Cross-Generational Reproductive Fitness Enforced by Microchimeric Maternal Cells

SummaryExposure to maternal tissue during in utero development imprints tolerance to immunologically foreign non-inherited maternal antigens (NIMA) that persists into adulthood. The biological advantage of this tolerance, conserved across mammalian species, remains unclear. Here, we show maternal cells that establish microchimerism in female offspring during development promote systemic accumulation of immune suppressive regulatory T cells (Tregs) with NIMA specificity. NIMA-specific Tregs expand during pregnancies sired by males expressing alloantigens with overlapping NIMA specificity, thereby averting fetal wastage triggered by prenatal infection and non-infectious disruptions of fetal tolerance. Therefore, exposure to NIMA selectively enhances reproductive success in second-generation females carrying embryos with overlapping paternally inherited antigens. These findings demonstrate that genetic fitness, canonically thought to be restricted to Mendelian inheritance, is enhanced in female placental mammals through vertically transferred maternal cells that promote conservation of NIMA and enforce cross-generational reproductive benefits

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Regulatory T Cell Suppressive Potency Dictates the Balance between Bacterial Proliferation and Clearance during Persistent Salmonella Infection

The pathogenesis of persistent infection is dictated by the balance between opposing immune activation and suppression signals. Herein, virulent Salmonella was used to explore the role and potential importance of Foxp3-expressing regulatory T cells in dictating the natural progression of persistent bacterial infection. Two distinct phases of persistent Salmonella infection are identified. In the first 3–4 weeks after infection, progressively increasing bacterial burden was associated with delayed effector T cell activation. Reciprocally, at later time points after infection, reductions in bacterial burden were associated with robust effector T cell activation. Using Foxp3 GFP reporter mice for ex vivo isolation of regulatory T cells, we demonstrate that the dichotomy in infection tempo between early and late time points is directly paralleled by drastic changes in Foxp3 + Treg suppressive potency. In complementary experiments using Foxp3 DTR mice, the significance of these shifts in Treg suppressive potency on infection outcome was verified by enumerating the relative impacts of regulatory T cell ablation on bacterial burden and effector T cell activation at early and late time points during persistent Salmonella infection. Moreover, Treg expression of CTLA-4 directly paralleled changes in suppressive potency, and the relative effects of Treg ablation could be largely recapitulated by CTLA-4 in vivo blockade. Together, these results demonstrate that dynami

<em>Listeria monocytogenes</em> Cytoplasmic Entry Induces Fetal Wastage by Disrupting Maternal Foxp3⁺ Regulatory T Cell-Sustained Fetal Tolerance

<div><p>Although the intracellular bacterium <em>Listeria monocytogenes</em> has an established predilection for disseminated infection during pregnancy that often results in spontaneous abortion or stillbirth, the specific host-pathogen interaction that dictates these disastrous complications remain incompletely defined. Herein, we demonstrate systemic maternal <em>Listeria</em> infection during pregnancy fractures fetal tolerance and triggers fetal wastage in a dose-dependent fashion. <em>Listeria</em> was recovered from the majority of concepti after high-dose infection illustrating the potential for <em>in utero</em> invasion. Interestingly with reduced inocula, fetal wastage occurred without direct placental or fetal invasion, and instead paralleled reductions in maternal Foxp3⁺ regulatory T cell suppressive potency with reciprocal expansion and activation of maternal fetal-specific effector T cells. Using mutants lacking virulence determinants required for <em>in utero</em> invasion, we establish <em>Listeria</em> cytoplasmic entry is essential for disrupting fetal tolerance that triggers maternal T cell-mediated fetal resorption. Thus, infection-induced reductions in maternal Foxp3⁺ regulatory T cell suppression with ensuing disruptions in fetal tolerance play critical roles in pathogenesis of immune-mediated fetal wastage.</p> </div

Proposed model for immune-mediated fetal wastage induced by prenatal infection that can occur with or without <i>in utero</i> pathogen invasion.

<p>After low dose infection during pregnancy, reductions in maternal regulatory T cell suppression unleash the activation of immune effectors enough to rapidly eliminate the pathogen. However, given the requirement for sustained expansion of maternal regulatory cell suppression in maintaining fetal tolerance, these reductions in suppressive potency also trigger immune-mediated fetal wastage. By comparison with higher dosage infection, blunted maternal regulatory T cell suppression that promotes immune activation does not eradicate infection as efficiently. In turn with ongoing disruption in fetal tolerance, remaining pathogen is drawn to inflammation at the uterine-placental interface that promotes invasion into the placental-fetal unit.</p

<i>Listeria monocytogenes</i> cytoplasmic entry disrupts fetal tolerance with infection during pregnancy.

<p>Representative FACS plots (top) and composite data (bottom) illustrating expansion and IFN-γ production by fetal-OVA-specific CD8 T cells among maternal splenocytes in mice impregnated by Actin-OVA males five days after LmΔactA (10⁷ CFUs) or LmΔLLOΔPLC (10⁸ CFUs) infection at midgestation. For IFN-γ production, cells were stimulated with OVA_257–264 peptide (black line) or no peptide controls (gray filled). Each data point represents results from an individual mouse combined from three independent experiments each with similar results.</p

<i>Listeria monocytogenes</i> infection triggers dose-dependent rates of fetal wastage during allogeneic pregnancy.

<p>(A) Number of live pups born with virulent WT Lm inoculated midgestation (E10.5) at the indicated dosages for pregnant C57Bl/6 females mated with Balb/c males. (B) Percent viable and resorbed fetuses five days after WT Lm infection at midgestation. (C) Percent viable and resorbed fetuses with recoverable Lm for the mice described in B. Each data point represents results from an individual mouse combined from three independent experiments each with similar results.</p