Multiple myeloma (MM) is a malignant cancer of plasma cells derived from bone marrow. MM accounts for approximately 2 percent of all newly diagnosed cancers, and it can be devastating due to its lack of a cure (J Clin Oncol 2010;28:1599-1605, SEER Cancer Statistics Review, 1975-2015, https://seer.cancer.gov/csr/1975_2015/).
In addition to standard chemotherapies, the current standard of care for MM includes five different targeted therapeutic treatment options: 1) immunomodulators; 2) proteasome inhibitors; 3) HDAC inhibitors; 4) monoclonal antibodies; and 5) hematopoietic stem-cell transplantation. These therapies prevent cancer growth and metastasis via targeting cancer-specific genes, proteins, and microenvironment.
While there have been advances made to these targeted therapeutic classes over the past 20 years, nearly all patients eventually develop resistance to these agents and even with existing therapies the 5-year overall survival rate for MM is just over 50 percent (Cancer Statistics Review, 1975-2015, https://seer.cancer.gov/csr/1975_2015/). This highlights a significant unmet need for novel targeted therapies for MM, particularly those with a different mechanism of action to the existing therapies.
In a recent study, researchers from Duke University, Durham, N.C., have identified a new combination therapy involving a sphingosine kinase 2 inhibitor, ABC294640 (opaganib), with a Bcl-2 inhibitor, ABT-199 (venetoclax), that shows synergistic anti-myeloma effects in preclinical testing (Cancer Med 2018; doi:101002/cam4.1543). Based upon distinct molecular mechanisms to the existing targeted therapies for MM, this early-stage study could be the foundational research that helps identify and validate a new class of targeted therapeutic agents for this devastating disease.
Sphingolipid metabolism is rapidly being recognized as being important in regulation of cancer cell signaling, as well as in the control of tumor suppression and survival (Nat Rev Cancer 2018; doi:10.1038/nrc.2017.96). Sphingosine kinases 1 and 2 (SK1 and SK2) phosphorylate sphingosine to sphingosine-1-phosphate (S1P), altering the balance of apoptotic sphingosine with anti-apoptotic S1P. Inhibition of SK2 helps to shift this balance towards increased cell death for malignant plasma cells.
Previous research from this team has shown that inhibition of SK2 using ABC294640 increases apoptosis of myeloma cells and suppresses tumor growth in a myeloma xenograft model (Blood 2014; 124:1915-1925). Additionally, these anti-cancer effects were seen with concomitant decreases in expression of myeloid cell leukemia 1 (Mcl-1) and c-Myc, two oncogenes with strong associations with MM in patients. Most importantly, ABC294640 did not affect the expression of B-cell lymphoma 2 (Bcl-2), an anti-apoptotic gene that plays a key role in cancer chemoresistance (Oncogene 2008;27:6398-6406, Adv Cancer Res 2018;137:37-75).
Bcl-2 was originally found to be involved in cancer through chromosomal translocations found in lymphomas (Science 1984;226(4678):1097-1099, Nature 1985;29;315(6017):340-343). Many lymphomas contain a t(14;18) chromosomal translocation, which places the Bcl-2 gene on chromosome 18 into juxtaposition with the immunoglobulin heavy-chain locus on chromosome 14. This causes much higher levels of Bcl-2 expression in B cells and subsequent accumulation of B cells creating a lymphoma not caused by increased cell division.
More recently other chromosomal translocations involving Bcl-2 have been identified, including a t(11;14) translocation that is found in about 15-20 percent of all myeloma patients (J Clin Oncol 2000;18:804, Blood 2002;99:3735-3741, Leuk Res 2013;37:1251-1257). Bcl-2 and other anti-apoptotic proteins such as Mcl-1 and Bcl-xL bind and sequester pro-apoptotic proteins, preventing them from inducing apoptosis in cancer cells. ABT-199, an inhibitor of Bcl-2, works to increase cancer cell death by subsequent release of pro-apoptotic factor Bim (Leukemia 2014;28:210). However, ABT-199 only shows anti-myeloma effects in cells that harbor the t(11;14) translocation, and rapid resistance to ABT-199 develops limiting its upside as a single agent targeted cancer therapy (Leukemia 2014;28:210).
Since ABC294640 induces apoptosis of myeloma cells and decreases Mcl-1 and c-Myc expression without affecting Bcl-2 while ABT-199 inhibits Bcl-2 without affecting Mcl-1 and Bcl-xL, the authors of this study hypothesized that a combination of the two treatments would lead to synergistic anti-myeloma effects. The authors also hoped that the combination therapy would help overcome some of the genetic limitations that each agent has in individual therapy as well as the development of resistance to individual therapy. The published findings support this hypothesis, showing for the first time that a combination of ABC294640 with ABT-199 has the potential to be a new class of targeted therapeutic agents for the treatment of MM.
This research team previously showed that ABC294640 decreased MM proliferation and had anti-myeloma efficacy in an in vivo mouse xenograft model (Blood 2014;124:1915-1925). The current study examined the combination of ABC294640 with ABT-199 in treatment of MM. Since ABT-199 has anti-myeloma activity limited expressly to cells with the t(11;14) translocation, the researchers tested the combination therapy in MM cell lines that lack this translocation. A significant synergistic anti-proliferative effect was seen with combination therapy at a moderate dose in all cell lines tested except the L363 cell line.
