A Phase I Clinical Trial with Ex Vivo Expanded Recipient Regulatory T cells in Living Donor Kidney Transplants.

There is considerable interest in therapeutic transfer of regulatory T cells (Tregs) for controlling aberrant immune responses. Initial clinical trials have shown the safety of Tregs in hematopoietic stem cell transplant recipients and subjects with juvenile diabetes. Our hypothesis is that infusion(s) of Tregs may induce transplant tolerance thus avoiding long-term use of toxic immunosuppressive agents that cause increased morbidity/mortality. Towards testing our hypothesis, we conducted a phase I dose escalation safety trial infusing billions of ex vivo expanded recipient polyclonal Tregs into living donor kidney transplant recipients. Despite variability in recipient's renal disease, our expansion protocol produced Tregs which met all release criteria, expressing >98% CD4+CD25+ with <1% CD8+ and CD19+ contamination. Our product displayed >80% FOXP3 expression with stable demethylation in the FOXP3 promoter. Functionally, expanded Tregs potently suppressed allogeneic responses and induced the generation of new Tregs in the recipient's allo-responders in vitro. Within recipients, expanded Tregs amplified circulating Treg levels in a sustained manner. Clinically, all doses of Treg therapy tested were safe with no adverse infusion related side effects, infections or rejection events up to two years post-transplant. This study provides the necessary safety data to advance Treg cell therapy to phase II efficacy trials.

Comparison of the TCR zeta-chain with the FcR gamma-chain in chimeric TCR constructs for T cell activation and apoptosis.

A promising strategy for cancer treatment is adoptive gene therapy/immunotherapy by genetically modifying T cells with a chimeric T cell receptor (cTCR). When transduced T cells (T-bodies) specifically bind to tumor antigens through cTCR, they will become cytotoxic T lymphocytes (CTL) and lyse the tumor cells in a non-major histocompatibility complex (MHC)-restricted manner. Both the FcR gamma-chain and the TCR zeta-chain have been used to construct such cTCR, and both have shown specific cytolytic functions against tumor cells. However, most researchers believe that the zeta-chain generates stronger cytolytic activities against tumor than the gamma-chain and therefore would be a better candidate for cTCR construction. On the other hand, because of the lack of costimulation signaling in such constructs, the T-body might cause activation-induced T cell death (AICD) when bound to tumor antigens. Therefore, one can argue that the gamma-chain might generate less AICD than the zeta-chain because the gamma-chain has only one immunoreceptor tyrosine-based activation motif (ITAM), and the cytolytic activities can be therefore recycled. Two cTCR, GAHgamma and GAHzeta, were constructed and evaluated for cytokine production, specific cytolytic function and AICD in T-bodies after exposure to tumor cells. Using EGP-2-positive LS174T colorectal carcinoma cells as targets, there was no substantial difference observed between a gamma-chain or zeta-chain as the T-body signaling moiety in terms of specific cytolytic functions and induced cytokine production. This paper also demonstrates that, in the absence of a costimulation system, tumor antigen may not trigger apoptosis of T cells transduced with a cTCR carrying either an FcR gamma-chain or a TCR zeta-chain. These observations challenge current ideas about the role of ITAM in T cell activation.

Immune Modulation in Cancer Patients After Adoptive Transfer of Anti-CD3/Anti-CD28-Costimulated T Cells-Phase I Clinical Trial.

Anti-CD3/anti-CD28 monoclonal antibody-coactivated T cells (COACTs) proliferate, secrete tumoricidal cytokines, and mediate non-major histocompatibility complex (MHC)-restricted cytotoxicity. This phase I study was done to determine the safety, maximum tolerated dose, technical limits of expansion, and modulation of immune functions in cancer patients given COACTs. Coactivated T cells were produced by stimulating peripheral blood mononuclear cells (PBMCs) with OKT3 anti-CD3 and 9.3 (anti-CD28)-coated beads in the presence of 100 IU interleukin (IL)-2 per milliliter for 14 days. The beads were removed after 4 days of culture. Ten courses of COACTs were given to eight patients with renal cell (1), ovarian (2), breast (1), and colorectal (4) carcinomas; two patients received two courses of COACTs. Patients were given up to 10 x 10 9 COACTs twice a week for 3 weeks without dose-limiting toxicities. Patients at the first and second dose levels received a mean total of 17.6 and 42.4 x 10 9 COACTs, respectively. After 14 days of culture, the COACTs contained a mean of 57.5% CD4 + cells and 42.5% CD8 + cells, exhibited non-MHC-restricted cytotoxicity, and produced significant amounts of interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, and granulocyte macrophage colony-stimulating factor (GM-CSF). Infusions were safe and induced measurable serum levels of IFNgamma, TNFalpha, and IL-4 in two patients. Peripheral blood mononuclear cells from patients who received COACTs secreted higher amounts of IFNgamma and GM-CSF on in vitro anti-CD3/anti-CD28 restimulation than PBMCs obtained before immunotherapy. The detection of cytokines in patient sera and enhanced in vitro production of cytokines by anti-CD3/anti-CD28-stimulated patient PBMCs after COACT infusions suggest that COACTs were modulating immune responses in cancer patients.