Tumor-infiltrating Lymphocytes Therapy Research Articles

Antitumor activity of T cells expressing a novel anti-folate receptor alpha (FOLR1) costimulatory antigen receptor (CoStAR) in a human xenograft murine solid tumor model and implications for in-human studies.

2535 Background: ITIL-306 is an autologous tumor-infiltrating lymphocyte (TIL) therapy that integrates T-cell receptor (TCR)-specific antigen recognition (Signal 1) with robust costimulation via the novel CoStAR transgene upon engagement with FOLR1 (Signal 2; Sukumaran, et al. JITC. 2021;9:198). Here, we assessed IL-2 independent effector function by anti-FOLR1 CoStAR T cells in vitro and evaluated activity in a novel, more representative murine solid tumor model. Methods: For in vitro studies, healthy donor T cells were manufactured to express anti-FOLR1 CoStAR or left nontransduced (NTD). Product cells were stimulated with a single (Day [D]0; ±IL-2) or serial (D0, 7, 14, 21; no IL-2) addition of target cells expressing membrane-anchored OKT3 (Signal 1) and FOLR1 (Signal 2). T-cell activation and proliferation were measured. For in vivo studies, the carcinoembryonic antigen (CEA)-positive (Signal 1) H508 cell line was engineered to express FOLR1 (Signal 2) and injected into 6 mice/group (D–21). NTD or healthy donor T cells expressing HLA-A*02-restricted anti-CEA TCR only, anti-FOLR1 CoStAR only, or dual TCR+CoStAR were given intravenously (D0), and compared with PBS control. Tumor growth, survival, and T-cell expansion were assessed in 2 donors to D96. Results: After single and serial stimulation, only anti-FOLR1 CoStAR T cells showed sustained proliferation without exogenous IL-2. PD-1 positivity was low ( < 10%) on anti-FOLR1 T cells up to D24. In vivo, T-cell persistence on D14 was increased in dual TCR+CoStAR vs anti-CEA TCR only mice ( P<.001) and all other groups ( P<.01-.001). Tumor growth up to D58 was significantly lower in dual TCR+CoStAR ( <.5 cm3) vs all other groups (each > 2 cm3; P<.0001). Survival was significantly longer in dual TCR+CoStAR ( P<.01) vs all other groups; only dual TCR+CoStAR mice were alive at D58, with 5/6 alive at D96. The second donor showed similar tumor control irrespective of exogenous IL-2 in responder mice, with > 50% alive at D96, and robust T-cell expansion in dual TCR+CoStAR mice. Conclusions: When combined with TCR-specific binding, CoStAR significantly enhanced T-cell proliferation, persistence, and antitumor activity in vivo vs TCR alone, resulting in tumor control and prolonged survival. Effects were not observed with CoStAR alone, underscoring that signaling through CoStAR alone does not induce T-cell effector function. The sustained proliferation of anti-FOLR1 CoStAR T cells without exogenous IL-2 support in vitro and in vivo supports a clinical TIL regimen free of high-dose IL-2. These results suggest that CoStAR may improve the clinical performance and benefit/risk profile of TILs, whereby fewer toxicities are expected with the removal of post-TIL IL-2 support. This will be explored in an upcoming first-in-human clinical study with ITIL-306.

A phase I trial of T-cell receptor gene therapy targeting KK-LC-1 for gastric, breast, cervical, lung and other KK-LC-1 positive epithelial cancers.

