Virtual memory T cells orchestrate extralymphoid responses conducive to resident memory

A primary immune response is initiated in secondary lymphoid organs. Virtual memory CD8+ T (TVM) cells are antigen-inexperienced T cells of a central memory phenotype, acquired through self-antigen–driven homeostatic proliferation. Unexpectedly, we find that TVM cells are composed of CCR2+ and CCR2− subsets that differentially elaborate a spectrum of effector- and memory-poised functions directly in the tissue. During a primary influenza infection, TVM cells rapidly infiltrate the lungs in the first day after infection and promote early viral control. TVM cells that recognize viral antigen are retained in the tissue, clonally expand independent of secondary lymphoid organs, and give rise to tissue-resident memory cells. By orchestrating an extralymphoid primary response, heterogenous TVM cells bridge innate reaction and adaptive memory directly in the infected tissue.

Adaptive immunity to human coronaviruses is widespread but low in magnitude

Endemic human coronaviruses (hCoV) circulate worldwide but cause minimal mortality. Although seroconversion to hCoV is near ubiquitous during childhood, little is known about hCoV-specific T cell memory in adults. We quantified CD4 T cell and antibody responses to hCoV spike antigens in 42 SARS-CoV-2 uninfected individuals. T cell responses were widespread within conventional memory and cTFH compartments but did not correlate with IgG titres. SARS-CoV-2 cross-reactive T cells were observed in 48% of participants and correlated with HKU1 memory. hCoV-specific T cells exhibited a CCR6+ central memory phenotype in the blood, but were enriched for frequency and CXCR3 expression in human lung draining lymph nodes. Overall, hCoV-specific humoral and cellular memory are independently maintained, with a shared phenotype existing among coronavirus-specific CD4 T cells. This understanding of endemic coronavirus immunity provides insight into the homeostatic maintenance of immune responses that are likely to be critical components of protection against SARS-CoV-2.

