Metformin Lowers The Mitochondrial Potential Of Lymphoma Cells and Its Use During Front-Line Rituximab-Based Chemo-Immunotherapy Improves The Clinical Outcome Of Diffuse Large B-Cell Lymphoma

Accumulating evidence has linked tumor progression to alterations in cellular metabolism, underscoring the therapeutic strategy of targeting metabolic pathways in cancer cells, including lymphoma. Cancer patients are known to exhibit metabolic deregulation such as the Warburg effect where cancer cells shift adenosine triphosphate (ATP) generation from oxidative phosphorylation to glycolysis. Metformin is an agonist of the adenosine monophosphate-activated protein kinase (AMPK) that plays a pivotal role in cellular metabolism and B-cell development. It has been demonstrated that AMPK inhibits the mammalian target of rapamycin (mTOR). Clinically, metformin improves outcome of diabetic patients with solid tumor malignancies, either alone or combined with chemotherapy, indicating its potential role on cancer therapy. The effects of metformin in B-cell malignancies are poorly characterized. We performed a retrospective analysis to evaluate the effects of metformin on the clinical outcome of diffuse large B-cell lymphoma patients undergoing standard chemo-immunotherapy (R+CHOP or R-DA-EPOCH) at our institute. In addition, we characterized the biological effects of metformin in lymphoma pre-clinical models. Using the RPCI tumor registry we identified 264 consecutive DLBCL patients treated with equivalent doses of rituximab and anthracycline-based therapy at our institution. Demographic, clinical, pharmacological and pathological characteristics were recorded. Differences in response rate, progression free survival (PFS) and overall survival (OS) were evaluated between non-diabetic, diabetic on metformin or diabetic on other glucose lowering agent DLBCL patients. To further evaluate the biological effects of metformin in B-cell lymphoma, a panel of rituximab chemotherapy-sensitive (RSCL) and -resistant (RRCL) cell lines were exposed to metformin. Changes in cell viability, apoptosis and cell cycle were determined via measuring ATP content and flow cytometry, respectively. Based on the use of metformin during front-line chemo-immunotherapy, DLBCL patients were classified as non-diabetic (N=215, 81.4%), diabetic on metformin (N=27, 10.2%) or diabetic on other glucose-lowering agents (N=22, 8.3%). Demographics and clinical characteristics were equally distributed between non-diabetics and diabetics. After receiving R-CHOP or R-CHOP-like treatment, there was no difference in OS (95.8m vs. 90.1m, p=0.950) and PFS (82m vs. 80.7m days, p=0.994) for non-diabetic vs. diabetic DLBCL patients. Diabetic patients on metformin had an improved PFS when compared to non-diabetic patients/diabetic patients on other glucose lowering agents (95m vs. 79m, P=0.047). A non-statistically significant difference in OS was also observed (101m vs. 93m days, P=0.102). An analysis of clinical outcomes between diabetic DLBCL patients demonstrated that the use of metformin during front-line chemo-immunotherapy resulted in an improved PFS (94m vs. 55.4m days, P=0.007) and OS (100m vs. 70.5m, P=0.031) when compared to other glucose lowering agents. In vitro exposure of RSCL, but not RRCL, resulted in a dose- and time-dependent cell death. In addition, metformin lowered the mitochondrial potential and induced G1 cell cycle arrest in RSCL but not in RRCL. Together, our data suggest that metformin has direct anti-tumor effects in RSCL but not in RRCL who are known to have a dysfunctional mitochondria and therefore greater resistance to chemotherapy. Perhaps related to its effects on the mitochondrial potential on lymphoma cells, concurrent use of metformin during front-line chemo-immunotherapy contributed to the improved outcomes observed in our diabetic DLBCL patients. A better understanding on the cellular effects of metformin in B-cell lymphoma is necessary in order to optimize its potential therapeutic activity against DLBCL; additional research data will be presented at the annual meeting (Research, in part, supported by The Eugene and Connie Corasanti Lymphoma Research Fund)