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BackgroundGene-modified autologous T cells expressing NY-ESO-1c259, an affinity-enhanced T-cell receptor (TCR) reactive against the NY-ESO-1-specific HLA-A*02-restricted peptide SLLMWITQC (NY-ESO-1 SPEAR T-cells; GSK 794), have demonstrated clinical activity in patients with advanced synovial sarcoma (SS). The factors contributing to gene-modified T-cell expansion and the changes within the tumor microenvironment (TME) following T-cell infusion remain unclear. These studies address the immunological mechanisms of response and resistance in patients with SS treated with NY-ESO-1 SPEAR T-cells.MethodsFour cohorts were included to evaluate antigen expression and preconditioning on efficacy. Clinical responses were assessed by RECIST v1.1. Engineered T-cell persistence was determined by qPCR. Serum cytokines were evaluated by immunoassay. Transcriptomic analyses and immunohistochemistry were performed on tumor biopsies from patients before and after T-cell infusion. Gene-modified T-cells were detected within the TME via an RNAish assay.ResultsResponses across cohorts were affected by preconditioning and intra-tumoral NY-ESO-1 expression. Of the 42 patients reported (data cut-off 4June2018), 1 patient had a complete response, 14 patients had partial responses, 24 patients had stable disease, and 3 patients had progressive disease. The magnitude of gene-modified T-cell expansion shortly after infusion was associated with response in patients with high intra-tumoral NY-ESO-1 expression. Patients receiving a fludarabine-containing conditioning regimen experienced increases in serum IL-7 and IL-15. Prior to infusion, the TME exhibited minimal leukocyte infiltration; CD163+ tumor-associated macrophages (TAMs) were the dominant population. Modest increases in intra-tumoral leukocytes (≤5%) were observed in a subset of subjects at approximately 8 weeks. Beyond 8 weeks post infusion, the TME was minimally infiltrated with a TAM-dominant leukocyte infiltrate. Tumor-associated antigens and antigen presentation did not significantly change within the tumor post-T-cell infusion. Finally, NY-ESO-1 SPEAR T cells trafficked to the TME and maintained cytotoxicity in a subset of patients.ConclusionsOur studies elucidate some factors that underpin response and resistance to NY-ESO-1 SPEAR T-cell therapy. From these data, we conclude that a lymphodepletion regimen containing high doses of fludarabine and cyclophosphamide is necessary for SPEAR T-cell persistence and efficacy. Furthermore, these data demonstrate that non-T-cell inflamed tumors, which are resistant to PD-1/PD-L1 inhibitors, can be treated with adoptive T-cell based immunotherapy.Trial registrationClinicalTrials.gov, NCT01343043 , Registered 27 April 2011.

We conducted a first-in-human study of intravenous delivery of a single dose of autologous T cells redirected to the epidermal growth factor receptor variant III (EGFRvIII) mutation by a chimeric antigen receptor (CAR). We report our findings on the first 10 recurrent glioblastoma (GBM) patients treated. We found that manufacturing and infusion of CAR-modified T cell (CART)-EGFRvIII cells are feasible and safe, without evidence of off-tumor toxicity or cytokine release syndrome. One patient has had residual stable disease for over 18 months of follow-up. All patients demonstrated detectable transient expansion of CART-EGFRvIII cells in peripheral blood. Seven patients had post-CART-EGFRvIII surgical intervention, which allowed for tissue-specific analysis of CART-EGFRvIII trafficking to the tumor, phenotyping of tumor-infiltrating T cells and the tumor microenvironment in situ, and analysis of post-therapy EGFRvIII target antigen expression. Imaging findings after CART immunotherapy were complex to interpret, further reinforcing the need for pathologic sampling in infused patients. We found trafficking of CART-EGFRvIII cells to regions of active GBM, with antigen decrease in five of these seven patients. In situ evaluation of the tumor environment demonstrated increased and robust expression of inhibitory molecules and infiltration by regulatory T cells after CART-EGFRvIII infusion, compared to pre-CART-EGFRvIII infusion tumor specimens. Our initial experience with CAR T cells in recurrent GBM suggests that although intravenous infusion results in on-target activity in the brain, overcoming the adaptive changes in the local tumor microenvironment and addressing the antigen heterogeneity may improve the efficacy of EGFRvIII-directed strategies in GBM.

