DNA Aptamer-Polymer Conjugates for Selective Targeting of Integrin α4β1 T-Lineage Cancers.

Selective therapeutic targeting of T-cell malignancies is difficult due to the shared lineage between healthy and malignant T cells. Current front-line chemotherapy for these cancers is largely nonspecific, resulting in frequent cases of relapsed/refractory disease. The development of targeting approaches for effectively treating T-cell leukemia and lymphoma thus remains a critical goal for the oncology field.

Intracerebroventricular B7-H3-targeting CAR T cells for diffuse intrinsic pontine glioma: a phase 1 trial.

Diffuse intrinsic pontine glioma (DIPG) is a fatal central nervous system (CNS) tumor that confers a median survival of 11 months. As B7-H3 is expressed on pediatric CNS tumors, we conducted BrainChild-03, a single-center, dose-escalation phase 1 clinical trial of repetitive intracerebroventricular (ICV) dosing of B7-H3-targeting chimeric antigen receptor T cells (B7-H3 CAR T cells) for children with recurrent or refractory CNS tumors and DIPG. Here we report results from Arm C, restricted to patients with DIPG.

Locoregional CAR T Cells for the Treatment of CNS Tumors in Children: Investigational Drug Service Pharmacy Activities.

Background: A major obstacle in translating the therapeutic potential of chimeric antigen receptor (CAR) T cells to children with central nervous system (CNS) tumors is the blood-brain barrier. To overcome this limitation, preclinical and clinical studies have supported the use of repeated, locoregional intracranial CAR T-cell delivery. However, there is limited literature available describing the process for the involvement of an investigational drug service (IDS) pharmacy, particularly in the setting of a children's hospital with outpatient dosing for CNS tumors.

Drug-regulated CD33-targeted CAR T cells control AML using clinically optimized rapamycin dosing.

Chimeric antigen receptor (CAR) designs that incorporate pharmacologic control are desirable; however, designs suitable for clinical translation are needed. We designed a fully human, rapamycin-regulated drug product for targeting CD33+ tumors called dimerizaing agent-regulated immunoreceptor complex (DARIC33). T cell products demonstrated target-specific and rapamycin-dependent cytokine release, transcriptional responses, cytotoxicity, and in vivo antileukemic activity in the presence of as little as 1 nM rapamycin.

Aptamer-Based Chromatographic Methods for Efficient and Economical Separation of Leukocyte Populations.

The manufacturing process of chimeric antigen receptor T cell therapies includes isolation systems that provide pure T cells. Current magnetic-activated cell sorting and immunoaffinity chromatography methods produce desired cells with high purity and yield but require expensive equipment and reagents and involve time-consuming incubation steps. Here, we demonstrate that aptamers can be employed in a continuous-flow resin platform for both depletion of monocytes and selection of CD8 T cells from peripheral blood mononuclear cells at low cost with high purity and throughput.

A stem cell epigenome is associated with primary nonresponse to CD19 CAR T cells in pediatric acute lymphoblastic leukemia.

CD19 chimeric antigen receptor T-cell therapy (CD19-CAR) has changed the treatment landscape and outcomes for patients with pre-B-cell acute lymphoblastic leukemia (B-ALL). Unfortunately, primary nonresponse (PNR), sustained CD19+ disease, and concurrent expansion of CD19-CAR occur in 20% of the patients and is associated with adverse outcomes. Although some failures may be attributable to CD19 loss, mechanisms of CD19-independent, leukemia-intrinsic resistance to CD19-CAR remain poorly understood.

Locoregional CAR T cells for children with CNS tumors: Clinical procedure and catheter safety.

Central nervous system (CNS) tumors are the most common solid malignancy in the pediatric population. Based on adoptive cellular therapy's clinical success against childhood leukemia and the preclinical efficacy against pediatric CNS tumors, chimeric antigen receptor (CAR) T cells offer hope of improving outcomes for recurrent tumors and universally fatal diseases such as diffuse intrinsic pontine glioma (DIPG). However, a major obstacle for tumors of the brain and spine is ineffective T cell chemotaxis to disease sites.

Murine CAR19 Tregs suppress acute graft-versus-host disease and maintain graft-versus-tumor responses.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) efficacy is complicated by graft-versus-host disease (GVHD), a leading cause of morbidity and mortality. Regulatory T cells (Tregs) have shown efficacy in preventing GVHD. However, high Treg doses are often required, necessitating substantial ex vivo or in vivo expansion that may diminish suppressor function.

FGFR4-Targeted Chimeric Antigen Receptors Combined with Anti-Myeloid Polypharmacy Effectively Treat Orthotopic Rhabdomyosarcoma.

Rhabdomyosarcoma (RMS) is the most common soft tissue cancer in children. Treatment outcomes, particularly for relapsed/refractory or metastatic disease, have not improved in decades. The current lack of novel therapies and low tumor mutational burden suggest that chimeric antigen receptor (CAR) T-cell therapy could be a promising approach to treating RMS.

Discovery of a Transferrin Receptor 1-Binding Aptamer and Its Application in Cancer Cell Depletion for Adoptive T-Cell Therapy Manufacturing.

The clinical manufacturing of chimeric antigen receptor (CAR) T cells includes cell selection, activation, gene transduction, and expansion. While the method of T-cell selection varies across companies, current methods do not actively eliminate the cancer cells in the patient's apheresis product from the healthy immune cells. Alarmingly, it has been found that transduction of a single leukemic B cell with the CAR gene can confer resistance to CAR T-cell therapy and lead to treatment failure.

Modified Manufacturing Process Modulates CD19CAR T-cell Engraftment Fitness and Leukemia-Free Survival in Pediatric and Young Adult Subjects.

