Identification and characterization of therapeutic molecules affecting expression levels of the tumor suppressor DAB2IP in cancer

In tumors, the reciprocal communication between malignant cells and non-transformed stromal cells involves a variety of signaling proteins and modulators that cooperate to control proliferation, migration and apoptosis. Among them, the tumor suppressor DAB2IP, a Ras-GAP and signaling adaptor protein, modulates signal transduction in response to several extracellular stimuli, negatively regulating multiple oncogenic pathways. Accordingly, the loss of DAB2IP in tumor cells fosters metastasis and enhances chemo- and radio-resistance. DAB2IP is rarely mutated in cancer but is frequently downregulated or inactivated by multiple mechanisms. Solid experimental evidences indicate that DAB2IP reactivation can reduce cancer aggressiveness in tumors driven by multiple different oncogenic mutations. In this regard, we showed that the ectopic overexpression of DAB2IP is sufficient to significantly affect the behavior of prostate cancer cells, possibly slowing tumor dissemination. All these evidences indicate DAB2IP as a strong target for anti-cancer therapy. Nevertheless, therapeutic approaches to increase DAB2IP function in cancer are still not available. Based on these observations, we performed a high-throughput screening with more than 1200 FDA- approved drugs to search for molecules that increase DAB2IP protein levels. Since detection of endogenous DAB2IP is technically difficult due to relatively low expression levels and the limitations of available antibodies, we exploited CRISPR/Cas9 gene editing to generate two prostate cancer cell models expressing endogenous DAB2IP fused to HiBiT, a peptide tag that enabled luminescence- based detection of protein levels in a sensitive and quantitative manner. Using this approach, we identified a set of candidate drugs able to increase DAB2IP levels. We focused our attention on the three more effective drugs: one antibacterial, one antileukemic and one antiasthmatic. Although not conclusive, functional experiments indicate that DAB2IP-upregulating drugs can inhibit some cancer-associated phenotypes, and that some of these effects are at least in part dependent on DAB2IP. These findings, if further confirmed, may suggest a potential repurposing of these drugs for solid cancers’ treatment, as support to current therapies.