Sensitizing Human Cancer Cells Through the Inhibition of NSMCE2

Abstract

Homologous recombination (HR) is a high-fidelity DNA repair pathway that is activated mainly by replication-associated DNA damage. DNA damaging agents that cause double strand breaks (DSBs) at the replication fork can be repaired by HR. NSMCE2, an E3 sumo ligase, regulates HR at damaged replication forks. NSMCE2, complexed with the structural maintenance of chromosomes (SMC) 5/6 complex, stimulates the SUMOylation of the SMC6 coiled-coil region, thus mediating HR. Cancers that have defects in their HR machinery are susceptible to chemotherapies that elicit DNA damage, and in particular, damage that affects DNA replication. Targeting regulators of HR could therefore sensitize HR-proficient cancers to standard therapies. We used in-silico protein modeling to generate a 3D model of the human NSMCE2 protein. We identified a deep pocket in NSMCE2 that is adjacent to an active-site zinc-binding region and twelve compounds that potentially could occupy that pocket. We sought to determine whether any of these compounds could inhibit NSMCE2 activity or function. We screened the compounds in the human osteosarcoma cell line (U2OS) for synergistic inhibition of cellular proliferation by the topoisomerase 1 inhibitor, camptothecin, and identified four that caused no cellular toxicity on their own whereas they were synergistic with camptothecin in limiting cell proliferation. One of the compounds, compound 3, showed a twelve-fold increase in inhibition of cellular proliferation in combination with camptothecin. We also were able to determine compound specificity to NSMCE2 using a mutation in HEK293T cells. We concluded using small molecule inhibitors of NSMCE2 could be useful in enhancing chemotherapy-mediated killing of HR-proficient cancer cells. Development of NSMCE2 inhibitors could also lead to adjuvant therapies that would allow for patients to receive lower doses of toxic topoisomerase poisons.