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Simple Summary: Natural Killer cells have shown promise to treat different malignancies. Several methods have been described to obtain fully activated NK cells for clinical use. Here, we use different cell culture media and different artificial antigen presenting cells to optimize a GMP compliant manufacturing method to obtain activated and expanded NK cells suitable for clinical use. Natural killer (NK) cells represent promising tools for cancer immunotherapy. We report the optimization of an NK cell activation–expansion process and its validation on clinical-scale. Methods: RPMI-1640, stem cell growth medium (SCGM), NK MACS and TexMACS were used as culture mediums. Activated and expanded NK cells (NKAE) were obtained by coculturing total peripheral blood mononuclear cells (PBMC) or CD45RA+ cells with irradiated K562mbIL15-41BBL or K562mbIL21-41BBL. Fold increase, NK cell purity, activation status, cytotoxicity and transcriptome profile were analyzed. Clinical-grade NKAE cells were manufactured in CliniMACS Prodigy. Results: NK MACS and TexMACs achieved the highest NK cell purity and lowest T cell contamination. Obtaining NKAE cells from CD45RA+ cells was feasible although PBMC yielded higher total cell numbers and NK cell purity than CD45RA+ cells. The highest fold expansion and NK purity were achieved by using PBMC and K562mbIL21-41BBL cells. However, no differences in activation and cytotoxicity were found when using either NK cell source or activating cell line. Transcriptome profile showed to be different between basal NK cells and NKAE cells expanded with K562mbIL21-41BBL or K562mbIL15-41BBL. Clinical-grade manufactured NKAE cells complied with the specifications from the Spanish Regulatory Agency. Conclusions: GMP-grade NK cells for clinical use can be obtained by using different starting cells and aAPC. [ABSTRACT FROM AUTHOR]

We have previously demonstrated the safety and feasibility of adoptive cell therapy with CD45RA− memory T cells containing severe acute respiratory syndrome coronavirus 2–specific T cells for patients with coronavirus disease 2019 from an unvaccinated donor who was chosen based on human leukocyte antigen compatibility and cellular response. In this study, we examined the durability of cellular and humoral immunity within CD45RA− memory T cells and the effect of dexamethasone, the current standard of care treatment, and interleukin-15, a cytokine critically involved in T-cell maintenance and survival. We performed a longitudinal analysis from previously severe acute respiratory syndrome coronavirus 2–infected and infection-naïve individuals covering 21 months from infection and 10 months after full vaccination with the BNT162b2 Pfizer/BioNTech vaccine. We observed that cellular responses are maintained over time. Humoral responses increased after vaccination but were gradually lost. In addition, dexamethasone did not alter cell functionality or proliferation of CD45RA- T cells, and interleukin-15 increased the memory T-cell activation state, regulatory T cell expression, and interferon gamma release. Our results suggest that the best donors for adoptive cell therapy would be recovered individuals and 2 months after vaccination, although further studies with larger cohorts would be needed to confirm this finding. Dexamethasone did not affect the characteristics of the memory T cells at a concentration used in the clinical practice and IL-15 showed a positive effect on SARS-CoV-2-specific CD45RA- T cells. [Display omitted] [ABSTRACT FROM AUTHOR]

Summary: Intestinal passenger T leukocytes are responsible for graft‐versus‐host disease (GvHD) in intestinal transplantation (ITx). We hypothesized that ex vivo fludarabine treatment of the bowel graft would diminish the risk of GvHD and improve overall survival post‐transplant. We performed isolated heterotopic small bowel transplantations from Lewis (LEW) to Brown Norway (BN) rat strains, which generated GvHD signs from the fourth day post‐transplant. These symptoms included rash, weight loss, piloerection, and diarrhea. The grafts of one of the experimental groups were immersed and sealed in cold Celsior preservation solution with 1000 µm fludarabine for 1 h, prior to its implantation into recipient animals. No histological signs of intestinal tissue alterations were observed after fludarabine treatment. Fludarabine‐treated bowel recipients showed significantly later and milder clinical signs of GvHD and reduced total donor cell chimerism, as determined by flow cytometry using strain‐specific anti‐HLA antibodies. Additionally, fludarabine treatment prolonged recipients' overall survival (13.5 days ± 0.3 days vs. 9.2 days ± 0.5). We conclude that active modification of the intestinal leukocyte composition is advantageous in our ITx animal model. Immunosuppression with fludarabine during the surgical procedure, which could be translated directly to the clinic, protects bowel recipients from GvHD and improves overall post‐transplant survival. [ABSTRACT FROM AUTHOR]

Chimeric Antigen Receptor (CAR) T-cell therapy has demonstrated high rates of response in recurrent B-cell Acute Lymphoblastic Leukemia in children and young adults. Despite this success, a fraction of patients' experience relapse after treatment. Relapse is often preceded by recovery of healthy B cells, which suggests loss or dysfunction of CAR T-cells in bone marrow. This site is harder to access, and thus is not monitored as frequently as peripheral blood. Understanding the interplay between B cells, leukemic cells, and CAR T-cells in bone marrow is paramount in ascertaining the causes of lack of response. In this paper, we put forward a mathematical model representing the interaction between constantly renewing B cells, CAR T-cells, and leukemic cells in the bone marrow. Our model accounts for the maturation dynamics of B cells and incorporates effector and memory CAR T-cells. The model provides a plausible description of the dynamics of the various cellular compartments in bone marrow after CAR T infusion. After exploration of the parameter space, we found that the dynamics of CAR T product and disease were independent of the dose injected, initial B-cell load, and leukemia burden. We also show theoretically the importance of CAR T product attributes in determining therapy outcome, and have studied a variety of possible response scenarios, including second dosage schemes. We conclude by setting out ideas for the refinement of the model. [ABSTRACT FROM AUTHOR]