In addition to demonstrating a synergistic decrease on cell proliferation with combination treatment, a similar synergistic effect was seen with induction of apoptosis in myeloma cell lines. Single agent treatment with ABC294640 and ABT-199 induced apoptotic cell death in 16-40 percent and 11-30 percent of cells, respectively. However, when given in combination, apoptotic cell death was increased to 60-72 percent.
The first in vitro experiment did not take tumor microenvironment into account, and myeloma cells reside within bone marrow, which plays an important role in MM growth, survival, and proliferation (Leukemia 2001;15:264, J Biol Chem 2017;292:4280-4292). To account for this, the authors tested a combination of ABC294640 and ABT-199 in myeloma cells lines co-cultured with bone marrow stromal cells. While combination therapy was unable to increase apoptosis in HS-5 cells alone, a strain of bone marrow stromal cells known to be protective against apoptosis in acute myeloid leukemia cells (Biomed Res 2018;29(5)), when HS-5 cells were co-cultured with MM cell line RPM8226 cells, the combination of ABC294640 and ABT-199 showed a significant induction of apoptosis.
In order to determine the mechanism by which the combination of ABC294640 and ABT-199 induces apoptosis, the authors measured several different molecular markers of cell death. First they looked at markers of apoptotic cell death such as cleavage of poly(ADP-ribose) polymerase (PARP) and pro-caspase-3, as well as caspase-9 expression (Toxicol Pathol 2007;35:495-516). The combination treatment enhanced downregulation of caspase-9 while also increasing accumulation of cleavage products of caspase-3 and PARP, indicating that apoptotic mechanisms were in fact responsible for MM cell death following treatment by ABC294640 and ABT-199.
Additional molecular markers were investigated in order to determine if apoptosis induced by each agent alone or by the combination of ABC294640 and ABT-199 was caused by intrinsic or extrinsic apoptotic pathways. While there were decreases in expression of anti-apoptotic markers such as Mcl-1 and Bcl-xL caused by ABC294640, the authors found no indication that either agent increases the expression of proapoptotic markers such as Bim, Bid, or Bak (J Biol Chem 2017;292:6478-6492).
In addition to decreasing expression of proteins that prevent apoptosis, it was also shown that ABT-199 could increase cleavage of Bax, a marker of mitochondrial-mediated apoptosis, as well as increase mitochondrial depolarization, a form of mitochondrial damage associated with extrinsic apoptotic mechanisms (Cancer Lett 2003;189:221-230, PLoS ONE 2011;6:e25976). Taken together, these data suggest that a combination of ABC294640 and ABT-199 can activate both intrinsic and extrinsic apoptotic pathways in MM cells.
Finally, to determine the in vivo anti-myeloma efficacy of ABC294640 and ABT-199 combination treatment the authors used a xenograft mouse model. MM tumors were induced in NSG mice with JJN3 cells followed by 20 days of treatment with either ABC294640 (50 mg/kg) or ABT-199 (50 mg/kg) alone or in combination. While the anti-tumor effect of each agent alone was moderate, the combination therapy showed a significantly greater anti-cancer effect. This study demonstrates the potent in vivo anti-MM effects of the combination therapy at well-tolerated doses.
Multiple myeloma continues to be one of the more devastating forms of cancer even with recent advances in targeted cancer therapies. There remains a significant unmet need for new effective therapeutic agents for MM, particularly ones that have a novel mechanism of action. Also, with the rapid development of resistance to certain classes of targeted cancer therapeutics, developing novel treatments that remain effective in patients for a longer time frame is integral to adding to our arsenal of cancer medicines.
Recent findings published in Cancer Medicine identified and characterized a dual-agent therapy consisting of an SK2 inhibitor, ABC294640, in combination with a Bcl-2 inhibitor, ABT-199, that shows strong anti-myeloma effects and potentially less chance to developing rapid resistance in patients. Since both are orally bioavailable small molecule inhibitors, there is a good precedent for them to be used as a combination therapy in humans, and both have been investigated as single agent therapeutics in active or recently completed clinical trials (NCT02757326 and NCT03552692, respectively).
Both SK2 and Bcl-2 are relatively novel targets for cancer therapies and both are involved in apoptosis, albeit with slightly different mechanisms of action (Blood 2014;124:1915-1925, Oncogene 2008;27:6398-6406, Adv Cancer Res 2018;137:37-75). The authors of the current study have elegantly demonstrated that ABC294640 is responsible for anti-myeloma apoptotic effects via extrinsic, cytoplasmic-associated apoptosis while ABT-199 works via intrinsic, mitochondrial-associated apoptosis. The combination of these agents leads to a synergistic anti-myeloma effect that is significantly greater than either one alone. Additionally, ABC294640 downregulated Mcl-1, cMyc, and Bcl-xL, which helps prevent development of resistance to ABT-199. These studies form a strong pre-clinical basis for an extended clinical evaluation of ABC294640 in combination with ABT-199 as a novel therapy for patients with multiple myeloma.
YUBIN KANG, MD, is Associate Professor of Medicine in the Division of Hematologic Malignancies & Cellular Therapy at Duke University Medical Center, Durham, N.C. PASUPATHI SUNDARAMOORTHY, PHD, is a Postdoctoral Associate in the Division of Hematologic Malignancies & Cellular Therapy at Duke University Medical Center.