GCS-100, a Galectin 3 Antagonist, Is a Novel Caspase-9 Apoptosis Activating Agent for the Treatment of Indolent B-Cell Malignancies.

Introduction: GCS-100 is a novel polysaccharide agent derived from natural citrus pectin with the ability to inhibit Galectin 3 (Gal-3). It has demonstrated anti-cancer activity and has been administered to patients with solid tumors in a Phase I study demonstrating a good safety profile and a lack of myelosuppression. It has been reported to have three major pharmacological effects: anti-metastatic, anti-angiogenic and pro-apoptotic. This study has investigated the mechanism of induction of apoptosis by GCS-100 for therapeutic benefit in indolent B-cell lymphomas.

Material and Methods: GCS-100 was tested in varying doses with B-cell lymphoma and leukaemia cell lines including the DoHH2 (t(14;18)), Ramos (high and low Bcl-2 expression), BV173 (t(9;22)) and K562 cell lines, primary patient CD19 selected CLL cells and normal B-cells in short term cultures. The addition of chemotherapy post GCS-100 was then undertaken to investigate enhanced anti-tumor activity. Apoptosis was assessed by a number of parameters including mitochondrial transition pore permeability (MTPP) by DiOC₆, caspase-8 and -9 activation by western analysis and membrane blebbing by MC540. Gal-3 expression was measured by western analysis and flow cytometry.

Results: The ability of GCS-100 to induce significant apoptosis in both malignant cell lines and primary patient CLL cells in vitro is clearly demonstrated with a minimal effect against normal B-cells (and stem cell cultures). The induced apoptotic pathway is the intrinsic mitochondrial and caspase-9 pathway. In addition, GCS-100 at low dosage will greatly enhance the apoptotic effect of chemotherapy, even in the presence of high levels of anti-apoptotic proteins such as Bcl-2. Immunoprecipitation has shown Bcl-2 binds to Gal-3. Gal-3 expression was increased in the malignant cells compared to the normal B-cells and could explain the selective activity.

Discussion: Gal-3 is a member of the b-galactoside-binding protein family that play significant roles in cellular interactions, cell growth, differentiation, resistance to apoptosis and loss of cell anchorage (anoikis). It shares the highly conserved BH1 functional receptor domain (NWGR) of the Bcl-2 gene family within its carbohydrate binding domain. The potential binding of GCS-100 to this region suggests this as its pro-apoptotic mechanism in B-cell malignancies. As shown in this study, normal B-cells with lower Gal-3 expression do not show significant apoptosis indicating GCS-100 is a selective anti-tumor agent. The possibility of using GCS-100 as an anti cancer agent, either alone or in combination with chemotherapy is suggested by this study. Its activity against indolent B-cell malignancies, such as follicular lymphoma (DoHH2 cell line) and CLL is particularly marked; a Phase 1 study is underway.