Differentiation of naive cord-blood T cells into CD19-specific cytolytic effectors for posttransplantation adoptive immunotherapy

Abstract

Disease relapse is a barrier to achieving therapeutic success after unrelated umbilical cord-blood transplantation (UCBT) for B-lineage acute lymphoblastic leukemia (B-ALL). While adoptive transfer of donor-derived tumor-specific T cells is a conceptually attractive approach to eliminating residual disease after allogeneic hematopoietic stem cell transplantation, adoptive immunotherapy after UCBT is constrained by the difficulty of generating antigen-specific T cells from functionally naive umbilical cord-blood (UCB)-derived T cells. Therefore, to generate T cells that recognize B-ALL, we have developed a chimeric immunoreceptor to redirect the specificity of T cells for CD19, a B-lineage antigen, and expressed this transgene in UCB-derived T cells. An ex vivo process, which is compliant with current good manufacturing practice for T-cell trials, has been developed to genetically modify and numerically expand UCB-derived T cells into CD19-specific effector cells. These are capable of CD19-restricted cytokine production and cytolysis in vitro, as well as mediating regression of CD19 ؉ tumor and being selectively eliminated in vivo. Moreover, time-lapse microscopy of the genetically modified T-cell clones revealed an ability to lyse CD19 ؉ tumor cells specifically and repetitively. These data provide the rationale for infusing UCB-derived CD19-specific T cells after UCBT to reduce the incidence of CD19 ؉ B-ALL relapse. IntroductionBanked unrelated umbilical cord blood (UCB) is source of hematopoietic stem cells for patients with B-lineage acute lymphoblastic leukemia (B-ALL). 1 However, despite maximally intensive preparative regimens, disease-relapse remains a significant cause of mortality after umbilical cord-blood transplantation (UCBT).Adoptive therapy after allogeneic hematopoietic stem cell transplantation (HSCT) with ex vivo-expanded donor-derived tumor-specific T cells might be used to augment the graft-versusleukemia (GVL)-effect, thereby reducing the incidence of leukemic relapse, without exacerbating graft-versus-host disease (GVHD). 2,3 While the feasibility of isolating, expanding, and infusing antigen-specific ␣␤ T-cell receptor (TCR) ϩ T cells from peripheral blood (PB) has been validated in animals and humans, 4-10 the naive function of neonatal T cells precludes a priori identification of resident tumor-specific T cells. Redirecting the specificity of T cells through enforced expression of antigenspecific immunoreceptors and differentiating UCB-derived T cells into cytotoxic T lymphocytes (CTLs) is one approach to overcoming this lack of endogenous tumor-specific T cells specific for desired targets. 11,12 We and others are developing adoptive immunotherapy platforms using single-chain chimeric immunoreceptors to redirect the specificity of primary human T cells and NK cells for cell-surface proteins expressed on tumor targets, such as the B-lineage-specific antigen CD19, a molecule expressed on normal and neoplastic B cells. [13][14][15][16][17][18][19][20][21][22] These chimeric immunoreceptor...

Ex vivo expansion of umbilical cord blood stem cells for transplantation: growing knowledge from the hematopoietic niche

“…This technique has been validated by timelapse microscopy in NOD/SCID mice. 151 Another adoptive immunotherapy technique designed to decrease the infectious complications would be to expand UCB T-cells using anti-CD3/anti-CD28-coated beads with subsequent expansion for reinfusion post transplant. 152 Both of these techniques will require validation in phase I trials.…”

Chimeric T-cell receptors: new challenges for targeted immunotherapy in hematologic malignancies

“…CD19-ChTCR-expressing UCBTderived T cells were capable of CD19-specific killing activity and cytokine secretion in vitro, as well as inducing regression of CD19 + tumors in NOD/SCID mice, and being selectively eliminated in vivo after administration of gancyclovir. 28 This approach has recently been extended to autologous cell therapy for patients affected by follicular lymphoma, showing that ex vivo cell expansion from cryopreserved cell banks was sufficient to produce doses of between 5×10 9 and 1×10 10 engineered T cells for each cycle of production. 29 This manufacturing strategy is therefore suitable for producing gene-manipulated T cells for phase I clinical trials in the context of B-cell lymphoid malignancies.…”

“…UCBT-derived T cells were efficiently redirected towards the CD19 + tumor target cells (including B-ALL) by electroporation with a plasmid encoding the CD19-ChTCR, also containing a suicide gene (the herpes virus 1 thymidine kinase, HSV-1 TK, gene). 28 Cells were efficiently expanded in relevant numbers for clinical intervention and afterwards cryopreserved for quality controls. CD19-ChTCR-expressing UCBTderived T cells were capable of CD19-specific killing activity and cytokine secretion in vitro, as well as inducing regression of CD19 + tumors in NOD/SCID mice, and being selectively eliminated in vivo after administration of gancyclovir.…”

“…[25][26][27][28][29] The initial construct was composed of a CD19-specific single-chain immunoglobulin extracellular targeting domain, fused to a CD3-zeta intracellular signaling domain. It was shown that CD19-redirected CTL were capable of potently killing primary B-ALL blasts and also of producing Th1 cytokines and were consistently proliferating after recognition of the targeted molecule.…”

Cord Blood Transplantation in Adults with Acute Leukemia

“…A clinical trial using CB derived multivirus specific CTLs for prevention and treatment of these virus infection in CB transplant is now underway. To generate CB derived T cells recognizing B-lineage ALL because GVT effects are largely mediated by CTLs, several researchers developed CB derived T cells are expanded and genetically modified to express CD19 chimeric antigen receptors (Serrano et al, 2006;Micklethwaite et al, 2010). The genetically modified T-cell clones revealed an ability to lyse CD19+ leukemic cells specifically and repetitively.…”