Anti-CD40 monoclonal antibody, clone 6D4 [Biotin]

Purpose: To investigate how induced tumor heterogeneity influences immune responses to radiotherapy with different proportions of mixed immune-responsive and unresponsive tumor cells in a triple-negative breast cancer model. It is hypothesized that studying the immune environment of mixed tumors and responses to radiotherapy could nominate immune active therapies to enhance immune responses after radiotherapy. Experimental Design: Evaluate efficacy and immune responses generated by radiotherapy in tumors with different proportions of immunologically responsive and unresponsive tumor cells. Then study the cellular responses and transcriptomic differences between the tumors to nominate immunotherapy combinations with radiotherapy and evaluate the combination. Results: The addition of the responsive cells to unresponsive tumors led to a greater than expected therapeutic response to radiotherapy with both innate and adaptive immune components. There was a distinct change in myeloid cells, greater inflammatory macrophage activity, and enhanced antigen presentation with responsive cells after radiotherapy. Because differences in matrix components, cell adhesion biology, and innate immune signaling correlated with myeloid cell response and phenotype, we hypothesized that radiotherapy combined with CD40 agonist antibody would sensitize unresponsive tumors. The combination therapy resulted in improved innate and adaptive immune response. Importantly, CD40 treatment increased tumor response to radiotherapy and protected against metastatic spread in a metastatic model. Conclusions: These data combined with transcriptomics from human patients support radiotherapy and myeloid cell targeting for immunologically cold tumors. The established study model presents opportunities to investigate the complex overlapping biologic mechanisms that limit immunotherapy and to implement radiotherapy with different immunotherapy combinations.

Multiple sclerosis (MS) is a demyelinating autoimmune disease of the central nervous system (CNS) mediated by autoreactive lymphocytes. The role of autoreactive lymphocytes in the CNS demyelination is well described, whereas very little is known about their role in remyelination during MS remission. In this study, we identified a new subpopulation of myelin-specific CD49d(+)CD154(+) lymphocytes presented in the peripheral blood of MS patients during remission, that proliferated in vitro in response to myelin peptides. These lymphocytes possessed the unique ability to migrate towards maturing oligodendrocyte precursor cells (OPCs) and synthetize proinflammatory chemokines/cytokines. The co-culture of maturing OPCs with myelin-specific CD49d(+)CD154(+) lymphocytes was characterized by the increase in proinflammatory chemokine/cytokine secretion that was not only a result of their cumulative effect of what OPCs and CD49d(+)CD154(+) lymphocytes produced alone. Moreover, maturing OPCs exposed to exogenous myelin peptides managed to induce CD40-CD154-dependent CD49d(+)CD154(+) lymphocyte proliferation. We confirmed, in vivo, the presence of CD49d(+)CD154(+) cells close to maturating OPCs and remyelinating plaque during disease remission in the MS mouse model (C57Bl/6 mice immunized with MOG(35-55)) by immunohistochemistry. Three weeks after an acute phase of experimental autoimmune encephalomyelitis, CD49d(+)/CD154(+) cells were found to be co-localized with O4(+) cells (oligodendrocyte progenitors) in the areas of remyelination identified by myelin basic protein (MBP) labelling. These data suggested that myelin-specific CD49d(+)CD154(+) lymphocytes present in the brain can interfere with remyelination mediated by oligodendrocytes probably as a result of establishing proinflammatory environment.