Human CD22-Transgenic, Primary Murine Lymphoma Challenges Immunotherapies in Organ-Specific Tumor Microenvironments

Targeted immunotherapies have greatly changed treatment of patients with B cell malignancies. To further enhance immunotherapies, research increasingly focuses on the tumor microenvironment (TME), which differs considerably by organ site. However, immunocompetent mouse models of disease to study immunotherapies targeting human molecules within organ-specific TME are surprisingly rare. We developed a myc-driven, primary murine lymphoma model expressing a human-mouse chimeric CD22 (h/mCD22). Stable engraftment of three distinct h/mCD22+ lymphoma was established after subcutaneous and systemic injection. However, only systemic lymphoma showed immune infiltration that reflected human disease. In this model, myeloid cells supported lymphoma growth and showed a phenotype of myeloid-derived suppressor cells. The human CD22-targeted immunotoxin Moxetumomab was highly active against h/mCD22+ lymphoma and similarly reduced infiltration of bone marrow and spleen of all three models up to 90-fold while efficacy against lymphoma in lymph nodes varied substantially, highlighting relevance of organ-specific TME. As in human aggressive lymphoma, anti-PD-L1 as monotherapy was not efficient. However, anti-PD-L1 enhanced efficacy of Moxetumomab suggesting potential for future clinical application. The novel model system of h/mCD22+ lymphoma provides a unique platform to test targeted immunotherapies and may be amenable for other human B cell targets such as CD19 and CD20.

P03.20 A murine, myc-driven lymphoma model expressing human CD22 enables testing of targeted therapies and their effects on tumor immune microenvironment

BackgroundThe tumor microenvironment (TME) is composed of various cell types which closely interact via cell cell contacts and cytokines leading to tumor promotion, immune cell inhibition and drug resistance. TME is increasingly recognized for its role in cancer immunotherapies. In B-cell malignancies, myeloid cells play a central role in supporting tumor growth and immune suppression (Roussel et al., 2017, Cancer Immunol Immunother). Despite the importance of a syngeneic TME, preclinical studies with novel drugs have mainly been performed in models lacking a functional immune system. Therefore, we developed an immune competent murine lymphoma model transgenic to human CD22 to study effects of targeted therapies on TME.Materials and MethodsA chimeric CD22 consisting of human extracellular and murine intracellular CD22 (h/mCD22) was introduced in BL6 mice (BL6h/mCD22). Crossbreeding with BL6λ-myc lead to spontaneous development of murine lymphoma that were serially transplanted. Tumor infiltration and TME was characterized by flow cytometry. Mice were treated with Moxetumomab pasudotox, a CD22 targeted immunotoxin and Doxorubicin.ResultsSpontaneously developed tumors in lymphoid organs from BL6h/mCD22 x λ-myc consist of a monomorphic population of h/mCD22+ murine B cells. Three primary lymphoma subclones were isolated from distinct mice and serially transplanted in syngeneic mice. Stable tumor growth was established after subcutaneous (sc) and intravenous (iv) injection. However, TME of sc tumors was infiltrated by less than 1% immune cells, while myc-driven lymphoma in humans usually show substantial immune infiltration. In contrast to sc tumors, systemically growing lymphoma in murine bone marrow (BM) are infiltrated by 30% myeloid cells and 1% T-cells and in murine spleen by 10% and 30%, respectively. Myeloid cells found in these tumors were shown to suppress T cell proliferation in vitro. To test functionality of the h/mCD22 transgene, lymphoma-bearing mice were treated with Moxetumomab, which reduced BM lymphoma infiltration by 20 to 100-fold and infiltration in spleen by 5 to 20-fold in the three lymphoma models. Effects of treatment on TME were analyzed after treatment with Doxorubicin which is known to activate myeloid cells in vivo. Compared to untreated controls, Doxorubicin increased CD11b+ cells in spleen by 1.5-fold. Among these cells, Ly6G+ granulocytic cells increased most substantially.ConclusionsWe established primary, myc-driven h/mCD22+ B-cell lymphoma which stably engraft in syngeneic mice with a TME mimicking myc-driven lymphoma in men. The model responds well to CD22-targeted therapy and Doxorubicin induces expected immunologic changes. Therefore, our unique model provides a platform to test CD22-targeting treatment strategies in an immune competent background.Disclosure InformationF. Gsottberger: None. C. Brandl: None. S. Petkovic: None. L. Nitschke: None. A. Mackensen: None. F. Müller: None.

Simulated microgravity-mediated reversion of murine lymphoma immune evasion

No human has returned to the moon since the end of the Apollo program 47 years ago, however, new missions are planned for an orbital outpost. Space radiation and the potential for cancer remain as important issues to the future of human space exploration. While improved shield technologies and protective biologicals are under development, little is known concerning the interaction between cancer cells and host immunity in microgravity. As a hallmark of cancer, tumor cells employ mechanisms of immune evasion to avoid elimination by protective CD4+ and CD8+ T cells. We showed that a murine lymphoma was able to produce a soluble factor that inhibited the function of dendritic cells in activating the CD4+ T cells. Culture of the lymphoma cells in simulated microgravity (SMG), and not Static conditions, restored the CD4+ T cell response and augmented CD8+ T cell-mediated destruction of the cancer cells in vitro and in vivo. Thus, SMG impaired the mechanism of tumor escape and rendered the cancer cells more susceptible to T cell-mediated elimination. The stress of microgravity may expose the most critical components of a tumor’s escape mechanism for astronaut protection and the generation of new cancer therapeutics for patients on Earth.