Background: ADP-A2M10 specific peptide enhanced affinity receptor (SPEAR) T cells (ADP-A2M10) are genetically engineered autologous T cells that express a high-affinity melanoma-associated antigen A10 (MAGE-A10)-specific T-cell receptor (TCR) targeting MAGE-A10 + tumors in the context of human leukocyte antigen (HLA)-A*02. ADP-0022-003 was a phase I dose-escalation trial that aimed to evaluate the safety and antitumor activity of ADP-A2M10 in non-small cell lung cancer (NSCLC) (NCT02592577)., Methods: Eligible patients were HLA-A*02 positive with advanced NSCLC expressing MAGE-A10. Patients underwent apheresis; T cells were isolated, transduced with a lentiviral vector containing the TCR targeting MAGE-A10, and expanded. Patients underwent lymphodepletion with varying doses/schedules of fludarabine and cyclophosphamide prior to receiving ADP-A2M10. ADP-A2M10 were administered at 0.08-0.12×10 9 (dose group 1), 0.5-1.2×10 9 (dose group 2), and 1.2-15×10 9 (dose group 3/expansion) transduced cells., Results: Eleven patients (male, n=6; female, n=5) with NSCLC (adenocarcinoma, n=8; squamous cell carcinoma, n=3) were treated. Five, three, and three patients received cells in dose group 1, dose group 2, and dose group 3/expansion, respectively. The most frequently reported grade ≥3 adverse events were lymphopenia (n=11), leukopenia (n=10), neutropenia (n=8), anemia (n=6), thrombocytopenia (n=5), and hyponatremia (n=5). Three patients presented with cytokine release syndrome (grades 1, 2, and 4, respectively). One patient received the highest dose of lymphodepletion (fludarabine 30 mg/m 2 on days -5 to -2 and cyclophosphamide 1800 mg/m 2 on days -5 to -4) prior to a second infusion of ADP-A2M10 and had a partial response, subsequently complicated by aplastic anemia and death. Responses included: partial response (after second infusion; one patient), stable disease (four patients), clinical or radiographic progressive disease (five patients), and not evaluable (one patient). ADP-A2M10 were detectable in peripheral blood and in tumor tissue. Peak persistence was higher in patients who received higher doses of ADP-A2M10., Conclusions: ADP-A2M10 demonstrated an acceptable safety profile and no evidence of toxicity related to off-target binding or alloreactivity. There was persistence of ADP-A2M10 in peripheral blood as well as ADP-A2M10 trafficking into the tumor. Given the discovery that MAGE-A10 and MAGE-A4 expression frequently overlap, this clinical program closed as trials with SPEAR T cells targeting MAGE-A4 are ongoing., Competing Interests: Competing interests: AG holds stock options in Adaptimmune. AJO has received research contracts from Alkermes, Antegene, Astellas, Checkmate, Gan & Lee, GlycoNex, InstilBio, Intensity, Istari, Kadmon, Kartos, NeoImmune, NGM, OncoSec, Sound Bio, SpringBank, Takeda; has received payment for advisory board roles from Merck, BMS, Pfizer, Takeda, Sanofi, Eisai. DB, DM, EN, JB, J-MN, JPS, MMP, NH, RB, SF are or were employees of Adaptimmune at the time of the study. GRB has received grants/contracts from Amgen, Bayer, Adaptimmune, Elelixis, Daiichi Sankyo, GlaxoSmithKline, Immatics, Immunocore, Incyte, Kite Pharma, Macrogenics, Torque, AstraZeneca, Bristol-Myers Squibb, Celgene, Genentech, MedImmune, Merck, Novartis, Roche, Xcovery, Tmunity Therapeutics, Regeneron, Beigene, Repertoire Immune