University of Kentucky Center for Cancer and Metabolism (Pilot Project - Dr. Xiuwei Yang)

The triple-negative breast cancer subtype (TNBC) (PR-, ER- and ErbB2-) remains one of the deadliest threats to women’s health, yet the underlying drivers are still poorly defined. One set of such promising drivers are integrins, a family of cell adhesion receptors, and their key signaling effector, focal adhesion kinase (FAK). These molecules are overexpressed or amplified in the TNBC patient cohort and promote tumorigenesis at the cellular and signaling levels. Intriguingly, these molecules also affect tumor growth and resistance to the current therapies in the context of the c- Myc oncogene. Given the pivotal role c-Myc plays in metabolism, this evidence implies that an integrin-FAK signaling axis (IFSA) plays a pro-metabolic role. Consistent with this notion, FAK and c-Myc appear to cooperatively promote glycolysis via activating lactate dehydrogenase A (LDHA), an enzyme producing oxidized Nicotinamide adenine dinucleotide (NAD+) and lactate. As a dominant driver of metabolic homeostasis, NAD+ is also synthesized through the nicotinamide phosphoribosyl transferase (NAMPT)-driven salvage pathway. A co-inhibition of FAK and NAMPT markedly impairs cell viability and glycolytic capacity. Together, these observations suggest a strong metabolic role of IFSA in breast cancer. Thus, we hypothesize that IFSA contributes to breast tumorigenesis by driving glycolysis and the NAD+ supply. This hypothesis will be tested in the following studies: Aim 1. Determine if IFSA drives glycolysis via regulation of LDHA activation. Based on functional and signaling links among IFSA, c-Myc and LDHA, we will evaluate a working model that IFSA promotes glycolysis by dictating expression and activity of LDHA. Accordingly, expression and activation of FAK and LDHA in TNBC cell lines will be manipulated using RNAi/CRISPR and mutagenesis techniques or chemical inhibitors. The effect on glycolysis will be quantified by measuring changes in extracellular acidification rate (ECAR) on the Seahorse analyzer, and key metabolites via the [U-13C]- Glucose]/NMR-based metabolomics technique. We anticipate a marked LDHA-dependent role of IFSA in cellular glycolysis and proliferation. Aim 2. Investigate therapeutic potential of co-targeting IFSA and NAMPT. Building on the close link of IFSA and NAMPT to cell growth and metabolism, we will investigate if IFSA acts synergistically with NAMPT to support tumor growth and the intracellular NAD+ supply. The FAK and NAMPT collaboration will be assessed by analyses of the effect of co-overexpressing or co-inhibiting these molecules on the NAD+ level and tumor growth. The clinical link between FAK/NAMPT and the TNBC malignancy will be evaluated by IHC analysis of patient biopsies. Overall, our project is expected to yield experimental and clinical evidence of the novel role IFSA has in breast cancer metabolism. The resulting data will serve as preclinical support for developing the IFSA-based synthetic lethal-type therapy for TNBC, and a crucial basis for launching a R01/R21 application.