Victoria Mcguinness Rimkunas

Survey of Tyrosine Kinase Signaling Reveals ROS Kinase Fusions in Human Cholangiocarcinoma

Cholangiocarcinoma, also known as bile duct cancer, is the second most common primary hepatic carcinoma with a median survival of less than 2 years. The molecular mechanisms underlying the development of this disease are not clear. To survey activated tyrosine kinases signaling in cholangiocarcinoma, we employed immunoaffinity profiling coupled to mass spectrometry and identified DDR1, EPHA2, EGFR, and ROS tyrosine kinases, along with over 1,000 tyrosine phosphorylation sites from about 750 different proteins in primary cholangiocarcinoma patients. Furthermore, we confirmed the presence of ROS kinase fusions in 8.7% (2 out of 23) of cholangiocarcinoma patients. Expression of the ROS fusions in 3T3 cells confers transforming ability both in vitro and in vivo, and is responsive to its kinase inhibitor. Our data demonstrate that ROS kinase is a promising candidate for a therapeutic target and for a diagnostic molecular marker in cholangiocarcinoma. The identification of ROS tyrosine kinase fusions in cholangiocarcinoma, along with the presence of other ROS kinase fusions in lung cancer and glioblastoma, suggests that a more broadly based screen for activated ROS kinase in cancer is warranted.

Analysis of Receptor Tyrosine Kinase ROS1-Positive Tumors in Non–Small Cell Lung Cancer: Identification of a FIG-ROS1 Fusion

Purpose: To deepen our understanding of mutant ROS1 expression, localization, and frequency in non–small cell lung cancer (NSCLC), we developed a highly specific and sensitive immunohistochemistry (IHC)-based assay that is useful for the detection of wild-type and mutant ROS1. Experimental Design: We analyzed 556 tumors with the ROS1 D4D6 rabbit monoclonal antibody IHC assay to assess ROS1 expression levels and localization. A subset of tumors was analyzed by FISH to determine the percentage of these tumors harboring ROS1 translocations. Using specific and sensitive IHC assays, we analyzed the expression of anaplastic lymphoma kinase (ALK), EGFR L858R, and EGFR E746-A750del mutations in a subset of lung tumors, including those expressing ROS1. Results: In our NSCLC cohort of Chinese patients, we identified 9 (1.6%) tumors expressing ROS1 and 22 (4.0%) tumors expressing ALK. FISH identified tumors with ALK or ROS1 rearrangements, and IHC alone was capable of detecting all cases with ALK and ROS1 rearrangements. ROS1 fusion partners were determined by reverse transcriptase PCR identifying CD74-ROS1, SLC34A2-ROS1, and FIG-ROS1 fusions. Some of the ALK and ROS1 rearranged tumors may also harbor coexisting EGFR mutations. Conclusions: NSCLC tumors with ROS1 rearrangements are uncommon in the Chinese population and represent a distinct entity of carcinomas. The ROS1 IHC assay described here is a valuable tool for identifying patients expressing mutant ROS1 and could be routinely applied in clinical practice to detect lung cancers that may be responsive to targeted therapies. Clin Cancer Res; 18(16); 4449–57. ©2012 AACR.

Dual Inhibition of IGF-1R and ErbB3 Enhances the Activity of Gemcitabine and Nab-Paclitaxel in Preclinical Models of Pancreatic Cancer

Purpose: Insulin-like growth factor receptor 1 (IGF-1R) is critically involved in pancreatic cancer pathophysiology, promoting cancer cell survival and therapeutic resistance. Assessment of IGF-1R inhibitors in combination with standard-of-care chemotherapy, however, failed to demonstrate significant clinical benefit. The aim of this work is to unravel mechanisms of resistance to IGF-1R inhibition in pancreatic cancer and develop novel strategies to improve the activity of standard-of-care therapies. Experimental Design: Growth factor screening in pancreatic cancer cell lines was performed to identify activators of prosurvival PI3K/AKT signaling. The prevalence of activating growth factors and their receptors was assessed in pancreatic cancer patient samples. Effects of a bispecific IGF-1R and ErbB3 targeting antibody on receptor expression, signaling, cancer cell viability and apoptosis, spheroid growth, and in vivo chemotherapy activity in pancreatic cancer xenograft models were determined. Results: Growth factor screening in pancreatic cancer cells revealed insulin-like growth factor 1 (IGF-1) and heregulin (HRG) as the most potent AKT activators. Both growth factors reduced pancreatic cancer cell sensitivity to gemcitabine or paclitaxel in spheroid growth assays. Istiratumab (MM-141), a novel bispecific antibody that blocks IGF-1R and ErbB3, restored the activity of paclitaxel and gemcitabine in the presence of IGF-1 and HRG in vitro. Dual IGF-1R/ErbB3 blocking enhanced chemosensitivity through inhibition of AKT phosphorylation and promotion of IGF-1R and ErbB3 degradation. Addition of istiratumab to gemcitabine and nab-paclitaxel improved chemotherapy activity in vivo. Conclusions: Our findings suggest a critical role for the HRG/ErbB3 axis and support the clinical exploration of dual IGF-1R/ErbB3 blocking in pancreatic cancer. Clin Cancer Res; 24(12); 2873–85. ©2018 AACR.

Abstract A19: Identification of Heregulin (HRG) expression as a driver of a difficult-to-treat cancer phenotype and development of a companion diagnostic for the HRG-ErbB3 targeting drug seribantumab

Heregulin (HRG) is the cognate ligand of the ErbB3 receptor and has been identified as a potent driver of a distinct tumor cell phenotype characterized by enhanced survival that appears to be inherently more resistant to standard of care therapies. Seribantumab (MM-121) is a fully human monoclonal antibody designed to block HRG from binding to ErbB3 and to prevent the establishment of HRG-driven cancer cell survival in response to cytotoxic therapies. Clinical evidence from randomized studies suggests that patients who receive standard therapy have shorter progression-free survival (PFS) when their tumors are positive for HRG RNA expression compared to patients receiving standard therapy with HRG-negative tumors. The addition of seribantumab was able to enhance the anti-tumor effect of these respective standard therapies and prolong PFS in patients with HRG-positive tumors. Profiling data indicates that positive HRG expression can be found in more than a third of most solid tumors identifying a high unmet medical need. In order to identify HRG-driven tumors with a more difficult-to-treat phenotype, an RNA in situ hybridization (ISH) diagnostic assay has been developed. Details will be presented illustrating the benefits of RNA ISH over alternate detection methods for HRG. This includes the visualization of tumor cell HRG expression with high sensitivity and specificity in small tissue specimens such as fine needle aspirates and core needle biopsies. This assay is currently being used to select patients for inclusion into diagnostic-driven clinical studies. Citation Format: Sara Mathews, Gregory Finn, Arthur J. Kudla, Victoria Rimkunas, Peter Laivins, Gavin MacBeath, Akos Czibere, Jason Baum. Identification of Heregulin (HRG) expression as a driver of a difficult-to-treat cancer phenotype and development of a companion diagnostic for the HRG-ErbB3 targeting drug seribantumab. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Targeting the Vulnerabilities of Cancer; May 16-19, 2016; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(1_Suppl):Abstract nr A19.