Abstract 4800: Targeting mitochondrial and cytosolic one-carbon metabolism in epithelial ovarian cancer via folate receptor alpha
Authors: Adrianne Wallace-Povirk, Carrie O’Connor, Aamod Dekhne, Zhanjun Hou, Md. Junayed Nayeen, Khushbu Shah, Aleem Gangjee, Larry Matherly
Journal: Cancer Research (2019)

Abstract Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Even though most patients initially respond to platinum-based therapy, the likelihood of disease reoccurrence is virtually 100%. Thus, there is an urgent need for new tumor-selective therapies for EOC. One such treatment option involves targeting tumors via folate receptor alpha (FRα) which is overexpressed in up to 90% of EOCs and shows increasing expression with stage and grade of disease. Our laboratory discovered novel 5-substituted pyrrolo[3,2-d]pyrimidine analogs (AGF291, AGF320, AGF347, AGF359 and AGF362) which inhibit mitochondrial one-carbon (C1) metabolism at serine hydroxymethyltransferase (SHMT) 2, with secondary inhibitions at cytosolic enzyme targets including those in de novo purine biosynthesis and SHMT1. Potent inhibition was seen toward isogenic Chinese hamster ovary (CHO) cell lines individually expressing FRα, the reduced folate carrier (RFC, ubiquitously expressed tissue folate transporter) and the proton-coupled folate transporter (PCFT, expressed in a limited number of normal tissues and inactive at normal pH, and several solid tumors at acidic pH including EOC), and with FRα-expressing tumor cells, including KB and IGROV1, a human EOC cell line. Inhibitory potencies were in order, AGF347 &amp;gt; AGF362 &amp;gt;&amp;gt; AGF291 = AGF320 = AGF359. Drug effects were substantially reduced with excess folic acid, confirming FRα-mediated drug uptake. Toward cisplatin resistant SKOV3, TOV112D and A2780 EOC cells, inhibition in the nanomolar range was detected with all compounds. Excess folic acid abrogated drug effects to varying degrees, suggesting significant uptake by the PCFT and/or the RFC, in addition to FRα. Short-term (5 minutes) cell uptake assays with the EOC cell lines and [3H]AGF347 confirmed transport by PCFT and RFC. With sustained exposures resulting in steady state AGF347 accumulations under both physiologic (pH 7.2) and acidic (pH 6.8, approximating the tumor microenvironment) conditions, FRα uptake predominated. [3H]AGF347 treatment of IGROV1 EOC resulted in substantial drug accumulation in both cytosol and mitochondria. Inhibition of cell proliferation for all analogs was reversed by addition of both glycine and adenosine, implicating C1 metabolism in mitochondria including SHMT2 and de novo purine biosynthesis in the cytosol as the targeted pathways; protection by 5-aminoimidazole-4-carboxamide (AICA) with or without glycine was incomplete, implying direct targeting of AICA ribonucleotide formyltransferase (AICARFTase), the second folate-dependent enzyme in purine biosynthesis. Our studies describe first-in-class FRα-selective compounds targeting mitochondrial and cytosolic C1 metabolism, with potent activity against EOC, including cisplatin resistant EOC. Citation Format: Adrianne Wallace-Povirk, Carrie O’Connor, Aamod Dekhne, Zhanjun Hou, Md. Junayed Nayeen, Khushbu Shah, Aleem Gangjee, Larry Matherly. Targeting mitochondrial and cytosolic one-carbon metabolism in epithelial ovarian cancer via folate receptor alpha [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4800.