Generation of faithful autochthonous mouse models of Mantle Cell Lymphoma

Abstract Introduction: MantleCell Lymphoma (MCL) is characterized by the translocation t(11;14) resulting in overexpression of Cyclin D1. Other frequently observed alterations include SOX11 overexpression (90% of patients), and aberrations of ATM (42-56%) and TP53 (16-31%). Despite recent treatment advances, patients frequently relapse. However, insights into MCL biology to inform novel treatments have been limited by a lack of MCL in vivo models. We herein generated and characterized genetically engineered mouse models recapitulating the heterogeneity of MCL. Methods: To mirror human disease in vivo, we used the CD19Cre (C) allele to induce the following B cell-specific alterations in a C57BL/6 background: Overexpression of Ccnd1 (D), harboring the point mutation p.T286Ato improve protein stabilization, and Sox11 (S) from the Rosa26 locus. Moreover, we employed LoxP-flanked conditional alleles for homozygous loss-of-function of Atm (AA) and Trp53 (PP). First, we performed analyses on up to 34 terminal animals per genotype to study survival and associated phenotypes by histology including immunohistochemistry (IHC), flow cytometry, as well as whole-exome (WES), whole-transcriptome (WTS) and BCR-sequencing (BCR-seq). Additionally, the evaluation of timed animals to study onset and longitudinal changes is currently ongoing. Results: B cell-specific Cyclin D1 overexpression (CD) results in inferior median survival of 84 weeks, compared to C control mice (median survival: not reached). At terminal endpoint, mice developed splenomegaly >400mg at a rate of 79%. Affected animals displayed systemic disease with lymphadenopathy, infiltration of blood and bone marrow, as well as non-lymphoid organs, such as liver or the gastrointestinal tract. Histology revealed disrupted splenic architecture and subtotal infiltration by pleomorphic to monomorphic cells with lymphoma-like morphology. IHC and flow-cytometry show high Ki67 and an MCL-like immunophenotype (CD19+CD5+/-IgM+CD23-CD138-). Importantly, BCR-seq revealed presence of oligo- or monoclonal disease without evidence of somatic hypermutation and class-switch-recombination, indicating a germinal-center (GC)-inexperienced lymphoma. Additionally, WES showed acquired genetic alterations commonly observed in human MCL, including non-synonymous mutations in Trp53 and Map3k14, as well as copy number variations affecting Card11 and Cdkn2a. Compared to this CD model, we observed accelerated lymphomagenesis with significantly shorter median survival times of 51 (AACD) and 19 (AACDPP) weeks, and higher lymphoma penetrance of 97% and 94%, respectively. AACD shows consistent phenotypic features to CD, with systemic infiltration by mainly monomorphic, blastoid-appearing lymphoma and an MCL-like phenotype (CD19+CD5+/-IgM+CD23-CD138-). Molecular analyses show high B-cell clonality with comparable genetic alterations as CD. Ultimately, we generated a Sox11-overexpressing mouse model (CS) to cover the main MCL hallmarks. These mice show splenomegaly development in 42% of cases and a median survival of 67 weeks. Splenic tissue is infiltrated by largely monoclonal lymphomas with an MCL-like immunophenotype (CD19+CD5+IgM+CD23-CD138-Ki67hi). Transcriptome analysis further revealed downregulation of GC-regulator Bcl-6, potentially abrogating GC formation and inducing pre-GC-derived lymphomagenesis. Additionally, we observe upregulation of SWI/SNF-subunit SMARCA4, which is associated with aggressive SOX11-positive MCL and highlights CS as a valuable tool to study the impact of SOX11 overexpression on MCL lymphomagenesis. Conclusion: Here, we show that B cell-specific Cyclin D1 overexpression results in MCL-like lymphomagenesis with high penetrance in vivo. Additional integration of Atm and/or Trp53 knock-out further accelerated lymphomagenesis and penetrance with features consistent with human MCL. Likewise, Sox11 overexpression alone drives MCL-like lymphomagenesis. Further in-depth characterization of these models, including CDS, is ongoing. In summary, we successfully established a range of novel in vivo models to accelerate preclinical and translational MCL research. Reflecting MCL hallmarks in lymphomagenesis and tumor microenvironment constitution, these are ideal tools to identify targetable vulnerabilities and develop novel rationale MCL treatments.