Principal Investigator: Andrew Yen

Co-PI: Praveen Sethupathy

DESCRIPTION (provided by applicant): 

The goal of the proposed studies is to extend the in vitro work of the previous granting cycle to in vivo studies. The FCF invited us to submit a proposal for a small extension of the completed grant. In the previous cycle we demonstrated that retinoic acid (RA) can inhibit the growth of FLC cells as well as ameliorate certain cellular, as well as molecular, traits typically associated with cancer transformation. In particular, we found that RA caused growth inhibition. The inhibition did not reflect apoptosis, but rather retardation of proliferation as measured by EdU incorporation during DNA replication. The cells emerging after RA treatment exhibited a different morphology; specifically, they are more substrate adherent. Consistent with acquiring anchorage dependence, the RA treated FLC cells grew in soft agar with fewer and smaller colonies. Cell cycle analysis indicated a general dilation of the cell cycle without classical G1/0 specificity. Consistent with this overall slowdown, ATP levels were attenuated in RA treated FLC cells. RA did not consistently or dramatically reduce DP expression at the RNA or protein levels. RNA-seq analysis revealed that other classic markers of FLC, including OAT, VCAN, and LINC00473, are significantly suppressed after RA treatment. Moreover, pathway analysis showed that substrates of PKA are among the most dramatically reduced after RA treatment. It is also notable that RA treatment primed FLC cells for a much more robust response to the pro-apoptotic compound, polyI:C. This result suggests that combination treatments of RA with other chemotherapeutic agents could be an attractive approach for FLC. Proteomics expression signatures comparing FLC tumor and non-transformed proximal cells showed prominent downregulation of vitamin A metabolizing molecules indicating a defect in synthesis or metabolism of RA. This suggests that pharmacological RA doses overcome this metabolic pathology seminal to disease and might be exploitable as a novel therapeutic modality. These studies used a PDX derived FLC cell line. We seek to extend this study to determine if the in vitro demonstration of principle would be borne out in vivo using PDX mice treated with retinoic acid. A limited trial is proposed that if successful would lead us to propose more comprehensive studies involving molecular mechanistic as well as more extensive animal trials.