The phosphonate group is a key pharmacophore in many antiviral, antimicrobial, and antineoplastic drugs. Due to its high polarity and short retention time, detecting and quantifying such phosphonate-containing drugs with LC/MS-based methods are challenging and require derivatization with hazardous reagents. Given the emerging importance of phosphonate-containing drugs, developing a practical, accessible, and safe method for their quantitation in pharmacokinetics (PK) studies is desirable. NMR-based methods are often employed in drug discovery but are seldom used for compound quantitation in PK studies. Here, we show that proton− phosphorous ( 1 H− 31 P) heteronuclear single quantum correlation (HSQC) NMR allows for the quantitation of the phosphonate-containing enolase inhibitor HEX in plasma and tissues at micromolar concentrations. Although mice were shown to rapidly clear HEX from circulation (over 95% in <1 h), the plasma half-life of HEX was more than 1 h in rats and nonhuman primates. This slower clearance rate affords a significantly higher exposure of HEX in rat models compared to that in mouse models while maintaining a favorable safety profile. Similar results were observed for the phosphonate-containing antibiotic, fosfomycin. Our study demonstrates the applicability of the 1 H− 31 P HSQC method to quantify phosphonate-containing drugs in complex biological samples and illustrates an important limitation of mice as preclinical model species for phosphonate-containing drugs.

A comparison of i.v., i.m. and s.c. administration erythromycin base in polyethylene glycol at 15 mg/kg and 30 mg/kg body weight was carried out in beef-type calves of approximately 200 kg body weight. Additional evaluations were carried out with oral administration of erythromycin phosphate and erythromycin stearate. Absorption of erythromycin was very slow by both the i.m. and s.c. routes of administration with a Kab of 0.0135 min-1 and 0.0185 min-1 for i.m. and 0.0032 min-1 and 0.0074 min-1 for s.c. at 15 mg/kg and 30 mg/kg, respectively. The bioavailability (32-42%) and peak serum concentrations were much lower with s.c. than with i.m. (60-65%) administration. The disposition of erythromycin administered i.v. appeared to be representative of dose-dependent kinetics rather than dose-independent first-order kinetics inasmuch as the elimination half-time (t1/2B) increased from 174.5 +/- 13 min for the 15 mg/kg dosage to 239 +/- 10.8 min with 30 mg/kg dosage. An acute apparent cardiovascular effect accompanied i.v. administration of erythromycin at 30 mg/kg dosage but not at 15 mg/kg. Severe diarrhea followed oral administration of either erythromycin phosphate or erythromycin stearate.

The phosphonate group is a key pharmacophore in many anti-viral, anti-microbial, and anti-neoplastic drugs. Due to its high polarity and short retention time, detecting and quantifying such phosphonate-containing drugs with LC/MS-based methods is challenging and requires derivatization with hazardous reagents. Given the emerging importance of phosphonate-containing drugs, developing a practical, accessible, and safe method for their quantitation in pharmacokinetics (PK) studies is desirable. NMR-based methods are often employed in drug discovery but are seldom used for compound quantitation in PK studies. Here, we show that proton-phosphorous (1H-31P) heteronuclear single quantum correlation (HSQC) NMR allows for quantitation of the phosphonate-containing enolase inhibitor HEX in plasma and tissue at micromolar concentrations. Although mice were shown to rapidly clear HEX from circulation (over 95% in <1 hr), the plasma half-life of HEX was more than 1hr in rats and nonhuman primates. This slower clearance rate affords a significantly higher exposure of HEX in rat models compared to mouse models while maintaining a favorable safety profile. Similar results were observed for the phosphonate-containing antibiotic, fosfomycin. Our study demonstrates the applicability of the 1H-31P HSQC method to quantify phosphonate-containing drugs in complex biological samples and illustrates an important limitation of mice as preclinical model species for phosphonate-containing drugs.

Homozygous deletion of MTAP occurs in about 15% of all human cancers, such as glioblastoma, pancreatic cancer, mesothelioma, urothelial bladder carcinoma, and lung squamous cell carcinoma. PRMT5 inhibitors show activity against MTAP-deleted cancer cells in culture and xenografts with a mechanism that relies on the significant elevation of the MTAP substrate, methylthioadenosine (MTA). Previously, we have shown that unlike cells in culture, MTA levels in MTAP-deleted primary human GBM tumors are not significantly higher than in MTAP-intact tumors. Therefore, combining the PRMT5 inhibitor with another drug may be required to increase the therapeutic window and clinical efficacy of a PRMT5 inhibitor in MTAP-deleted patients. Here, we identified a natural small molecular chemical compound with a good safety profile that synergizes with a PRMT5-MTA complex inhibitor, which boosts the efficacy of MTAP-deleted selective cell killing in the presence of MTA sequestering cells. This combination therapy significantly increases the potency of PRMT5 inhibitor treatment in MTAP-deleted cells across various tumor cell lines and lowers the IC50 of PRMT5 inhibitor treatment. In vivo, PRMT5 inhibitor combination treatment leads to smaller tumor volumes in MTAP-deleted CDX tumors (U87, glioma cell line) compared to PRMT5 inhibitor monotherapy. In summary, our proposed combination therapy of PRMT5 inhibition with a natural compound may increase the therapeutic window and clinical efficacy of PRMT5 inhibitors leading to better treatment options for patients harboring MTAP-deleted cancer. Citation Format: Yasaman Barekatain, Kyle LaBella, Hikaru Sugimoto, Kristen Harris, Sunada Khadka, Florian Muller, Raghu Kalluri. PRMT5 inhibition synergizes with a natural small molecule compound to kill MTAP-deleted cells and suppress tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1073.