]. Furthermore, doses higher than 40% of total caloric intake can be poorly tolerated in humans with diarrhea and nausea. In GLUT1-DS, while we aimed for a standard dose of 30% to 35% of caloric intake, some patients seemed to do well under lower doses of 25% to 30% (personal unpublished data). Hence, titration studies will be very useful to better determine patients' individual requirements.

Fig. 1 .

 1 Fig.1. Metabolism of triheptanoin. Triheptanoin is split into glycerol and three molecules of heptanoate in the gut. In the liver, glycerol is converted to pyruvate, and heptanoate can be further metabolized to C4-ketone bodies (3-hydroxybutyrate and acetoacetate) and C5ketone bodies (3-hydroxypentanoate and 3-ketopentanoate). In the

ACKNOWLEDGMENTS

The author wishes to thank Jean-Marie Saudubray and Daniel Rabier (pyruvate carboxylase deficiency), Raphael Schiffmann and Emmanuel Roze (GLUT1 deficiency syndrome), and Alexandra Durr and Pierre-Gilles Henry (Huntington disease) for their active collaboration in treating patients with rare diseases. This work would not have been possible without the contribution of enthusiastic clinical research coordinators (C eline Jauffret, Daisy Rinaldi, and Marie-Pierre Luton) and dedicated dietitians (Mary Wallace, Samantha Caillet, and Mariana Atencio). The author is also grateful for the trust and support from patients and their families over the past 14 years.

Supported by grants from Inserm, Ipsen, Ultragenyx Pharmaceuticals, and the program "Investissements d'avenir" ANR-10-IAIHU-06.

CONFLICT OF INTEREST STATEMENT

Dr. Mochel holds a patent on the use of triheptanoin in Huntington disease (WO2008068230) and GLUT1-DS (WO2014093901). Dr Mochel received research support from Ultragenyx and Ipsen, and an honorarium on an advisory board from Ultragenyx.


Chandan Yadav

Marina Deng
email: marina.deng@ims-bordeaux.fr

Magali De Matos

Sebastien Fregonese

Thomas Zimmer

Importance of complete characterization setup on onwafer TRL calibration in sub-THz range

Keywords: Test-Structures, TRL calibration, characterization, EM simulation, HFSS, Sub-THz, SPICE

In this paper, we present the effect of different sub-mm and mm-wave probe geometry and topology on the measurement results of dedicated test-structures calibrated with on-wafer TRL. These results are compared against 3D EM simulation of the intrinsic test-structures. To analyze difference between the measured and intrinsic EM simulation results, onwafer TRL calibration performed on EM simulation results of a dedicated test-structure is also presented.

I. INTRODUCTION

With time, millimeter-wave and sub-millimeter wave are found suitable for diverse applications [START_REF] Crowe | Terahertz RF Electronics and System Integration [Scanning the Issue[END_REF], [START_REF] Tonouchi | Cutting-edge terahertz technology[END_REF] and require extensive research focus from high speed integrated devices to the system level design for full utilization of these frequency bands. To design a system working in the mm-and sub-mm range, development of an accurate SPICE model of the elementary devices is required. In development of a robust and precise SPICE model for the state-of-the-art high speed devices, its validation against device experimental data is indispensable. Accuracy of the used experimental data is essential to extract correct values of the compact model parameters and to use them further for circuit design. To obtain accurate experimental data, design of test-structures, de-embedding/calibration methods along with characterization techniques play a very important role at the sub-THz frequencies [START_REF] Mangan | De-embedding transmission line measurements for accurate modeling of IC designs[END_REF][START_REF] Issaoun | On the Deembedding Issue of CMOS Multigigahertz Measurements[END_REF][START_REF] Manuel | New 3D-TRL structures for on-wafer calibration For high frequency Sparameter measurement[END_REF].

