Patients with angioimmunoblastic T-cell lymphoma (AITL) and other peripheral T-cell lymphomas that harbor features of follicular helper T (T) cells have a very poor prognosis. These lymphomas commonly present with paraneoplastic autoimmunity and lymphopenia. RhoA G17V mutation is present in 60% of T-like lymphomas, but its role in tumorigenesis is poorly understood. We generated transgenic mice that express RhoA G17V under the control of murine CD4 regulatory elements at levels comparable to a heterozygous mutation (tgRhoA mice). These mice had markedly reduced naive T cells but relatively increased T-cell populations. Surprisingly, naive CD4 T cells expressing RhoA G17V were hyperreactive to T-cell receptor stimulation. All tgRhoA mice developed autoimmunity that included a cellular infiltrate within ears and tails that was recapitulated in wild-type (WT) recipients after bone marrow transplantation. Older tgRhoA mice developed elevated serum titers of anti-double-stranded DNA antibodies and renal immune complex deposition. RhoA G17V mice crossed with ; Vav-Cre mice, which delete throughout the hematopoietic compartment, developed T-cell lymphomas that retained histologic and immunophenotypic features of AITL and had transcriptional signatures enriched for mechanistic target of rapamycin (mTOR)-associated genes. Transplanted tumors were responsive to the mTOR inhibitor everolimus, providing a possible strategy for targeting RhoA G17V. Taken together, these data indicate that RhoA G17V contributes to both neoplastic and paraneoplastic phenotypes similar to those observed in patients with T lymphomas.
The RhoA G17V mutation occurs in 60% of tumors from patients with Angioimmunoblastic T-cell Lymphomas (AITL) as well as in a subset of Peripheral T-cell Lymphoma, not otherwise specified (PTCL-NOS). The non-malignant counterparts of AITL cells are posited to be follicular-helper T-cells (TFH), and RhoA G17V mutation is strongly associated with a TFH gene expression program in both AITL and PTCL-NOS cases. The G17V substitution is predicted to lock the small GTPase RhoA in an inactive conformation, generating a protein that can bind guanine exchange factors more avidly than unmutated RhoA, likely producing neomorphic effects by inhibiting other small GTPases and G-coupled protein receptors. We have generated mice that express RhoA G17V under the control of regulatory elements from the murine CD4 locus, including two separate founder lines with essentially identical developmental and functional phenotypes. These mice demonstrate a severe reduction in thymic and splenic T cells beginning at the thymic CD4/CD8 double-positive stage, consistent with effects reported from loss-of-function (LOF) RhoA models. RhoA G17V mice also show distinctive effects in peripheral T-cell compartments, including a marked reduction in splenic naïve T-cells with a relative increase in the proportion of activated and/or memory T-cells among those that remain. Notably, there is an increase in the proportion of TFHcells among RhoA G17V mice, suggesting that the mutant protein skews differentiation towards this lineage. Also, unlike reports from RhoA LOF models, we observe increased activation of RhoA G17V T cells when sub-optimal T-cell receptor (TCR)-pathway stimulation is performed with CD3 crosslinking, consistent with a neomorphic activity. We have also observed a striking inflammatory tail fibrosis that occurs with essentially complete penetrance by 8-10 weeks of age in RhoA G17V mice but not in littermate controls. Development of fibrosis is preceded by a cellular infiltrate of hematopoietic cells. including a prominent population of CD4+cells, suggesting that CD4-specific RhoA G17V expression promotes autoimmunity. Finally, we crossed RhoA G17V mice with mice lacking expression of TET2 in the hematopoietic compartment (via Vav-Cre-mediated deletion) and the ovalbumin-specific OT-II transgenic TCR. These mice developed spontaneous T-cell lymphomagenesis comprised of Bcl6-expressing CD4 cells and involving the liver, spleen, and peripheral lymph nodes. Further immunization of these mice with NP40-ovalbumin and alum resulted in Bcl-6-expressing T cell lymphomas within 1-4 months after initial immunization. Thus, the same genotype observed in >50% of AITLs (i.e., RhoA G17V/TET2 loss), combined with TCR stimulation rapidly produces T cell lymphomas with TFHphenotypes. In conclusion, T-cells with RhoA G17V mutations may overcome regulatory pathways before and after malignant transformation, including those that control activation and prevent autoreactivity. These phenotypes also provide a platform from which to determine the specific biochemical effects mediated by RhoA G17V that contribute to T-cell hyperactivation and transformation. Disclosures Weinstock: Novartis: Consultancy, Research Funding.
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