TPS2678 Background: T cell receptor (TCR)-T cell therapy is an emerging cancer treatment strategy. Thus far, demonstration of clinical activity has been limited to a subset of solid tumors including melanoma, synovial cell sarcoma and HPV-associated cancer. It is estimated that metastatic epithelial cancers are responsible for approximately 90% of cancer deaths in the United States. The estimated 600,000 cancer deaths each year is driven largely by lung adeno- and squamous cell carcinoma and invasive breast cancer, which account for approximately 30% of all cancer-related deaths. Kita-Kyushu Lung Cancer Antigen 1 (KK-LC-1) is a cancer germline antigen with expression restricted to germ cells in adults and certain epithelial cancers including lung, breast, gastric and cervical. We identified a KK-LC-1 TCR from the tumor-infiltrating lymphocytes (TIL) of a patient with cervical cancer who had a complete tumor response to TIL therapy. The KK-LC-1 TCR was the most dominate clonotype in the infused TIL product and persisted in the peripheral blood following infusion, suggesting that it may have contributed to cancer regression in this patient. Methods: We are conducting a phase I cell dose escalation trial to test the safety and efficacy of KK-LC-1 TCR-T cell therapy in patients with metastatic KK-LC-1 positive epithelial cancer. Patients receive a lymphocyte depleting conditioning regimen followed by a one-time infusion of genetically engineered T cells expressing the KK-LC-1 TCR (KK-LC-1 TCR-T cells) and high-dose systemic aldesleukin. KK-LC-1 positivity is determined by RNAscope assay measuring the percentage of cancer cells expressing CT83 (gene encoding KK-LC-1) with a percentage of 25 or greater considered positive. Main inclusion criteria include HLA-A*01:01 allele, prior first-line therapy, ECOG of 0 or 1 and adequate organ and hematologic function. Main exclusion criteria include active major medical illness of the cardiovascular, respiratory or immune system, primary or secondary immunodeficiency and autoimmune disease. Participants will be entered in sequential dose levels and receive escalating doses of cells beginning at 1x108 and ending with 6x1010. Dose-limiting toxicities will be assessed during the first 30 days of cell infusion. The primary objective is to determine the maximally tolerated dose of KK-LC-1 TCR-T cells. Exploratory objectives include assessing the safety and efficacy of KK-LC-1 TCR-T cells and to conduct immunologic studies to understand and improve the administered treatment. Clinical trial information: NCT05035407.

A T cell resilience model associated with response to immunotherapy in multiple tumor types.

Despite breakthroughs in cancer immunotherapy, most tumor-reactive T cells cannot persist in solid tumors due to an immunosuppressive environment. We developed Tres (tumor-resilient T cell), a computational model utilizing single-cell transcriptomic data to identify signatures of T cells that are resilient to immunosuppressive signals, such as transforming growth factor-β1, tumor necrosis factor-related apoptosis-inducing ligand and prostaglandin E2. Tres reliably predicts clinical responses to immunotherapy in melanoma, lung cancer, triple-negative breast cancer and B cell malignancies using bulk T cell transcriptomic data from pre-treatment tumors from patients who received immune-checkpoint inhibitors (n = 38), infusion products for chimeric antigen receptor T cell therapies (n = 34) and pre-manufacture samples for chimeric antigen receptor T cell or tumor-infiltrating lymphocyte therapies (n = 84). Further, Tres identified FIBP, whose functions are largely unknown, as the top negative marker of tumor-resilient T cells across many solid tumor types. FIBP knockouts in murine and human donor CD8+ T cells significantly enhanced T cell-mediated cancer killing in in vitro co-cultures. Further, Fibp knockout in murine T cells potentiated the in vivo efficacy of adoptive cell transfer in the B16 tumor model. Fibp knockout T cells exhibit reduced cholesterol metabolism, which inhibits effector T cell function. These results demonstrate the utility of Tres in identifying biomarkers of T cell effectiveness and potential therapeutic targets for immunotherapies in solid tumors.

Enriching for tumor-reactive CD8 TIL (AGX148) leads to effective tumor clearance in a patient-derived xenograph model

Abstract Adoptive Cell Therapy (ACT) with Tumor Infiltrating Lymphocytes (TIL) can induce durable clinical responses in a subset of patients with melanoma; however, the efficacy of standard bulk TIL therapy can be limited. We hypothesize that the therapeutic efficacy of TIL ACT is dependent on the frequency and functionality of tumor-reactive T cells when infused back into patients. Our approach enhances the activity of the ACT product by enriching for the tumor-reactive CD8 TIL prior to ex vivo expansion by sorting on cells that co-express CD103 and CD39. The CD103/CD39 double positive (DP) CD8 TIL can be found in most human solid tumors but at varying frequencies in any given patient. Surgically resected human tumor samples were provided through a collaboration with the EACRI. Tumor samples were flow sorted and expanded ex vivo and patient-matched autologous tumor lines were generated. Tumors were then cocultured with the expanded autologous ACT product (AGX148) in vitro or used in xenograph mouse experiments. The AGX148 TIL products are effective at recognizing and killing autologous tumor cells, in vitro. Upon adoptive transfer into patient-derived tumor-bearing huIL-2 NOG mice, the transferred human T cells increased in frequency, migrated to the tumor, became activated in the tumor and recognized/destroyed established tumors. Interestingly there was little to no in vivo activity of CD8 TIL that did not express these two markers. We have generated a potent therapeutic ACT TIL product (AGX148) through the selective isolation and expansion of tumor-reactive T cells. This TIL product effectively induces durable curative responses in hosts harboring autologous tumors providing rationale for clinical investigation.

