Donor Memory-like NK cells Persist and Induce Remissions in Pediatric Patients with Relapsed AML after Transplant

Pediatric and young adult (YA) patients with acute myeloid leukemia (AML) who relapse after allogeneic hematopoietic cell transplantation (HCT) have extremely poor prognosis. Standard salvage chemotherapy and donor lymphocyte infusions (DLI) have little curative potential. Previous studies showed that natural killer (NK) cells can be stimulated ex vivo with interleukin-12 (IL-12), IL-15, and IL-18 to generate memory-like (ML) NK cells with enhanced anti-leukemia responses. We treated nine pediatric/YA patients with post-HCT relapsed AML with donor ML NK cells on a phase I trial. Patients received fludarabine, cytarabine and filgrastim followed two weeks later by infusion of DLI and ML NK cells from the original HCT donor. ML NK cells were successfully generated from haploidentical, matched-related and matched-unrelated donors. Following infusion, donor-derived ML NK cells expanded and maintained ML multidimensional mass cytometry phenotype for over 3 months. Furthermore, ML NK cells exhibited persistent functional responses as evidenced by leukemia-triggered IFN-g production. Following DLI and ML NK cell adoptive transfer, 4 of 8 evaluable patients achieved complete remission at day 28. Two patients maintained a durable remission for over 3 months with one patient in remission for greater than two years. No significant toxicity was experienced. This study demonstrates that in a compatible immune environment post-HCT, donor ML NK cells robustly expand and persist with potent anti-leukemic activity in the absence of exogenous cytokines. ML NK cells in combination with DLI present a novel immunotherapy platform for AML that has relapsed after allogeneic HCT. This trial was registered at www.clinicaltrials.gov as #NCT03068819.

B cell residency but not T cell–independent IgA switching in the gut requires innate lymphoid cells

Immunoglobulin A (IgA)–producing plasma cells derived from conventional B cells in the gut play an important role in maintaining the homeostasis of gut flora. Both T cell–dependent and T cell–independent IgA class switching occurs in the lymphoid structures in the gut, whose formation depends on lymphoid tissue inducers (LTis), a subset of innate lymphoid cells (ILCs). However, our knowledge on the functions of non-LTi helper-like ILCs, the innate counter parts of CD4 T helper cells, in promoting IgA production is still limited. By cell adoptive transfer and utilizing a unique mouse strain, we demonstrated that the generation of IgA-producing plasma cells from B cells in the gut occurred efficiently in the absence of both T cells and helper-like ILCs and without engaging TGF-β signaling. Nevertheless, B cell recruitment and/or retention in the gut required functional NKp46−CCR6+ LTis. Therefore, while CCR6+ LTis contribute to the accumulation of B cells in the gut through inducing lymphoid structure formation, helper-like ILCs are not essential for the T cell–independent generation of IgA-producing plasma cells.

Is It Feasible to Use CMV-Specific T-Cell Adoptive Transfer as Treatment Against Infection in SOT Recipients?

During the last decade, many studies have demonstrated the role of CMV specific T-cell immune response on controlling CMV replication and dissemination. In fact, it is well established that transplanted patients lacking CMV-specific T-cell immunity have an increased occurrence of CMV replication episodes and CMV-related complications. In this context, the use of adoptive transfer of CMV-specific T-cells has been widely investigated and applied to Hematopoietic Stem Cell Transplant patients and may be useful as a therapeutic alternative, to reconstitute the CMV specific T-cell response and to control CMV viremia in patients receiving a transplantation. However, only few authors have explored the use of T-cell adoptive transfer in SOT recipients. We propose a novel review in which we provide an overview of the impact of using CMV-specific T-cell adoptive transfer on the control of CMV infection in SOT recipients, the different approaches to stimulate, isolate and expand CMV-specific T-cells developed over the years and a discussion of the possible use of CMV adoptive cellular therapy in this SOT population. Given the timeliness and importance of this topic, we believe that such an analysis will provide important insights into CMV infection and its treatment/prevention.

Lack of Effect of Murine Norovirus Infection on the CD4+ CD45RBhigh T-cell Adoptive Transfer Mouse Model of Inflammatory Bowel Disease

Murine norovirus (MNV) infection is highly prevalent in laboratory mice. Although MNV infection does not typically induce clinical disease in most laboratory mice, infection may nonetheless affect mouse models of disease by altering immune responses. We previously reported that MNV altered the bacterial-induced mouse model of inflammatory bowel disease (IBD) using Helicobacter-infected Mdr1a–/– mice. Therefore, we hypothesized that MNV infection would exacerbate another mouse model of IBD, the T-cell adoptive transfer (AT) model. In this model, Helicobacter infection is used to accelerate the progression of IBD induced by AT of naïve CD4+CD45RBhigh T cells into B6.129S7- Rag1tm1Mom/J (Rag1–/–) mice. We evaluated the effects of MNV infection in both Helicobacter-accelerated as well as Helicobacter-free AT models. In our studies, Helicobacter-infected Rag1–/– mice that received CD4+CD45RBhigh T cells through AT rapidly developed weight loss and typhlocolitis; MNV infection had no effect on disease severity or rate of progression. In the absence of Helicobacter infection, progression of IBD caused by AT of CD4+CD45RBhigh T cells was slower and typhlocolitis was less severe; this inflammation likewise was unaltered by MNV infection. These results indicate that MNV infection does not alter IBD progression and severity in the CD4+CD45RBhigh T-cell AT model in Rag1–/– mice.

