UC Riverside

Proliferating cell nuclear antigen (PCNA) acts as a vital scaffold to mediate DNA metabolism and cell survival mechanisms. The identification by our collaborators at the City of Hope medical center of a cancer associated isoform of PCNA, caPCNA, provided a means for the development of anti-cancer therapeutics that are selective to cancer cells. Our collaborators also developed a selective caPCNA inhibitor, AOH1160, that induced cancer cell apoptosis at nanomolar concentrations in cellulo and in mouse in vivo models, without effecting non-malignant cells. Yet, this AOH1160 molecule was not metabolically stable, and required further optimization to develop a more drug-like lead. We hypothesized that structural-based drug design (SBDD) techniques could be used to characterize the AOH1160 scaffold binding interactions at atomic resolutions and help guide future caPCNA inhibitor design. Thus far, we confirmed that the city of Hope AOH1160 series of compounds bind to recombinant PCNA protein. Furthermore, through our discovery of novel recombinant PCNA crystallization conditions, we have determined three co-crystal structures of caPCNA:AOH1160 analogs. This includes the recently developed analog AOH1996, which has superior metabolic stability as compared to the AOH1160 parent molecule, while maintaining its potency, and it has now entered Phase-1 clinical trials for breast cancer therapy. However, there is a ~90% failure rate of novel chemical entities heading into the clinic, with solid tumors being a particularly challenging area for drug discovery and development. Thus, we are continuing to investigate analogs around this scaffold, as well as aiming to discover and develop PIP-box selective inhibitors that are in different areas of chemical space. Our binding and structural studies have identified a novel PIP-box binding molecule, B05, that has approximately 20 times greater potency in cellulo as compared to the AOH1996 scaffold. In addition, we have identified a pool of promising fragment binders through our rapid binding assays to funnel into fragment-based lead design (FBLD) studies that aim to identify advantageous chemistries to be utilize in lead design. A combination of in silico docking and with fragment screening led to our discovery of new caPCNA binders with novel chemical scaffolds, which we are exploring further. Also, we have recently identified novel apo-PCNA crystallization conditions that diffract to much higher resolutions than previously, which likely allows for the unambiguous characterization of fragment binders of interest by X-ray crystallography. Overall, we provided the first determination that the City of Hope AOH series of compounds are PCNA binders, and our successful structural determinations of AOH1160 analogs, together with our new B05 analog, are currently enabling a new generation of caPCNA inhibitors to be developed with the goal of enhanced clinical utility.