Chelsea Hope

Tumoricidal Effects of Macrophage-Activating Immunotherapy in a Murine Model of Relapsed/Refractory Multiple Myeloma.

Jensen and colleagues report that inhibition of innate immunity checkpoint TPL2 kinase signaling potentiates the efficacy of anti–CD40-based immunotherapy, which expands M1-polarized macrophages in the bone marrow, prolonging survival in an immunocompetent, transplant-based preclinical model of relapsed/refractory multiple myeloma. Myeloma remains a virtually incurable malignancy. The inevitable evolution of multidrug-resistant clones and widespread clonal heterogeneity limit the potential of traditional and novel therapies to eliminate minimal residual disease (MRD), a reliable harbinger of relapse. Here, we show potent anti-myeloma activity of macrophage-activating immunotherapy (αCD40+CpG) that resulted in prolongation of progression-free survival (PFS) and overall survival (OS) in an immunocompetent, preclinically validated, transplant-based model of multidrug-resistant, relapsed/refractory myeloma (t-Vκ*MYC). αCD40+CpG was effective in vivo in the absence of cytolytic natural killer, T, or B cells and resulted in expansion of M1-polarized (cytolytic/tumoricidal) macrophages in the bone marrow. Moreover, we show that concurrent loss/inhibition of Tpl2 kinase (Cot, Map3k8), a MAP3K that is recruited to activated CD40 complex and regulates macrophage activation/cytokine production, potentiated direct, ex vivo anti-myeloma tumoricidal activity of αCD40+CpG–activated macrophages, promoted production of antitumor cytokine IL12 in vitro and in vivo, and synergized with αCD40+CpG to further prolong PFS and OS in vivo. Our results support the combination of αCD40-based macrophage activation and TPL2 inhibition for myeloma immunotherapy. We propose that αCD40-mediated activation of innate antitumor immunity may be a promising approach to control/eradicate MRD following cytoreduction with traditional or novel anti-myeloma therapies. Cancer Immunol Res; 3(8); 881–90. ©2015 AACR.

Versican-Derived Matrikines Regulate Batf3–Dendritic Cell Differentiation and Promote T Cell Infiltration in Colorectal Cancer

Colorectal cancer originates within immunologically complex microenvironments. To date, the benefits of immunotherapy have been modest, except in neoantigen-laden mismatch repair–deficient tumors. Approaches to enhance tumor-infiltrating lymphocytes in the tumor bed may substantially augment clinical immunotherapy responses. In this article, we report that proteolysis of the tolerogenic matrix proteoglycan versican (VCAN) strongly correlated with CD8+ T cell infiltration in colorectal cancer, regardless of mismatch repair status. Tumors displaying active VCAN proteolysis and low total VCAN were associated with robust (10-fold) CD8+ T cell infiltration. Tumor-intrinsic WNT pathway activation was associated with CD8+ T cell exclusion and VCAN accumulation. In addition to regulating VCAN levels at the tumor site, VCAN proteolysis results in the generation of bioactive fragments with novel functions (VCAN-derived matrikines). Versikine, a VCAN-derived matrikine, enhanced the generation of CD103+CD11chiMHCIIhi conventional dendritic cells (cDCs) from Flt3L-mobilized primary bone marrow–derived progenitors, suggesting that VCAN proteolysis may promote differentiation of tumor-seeding DC precursors toward IRF8- and BATF3-expressing cDCs. Intratumoral BATF3-dependent DCs are critical determinants for T cell antitumor immunity, effector T cell trafficking to the tumor site, and response to immunotherapies. Our findings provide a rationale for testing VCAN proteolysis as a predictive and/or prognostic immune biomarker and VCAN-derived matrikines as novel immunotherapy agents.