Decrease of Tumor-infiltrating Regulatory T Cells Using Pentoxifylline: An Ex Vivo Analysis in Triple-negative Breast Cancer Mouse Model.

Triple-negative breast cancer (TNBC) is the most aggressive type of BC with the highest percentage of tumor-infiltrating lymphocytes (TILs). Hence, TIL therapy is considered a promising approach to target TNBC. Depletion of regulatory T cells (Tregs) in TILs can improve the antitumor function of TIL therapy. Pentoxifylline (PTXF) is a xanthine derivative that can modulate the nuclear factor kappa B (NF-κB) signaling and probably affect the Treg proportion in TILs. We aimed to evaluate the ex vivo effect of PTXF on the proportion of Treg cells in the TILs derived from a mouse model of TNBC. The 4T1 cells were inoculated subcutaneously to BALB/c mice to induce TNBC. TILs were isolated from tumor tissue by enzymatic digestion and cultured alone or with 4T1 cells for 24, 48, and 72 h in the presence of interleukin (IL)-2 and different concentrations of PTXF. The toxicity of PTXF and its effects on Tregs proportion as well as cytokine production was evaluated using MTT assay, flow cytometry, and ELISA, respectively. PTXF had no significant impact on the viability of TILs. Both 500 and 1000 mg/mL of PTXF decreased the proportion of Tregs in a dose-dependent manner. The level of interferon-g and tumor growth factor-b in TILs supernatant was increased and decreased, respectively. Our data suggest that ex vivo treatment of TILs with pentoxifylline could decrease the proportion of Tregs in the conventional IL-2-mediated TIL expansion and change the cytokine balance of TILs in favor of antitumor immune response.

A TUMOR-IMMUNE MODEL WITH MIXED IMMUNOTHERAPY AND CHEMOTHERAPY: QUALITATIVE ANALYSIS AND OPTIMAL CONTROl

We develop a mathematical model of tumor-immune interactions, including six populations (tumor cells, CD8[Formula: see text]T cells, natural killer (NK) cells, dendritic cells, helper T cells, cytokine interleukin-12 (IL-12)) and three potential treatments (chemotherapy, Tumor-infiltrating lymphocyte (TIL) therapy and IL-12 therapy). We characterize the dynamics of our model without treatment through stability and sensitivity analysis, which provides a broad understanding of the long-term qualitative behavior. To find the best combination of the chemo-immunotherapy regimens to eliminate tumors, we formulate an optimal control problem with path constraints of total drug dose and solve it numerically with the optimal control software Pyomo. We also simulate the scenarios of traditional treatment protocols as a comparison and find that our optimal treatment strategies have a better therapeutic effect. In addition, numerical simulation results show that IL-12 therapy is a good adjunctive therapy and has a high potential for inhibiting a large tumor in combination with other therapy. In most cases, combination therapy is more effective than a single treatment.

Tumor-Infiltrating T Cells Can Be ExpandedSuccessfully from Primary Uveal Melanoma after Separation from Their TumorEnvironment.

PurposeTo evaluate whether expanded tumor-infiltrating lymphocytes (TILs) can be obtained from primary uveal melanoma (UM) for potential use as adjuvant treatment in patients at risk of developing metastatic disease.DesignExperimental research study.ParticipantsFreshly obtained primary UM from 30 patients.MethodsThree different methods were used to expand TILs: (1) direct culture from small fragments of fresh tumor tissue, (2) single-cell tissue preparation by enzymatic digestion and subsequent enrichment of mononuclear cells, and (3) selection of CD3+ T cells using magnetic beads. Surface expression of costimulatory and inhibitory T-cell markers and T-cell reactivity against autologous tumor cells was assessed. Clinical, histopathologic, genetic, and immunologic characteristics of the tumors were compared with the capacity to expand TILs and with their reactivity against autologous tumor cells.Main Outcome MeasuresThe feasibility of expanding TILs from primary UM, testing their reactivity to autologous UM cells, and evaluating the impact of an immunomodulatory environment.ResultsDirect culture of tumor parts led to successful TIL culture in 4 of 22 tumors (18%), enrichment of mononuclear cells gave rise to TILs in 5 of 12 tumors (42%), while preselection of CD3+ T cells with magnetic beads resulted in TIL expansion in 17 of 25 tumors (68%). In 8 of 17 tumors (47%), the TIL cultures comprised UM-reactive T cells. The presence of UM-reactive T cells among TILs was not related to clinical, histologic, genetic, or immunological tumor characteristics. Interestingly, RNA-Seq analysis showed that approximately half of the UM tumors displayed an increased expression of immunomodulatory molecules related to T-cell suppression, such as galectin 3, programmed death-ligand 1, cytotoxic T-lymphocyte-associated protein 4, indoleamine 2,3-dioxygenase 1, and lymphocyte activating 3, potentially explaining why T cells require optimal removal of tumor components for expansion.ConclusionsThe need to separate TILs from their tumor microenvironment for their successful expansion and the presence of UM-reactive T cells among TILs suggests that these UM-reactive T cells are strongly suppressed in vivo and that UM is immunogenic. These findings indicate that adoptive TIL therapy could be an option as an adjuvant treatment in primary UM patients at high risk of developing metastatic disease.