Calibration on Impedance Standard Substrate (ISS)standard combined with Open-Short de-embedding method is an industry de-facto standard but it becomes less accurate above 30 to 40 GHz [START_REF] Potereau | Limitations of On-Wafer Calibration and De-embedding Methods in the Sub-THz Range[END_REF][START_REF] Raya | Méthodes de caractérisation en hautes fréquences des technologies de circuits intégrés en silicium dédiées aux applications Téra hertz et sub-Téra hertz[END_REF]. More advanced de-embedding methods can be adopted to improve the accuracy [START_REF] Pourchon | From measurement to intrinsic device characteristics: Test structures and parasitic determination[END_REF][START_REF] Derrier | State-of-the-art and future perspectives in calibration and de-embedding techniques for characterization of advanced SiGe HBTs featuring sub-THz fT/fMAX[END_REF][START_REF] Potéreau | Meander type transmission line design for on-wafer TRL calibration[END_REF] in cost of the complexity. To reduce reliance from less accurate offwafer calibrations, to improve accuracy and repeatability of the measured data, on-wafer calibration technique enabling accurate characterization of high speed devices in sub-THz frequency range is highly desirable [START_REF] Rumiantsev | Comparison of onwafer multiline TRL and LRM+ calibrations for RF CMOS applications[END_REF]. In this work, we have adopted thru-reflect-line (TRL) method for the on-wafer calibration of device-under-test (DUT), which is the reference for on-wafer measurements at sub-mm wave and mm wave frequencies [START_REF] Williams | Calibration-Kit Design for Millimeter-Wave Silicon Integrated Circuits[END_REF].

II. ON-WAFER CALIBRATION AND VERIFICATION STANDARDS

In order to perform on-wafer TRL calibrations, STMicroelectronics BiCMOS55 (B55) technology is used to design and fabricate all the test-structures [START_REF] Chevalier | A 55 nm triple gate oxide 9 metal layers SiGe BiCMOS technology featuring 320 GHz fT / 370 GHz fMAX HBT and high-Q millimeter-wave passives[END_REF]. The developed test-structures consist of nine metallic layers (copper in M1-M8 + Aluminum layer on top of M8 to realize contact pads) in back-end-of-line (BEOL). Our test-structures are compatible to perform characterization using 50 µm pitch probes and 100 µm pitch probes. In this work, used on-wafer test-structures (see Fig. 1 -Fig. 3) are as follows:

 condition is assigned on the symmetry plane to obtain solution equivalent to complete structure. To perform EM simulation on intrinsic structures, only intrinsic parts of structures are taken into account (e.g. see intrinsic structure of Open-M1 shown in Fig. 4 in comparison to its complete structure shown in Fig. 3).

IV. RESULTS AND DISCUSSION

Using fabricated test-standards on the silicon wafer, first onwafer TRL calibration is performed and the reference plane is -ved. In particular, the combination of Picoprobe with 50 µm pitch and TRL on-wafer calibration seems to be not sufficient for accurate representation of the DUT's intrinsic behavior.

The magnitude and phase of transmission coefficient S21 of a meander-line is shown in Fig. 9 and Fig. 10 up 220 GHz. For this DUT, the different measurement configurations give similar results except in the low frequency range for the Picoprobe with 100 µm pitch. A coupling with neighboring structures may be the origin of this non-ideal behavior. Inductance of the Short-M1 w.r.t. frequency is shown in Fig. 11. From 50 to 110 GHz, the Picoprobe with 100 µm pitch, and from 140-220 GHz, the Cascade-probe with 100 µm pitch, are matching well the theoretically predicted intrinsic DUT characteristic. In order to analyze differences between the intrinsic HFSS simulation results and measured data, we prepared 100 μm pitch probe model in HFSS based on the coplanar + coaxial topology (similar to Picoprobe with 100 µm pitch). Using the probe model, 3D EM simulations with and without neighboring structures are carried out on test-structures shown in Figs. 123. After performing on-wafer TRL calibration on EM simulation data, magnitude of reflection coefficient S11 of Open-M1 is shown in Fig. 12. From Fig. 12, it can be observed that HFSS probe model gives different results than the intrinsic simulation and upto some extend mimic behavior of measurement data of Fig. 7. From Fig. 12, it can also be observed that measurement results are more prone to probe design and presence of neighboring structures around DUTs at higher frequencies. Therefore, effects predicted by calibrated EM simulation results shown in Fig. 12 could be the possible reasons behind difference between the intrinsic simulation and measured data visible in Fig. 7 -Fig. 11. In addition, these results also show importance of the complete characterization setup in the on-wafer TRL calibration.