Extracellular matrix and the myeloid‐in‐myeloma compartment: balancing tolerogenic and immunogenic inflammation in the myeloma niche

The last 10–15 years have witnessed a revolution in treating multiple myeloma, an incurable cancer of Ab‐producing plasma cells. Advances in myeloma therapy were ushered in by novel agents that remodel the myeloma immune microenvironment. The first generation of novel agents included immunomodulatory drugs (thalidomide analogs) and proteasome inhibitors that target crucial pathways that regulate immunity and inflammation, such as NF‐κB. This paradigm continued with the recent regulatory approval of mAbs (elotuzumab, daratumumab) that impact both tumor cells and associated immune cells. Moreover, recent clinical data support checkpoint inhibition immunotherapy in myeloma. With the success of these agents has come the growing realization that the myeloid infiltrate in myeloma lesions—what we collectively call the myeloid‐in‐myeloma compartment—variably sustains or deters tumor cells by shaping the inflammatory milieu of the myeloma niche and by promoting or antagonizing immune‐modulating therapies. The myeloid‐in‐myeloma compartment includes myeloma‐associated macrophages and granulocytes, dendritic cells, and myeloid‐derived‐suppressor cells. These cell types reflect variable states of differentiation and activation of tumor‐infiltrating cells derived from resident myeloid progenitors in the bone marrow—the canonical myeloma niche—or myeloid cells that seed both canonical and extramedullary, noncanonical niches. Myeloma‐infiltrating myeloid cells engage in crosstalk with extracellular matrix components, stromal cells, and tumor cells. This complex regulation determines the composition, activation state, and maturation of the myeloid‐in‐myeloma compartment as well as the balance between immunogenic and tolerogenic inflammation in the niche. Redressing this balance may be a crucial determinant for the success of antimyeloma immunotherapies.

MAP3K8 kinase regulates myeloma growth by cell-autonomous and non-autonomous mechanisms involving myeloma-associated monocytes/macrophages

Benefit from cytotoxic therapy in myeloma may be limited by the persistence of residual tumour cells within protective niches. We have previously shown that monocytes/macrophages acquire a proinflammatory transcriptional profile in the myeloma microenvironment. Here we report constitutive activation of MAP3K8 kinase‐dependent pathways that regulate the magnitude and extent of inflammatory activity of monocytes/macrophages within myeloma niches. In myeloma tumour cells, MAP3K8 acts as mitogen‐induced MAP3K in mitosis and is required for TNFα‐mediated ERK activation. Pharmacological MAP3K8 inhibition results in dose‐dependent, tumour cell‐autonomous apoptosis despite contact with primary stroma. MAP3K8 blockade may disrupt crucial macrophage‐tumour cell interactions within myeloma niches.

Versican Proteolysis Predicts Immune Effector Infiltration And Post-Transplant Survival In Myeloma

High-dose alkylator-based conditioning followed by autologous stem-cell transplantation (ASCT) is a therapeutic mainstay for eligible patients with multiple myeloma. However, post-transplant relapses are common and prognostic biomarkers are scarce. Relapses are characterized by the influx of regulatory myeloid cells and dysfunctional T effectors. We have shown that myeloma-infiltrating myeloid cells produce versican (VCAN), a large matrix proteoglycan with tolerogenic activities. VCAN proteolysis by a-disintegrin-and-metalloproteinase-with-thrombospondin-motifs (ADAMTS) proteases generates versikine, a bioactive fragment (“matrikine”) that regulates Batf3-dendritic cells, known to control CD8+-attracting chemokine networks. Here we demonstrate that intense VCAN proteolysis predicts CD8+ infiltration post-transplant and paradoxically portends significantly inferior survival outcomes. Our data suggest that VCAN proteolysis promotes the influx of CD8+ effectors that are rendered overwhelmingly dysfunctional and/or frankly immunoregulatory (CD8+ Treg) at the tumor site. Thus, complex immunosuppressive circuits orchestrated through VCAN accumulation and turnover generate conditions favorable for myeloma tumor regrowth and point to a readily-assayed biomarker to identify the patients at risk for relapse and early death. The dismal outcomes associated with VCAN proteolysis may be rationally overcome through immunotherapies such as checkpoint inhibition (e.g., anti-TIGIT), tumor vaccines or anti-myeloid (e.g., anti-CSF-1R) approaches.

