Pilot: Targeting NNMT to Overcome Chemoresistance in Triple Negative Breast Cancer

PROJECT SUMMARY Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, with high rates of recurrence, metastasis, and mortality. Current treatment regimens are mostly palliative and generally fail to halt disease progression. Mounting evidence indicates that TNBC cells underwent a metabolic rewiring that endows them with increased cancer stem cell (CSC)-like properties that can withstand the increased oxidative stresses encountered during metastasis and drug treatment. Therefore, it is imperative to understand how TNBC cells elicit antioxidative defense mechanisms to neutralize ROS and mitigate oxidative damages without compromise to their increased metabolism and cancer stem cell (CSC) properties. Nicotinamide adenine dinucleotide (NADH & NADPH), defined as the backbone of “cellular redox code”, is the first line of defense machinery to mitigate reactive oxygen species (ROS), metabolic byproducts that are associated with aging, diabetes and cancer. However, it remains a mystery how TNBC cells boost NAD+ levels in order to rejuvenate their oncogenic potentials. Recently, we found that nicotinamide N-methyltransferase (NNMT), a key enzyme in the NAD+ salvage pathway with unclear functions, is upregulated in TNBC. Specifically, we discovered that: 1) NNMT is expressed highly and specifically in TNBC; 2) robust NNMT upregulation is concomitant with increased NAM uptake and NAD(P)+ biosynthesis in TNBC cells under metabolic stress; 3) NNMT-KO increases ROS levels in TNBC cells and confer them with sensitivity to H2O2 and chemotherapeutic agent; and 4) NNMT-KO inhibits TNBC cell growth in vitro and suppresses tumor growth in animal model. Based on these findings, we hypothesize that NNMT is a pivotal factor that enhances NAM uptake and promotes NAD+ synthesis, which is essential to scavenge the increased ROS and maintain CSC characteristics under metabolic stress and drug treatments. To test this hypothesis, two specific aims are proposed. Aim 1 will define whether and how NNMT boosts NAD(P)+ synthesis under metabolic stress in TNBC. We will: 1) determine how NNMT upregulation promotes NAD(P)+ synthesis; 2) delineate how NNMT regulates cellular metabolism (such as glycolysis and TCA cycle) under metabolic stress; 3) define whether NNMT is required to maintain SIRT enzyme activities. Aim 2 will determine how NNMT regulates CSC properties and contributes to drug resistance in TNBC. We will: 1) determine how NNMT promotes breast CSC properties and contributes to chemo-drug resistance in TNBC; 2) delineate how NNMT mitigates ROS upsurge in TNBC cells under metabolic stress; and 3) define the function of NNMT in TNBC. Our findings highlight NNMT as an attractive therapeutic target in TNBC treatment. The impact of this project will be significant because it will provide a novel mechanism-guided therapeutic strategy to overcome hemotherapy resistance and enhance the effectiveness of TNBC therapy by unraveling a targetable metabolic vulnerability. Support from COBRE pilot grant will generate key data to apply for a competitive NIH R01 grant.