V. CONCLUSION

In conclusion, we want to point out that considering intrinsic simulation data as a reference, we observe that after on-wafer TRL calibration the measurements of dedicated test-structures using different probes topologies (i.e. based on coplanar and microstrip) give different results. It is interesting to see that after on-wafer TRL calibration the measurements done on the same DUT with same probe topology but with different pitch also shows variation in results as frequency increases. These results show the importance to take into account the complete characterization setup when performing on-wafer TRL calibration. In particular, depending on the design of the teststructures together with the probe geometry, on-wafer TRL calibration is not sufficient for some particular cases and coupling correction becomes mandatory.

Fig 1 :Fig 2 :

 12 Fig 1: 3D view of THRU, OPEN and SHORT standards used in on-wafer TRL calibration are shown in panel (a), (b), and (c), respectively. Length of the THRU standard is 35 µm.

Fig 3 :

 3 Fig 3: 3D view of the Open-M1 where metal-1 (M1) does not have connection with ground as shown in the enlarged view.

Fig 4 :

 4 Fig 4: 3D view of intrinsic structure of Open-M1. Complete structure of Open-M1 including signal and ground pads is shown in Fig. 3.

Fig 5 :

 5 Fig 5: Image of the Picoprobe 50 µm pitch with waveguide used in G-band (140 GHz-220 GHz) measurement. Note that two other Picoprobes are also used in measurement but their images are not shown.

Fig 6 :

 6 Fig 6: Image of the Cascade Infinity probe used in G-band (140 GHz -220 GHz) measurement.

Fig 7 :

 7 Fig 7: Magnitude of reflection coefficient S11 of Open-M1 w.r.t. frequency. Symbols are used to represent the measurement data and line is used for representation of intrinsic simulation performed in HFSS.

Fig 8 :

 8 Fig 8: Phase of S11 of Open-M1 w.r.t. frequency. Symbols are used to represent the measurement data and line is used for representation of intrinsic simulation performed in HFSS.

Fig 9 :

 9 Fig 9: Magnitude of transmission coefficient S21 of meander-line w.r.t. frequency. Symbols are used to represent the measurement data and line is used for representation of intrinsic simulation.

Fig 10 :

 10 Fig 10: Phase of transmission coefficient S21 of Meander-LINE w.r.t. frequency. Symbols are used to represent the measurement data and line is used for representation of intrinsic simulation performed in HFSS.

Fig 11 :

 11 Fig 11: Inductance of the Short-M1 with respect to frequency. Symbols are used to represent the measurement data and line is used for representation of intrinsic simulation performed in HFSS.

Fig 12 :

 12 Fig 12: Comparison of reflection coefficient S11 of Open-M1 w.r.t. frequency. These results are obtained from EM simulation of intrinsic structure and onwafer calibration performed on EM simulation of complete structure. Differences between intrinsic simulation and simulation using probe model has very similar pattern as observed in Fig. 7 between the measured data and intrinsic simulation. Simulation with and without neighboring structure around DUT indicates surrounding condition can impact the measurement.

ACKNOWLEDGMENT

This work is partly funded by the French Nouvelle-Aquitaine Authorities through the SUBTILE and FAST project. The authors also acknowledge financial support from the EU under Project Taranto (No. 737454). The authors would like to thank STMicroelectronics for supplying Silicon wafer.

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