Prosurvival and Immunosuppressive Functions of CD28 in the Myeloma Tumor Microenvironment

Abstract

Abstract 473Multiple myeloma (MM) is the second most common hematologic malignancy and remains incurable for most patients. Myeloma cells are the transformed counterpart of the normal, bone marrow-resident long lived plasma cells (LLPC) that can survive for years to decades and are also responsible for long-term production of protective antibody titers. Critical interactions between MM and their bone marrow stromal cells (BMSC), that are important for their long term survival and chemotherapy resistance (in MM) identifies potential therapeutic targets, and are often the target for many IMIDs (thalidomide, lenalidomide). However, the specific molecular and cellular components of these interactions remain poorly characterized.These interactions directly transduce pro-survival signals to the myeloma cells as well as induce niche production of supportive soluble factors, the prototypic example being MM induction of stromal IL-6 - a key pro-MM survival cytokine. Despite this importance, the specific molecular and cellular components involved in these interactions remain poorly characterized. We have previously shown that CD28 expressed on human myeloma cells directly transduces a survival signal to MM cells upon binding its CD80/CD86 ligands on conventional (myeloid) dendritic cells (DC), and that DC preferentially co-localize with myeloma cells in the patient bone marrow niche. In our previous presentation at ASH (2010), we showed that myeloma cells interact with DCs to produce immunosuppressive factors such as IDO, and protects myeloma cells against cell death via a CD28-B7 mediated interaction. We now show that DC-IDO not only suppresses T-cell proliferation in invitro assays, but also contributes to the immunosuppressive milieu by inducing naïve T-cells to form T-regs (Fig 1). We hypothesize that while IDO activity by itself can suppress T-cell proliferation and induce T-cell growth arrest and apoptosis, the generation of T-regs by these immunosuppressive DCs (previous studies have shown a close association of T-regs with myeloma cells) form two facets of an immunosuppressive defense that myeloma cells mount against the body’s anti-myeloma immune response. [Display omitted] As we have shown in our earlier presentations at ASH, on the myeloma side, the activation of CD28 induces pro-survival responses that can be extinguished by blocking CD28-B7 interactions between myeloma cells and DCs. Now we show that CD28 activation is accompanied by rapid tyrosine phosphorylation of CD28, association of p85 (PI3K), activation of Vav-1 and increase in CD28 associated tyrosine kinase activity, as shown by immunoprecipitation, western and kinase activity assays. Our data suggests a role for SLP76 downstream of Vav1 in CD28 mediated survival of myeloma cells. Immunoassays with protein extracts from myeloma cells that were previously co-cultured with DCs and isolated using positive magnetic selection show a decrease in BLIMP1 expression (Fig 2) that correlates with published data by other groups that indicates that DC mediated increase in myeloma clonogenicity/tumorigenicity is accompanied by increases in BCL6 (a negative regulator of BLIMP1). Blocking CD28-B7 interactions between myeloma and dendritic cells reversed this decrease in BLIMP1 expression. The implications of this to myeloma survival is currently under study in our lab. [Display omitted] In summary, we propose that CD28 expressed by myeloma cells serves as a central molecular bridge within a complex and integrated cellular and soluble factor microenvironment necessary for MM cell survival. CD28 directly delivers a pro-survival signal to the myeloma cell, and by ligating CD80/CD86 on conventional DC backsignals to these stromal cells to elicit immunosuppressive enzyme IDO and inducing immunosuppressive T-regulatory cells. Although undoubtedly incomplete, this model begins to point to novel therapeutic targets for the treatment of multiple myeloma. Disclosures:No relevant conflicts of interest to declare.