Cytokine-Induced Memory-like (ML) NK Cells Persist for > 2 Months Following Adoptive Transfer into Leukemia Patients with a MHC-Compatible Hematopoietic Cell Transplant (HCT)

Natural killer (NK) cells exhibit innate memory or memory-like responses following stimulation with haptens, viruses, or cytokines. Human memory-like (ML) NK cells differentiate following a short-term activation with IL-12, IL-15, and IL-18, and have increased anti-tumor activity against AML and other cancers in vitro, in xenograft models, and in the first-in human phase 1 clinical trial of ML NK cells in AML (PMID27655849). In this trial, CR/Cri was observed in >50% of patients treated, and mass cytometry revealed a unique multi-dimensional phenotype of in vivo differentiated ML NK cells that was confirmed using donor-specific HLA markers. Although adoptively transferred MHC-haploidentical ML NK cells expanded and differentiated over 2-3 weeks, these cells were eliminated by recipient allogeneic immune responses, a challenge observed with all allogeneic lymphocyte therapies. The immune rejection observed in the allogeneic setting precluded following ML NK phenotype, persistence, and function long-term in these patients. We hypothesized that ML NK cells could persist longer than 2-3 weeks in an MHC-compatible setting, and thus be able to assess ML NK cells durability. To test this idea, a phase 2 clinical trial was designed for relapsed/refractory AML patients, who receive a reduced-intensity HLA-haploidentical hematopoietic cell transplant (HCT), followed by same-donor ML NK cell adoptive transfer at day 7, with 2 weeks of IL-15/N-803 support (NCT02782546). Using mass cytometry, ML NK cells were confirmed as distinct from conventional NK cells, CD56hi/NKp30hi/CD62Lhi/KIR+/NKG2A+/CD57+/-, by viSNE analysis, and clearly inconsistent with immature NK cells arising from the HCT graft (CD56bright/KIR-/CD57-). ML NK cells persisted in patients for at least 2 months (n=5) following adoptive transfer, and constituted 20-50% of total NK cells at day 60 (n=3, 206±97 cells/μl; mean±SEM; peak ML NK cells = 751-1106 cells/µl, D21-D28, n=5). These ML NK cells appeared highly functional (56±8% IFN-γ+, 20±3% TNF+, 41±7% CD107a+) when stimulated with tumor targets immediately ex vivo on study day 28 (n=7). Unsupervised clustering of scRNA-seq from patient samples acquired 14-60 days after ML NK cell adoptive transfer identified a subset of NK cells transcriptionally distinct from conventional CD56bright and CD56dim. This NK cell population was the majority of NK cells at study Day 21 and remained identifiable 2 months post-transfer. In agreement with the mass cytometry data, these NK cells expressed high levels of KIRs with scRNA-seq analysis uncovering novel transcriptional changes in granzyme M, perforin, KLRG1, and IFNG suggesting ML NK cells represent a mature, activated NK cell subset distinct from conventional NK cells arising from the graft. In addition, scRNAseq analysis identified high expression of the transcription factor RUNX3, a potential regulator of ML NK cell phenotype in vivo. In conclusion, a single infusion of ML NK cells resulted in a durable population of highly functional NK cells, as evidenced by multi-dimensional analyses using mass cytometry and scRNA-sequencing. These studies provide evidence that ML NK cell therapy in the MHC-compatible setting overcomes persistence barriers and provide a platform for innovation in NK cell therapeutics. Disclosures Cashen: Celgene: Other: Speaker's Bureau; Seattle Genetics: Other: Speaker's Bureau; Novartis: Other: Speaker's Bureau. Fehniger:Cyto-Sen Therapeutics: Consultancy; Horizon Pharma PLC: Other: Consultancy (Spouse).

Male sex chromosomal complement exacerbates the pathogenicity of Th17 cells in a chronic model of CNS autoimmunity

SummarySex differences in the incidence and severity of multiple sclerosis (MS) have long been recognized. However, the underlying cellular and molecular mechanisms for why male sex is associated with more aggressive and debilitating disease remain poorly defined. Using an T cell adoptive transfer model of chronic EAE, we find that male Th17 cells induced disease of increased severity relative to female Th17 cells, irrespective of whether transferred to male or female recipients. Throughout the disease course, a greater frequency of male Th17 cells produced the heterodox cytokine IFNγ, a hallmark of pathogenic Th17 responses. Intriguingly, sex chromosomal complement, and not hormones, were responsible for the increased pathogenicity of male Th17 cells and an X-linked immune regulator, Jarid1c, was downregulated in both pathogenic male Th17 and CD4+ T cells from men with MS. Together, our data indicate that male sex critical regulates Th17 cell plasticity and pathogenicity via sex chromosomal complement.

Inhibition of CRTH2-mediated Th2 activation attenuates pulmonary hypertension in mice

Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by progressive pulmonary artery (PA) remodeling. T helper 2 cell (Th2) immune response is involved in PA remodeling during PAH progression. Here, we found that CRTH2 (chemoattractant receptor homologous molecule expressed on Th2 cell) expression was up-regulated in circulating CD3+CD4+ T cells in patients with idiopathic PAH and in rodent PAH models. CRTH2 disruption dramatically ameliorated PA remodeling and pulmonary hypertension in different PAH mouse models. CRTH2 deficiency suppressed Th2 activation, including IL-4 and IL-13 secretion. Both CRTH2+/+ bone marrow reconstitution and CRTH2+/+ CD4+ T cell adoptive transfer deteriorated hypoxia + ovalbumin–induced PAH in CRTH2−/− mice, which was reversed by dual neutralization of IL-4 and IL-13. CRTH2 inhibition alleviated established PAH in mice by repressing Th2 activity. In culture, CRTH2 activation in Th2 cells promoted pulmonary arterial smooth muscle cell proliferation through activation of STAT6. These results demonstrate the critical role of CRTH2-mediated Th2 response in PAH pathogenesis and highlight the CRTH2 receptor as a potential therapeutic target for PAH.