ujjwal suwal

High-grade serous ovarian cancer (HGSOC) is deadliest of gynecological malignancies. Ovarian tumor cells can directly spread throughout the peritoneal space as single cells that grow into tumorspheres concomitant with the generation of malignant ascites that contains growth factors and extracellular matrix (ECM) proteins. Greater than 70% of advanced stage HGSOC patients exhibit gains or amplifications for the Focal Adhesion Kinase (FAK) gene, a tyrosine kinase that is activated by ECM proteins. In other solid tumors, changes in ECM and associated proteins (termed the Matrisome) are generated in part by tumor associated fibroblasts. Herein we compared transcriptional (RNA sequencing) and proteomic (mass spectrometry) changes using two ascites generating models: MOVCAR, a SV40 T antigen under control of the Mullerian inhibitory substance type II receptor; and KMF, a spontaneously aggressive ID8 derivative containing KRas, Myc, and FAK gene amplifications. FAK inactivation was generated by crossing MOVCAR with FAK floxed mice or CRISPR to disrupt FAK expression in KMF. Comparisons were made with FAK-null or cells re-expressing FAK wildtype or a FAK kinase-inactive point mutant (K454R) by lentiviral transduction with tumor cells grown as spheroids in vitro or harvested after intraperitoneal tumor growth in mice. FAK expression and activity were required for optimal tumor growth in mice and a common set of differentially expressed genes were identified upon FAK loss and re-expression. Gene ontology analyses revealed significant differences in Matrisome, morphogenesis, and epithelial proliferation within spheroids and tumors lacking FAK. Parallel protein mass spectrometry analyses revealed that thirty-four Matrisome proteins were down regulated (> log2 fold change) including collagens -1, -3 and -5, fibulin-1 and -2, elastin, tenascin, and fibronectin. Several of these proteins were regulated by FAK expression and not necessarily intrinsic FAK activity. Bioinformatic analysis of the TCGA ovarian tumor database showed that FAK, collagen-3 and collagen-5 expression negatively correlate with patient survival. As FAK is highly tyrosine phosphorylated in tumor spheroids isolated from patients and from our mouse models, our results support the hypothesis that elevated tumor-intrinsic FAK expression results in increased ECM production that can feed-forward to drive increased FAK activity. These results may impact the clinical testing of small molecule inhibitors of FAK activity. Citation Format: Marjaana Ojalill, Joanna Zhang, Ujjwal Suwal, Pekka Rappu, Jyrki Heino, Denise C. Connolly, Dwayne G. Stupack, David D. Schlaepfer. Ovarian tumor-intrinsic FAK-dependent regulation of the matrisome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2363.

High-grade serous ovarian cancer (HGSOC) is deadliest of gynecological malignancies. Ovarian tumor cells can directly spread throughout the peritoneal space as single cells that grow into tumorspheres concomitant with the generation of malignant ascites that contains growth factors and extracellular matrix (ECM) proteins. Greater than 70% of advanced stage HGSOC patients exhibit gains or amplifications for the Focal Adhesion Kinase (FAK) gene, a tyrosine kinase that is activated by ECM proteins. In other solid tumors, changes in ECM and associated proteins (termed the Matrisome) are generated in part by tumor associated fibroblasts. Herein we compared transcriptional (RNA sequencing) and proteomic (mass spectrometry) changes using two ascites generating models: MOVCAR, a SV40 T antigen under control of the Mullerian inhibitory substance type II receptor; and KMF, a spontaneously aggressive ID8 derivative containing KRas, Myc, and FAK gene amplifications. FAK inactivation was generat...

IntroductionProtein arginine deiminases (PADs) are intracellular enzymes that may, especially in pathological conditions, also citrullinate extracellular substrates, including matrisome proteins such as structural proteins in extracellular matrix (ECM). PADs are abundantly expressed in human cancer cells. Citrullination of matrisome proteins has been reported in colon cancer but the phenomenon has never been systematically studied.MethodsTo gain a broader view of citrullination of matrisome proteins in cancer, we analyzed cancer proteomics data sets in 3 public databases for citrullinated matrisome proteins. In addition, we used three-dimensional cell cocultures of fibroblasts and cancer cells and analyzed citrullination of ECM.Results and discussionOur new analysis indicate that citrullination of ECM occurs in human cancer, and there is a significant variation between tumors. Most frequently citrullinated proteins included fibrinogen and fibronectin, which are typically citrullinat...