(T1030-04-27) Identification of FDA-Approved Drugs Targeting Malonyl Co-A Decarboxylase for the Treatment of Obesity

Purpose: Obesity is one of the challenging diseases associated with the development of cardiometabolic disorders. Hyperglycemia and insulin resistance are common complications associated with obesity that occur due to the elevation of circulating fatty acid and reduction in glucose utilization in different organs. Malonyl Co-A is regulated in fatty acid synthesis produced by acetyl Co-A carboxylase. Consequently, malonyl Co-A will be metabolized by fatty acid synthase (FAS) and incorporated into long-chain fatty acids. Additionally, malonyl Co-A decarboxylase (MCD) catalyzes the decarboxylation of malonyl Co-A to produce acetyl Co-A, which stimulates the mitochondrial uptake of free fatty acids for β-oxidation. This will relieve the malonyl-CoA-mediated inhibition of carnitine palmitoyl transferase (CPT1), the rate-limiting enzyme in fatty acid oxidation. Additionally, Malonyl Co-A has a role in the regulation of food intake through its actions in the central nervous system and signaling satiety. Several cellular and pre-clinical studies showed that inhibition of MCD can inhibit fatty acid oxidation through malonyl Co-A inhibitory effect for CPT 1. This will shift energy metabolism in cardiometabolic disorders in addition to the reduction of food intake so could be an approach for treating obesity and associated complications. However, up till now, there are no FDA-approved MCD inhibitors.

Purpose: Using a structure-based drug repurposing approach to identify FDA-approved drug(s) targeting MCD covalently to alleviate obesity and associated complications. In addition to, In-vivo phenotypic screening for the top candidates against obesity, hyperglycemia, and insulin resistance.

Methods: We applied a structure-based high throughput virtual screening to identify FDA-approved drug(s) to target MCD covalently. This was coupled with biochemical validation of the top hits along with intact mass spectroscopy and thermal shift assay for human MCD protein. Eventually, our lead compound(s) were tested in vivo for diet-induced obesity mouse models. This was followed by an assessment of energy expenditure, food intake, body composition, lipid profiling, oral glucose, and insulin test. Further mechanistic investigations were carried out on different isolated tissues using western blot analysis and LC/MS

Results: Our in silico molecular modeling experiments revealed potential candidates targeting the MCD catalytic dyad of Ser329 and His423 via hydrogen bonds and hydrophobic-hydrophobic interactions covalently. The top 20 candidates were selected for biochemical validation for their activity against MCD at 10µM. This was followed by IC50 profiling for 2 promising MCD inhibitors Comp. (A) and Comp. (B) that showed IC50 at 5.20 ± 0.70 µM and 2.07 ± 0.34 µM, respectively. Our intact mass spectroscopic analysis revealed that when either comp. (A) or comp. (B) incubated with MCD were nearly completely covalently bound. Thermal shift analysis showed that adding comp. (A) to MCD resulting in a ΔTm (melting temperature) shift of 12.75 ^oC compared to DMSO control. Comp. (A) was administrated subcutaneously at the dose of 10 mg/kg (equivalent to the human daily dose) in a diet-induced obesity mice model for 30 days. That led to a significant decrease in body weight, fat-to-lean ratio, blood glucose, and insulin levels in the treated group compared to the control group receiving vehicle.

Conclusion: Our study offers the first-in-class FDA-approved covalent MCD inhibitors to alleviate obesity and associated complications.

References: Al Batran, R. et al.,. (2020). Cell Metabolism, 31(5), 909-919.
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Wang, W. et al., (2019). JACC: Basic to Translational Science, 4(3), 385-400.

Acknowledgements:This work is supported by start-up funds from the HSC research office and SOP. We thank Openeye for a free academic license

Figure 1. Identification of FDA-approved drugs targeting human malonyl Co-A decarboxylase covalently. (A) Crystal structure of hMCD (PDB ID:2YGW) highlighting the catalytic dyad of Ser329 and His423. (B) Binding Modes of Comp. (A) and Comp. (B) to the catalytic binding domain of MCD. (C) Biochemical evaluation of the top 20 hits against hMCD at 10 µM. (D) IC50 profiling of comp. A and B against hMCD. (E) Intact mass spectroscopy assessment for the covalent binding of Comp. A and B with hMCD. (F) Thermal shift assay for the binding affinity of comp. A with hMCD.

Figure 2. Representative experiments for in-vivo evaluation of Comp. (A) in the diet-induced obesity mouse model. (A) Schematic for in vivo administration of Comp. (A) in Diet-induced obesity mouse model. (B) body weight measurement over 30 days of administration of vehicle and comp. A subcutaneous at 2 dose levels for high fat diet-induced obesity mouse model. (C) Blood glucose levels for the vehicle, comp. A (5 mg/kg), and comp. A (10 mg/kg) in the high fat diet-induced obesity mouse model.