Mandy L. Ford

Background.: Costimulation blockade-based immunosuppression with belatacept confers improved renal function compared with calcineurin inhibitors, but comes at a cost of increased incidence and severity of early acute rejection. Strategies to limit costimulation blockade-resistant immune cells are needed. We have previously shown that the coinhibitory receptor T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domains (TIGIT) is expressed on memory T-cell subsets associated with costimulation blockade-resistant rejection in humans. Here, we aimed to determine the functional impact of TIGIT agonism on donor-reactive memory T-cell responses in the presence of costimulation blockade in a murine model of transplantation. Methods.: We used an experimental transplant model in which murine recipients possess donor-reactive CD8 ⁺ T cells before transplantation and were treated with either cytotoxic T-lymphocyte antigen 4 immunoglobulin (CTLA-4Ig) alone or CTLA-4Ig in the presence of an agonistic anti-TIGIT antibody. Results.: Results demonstrated that a significant prolongation in graft survival was observed in mice treated with CTLA-4Ig plus the TIGIT agonist compared with CTLA-4Ig alone. Furthermore, the number of graft-infiltrating donor-reactive CD8 ⁺ T cells was significantly decreased, and the number of splenic forkhead box P3 ⁺ CD4 ⁺ regulatory T cells was significantly increased, in animals treated with CTLA-4Ig + TIGIT agonist compared with CTLA-4Ig alone. Finally, CTLA-4Ig + TIGIT agonist treatment resulted in the suppression of inflammatory transcriptional programs within donor-reactive CD8 ⁺ memory T cells. Conclusions.: These data suggest that agonism of the TIGIT coinhibitory pathway may effectively inhibit costimulation blockade-resistant memory T-cell responses.

We investigated whether destroying malignant cells and the associated tumor microenvironment (TME) by focal gene therapy would broaden immune checkpoint inhibitor (ICI) effectiveness. We show that ICI antitumor activity against syngeneic (murine) triple-negative breast cancer (TNBC) was augmented when a therapeutic transgene (purine nucleoside phosphorylase , referred to here as E . coli PNP) was used to cleave fludarabine (2-fluoro-arabinofuranosyl adenine) to the anticancer purine base, 2-fluoroadenine (F-Ade). We also established strong repression of anatomically distant, non-PNP-expressing tumors being treated by the same strategy. TNBC cytoreduction was associated with decreased intratumoral PD1 ⁺ Tregs, increased granzyme B ⁺ NK cells, elevated MKI67 ⁺ T8 cells, and rapid immune clearance. Because F-Ade works by a mechanism that destroys quiescent neoplastic and supporting cells in the microenvironment, and since resistance to ICIs depends upon an intact TME, tumor killing by this approach offers a means to sensitize refractory malignancies to immune ablation and points to broad applicability against numerous cancer subtypes.

CD154 is a promising target for immunosuppression in transplantation, autoimmunity, and inflammatory diseases. We previously identified CD11b as a novel alternative receptor for CD154 during alloimmunity. However, the impact of specific CD154:CD11b blockade on immune responses to infection has not been well characterized. Here, we have shown that in contrast with its immunosuppressive effect on graft-specific CD8 ⁺ T cells, CD154:CD11b blockade unexpectedly improved both the quantity and quality of murine herpesvirus-68–specific CD8 ⁺ T cells as measured by an increase in tetramer-positive KLRG1 ^lo CD127 ^hi memory precursor effector cells. The differential effect of CD154:CD11b blockade on graft- versus virus-specific CD8 ⁺ T cells was underpinned by differences in phosphorylated S6 downstream of mTOR complex 1; however, differential expression of key transcription factors Eomes and TCF-1 was dictated by the type of antigen stimulus. These data demonstrate that priming conditions play an important role in determining the outcome of immunotherapy and suggest that specific inhibition of CD154:CD11b interactions could be effective for suppressing alloimmune responses while maintaining protective immunity to minimize infectious complications following transplantation.

While the inhibitory receptor FcγRIIB has been shown to be upregulated on activated CD8 ⁺ T cells in both mice and humans, its effect on T cell fate during infection has not been fully elucidated. We identified an increase in FcγRIIB-expressing CD8 ⁺ T cells in patients with COVID-19 relative to healthy controls as well as in mouse models of viral infection. Despite its well-known role as an Fc receptor, FcγRIIB also ligates the immunosuppressive cytokine Fgl2, resulting in CD8 ⁺ T cell apoptosis. Both chronic LCMV infection in mice and COVID-19 in humans resulted in a significant increase in plasma Fgl2. Transfer of CD8 ⁺ T cells into a Fgl2-replete, but not Fgl2-devoid, environment resulted in elimination of FcγRIIB ⁺ , but not FcγRIIB ^– , CD8 ⁺ T cells. Similarly, plasma Fgl2 was directly proportional to CD8 ⁺ T cell lymphopenia in patients with COVID-19. RNA-Seq analysis demonstrated that Fgl2 was produced by murine virus–specific CD8 ⁺ T cells, with an increase in Fgl2 in CD8 ⁺ T cells elicited during chronic versus acute viral infection. Fgl2 was also upregulated in CD8 ⁺ T cells from patients with COVID-19 versus healthy controls. In summary, CD8 ⁺ T cell production of Fgl2 during viral infection underpinned an FcγRIIB-mediated loss of CD8 ⁺ T cell immunity in both mice and humans.

Cancer immunotherapy has emerged as a promising therapeutic modality but heterogeneity in patient responsiveness remains. Thus, greater understanding of the immunologic factors that dictate response to immunotherapy is critical to improve patient outcomes. Here, we show that fibrinogen-like protein 2 (Fgl2) is elevated in the setting of melanoma in humans and mice and plays a functional role in inhibiting the CD8 ⁺ T cell response. Surprisingly, the tumor itself is not the major cellular source of Fgl2. Instead, we found that macrophage-secreted Fgl2 dampens the CD8 ⁺ T cell response through binding and apoptosis of FcγRIIB ⁺ CD8 ⁺ T cells. This regulation was CD8 ⁺ T cell autonomous and not via an antigen-presenting cell intermediary, as absence of Fcgr2b from the CD8 ⁺ T cells rendered T cells insensitive to Fgl2 regulation. Fgl2 is robustly expressed by macrophages in 10 cancer types in humans and in 6 syngeneic tumor models in mice, underscoring the clinical relevance of Fgl2 as a therapeutic target to promote T cell activity and improve patient immunotherapeutic response.