Medicines, Verastem; consulting fees from Abbvie, Adicet, Amgen, Ariad, Bayer, Clovis Oncology, AstraZeneca, Bristol-Myers Squibb, Celgene, Daiichi Sankyo, Instil Bio, Genentech, Gilead, Lilly, Janssen, MedImmune, Merck, Novartis, Roche, Tyme Oncology, Xcovery, Virogin Biotech, Maverick Therapeutics; has participated on a Data Safety Monitoring Board or Advisory Board for Virogen Biotech, Maverick Therapeutics; holds stock or stock options in Virogin Biotech; and has other financial or non-financial interests in Johnson & Johnson/Janssen (immediate family member employed). JG has been a compensated consultant or received honoraria from Bristol-Myers Squibb, Genentech, Ariad/Takeda, Loxo, Pfizer, Incyte, Novartis, Merck, Agios, Amgen, Jounce, Karyopharm, GlydeBio, Mirati, AstraZeneca, Regeneron, Oncorus, Helsinn, Array, and Clovis Oncology; has been an employee (immediate family member) with equity in Ironwood Pharmaceuticals; has received research funding from Novartis, Genentech/Roche, and Ariad/Takeda; has received institutional research support from Tesaro, Moderna, Blueprint, Scholar Rock, BMS, Array, Adaptimmune, Novartis, Genentech/Roche, Alexo and Merck. JVH has received grants/contracts from AstraZeneca, GlaxoSmithKline and Spectrum; holds royalty/licenses in Spectrum; has received consulting fees from AstraZeneca, Boehringer-Ingelheim, Catalyst, Genentech, GlaxoSmithKline, Guardant Health, Foundation medicine, Hengrui Therapeutics, Eli Lilly, Novartis, Spectrum, EMD Serono, Sanofi, Takeda, Mirati Therapeutics, BMS, BrightPath Biotherapeutics, Janssen Global Services, Nexus Health Systems, EMD Serono, Pneuma Respiratory, Kairos Venture Investments, Roche, Leads Biolabs, RefleXion; has received honoraria from Medlinker, Peerview, Nexus Health Medicine, Targeted Oncology, MJH Events; has received support for attending meetings from IASLC Targeted Therapies, IASLC World Conference on Lung Cancer; has been in a leadership or fiduciary role in other board, society, committee or advocacy group, paid or unpaid for Mechanisms of Cancer Therapeutics-1 (MCT1) (Study Section – Chair). MJ has received research funding from AbbVie, Acerta, Adaptimmune, Apexigen, Array BioPharma, AstraZeneca, Atreca, BeiGene, Boehringer Ingelheim, Checkpoint Therapeutics, Corvus Pharmaceuticals, CytomX, Daiichi Sankyo, Dynavax, Lilly, EMD Serono, Genentech/Roche, Genmab, Genocea Biosciences, GlaxoSmithKline, Gritstone Oncology, Guardant Health, Hengrui Therapeutics, Immunocore, Incyte, Janssen, Jounce Therapeutics, Kadmon Pharmaceuticals, Loxo Oncology, Lycera, Merck, Mirati Therapeutics, Neovia Oncology, Novartis, OncoMed Pharmaceuticals, Pfizer, Regeneron Pharmaceuticals, Sanofi, Seven and Eight Biopharmaceuticals, Shattuck Labs, Stem CentRx, Syndax Pharmaceuticals, Takeda Pharmaceuticals, Tarveda, University of Michigan, WindMIL, TCR2 Therapeutics, Arcus Biosciences, Ribon Therapeutics, Amgen; has held a consulting/advisory role (spouse) for Astellas, Otsuka Pharmaceuticals; has held a consulting/advisory role (self) for AbbVie, Achilles Therapeutics, AstraZeneca, Atreca, Boehringer Ingelheim, Calithera Biosciences, Genentech, GlaxoSmithKline, Gritstone Oncology, Guardant Health, Incyte, Janssen, Lilly, Loxo Oncology, Merck, Mirati Therapeutics, Novartis, Pfizer, Ribon Therapeutics, Sanofi, Association of Community Cancer Center; has received food/beverage/travel