Small-Molecule Targeting of Proliferating Cell Nuclear Antigen Chromatin Association Inhibits Tumor Cell Growth

Abstract

Proliferating cell nuclear antigen (PCNA), a potential anticancer target, forms a homotrimer and is required for DNA replication and numerous other cellular processes. The purpose of this study was to identify novel small molecules that modulate PCNA activity to affect tumor cell proliferation. An in silico screen of a compound library against a crystal structure of PCNA and a subsequent structural similarity search of the ZINC chemical database were carried out to derive relevant docking partners. Nine compounds, termed PCNA inhibitors (PCNA-Is), were selected for further characterization. PCNA-I1 selectively bound to PCNA trimers with a dissociation constant (K d ) of ϳ0.2 to 0.4 M. PCNA-Is promoted the formation of SDSrefractory PCNA trimers. PCNA-I1 dose-and time-dependently reduced the chromatin-associated PCNA in cells. Consistent with its effects on PCNA trimer stabilization, PCNA-I1 inhibited the growth of tumor cells of various tissue types with an IC 50 of ϳ0.2 M, whereas it affected the growth of nontransformed cells at significantly higher concentrations (IC 50 , ϳ1.6 M). Moreover, uptake of BrdU was dose-dependently reduced in cells treated with PCNA-I1. Mechanistically the PCNA-Is mimicked the effect of PCNA knockdown by siRNA, inducing cancer cell arrest at both the S and G 2 /M phases. Thus, we have identified a class of compounds that can directly bind to PCNA, stabilize PCNA trimers, reduce PCNA association with chromatin, and inhibit tumor cell growth by inducing a cell cycle arrest. They are valuable tools in studying PCNA function and may be useful for future PCNA-targeted cancer therapy.

Deviating the level of proliferating cell nuclear antigen inTrypanosoma bruceielicits distinct mechanisms for inhibiting proliferation and cell cycle progression

“…50 This study demonstrates that depleting TbPCNA by RNAi prevented G2 cell cycle progression in T. brucei. Such a phenotype is consistent with the S and G2/M phase arrest observed after PCNA was depleted in human cell by siRNA 51,52 and demonstrates an evolutionarily conserved function of PCNA.…”

“…This has fundamentally restricted the types of studies for chemotherapeutic interventions of PCNA to human proliferative diseases such as cancer, arthritis and nephritis. 52,[56][57][58][59] The results of this study validate TbPCNA as a viable target for therapeutic intervention against African trypanosomiasis and broaden the categories of where PCNA therapeutics may be beneficial to infectious disease research. Future studies will elucidate mechanisms of TbPCNA regulation in T. brucei.…”

“…Information gained from such studies can also reveal how deregulating TbPCNA triggers G2/M arrest. This information in combination with that obtained from several novel classes of PCNA inhibitors 52,59 may lead to the development of innovative therapies that target components of the T. brucei DNA replication machinery.…”

Antitumor Effects of a Novel Small Molecule Targeting PCNA Chromatin Association in Prostate Cancer

“…Previously, we performed an in silico screen of a compound library against a crystal structure of human PCNA and functional assays, these studies led to identification of nine compounds named as PCNA-inhibitors (PCNA-Is). These PCNA-Is bind directly to PCNA trimers, stabilize PCNA homotrimers structure, reduce PCNA association with chromatin, and attenuate DNA replication, and selectively inhibit growth of tumor cells of various tissue origins with IC 50 values in the nanomolar range (29). Of those nine compounds, PCNA-I1 was the most potent.…”

Selective inhibition T2AA for human PCNA over PCNA homolog in Trypanosoma brucei

“…The PIP-box is represented by the consensus sequence (Qxxx) where = (hydrophobic residues) and = (aromatic residues) that determines how well the motif can pack into the hydrophobic pocket formed by the interdomain-connecting loop and central loop of PCNA (Bruning and Shamoo, 2004;Gulbis et al, 1996;Moldovan et al, 2007). Basic knowledge about PCNA/PIP-box interactions has translated to the pre-clinical proof-of-mechanism for the inhibitor T2AA to therapeutically target HsPCNA in cancer cells (Actis et al, 2013;Inoue et al, 2014;Punchihewa et al, 2012;Tan et al, 2012). There is a homolog of PCNA in T. brucei, TbPCNA, which is regulated through the cell cycle, and which is located at the nuclear periphery (Kaufmann et al, 2012).…”