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Introduction. Chronic graft-versus-host disease (cGVHD) can manifest as a complication in patients following allogeneic hematopoietic stem cell transplant resulting in morbidity and mortality. Effective treatment strategies for cGVHD are currently lacking. Ibrutinib, an irreversible BTK inhibitor with activity against several other Tec-family kinases, has been clinically developed for cGVHD treatment due to regulation of pathogenetic B cells and T-cell subsets. Vecabrutinib is a more selective, reversible inhibitor of BTK with a distinct kinase domain interaction, which could result in differentiated safety and activity profiles compared to ibrutinib. Vecabrutinib has the capacity to overcome the C481S mutation that mediates resistance to ibrutinib. Vecabrutinib also also demonstrates activity against ITK, which is expressed in T cells. In this study, we investigated the activity and immune modulation of vecabrutinib treatment in a murine model of sclerodermatous cGVHD. Ibrutinib was utilized for comparison. Methods. A murine model of sclerodermatous cGVHD was initiated by adoptive transfer of T-cell depleted bone marrow plus whole splenocytes from B10.D2 donors into BALB/c recipients that were previously subjected to sub-lethal irradiation. Total bone marrow and irradiation only groups were included as controls. Mice with established cGVHD characterized by weight loss and skin irritation symptoms were treated with vecabrutinib once daily at 50mg/kg by oral gavage or ibrutinib at 30mg/kg in drinking water 5 days per week for approximately 3 weeks beginning on day 27 post-adoptive transfer and ending on day 45 (n=10 mice per group). Body weight and clinical symptoms (appearance, activity, skin symptoms, diarrhea, conjunctivitis) were measured throughout the study. Immunophenotyping for B cells and T cells was performed on spleens collected from euthanized mice by flow cytometry at two timepoints (day 40 during treatment and on day 76 post-treatment). Levels of circulating immunoglobulins were measured by multiplex cytokine assay at both timepoints. Results. Clinical symptoms, including appearance, activity, skin irritation, redness, alopecia and diarrhea were significantly reduced in vecabrutinib- and ibrutinib-treated groups. Furthermore, there was a trend toward a more favorable clinical score overall for the vecabrutinib-treated group, however no statistical difference was observed compared to ibrutinib possibly due to small sample size. Weight loss was slightly elevated during vecabrutinib and ibrutinib treatment compared to vehicle (trend), however mice recovered body weight post-treatment and continued to maintain benefit. On day 40 (during treatment) and day 76 (post-treatment) groups of mice were euthanized for immunophenotyping analysis utilizing a 22-color flow cytometry panel. During treatment, both vecabrutinib and ibrutinib-treated mice retained total B cell numbers but exhibited reduced B-cell activation, proliferation, and number of B220+ CD138+ plasma cells. In addition, B cells secreted less IL-10, and IL-4/5. Expression of antigen-presentation molecules CD80 and CD86 on B cells were unchanged. Total CD3+ T cells, activated and proliferating CD4+ and CD8+ T cells, and cytotoxic granzyme-B+ CD8+ T cells were reduced in treated mice. Interestingly, CD4+ CD25+ FoxP3+ Treg number, expression of PD-1 on Tregs and CD4+ CXCR5+ PD-1+ T follicular helper cells were also reduced in both treatment groups. In addition, there were globally reduced numbers of cytokine-secreting CD4+ cells but no differences in Th1/Th2 or Th17/Treg ratios were observed. Post-treatment, proliferation and cytokine secretion of B cells and T cells were still lowered but less impaired than during treatment. Tregs and PD-1 expression were still reduced post-treatment. Finally, circulating levels of IgA were reduced during and post treatment in vecabrutinib-treated mice compared to vehicle, while IgG1, IgG2b were reduced in both treated groups. No changes in IgM levels were observed in either treatment group. In conclusion, vecabrutinib treatment demonstrated efficacy and beneficially regulated B cell and T cell immune subsets in a preclinical murine model of sclerodermatous cGVHD. Studies to further evaluate differences between vecabrutinib and ibrutinib treatment are ongoing. Figure 1 Figure 1. Disclosures Fox: Sunesis Pharmaceuticals: Current Employment. Taverna: Sunesis Pharmaceuticals: Current Employment. Pinilla Ibarz: Sellas: Other: ), patents/royalties/other intellectual property; MEI, Sunesis: Research Funding; AbbVie, Janssen, AstraZeneca, Takeda: Speakers Bureau; AbbVie, Janssen, AstraZeneca, Novartis, TG Therapeutics, Takeda: Consultancy, Other: Advisory.