T cells modified to express a chimeric antigen receptor (CAR) targeting CD19 can induce potent and sustained responses in children with relapsed/refractory acute lymphoblastic leukemia (ALL). The durability of remission is related to the length of time the CAR T cells persist. Efforts to understand differences in persistence have focused on the CAR construct, in particular the costimulatory signaling module of the chimeric receptor.

B7-H3 Specific CAR T Cells for the Naturally Occurring, Spontaneous Canine Sarcoma Model.

One obstacle for human solid tumor immunotherapy research is the lack of clinically relevant animal models. In this study, we sought to establish a chimeric antigen receptor (CAR) T-cell treatment model for naturally occurring canine sarcomas as a model for human CAR T-cell therapy. Canine CARs specific for B7-H3 were constructed using a single-chain variable fragment derived from the human B7-H3-specific antibody MGA271, which we confirmed to be cross-reactive with canine B7-H3.

Off-the-shelf, steroid-resistant, IL13Rα2-specific CAR T cells for treatment of glioblastoma.

Background: Wide-spread application of chimeric antigen receptor (CAR) T cell therapy for cancer is limited by the current use of autologous CAR T cells necessitating the manufacture of individualized therapeutic products for each patient. To address this challenge, we have generated an off-the-shelf, allogeneic CAR T cell product for the treatment of glioblastoma (GBM), and present here the feasibility, safety, and therapeutic potential of this approach.

Methods: We generated for clinical use a healthy-donor derived IL13Rα2-targeted CAR+ (IL13-zetakine+) cytolytic T-lymphocyte (CTL) product genetically engineered using zinc finger nucleases (ZFNs) to permanently disrupt the glucocorticoid receptor (GR) (GRm13Z40-2) and endow resistance to glucocorticoid treatment.

Arming Immune Cell Therapeutics with Polymeric Prodrugs.

Engineered immune cells are an exciting therapeutic modality, which survey and attack tumors. Backpacking strategies exploit cell targeting capabilities for delivery of drugs to combat tumors and their immune-suppressive environments. Here, a new platform for arming cell therapeutics through dual receptor and polymeric prodrug engineering is developed.

Hematopoietic Cell Transplantation after CD19 Chimeric Antigen Receptor T Cell-Induced Acute Lymphoblastic Lymphoma Remission Confers a Leukemia-Free Survival Advantage.

Consolidative hematopoietic cell transplantation (HCT) after CD19 chimeric antigen receptor (CAR) T cell therapy is frequently performed for patients with refractory/ relapsed B cell acute lymphoblastic leukemia (B-ALL). However, there is controversy regarding the role of HCT following remission attainment. We evaluated the effect of consolidative HCT on leukemia-free survival (LFS) in pediatric and young adult subjects following CD19 CAR T cell induced remission.

Locoregional infusion of HER2-specific CAR T cells in children and young adults with recurrent or refractory CNS tumors: an interim analysis.

Locoregional delivery of chimeric antigen receptor (CAR) T cells has resulted in objective responses in adults with glioblastoma, but the feasibility and tolerability of this approach is yet to be evaluated for pediatric central nervous system (CNS) tumors. Here we show that engineering of a medium-length CAR spacer enhances the therapeutic efficacy of human erb-b2 receptor tyrosine kinase 2 (HER2)-specific CAR T cells in an orthotopic xenograft medulloblastoma model. We translated these findings into BrainChild-01 ( NCT03500991 ), an ongoing phase 1 clinical trial at Seattle Children's evaluating repetitive locoregional dosing of these HER2-specific CAR T cells to children and young adults with recurrent/refractory CNS tumors, including diffuse midline glioma.

Rationally Designed Transgene-Encoded Cell-Surface Polypeptide Tag for Multiplexed Programming of CAR T-cell Synthetic Outputs.

Synthetic immunology, as exemplified by chimeric antigen receptor (CAR) T-cell immunotherapy, has transformed the treatment of relapsed/refractory B cell-lineage malignancies. However, there are substantial barriers-including limited tumor homing, lack of retention of function within a suppressive tumor microenvironment, and antigen heterogeneity/escape-to using this technology to effectively treat solid tumors. A multiplexed engineering approach is needed to equip effector T cells with synthetic countermeasures to overcome these barriers.

A CAR RNA FISH assay to study functional and spatial heterogeneity of chimeric antigen receptor T cells in tissue.

Chimeric antigen receptor (CAR) T cells are engineered cells used in cancer therapy and are studied to treat infectious diseases. Trafficking and persistence of CAR T cells is an important requirement for efficacy to target cancer. Here, we describe a CAR RNA FISH histo-cytometry platform combined with a random reaction seed image analysis algorithm to quantitate spatial distribution and in vivo functional activity of a CAR T cell population at a single cell resolution for preclinical models.

Optimized serum stability and specificity of an αvβ6 integrin-binding peptide for tumor targeting.

The integrin αvβ6 is an antigen expressed at low levels in healthy tissue but upregulated during tumorigenesis, which makes it a promising target for cancer imaging and therapy. A20FMDV2 is a 20-mer peptide derived from the foot-and-mouth disease virus that exhibits nanomolar and selective affinity for αvβ6 versus other integrins. Despite this selectivity, A20FMDV2 has had limited success in imaging and treating αvβ6 tumors in vivo because of its poor serum stability.

CD28 Co-Stimulus Achieves Superior CAR T Cell Effector Function against Solid Tumors Than 4-1BB Co-Stimulus.

Spacer or co-stimulatory components in chimeric antigen receptor (CAR) design influence CAR T cell effector function. Few preclinical mouse models optimally support CAR candidate pre-selection for clinical development. Here we use a model in which murine CAR T cells can be exploited with human tumor xenografts.