Deploying myeloid cells against myeloma.

ABSTRACT Myeloma remains incurable despite recent therapeutic advances. We propose that minimal residual disease following cytoreductive therapy may be controlled through re-education of myeloid cells to elicit tumoricidal activity. We review work from our laboratory and others highlighting aspects of macrophage-myeloma cell crosstalk as well as strategies for therapeutic macrophage reprogramming.

Abstract 1169: TPL2 kinase regulates the inflammatory milieu of the myeloma niche

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Early-stage myeloma tumor cells are critically dependent on paracrine cytokine support originating from their bone marrow microenvironment whereas advanced-stage myeloma tumor cells may elaborate autocrine cytokine production mechanisms and/or cell-autonomous mutations in critical downstream signaling pathways (e.g., NFκB pathway). However, the molecular mechanisms underpinning the paracrine network in myeloma are unclear. The pro-inflammatory cytokine IL-1β has emerged as a major link between inflammation and cancer and has been validated as a therapeutic target in high-risk monoclonal gammopathy, the precursor to myeloma, as well as early-stage myeloma. To determine the source of IL-1β production in early-stage myeloma we analyzed paired purified cell fractions obtained from each of 5 patients at diagnosis: CD138+ tumor cells, CD14+ bone marrow-resident monocyte/macrophages as well as bone marrow-derived stromal/mesenchymal cells (BM-MSC). We found that in all cases, monocytes/macrophages were the predominant IL-1β synthesizer cell type in the myeloma microenvironment. Cytokine production by activated macrophages is controlled by the MAP3 kinase, TPL2 (Cot, MAP3K8). We have previously detected constitutive activation of TPL2-regulated signaling pathways in human myeloma-associated macrophages but its precise functional consequences have been unclear. To dissect the relevant mechanisms, we ablated Tpl2 in the genetically-engineered myeloma in vivo model, Vκ*MYC. Vκ*MYC animals activate MYC sporadically in B lymphocytes participating in germinal center reactions and develop a disease analogous to human multiple myeloma with production of paraprotein (monoclonal immunoglobulin), plasma cell infiltration of the bone marrow as well as end-organ damage (“myeloma kidney”, osteolytic lesions). Vκ*MYC+/Tpl2-null animals developed myeloma with a significantly prolonged latency and the disease burden was lower at all timepoints tested compared to controls. Analysis of monocytic cells from myeloma lesions showed that loss of Tpl2 did not result in macrophage repolarization to an unopposed M1 (tumoricidal/cytotoxic) phenotype. Instead, Tpl2-null myeloma-associated monocytes/macrophages exhibited severe defects in production of inflammatory cytokines, predominantly IL-1β, but also IL-6. Tpl2 loss did not have discernible impacts on tumor cell-autonomous growth and survival. Our results suggest that monocytes/macrophages and TPL2 kinase activity play a central role in orchestrating the inflammatory milieu of the myeloma niche. Disruption of the myeloma cytokine network through pharmacologic TPL2 kinase inhibition could provide novel therapeutic opportunity by interfering with the co-ordinate regulation of key pro-myeloma inflammatory cytokines through a targeted approach. Citation Format: Chelsea Hope, Samuel J. Ollar, Erika Heninger, Jeffrey L. Jensen, Ellen Hebron, Jaehyup Kim, Ioanna Maroulakou, Shigeki Miyamoto, Natalie Callander, Peiman Hematti, Marta Chesi, P. Leif Bergsagel, Fotis Asimakopoulos. TPL2 kinase regulates the inflammatory milieu of the myeloma niche. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1169. doi:10.1158/1538-7445.AM2014-1169