The gene product of tumour suppressors are involved in signal transduction pathways that regulate cell movement and orientation, cell cycle progression, detect DNA damage and initiate DNA repair, autophagy, the regulation of apoptotic processes, epigenetic regulation of gene expression and protein function, and maintenance of metabolic balance of the cell.
Liver kinase B1 (LKB1) is a multi-tasking tumour suppressor serine-threonine kinase that is localized to both the cytoplasm and nucleus. LKB1s first interacting partners was identified by Dr. Marignani while a post-doctoral fellow at Harvard. Using in vitro expression cloning (IVEC) strategy she developed along with another PDF, LKB1 was discovered to bind to the SWI-SNF family member SMARCA4 (BRG1). Although a kinase, LKB1s catalytic activity was not required for binding, nor did LKB1 phosphorylate SMARCA4. IVEC, was also used to discover a new binding partner for 14-3-3; TAZ.
As a kinase, LKB1 catalytic activity is enhanced when in complex with pseudokinase STRAD and adaptor protein MO25, allowing for the phosphorylation and activation of AMP protein kinase (AMPK) on threonine 172 (Marignani et al 2007). When cellular AMP levels are elevated, AMP binds to the pseudosubstrate sequence within myristoylated AMPK, thereby enhancing the substrate readiness of AMPK to be phosphorylated by LKB1. Changes in the ratio of AMP/ATP can occur in response to metabolic stressors that perturb energy balance; for example, nutrient availability, glucose deprivation, hypoxia, ischemia, inhibition of glycolysis, tricarboxylic acid cycle or oxidative phosphorylation. Once activated, AMPK functions as a negative regulator of mTOR, a serine/threonine kinase that is the catalytic component of two distinct signalling pathways, mTOR complex 1 (mTORC1) and mTORC2. Both complexes are activated in response to growth factor signals mediated by PI3K, Ras and extracellular signal regulated protein kinase (ERK) pathways, and the availability of nutrients within a cell. TORC1 is primarily involved in regulating ribosomal biosynthesis and protein synthesis while TORC2 is primarily involved in the regulation of actin reorganization.
Our lab is interested in understanding how loss of LKB1 tumour suppressor function leads to disease. Using animal models; transgenic Cre-lox systems, Cre-dependent Cas9 mice, patient derived xenografts (PDX), primary cells, and cell lines in conjunction with “omic” technologies, we are identifying and characterizing new functions for LKB1 and oncogenic LKB1 (Scott et al Cancer Research 2007, Nath-Sain, Molecular Biology of the Cell 2009 in development, genome stability, tumour initiation, metabolism, epigenetics, immune biology and cancer stem cell biology (Andrade-Vieira et al Plos One 2013, Andrade-Vieira, et al, Cancer Biology and Therapy 2013, Vila-Leahy et al OncoImmunology 2016, Sengupta S et al Oncogene 2017).
The Marignani team generated a new mouse model that mimics stochastic development of HER2+ breast cancer. In this model, Lkb1-/-/NIC mice were engineered by disrupting mammary Lkb1 expression while simultaneously activating the expression of ErbB2 (NI) and Cre recombinase (C) under the constitutive MMTV promoter, referred to NIC mice. The Lkb1-/-/NIC mice showed reduced latency of tumour formation compared with NIC mice. Analysis of Lkb1-/-/NIC mammary tumours revealed the Warburg Effect; elevated ATP/reduced AMP levels, changes in metabolic enzymes and metabolites, along with hyperactivation of mTOR (both mTORC1 and mTORC2) pathways. The outcome of this study was the production of a new mouse model of Her2+ breast cancer that is hyperactive for mTOR and metabolically active.
We conducted pre-clinical drug studies that targeted hyperactive mTOR and the Warburg effect in our mouse model of breast cancer (Andrade-Vieira et al Plos One 2013). Treatment of Lkb1-/-NIC mice with AZD8055 and 2-DG mono-therapies significantly reduced mammary gland tumourigenesis by inhibition of mTOR pathways and glycolytic metabolism; however simultaneous inhibition of these pathways with AZD8055/2-DG combination was significantly more effective at reducing tumour volume and burden (Andrade-Vieira et al Oncotarget 2014). At the molecular level, combination treatments inhibited mTOR activity, aberrant mitochondria function and blocked MAPK pro-survival signalling pathways that are responsible for activation of the ERK-p90RSK feedback loop that drive breast cancer relapse.
Lung cancer mortality rates are the highest amongst all cancers. In Canada, lung cancer accounts for approximately 26.1% of cancer-related deaths. Surprisingly, the incidence of cancers in never-smokers is on the rise, representing approximately 25% of all lung cancers, with a greater incidence in women than in men. There is evidence that lung cancer in never-smokers is distinct enough from lung cancer in smokers from a molecular and epidemiological perspective to be considered a separate entity. Despite the difference in the etiology of lung cancers between never-smokers and smokers, lung cancer remains the leading cause of cancer-related deaths amongst Canadians.
My research team is applying CRISPR-Cas9 technology in vivo to characterize temporal development of lung cancer in mice. These studies complement our experiments that characterize the molecular heterogeneity of resected human lung cancers.