expenses from Abbvie, Astellas, AstraZeneca, Boehringer Ingelheim, Clovis, Daiichi Sankyo, EMD Serono, Bristol Myers Squibb, Exelixis, Genentech/Roche, Incyte, Merck, Pfizer, Sysmex Inostics, Vapotherm, Janssen, Lilly, Novartis, Sanofi. MJE has received research funding from GlaxoSmithKline, Mersana, Amgen, Windmil, Nektar, Apexigen, Adaptimmune; has received consulting fees from Kanaph, Flame; has received honoraria from Sanofi; has been a member of DSMB for AstraZeneca, Seattle Genetics, GlaxoSmithKline, Takeda; has been a consultant/on an advisory board for Windmil, Regeneron, Syndax; has been the Chair, Scientific Advisory board for Lung Cancer Foundation of America; has been the Deputy Editor of 'Lung Cancer'; has received stock options from Biomarker strategies; Creatv Biotech. MJF has been a consultant for Arcellx, BMS, Novartis, Kite, Iovance. PMF is an employee of Adaptimmune; holds stock in Adaptimmune and Bristol-Myers Squib; has received compensation for travel and congress meetings. RG has received consulting fees from BMS, Abbvie, Geneplus; has participated on a Data Safety Monitoring Board or Advisory Board for Roche Genentech. SD has been a consultant for Jacobio pharmaceuticals (without financial compensation). TH was an employee of Adaptimmune at the time of the study; has been a consultant for Adaptimmune. VKL has been in a consulting or advisory role for Takeda, Bristol-Myers Squibb, Seattle Genetics; has received research funding from GlaxoSmithKline, Bristol-Myers Squibb, Guardant Health, Takeda. VM has received consulting fees from Roche, Bayer, Pieris, BMS, Janssen and Basilea. BDdS, CB, ZW have nothing to declare., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

CCR5 is a G protein-coupled receptor for RANTES, MIP-1alpha, MIP-1beta, and MCP-2 that functions as the front line coreceptor for human immunodeficiency virus type 1 infection. To elucidate the mechanism for CCR5 activation, this coreceptor was expressed in yeast coupled to the pheromone response pathway and a constitutively active mutant (CAM) was derived by random mutagenesis. Conversion of Thr-82 in the highly conserved TXP motif in transmembrane helix 2 to Pro, His, Tyr, Arg, or Lys conferred autonomous signaling activity in yeast and mammalian cells. This substitution also imparted constitutive signaling to CCR2 in yeast and mammalian cells, but not CCR1, CCR3, CCR4, CXCR2, or CXCR4. The CCR5-CAM, but not the CCR2-CAM had a reduction in ligand binding affinity. Whereas the amplitude of calcium mobilization induced by RANTES stimulation was lower in the CCR5-CAM than the wild-type (WT) receptor, MCP-1 induced a higher signal in the CCR2-CAM than in CCR2-WT. The chemotactic response of CCR5-CAM(T82P) to RANTES was similar to that of CCR5-WT, but CCR5-CAM(T82K) was dramatically decreased. The chemotactic response of CCR2-WT and CCR2-CAM(T94K) were similar. These findings extend insight into the role of the TXP motif in the mechanism for CCR5 signaling. CCR2, the receptor most closely genetically related to CCR5, shared a similar signaling mechanism, but other receptors containing the TXP motif did not. The expression of CCR5 and CCR2 in yeast and the availability of variants with autonomous signaling represent critical tools for characterizing receptor antagonists and developing approaches to block their role in human diseases.