Discovery of Potent and Selective Methylenephosphonic Acid CD73 Inhibitors

Abstract

Solid tumors are often associated with high levels of extracellular ATP. Ectonucleotidases catalyze the sequential hydrolysis of ATP to adenosine, which potently suppresses T-cell and NK-cell functions via the adenosine receptors (A2a and A2b). The ectonucleotidase CD73 catalyzes the conversion of AMP to adenosine. Thus, increased CD73 enzymatic activity in the tumor microenvironment is a potential mechanism for tumor immune evasion and has been associated with poor prognosis in the clinic. CD73 inhibition is anticipated to restore immune function by skirting this major mechanism of adenosine generation. We have developed a series of potent and selective methylenephosphonic acid CD73 inhibitors via a structure-based design. Key binding interactions of the known inhibitor adenosine-5′-(α,β-methylene)­diphosphate (AMPCP) with hCD73 provided the foundation for our early designs. The structure–activity relationship study guided by this structure-based design led to the discovery of 4a, which exhibits excellent potency against CD73, exquisite selectivity against related ectonucleotidases, and a favorable pharmacokinetic profile.

Targeting Metabolism of Extracellular Nucleotides via Inhibition of Ectonucleotidases CD73 and CD39

“…Previous X-ray structural data clearly showed that the C6-NH 2 group of 5 was oriented toward a solvent-exposed area, which provided ample opportunity for derivatization. [56][57][58]60,61 Indeed, improved potency was observed upon substitution of the C6 position of the nucleobase with an array of alkyl-and benzylamines, consistent with previous observations by Muller and co-workers 45,72−74 (selected examples are shown in Table 3). Compounds derived from β-fluoroarabinose generally exhibited favorable PK profiles, with long half-lives and low plasma clearance in rats (e.g., 29, CL = 0.021 L h −1 kg −1 ).…”

“…Selected examples are highlighted in Figure 3, including AB680 (6), 56−59 a remarkably potent and selective inhibitor developed by Arcus Biosciences that was the first small-molecule CD73 inhibitor to enter clinical trials. A variety of nucleoside-derived compounds have been reported by researchers across academia and industry, including by Muller and co-workers (e.g., 7− 10), 45,[72][73][74][75][76]80 Peloton Therapeutics (11), 66 Arcus Biosciences (12), 60,61 ORIC Pharmaceuticals (13), 64,65 Calithera Biosciences ( 14), 52 and Vitae Pharmaceuticals/Boehringer Ingelheim (15), 54 among others. 67 Guided by the X-ray cocrystal structure of 5 and CD73, Muller and co-workers designed and synthesized a wide variety of AMPCP analogs with improved potency and metabolic stability, including C2-chloro analog 7 (Figure 3) 72,76 and PSB-12379 (8), which bears a benzylamine substituent at C6 of the nucleobase and displayed a K i of 2.21 ± 0.40 nM vs hCD73.…”

“…56 Initially, they envisioned replacing the highly acidic PCP group of AMPCP-based inhibitors, as its physicochemical properties presented challenges for achieving oral bioavailability. While selected monophosphonic acid analogs, such as 12 (Figure 3), 60,61 did retain nanomolar potency, most attempts to replace the PCP group with less acidic moieties led to a loss of potency against CD73. 57,58 The apparent necessity of the highly ionizable PCP group, coupled with a dearth of viable prodrug strategies, directed their ensuing discovery program toward the development of a molecule that would be suitable for intravenous (iv) administration, i.e., a potent compound with excellent pharmacokinetic (PK) properties, including very low clearance (CL) and a long circulating half-life (t 1/2 ).…”

“…Monophosphonic Acid CD73 Inhibitors. Researchers at multiple organizations, including Arcus Biosciences, 55,60,61 Vitae Pharmaceuticals, 54 and ORIC Pharmaceuticals, 63−65 have evaluated nucleoside-derived methylene monophosphonic acids as CD73 inhibitors. Studies described by Arcus Biosciences showed that the SAR within this scaffold is in close alignment with that of bisphosphonic acid inhibitors, such as AB680.…”

A Novel CD73 Inhibitor SHR170008 Suppresses Adenosine in Tumor and Enhances Anti-Tumor Activity with PD-1 Blockade in a Mouse Model of Breast Cancer

“… 25 The crystal structure of CD73 in complex with AMPCP (accession code: 4H2I) and small-molecule antagonist (accession code: 6YE2) were downloaded from PDB. 26 , 27 The co-crystal ligand (accession code: 6YE2) was modified to SHR170008. By selecting SHR170008 and the surrounding residues within 4.5Å, iterative energy minimizations with gradually increased tether deviations were performed to relax the clash between the ligand and protein while forming multiple interactions between ligand and protein.…”