ROR1 − a druggable target : preclinical studies of ROR1 and combinatorial partners in malignancies

Abstract: Cancer cells are masters of adaption, and their ability to sustain proliferative signaling presents a complicated challenge to cancer treatment. Therefore, novel drugs that attack key drivers of cancer cell growth are urgently needed. Targeted cancer therapy aims to selectively kill tumor cells and block their growth by interfering with specific molecular survival mechanisms. Tyrosine kinase orphan receptor 1 (ROR1) is abundantly expressed in numerous human cancers, whereas it has lower expression in most normal adult tissues. ROR1 is essential for oncogenesis and tumor cell survival. Based on these characteristics, ROR1 is considered to be an attractive target for a targeted drug strategy. The aim of the thesis was to understand how ROR1 activity is regulated in malignancies and identify the mechanism of action (MOA) of the small molecule inhibitor of ROR1 in killing tumor cells. The first study investigated the in vitro activity of the first-in-class small molecule ROR1 inhibitor (KAN0439834) against a number of human pancreatic carcinoma cell lines. The result showed that the combination of KAN0439834 with ibrutinib or erlotinib had a significant additive apoptosis effect, which supports further investigation as both drugs are currently in clinical trials concerning pancreatic carcinoma. The second study investigated the functional and clinical inhibitory properties of ROR1 in diffuse large B-cell lymphoma (DLBCL). Our data show that a second-generation small-molecule of ROR1 inhibitor (KAN0441571C) in combination with venetoclax achieves the desired outcome in apoptosis of DLBCL tumor cells. Furthermore, KAN0441571C caused significant tumor reduction in zebrafish transplanted with a ROR1+ DLBCL cell line. The third study assessed the result of KAN0441571C treatment in chronic lymphocytic leukemia (CLL) cells from six patients collected before and after the acquisition of resistance to ibrutinib. The result showed an induction of apoptosis in both ibrutinib-sensitive and -resistant CLL cells by ROR1 inhibitor. However, the combination of ROR1 inhibitor and venetoclax had a synergistic cell death in ibrutinib-resistant cells. The fourth study examined the inhibition of ROR1 in human MCL cell lines and primary MCL cells. We described a combinatorial approach of KAN0441571C with ibrutinib, venetoclax, idelalisib, everolimus, and bendamustine which had an additive or mostly synergistic apoptotic effect in MCL cells. The fifth study investigated the clinical and functional expression of ROR1 in non-small cell lung cancer (NSCLC) cases and the outcomes of ROR1 inhibition in NSCLC cell lines. Dephosphorylation of ROR1 by KAN0441571C results in time- and dose-dependent apoptosis of NSCLC cell lines. In addition, targeting NSCLC cells by using ROR1 and EGFR inhibitors showed synergistic or additive effects on lung cancer cells. In conclusion, these studies reveal novel functions for ROR1 tyrosine kinase in cancer cells and demonstrate that blocking its function by a small molecule ROR1 inhibitor results in a cancer-specific apoptotic effect, both alone and in combination with conventional therapies, which could lead to more effective cancer therapy.