Loss of TET2 Uncouples Proliferative and Effector Functions in CAR T Cells

Chimeric antigen receptor (CAR) T cells have opened a new paradigm for the treatment of leukemia and lymphoma. Their production, however, is laborious, requiring tens of millions of CAR T cells per infusion. This constraint could be significantly alleviated if safe and more efficacious T cells could be generated. In a patient with chronic lymphocytic leukemia, treated with anti-CD19 CAR T cells, a recent report described the emergence of a single T cell clone that at its expansion peak accounted for 94% of circulating CAR T cells, coinciding with the development of cytokine release syndrome and tumor regression (Fraietta et. al. Nature 2018). Insertional mutagenesis in this T cell had disrupted an allele of TET2, an epigenetic regulator mediating the oxidation of 5-methylcytosine. The other allele appeared to bear an inherited hypomorphic variant, resulting in the near complete loss of TET2 function in this clone. To understand the mechanisms accounting for this chance clinical finding, we investigated the effect of TET2 loss in human T cells engineered to express different chimeric receptors. Using CRISPR/Cas9, we edited TET2 in T cells engineered to express a CD19-specific second-generation CAR encompassing the costimulatory domain of either CD28 or 4-1BB (Rv-1928z and Rv-19BBz). TET2 disruption enhanced the in vivo anti-tumor activity of Rv-19BBz but not Rv-1928z CAR T cells tested under stress test conditions using limiting CAR T cell doses (as previously described in a human B cell acute lymphoblastic leukemia (B-ALL) NALM6 model, Zhao et. al. Cancer Cell 2015). Since Rv-1928z induces potent effector differentiation but limited persistence compared to Rv-19BBz, we hypothesized that loss of TET2 could amplify the expansion and persistence of 4-1BB-costimulated T cells but not override the differentiation program imparted by Rv-1928z. To test this hypothesis, we utilized two orthogonal approaches known to limit exhaustion and increase persistence of CD28-costimulated CAR T cells, Rv-1928z co-expressed with 4-1BB ligand (Rv-1928z-41BBL) and 1928z driven by the TRAC promoter (TRAC-1928z). Disruption of TET2 enhanced the anti-tumor efficacy of both these CAR T cells and promoted acquisition of a central memory phenotype. However, over time (50-200 days), TET2-edited TRAC-1928z and Rv-1928z-41BBL attained a hyper-proliferative phenotype ultimately requiring euthanasia due to splenomegaly and extensive CAR T cell accumulation in various organs. Post-mortem analysis found no evidence of NALM6 in these mice. This was in contrast to stress test studies with Rv-1928z and Rv-19BBz where most mice succumbed to NALM6 progression. These observations established an essential role for CAR signaling in determining the phenotypic outcome of TET2 loss in T cells. To examine the long-term effects of TET2 disruption in the context of all 4 receptors, we treated human B-ALL bearing mice with curative doses of all 4 CAR T cells and followed them for up to 200 days. We found that all 4 CAR expressing TET2-edited T cells could eventually attain a hyper-proliferative phenotype, but with varying frequency depending on the CAR design (Rv-1928z-41BBL and TRAC-1928z > Rv-19BBz > Rv-1928z). To assess their effector function, NALM6-bearing mice were infused with adoptively transferred hyper-proliferative TET2-edited CAR T cells. Strikingly, these T cells were unable to elicit any tumor control, despite their maintaining a central memory phenotype as assessed by flow cytometry. This loss of effector function was observed for all 4 CAR T cell types, suggesting a discrepancy between function and flow cytometric phenotype. Transcriptional, methylation and genome accessibility studies revealed a unique T cell state wherein the proliferative program is uncoupled from effector response. We identified a unique transcriptional and epigenetic signature that is manifested in a loss of effector function while maintaining robust proliferation. This state stands in contrast to the classically described T cell exhaustion state where loss of effector function is preceded by loss of proliferative ability. TET2 disruption thus promotes a CAR T cell proliferative program that depends on the CAR design but does not in itself enhance anti-tumor activity. Disclosures Sadelain: Atara: Patents & Royalties, Research Funding; Mnemo: Patents & Royalties; Minerva: Other: Biotechnologies , Patents & Royalties; Fate Therapeutics: Patents & Royalties, Research Funding; Takeda: Patents & Royalties, Research Funding.

Breaking Bottlenecks for the TCR Therapy of Cancer

Immune checkpoint inhibitors have redefined the treatment of cancer, but their efficacy depends critically on the presence of sufficient tumor-specific lymphocytes, and cellular immunotherapies develop rapidly to fill this gap. The paucity of suitable extracellular and tumor-associated antigens in solid cancers necessitates the use of neoantigen-directed T-cell-receptor (TCR)-engineered cells, while prevention of tumor evasion requires combined targeting of multiple neoepitopes. These can be currently identified within 2 weeks by combining cutting-edge next-generation sequencing with bioinformatic pipelines and used to select tumor-reactive TCRs in a high-throughput manner for expeditious scalable non-viral gene editing of autologous or allogeneic lymphocytes. “Young” cells with a naive, memory stem or central memory phenotype can be additionally armored with “next-generation” features against exhaustion and the immunosuppressive tumor microenvironment, where they wander after reinfusion to attack heavily pretreated and hitherto hopeless neoplasms. Facilitated by major technological breakthroughs in critical manufacturing steps, based on a solid preclinical rationale, and backed by rapidly accumulating evidence, TCR therapies break one bottleneck after the other and hold the promise to become the next immuno-oncological revolution.

Systematic examination of T cell responses to SARS-CoV-2 versus influenza virus reveals distinct inflammatory profile

