The Role of the MLK Family in Cancer and Resistance to Targeted Therapies

Abstract

The mixed lineage kinases (MLKs) are a family of MAP3Ks that consists of three subfamilies: the MLK subfamily, comprising MLK1-4; the dual leucine zipper-bearing kinases (DLKs), composed of DLK and leucine zipper-bearing kinase (LZK); and the zipper sterile-alpha-motif kinases (ZAKalpha and ZAKβ). Alterations in MLK proteins have been found in various cancers, including MLK1 mutations in melanoma, and copy number gain of LZK in squamous cell carcinoma (SCC). However, the role of these kinases in cancer has not been well defined. This project has focused on evaluating the functional consequences of MLK mutation and overexpression in cancer.The first part of this thesis concerns the role of MLK1-4 in vemurafenib resistance in melanoma. MLKs phosphorylate MKK4/7 and MKK3/6 to activate the JNK and p38 pathways, respectively. We found that MLK1-4 can also directly phosphorylate MEK to activate the ERK pathway, enabling them to mediate resistance to the BRAF inhibitor vemurafenib in melanoma. Overexpression of MLK1-4 reactivated ERK, improved survival, and rescued cells from apoptosis induced by RAF inhibitor treatment. Increased expression of MLK1-4 was identified in patients who developed resistance to vemurafenib, and expression of MLK1 could mediate resistance in a xenograft model. Additionally, activating mutations were found in MLK1 in melanoma patients and could potentially confer de novo resistance to BRAF inhibitors.In the second part of this thesis, the role of LZK in SCC is explored. LZK resides on chromosome 3q, which is subject to copy number gain in ~80% of head and neck squamous cell carcinomas (HNSCC), as well as other squamous cancers. HNSCC accounts for ~600,000 cases and 300,000 deaths per year worldwide. Analysis of HNSCC cell lines with copy number gain of 3q revealed that LZK mRNA and protein levels were increased compared with control cell lines. Knockdown of LZK by siRNA reduced cell viability and survival in two HNSCC cell lines, but not control cells. Knockdown by inducible shRNA reduced survival, proliferation, and colony forming ability of HNSCC lines with 3q gain, but not control lines without copy number gain of the gene. Further work needs to be completed to determine the exact mechanism of this effect, but initial data indicated that regulation of AKT or of gain-of-function p53 by LZK maintains cancer cell proliferation. This study needs to be extended to a wider range of cell lines, but these results suggest that copy number gain of LZK can play a pro-proliferative role in HNSCC.

Date of Award	1 Aug 2016
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Donald Ogilvie (Supervisor) & John Brognard (Supervisor)