Next-Generation Cell-Active Inhibitors of the Undrugged Oncogenic PTP4A3 Phosphatase

Area of Science:

• Biochemistry and Molecular Biology
• Cancer Research
• Medicinal Chemistry

Background:

• Oncogenic protein tyrosine phosphatases (PTPs) are frequently overexpressed in human cancers, presenting significant therapeutic challenges.
• The PTP4A phosphatase family plays a crucial role in fundamental malignant processes, making it a key target for cancer drug development.
• Existing PTP inhibitors face challenges, necessitating the exploration of novel chemotypes and mechanisms of action.

Purpose of the Study:

• To elucidate the mechanism of action of the novel PTP4A inhibitor JMS-053 and investigate its chemotype, with a focus on the role of oxidation.
• To synthesize and evaluate new analogs of JMS-053 for improved specificity and efficacy against PTP4A phosphatases.
• To assess the cytotoxic effects of these inhibitors on relevant cancer cell models and determine their dependence on PTP4A expression.

Main Methods:

• Synthesis and characterization of JMS-053 analogs, including 7-imino-5-methyl-2-phenylthieno[3,2-c]pyridine-4,6(5H,7H)-dione (NRT-870-59).
• In vitro inhibition assays using PTP4A3 and its mutants, as well as CDC25B, to determine specificity.
• Cytotoxicity assays using three-dimensional spheroid cultures of human ovarian and breast cancer cells, and colony formation assays.
• Assessment of reactive oxygen species (ROS) generation using in vitro and cellular assays.
• Mass spectrometry to detect potential oxidation of the catalytic cysteine residue.
• Gene expression profiling to analyze cellular responses to inhibitor treatment.

Main Results:

• JMS-053 and its analogs demonstrated potent in vitro inhibition of PTP4A3.
• NRT-870-59 exhibited unique specificity for PTP4A3 over a PTP4A3 mutant and CDC25B, acting as a reversible inhibitor.
• All tested thienopyridinediones showed cytotoxicity against ovarian and breast cancer spheroids, with inhibition of colony formation dependent on PTP4A3 expression.
• JMS-053 and NRT-870-59 did not generate significant ROS in vitro or in cancer cells, and mass spectrometry confirmed no oxidation of PTP4A3's catalytic cysteine.
• Gene expression profiling indicated that JMS-053 treatment phenocopied PTP4A3 loss without inducing oxidative stress.

Conclusions:

• Selective small molecule inhibition of PTP4A phosphatases is achievable with the thienopyridinedione chemotype.
• These novel inhibitors are effective against ovarian and breast cancer cells and do not rely on ROS generation, suggesting a favorable safety profile.
• The findings support the further investigation of this class of reversible PTP4A inhibitors as potential lead compounds for cancer therapy.