There is a pressing need for an in-depth understanding of immunity to SARS-CoV-2. Here we investigated T cell recall responses to fully glycosylated Spike trimer, recombinant N protein as well as to S, N, M and E peptide pools in the early convalescent phase. All subjects showed SARS-CoV-2-specific T cell responses to at least one antigen. SARS-CoV-2-specific CD4+ T cells were primarily of the central memory phenotype and exhibited a lower IFN-[gamma] to TNF-[alpha] ratio compared to influenza-specific responses of the same donors, independent of disease severity. SARS-CoV-2-specific T cells were less multifunctional than influenza-specific T cells, particularly in severe cases, potentially suggesting exhaustion. High IL-10 production was noted in response to N protein, possibly contributing to immunosuppression, with potential implications for vaccine design. We observed granzyme B+/IFN-[gamma] CD4+ and CD8+ proliferative responses to peptide pools in most individuals, with CD4+ responses predominating over CD8+ responses. Peripheral T follicular helper responses to S or N strongly correlated with serum neutralization assays as well as RBD-specific IgA. Overall, T cell responses to SARS-CoV-2 are robust, however, CD4+ Th1 responses predominate over CD8+ responses and are more inflammatory with a weaker Tfh response than influenza-specific CD4+ responses, potentially contributing to COVID-19 disease.

Rapid GMP-compliant expansion of SARS-CoV-2-specific T cells from convalescent donors for use as an allogeneic cell therapy for COVID-19

COVID-19 disease caused by the SARS-CoV-2 virus is characterized by dysregulation of effector T cells and accumulation of exhausted T cells. T cell responses to viruses can be corrected by adoptive cellular therapy using donor-derived virus-specific T cells. Here we show that SARS-CoV-2-exposed blood donations contain CD4 and CD8 memory T cells specific for SARS-CoV-2 spike, nucleocapsid and membrane antigens. These peptides can be used to isolate virus-specific T cells in a GMP-compliant process. These T cells can be rapidly expanded using GMP-compliant reagents for use as a therapeutic product. Memory and effector phenotypes are present in the selected virus-specific T cells, but our method rapidly expands the desirable central memory phenotype. A manufacturing yield ranging from 1010 to 1011 T cells can be obtained within 21 days culture. Thus, multiple therapeutic doses of virus-specific T cells can be rapidly generated from convalescent donors for treatment of COVID-19 patientsOne Sentence SummaryCD4+ and CD8+ T cells specific for SARS-CoV-2 can be isolated from convalescent donors and rapidly expanded to therapeutic doses at GMP standard, maintaining the desired central memory phenotype required for protective immune responses against severe COVID-19 infections.

Disease stage-specific pathogenicity of CD138 (syndecan 1)-expressing T cells in systemic lupus erythematosus

ABSTRACTObjectiveTo identify and characterize CD138 (syndecan 1)-expressing T cells in SLE-prone mice.MethodsWe characterized CD138-expressing T cells in MRL/Lpr mice by flow cytometry assay and by gene analysis. Functional properties of TCRβ+CD138+ cells were assessed either by activating through TCR or by co-incubating with purified B cells in the presence of auto-antigens. Purified TCRβ+CD138+ cells were adoptively transferred into MRL/Lpr mice and lupus disease was assessed by measuring serum auto-antibodies, proteinuria and by histopathological evaluation of kidney.ResultsWe found that the frequency of TCRβ+CD138+ cells was significantly higher in MRL/Lpr mice than in wild-type MRL mice (p < 0.01), and the increase in their numbers correlated with disease severity. Majority of the TCRβ+CD138+ cells were CD4 and CD8 double-negative and 20% were CD4. Compared to TCRβ+CD138− cells, TCRβ+CD138+ cells exhibited central memory phenotype with reduced ability to proliferate, and produce the cytokines IFNγ and IL-17. When co-cultured with B cells, the ability of TCRβ+CD138+ cells to promote plasma cell formation and autoreactive antibody production was dependent on the presence of auto-antigens and CD4 co-receptor expression. Surprisingly, adoptively transferred TCRβ+CD138+ cells slowed down the disease progression in young MRL/Lpr mice but had the opposite effect when DNA was co-administered or when TCRβ+CD138+ cells were transferred into older MRL/Lpr mice with established disease.ConclusionHere, we provided evidence for the pathogenic role of CD138-expressing T cells when auto-antigens are exposed to immune system. Thus, monitoring the changes in TCRβ+CD138+ cell-frequency may serve as a tool to assess SLE severity. Moreover, CD138-expressing T cells may be targeted to alleviate lupus progression.