Targeting Galectin As A Strategy To Improve BH3 Mimetic Efficacy For The Therapy Of Acute Myeloid Leukemia

Abstract

Background: Galectins are a family of glycan binding proteins that regulate cell survival by mechanisms including apoptosis, cell cycle, and mRNA processing. Galectin-3 (LGALS3) is unique among galectins by having an N terminal region of roughly 130 amino acids that allows for multimerization and binding to other proteins independent of carbohydrate binding. In addition to promoting BCL2 gene expression and mitochondrial integrity, LGALS3 (along with LGALS1) positively regulates RAS signaling and thus the stability of proteins dependent on ERK phosphorylation such as MCL-1. The pro-survival functions of LGALS3 and other galectins suggest that their targeting could be therapeutic for cancers including AML. Indeed, galectin 3 (LGALS3) expression is a predictor of poor prognosis in acute myeloid leukemia (AML), as reported by Cheng and colleagues (Blood 2013). Current strategies to eradicate leukemia cells involve use of agents targeting BCL2 family members such a BH3 mimetics ABT-7373 and ABT-199. MCL-1 has been implicated as a resistance factor against BH3 mimetic killing and unfortunately an effective anti-MCL-1 drug is currently not available for use in the clinic. Alternatively, a strategy to target MCL-1 indirectly by suppressing Galectin 3 could improve BH3 mimetic killing of leukemia cells. In the current study we examine the effect of Galectin 3 suppression by shRNA or with a broad Galectin inhibitor on BH3 mimetic induced apoptosis in AML cell lines OCI-AML3 and THP1. Study Design: GCS-100 (kind gift from George Tidmarsh; La Jolla Pharmaceutical, San Diego, CA), is now in a Phase II clinical trial for chronic kidney disease. The drug was used in combination with BH3 mimetic ABT-737 (which targets BCL2 and BCL-XL at low dose). Galectin 3 was suppressed in both AML cell lines using lentiviral shRNA. Effect of GCS-100/ABT-737 combination was tested in an in vitro model of the tumor microenvironment using mesenchymal stem cells (MSC). Cell growth and induction of apoptosis was measured by flow cytometry. Analysis of protein expression and signaling was assessed by immunoblot analysis. Level of gene expression of various BCL2 family members was measured by qRT-PCR. Results: As a single agent GCS-100 suppresses the growth of AML cell lines OCI-AML3, THP-1, and HL60. The drug potently suppresses phosphorylation of AKT and ERK and reduces protein expression of BCL2 and MCL-1. We found that GCS-100 potently synergized with ABT-737 to kill OCI-AML3 cells: while 1 uM ABT-737 or 125 ug/mL GCS-100 reduced total viable cells by w 30% and induced apoptosis in 80% and > 60% reduction of viable cells, respectively, despite the presence of MSC when treated with 250 ug/mL GCS-100 for 72 hours. In addition, GCS-100 was found to block adhesion of OCI-AML3 cells to MSC suggesting that GCS-100 could be effective preventing leukemia cell contact with MSC. Conclusions: Our findings suggest that GCS-100 can induce apoptosis in AML cells as a single agent or in combination with the BH3 mimetic ABT-737. The agent is effective even in the presence of MSC suggesting it could be efficacious in the leukemia niche. These findings suggest GCS-100 could be effective for AML therapy.