Building a CAR-Treg: Going from the basic to the luxury model.

Regulatory T cells (Tregs) control immune homeostasis and prevent exacerbated immune responses, and can be used as cell therapy to dampen a variety of autoimmune or autoinflammatory responses. Treg therapy is significantly more effective if the cells are antigen-specific. One way to re-direct the specificity of Tregs is to engineer them to express a Chimeric Antigen Receptor (CAR). Proof-of-concept studies have shown the potential for "basic" models of CAR-Tregs to be used as cellular therapy in autoimmunity, organ transplantation and hematopoietic stem cell transplantation. In parallel, work in the context of cancer has significantly advanced knowledge of how to optimise CAR-T cell structure and function for more precise and potent function. In this review, we summarize the current state of knowledge about important considerations when generating CAR-Tregs. We also extrapolate from emerging findings with CAR-T cells about strategies to further improve CAR-Treg function, creating "luxury" models with refined activity.

Functional effects of chimeric antigen receptor co-receptor signaling domains in human regulatory T cells.

Antigen-specific regulatory T cells (Tregs) engineered with chimeric antigen receptors (CARs) are a potent immunosuppressive cellular therapy in multiple disease models and could overcome shortcomings of polyclonal Treg therapy. CAR therapy was initially developed with conventional T cells, which have different signaling requirements than do Tregs To date, most of the CAR Treg studies used second-generation CARs, encoding a CD28 or 4-1BB co-receptor signaling domain and CD3ζ, but it was not known if this CAR design was optimal for Tregs Using a human leukocyte antigen-A2-specific CAR platform and human Tregs, we compared 10 CARs with different co-receptor signaling domains and systematically tested their function and CAR-stimulated gene expression profile. Tregs expressing a CAR encoding CD28wt were markedly superior to all other CARs tested in an in vivo model of graft-versus-host disease. In vitro assays revealed stable expression of Helios and an ability to suppress CD80 expression on dendritic cells as key in vitro predictors of in vivo function. This comprehensive study of CAR signaling domain variants in Tregs can be leveraged to optimize CAR design for use in antigen-specific Treg therapy.

New signatures of poor CD4 cell recovery after suppressive antiretroviral therapy in HIV-1-infected individuals: involvement of miR-192, IL-6, sCD14 and miR-144.

Up to 40% of newly diagnosed cases of HIV-1 infection are late diagnoses, with a profound decrease in CD4 cell counts in many cases. One-third of these individuals do not achieve optimal CD4 cell recovery (OR) after suppressive antiretroviral treatment (ART). This retrospective/longitudinal study of poor recovery (PR) included 79 HIV-1-infected individuals with CD4 count <200 cells/mm3 (25 PR and 54 OR) before ART. After suppressive ART, 21 PR and 24 OR individuals were further analysed, including paired samples. Selected miRs and plasma inflammatory markers were determined to investigate their potential predictive/diagnostic value for poor recovery. miR-192, IL-6 and sCD14 were independently associated with CD4 recovery before ART (p = 0.031, p = 0.007, and p = 0.008, respectively). The combination of these three factors returned a good discrimination (predictive value for PR) value of 0.841 (AUC, p < 0.001). After suppressive ART, miR-144 was independently associated with CD4 recovery (p = 0.017), showing a moderate discrimination value of 0.730 (AUC, p = 0.008) for PR. Our study provides new evidence on the relationship between miRs and HIV-1 infection that could help improve the management of individuals at HIV-1 diagnosis. These miRs and cytokines signature sets provide novel tools to predict CD4 cell recovery and its progression after ART.

Glutaminolysis and lipoproteins are key factors in late immune recovery in successfully treated HIV-infected patients.

The immunological, biochemical and molecular mechanisms associated with poor immune recovery are far from known, and metabolomic profiling offers additional value to traditional soluble markers. Here, we present novel and relevant data that could contribute to better understanding of the molecular mechanisms preceding a discordant response and HIV progression under suppressive combined antiretroviral therapy (cART). Integrated data from nuclear magnetic resonance (NMR)-based lipoprotein profiles, mass spectrometry (MS)-based metabolomics and soluble plasma biomarkers help to build prognostic and immunological progression tools that enable the differentiation of HIV-infected subjects based on their immune recovery status after 96 weeks of suppressive cART. The metabolomic signature of ART-naïve HIV subjects with a subsequent late immune recovery is the expression of pro-inflammatory molecules and glutaminolysis, which is likely related to elevate T-cell turnover in these patients. The knowledge about how these metabolic pathways are interconnected and regulated provides new targets for future therapeutic interventions not only in HIV infection but also in other metabolic disorders such as human cancers where glutaminolysis is the alternative pathway for energy production in tumor cells to meet their requirement of rapid proliferation.

Higher levels of IL-6, CD4 turnover and Treg frequency are already present before cART in HIV-infected subjects with later low CD4 recovery.

Immunological characterization of HIV-infected subjects with low CD4-recovery (LR-subjects) has been extensively performed after a variable period of combined antiretroviral therapy (cART). We now explore immunological alterations present before the cART onset. In a case-control study, we selected pre-cART samples of HIV-subjects with and without low CD4-recovery after cART (n = 21 per group). CD4 T-cell activation, senescence and exhaustion related markers were not found specifically altered before cART initiation. On the other hand, we found that LR-subjects before cART already showed increased levels of IL6 (p = 0.009) and increased frequencies of Ki67+CD4+ T-cells (p = 0.026), CD45RA-CD27+CD4+ T-cells (p = 0.008) and Treg (p = 0.001), as well as increased expression of CD95 and CD127 on CD4 T-cells (p = 0.016; p = 0.032, respectively). These parameters characterize the immunological damage in LR-subjects before the cART onset and could be associated to the mechanisms hindering the subsequent CD4 recovery.

CCR5+ CD8 T-cell levels and monocyte activation precede the onset of acute coronary syndrome in HIV-infected patients on antiretroviral therapy.

Acute coronary syndrome (ACS) is nowadays one of the leading causes of morbid-mortality in HIV-infected population, but innate and adaptive immune mechanisms preceding this event are unknown. In this work we comprehensively and longitudinally observed, by multiparametric flow cytometry and following a case-control design, increased CCR5+CD8+ T-cells levels and monocytes expressing activation and adhesion markers in HIV-infected patients who are going to suffer ACS. In addition, we found direct associations between activated CD8+ T-cells and myeloid cells that were only statistically significant in the group of patients with ACS and in the follow up time point just before the ACS. Our data highlight the important role of CCR5 in the onset of ACS and suggest this receptor as a marker of cardiovascular risk and potential therapeutic target to prevent the development of such non-AIDS-related event in HIV-infected patients.