Genomic Correlates of Outcome in Tumor-Infiltrating Lymphocyte Therapy for Metastatic Melanoma.

Adoptive cell therapy (ACT) of tumor-infiltrating lymphocytes (TIL) historically yields a 40%-50% response rate in metastatic melanoma. However, the determinants of outcome are largely unknown. We investigated tumor-based genomic correlates of overall survival (OS), progression-free survival (PFS), and response to therapy by interrogating tumor samples initially collected to generate TIL infusion products. Whole-exome sequencing (WES) data from 64 samples indicated a positive correlation between neoantigen load and OS, but not PFS or response to therapy. RNA sequencing analysis of 34 samples showed that expression of PDE1C, RTKN2, and NGFR was enriched in responders who had improved PFS and OS. In contrast, the expression of ELFN1 was enriched in patients with unfavorable response, poor PFS and OS, whereas enhanced methylation of ELFN1 was observed in patients with favorable outcomes. Expression of ELFN1, NGFR, and PDE1C was mainly found in cancer-associated fibroblasts and endothelial cells in tumor tissues across different cancer types in publicly available single-cell RNA sequencing datasets, suggesting a role for elements of the tumor microenvironment in defining the outcome of TIL therapy. Our findings suggest that transcriptional features of melanomas correlate with outcomes after TIL therapy and may provide candidates to guide patient selection.

Investigating the Potential of Isolating and Expanding Tumour-Infiltrating Lymphocytes from Adult Sarcoma.

Simple SummarySarcomas are rare cancers that arise from connective tissue. There are more than 50 subtypes, many of which are associated with a high risk of metastasis and poor prognosis. Subtype-specific treatment is limited and conventional treatment for advanced disease has varying effects across individuals and tumour subtypes. Adoptive cell therapy shows potential to provide more personalized treatment; this study aims to explore the potential of using tumour-infiltrating lymphocytes (TIL) to treat sarcoma. We optimized a sarcoma-specific expansion protocol and successfully expanded TILs from 54 of 92 sarcoma specimens. We characterized primarily CD4+ and CD8+ T-cells in the expanded TIL cultures and demonstrated their reactivity to general stimuli. Although sarcomas in general do not have abundant lymphocytic infiltration, our expansion protocol allowed for successful expansions of viable and reactive lymphocytes, thus showing the prospects of adopting TIL therapy in sarcoma.Sarcomas are a heterogeneous group of mesenchymal neoplasms, many of which are associated with a high risk of metastasis and poor prognosis. Conventional chemotherapy and targeted therapies have varying effects across individuals and tumour subtypes. The current therapies frequently provide limited clinical benefit; hence, more effective treatments are urgently needed. Recent advances in immunotherapy, such as checkpoint inhibition or adoptive cell therapy (ACT), show potential in increasing efficacy by providing a more personalized treatment. Therapy with tumour-infiltrating lymphocytes (TILs) is an emerging field in immunotherapy. Here, we collected 190 sarcoma tumour specimens from patients without pre-operative adjuvant treatment in order to isolate TILs. We compared different methods of TIL expansion and optimized a protocol specifically for efficacy in culturing TILs from sarcoma. The expanded TIL populations were characterized by flow cytometry analysis using CD3, CD4, CD8, CD14, CD19 and CD56 markers. The TIL populations were non-specifically stimulated to establish TIL reactivity. Through an optimized expansion protocol, TILs were isolated and cultured from 54 of 92 primary sarcoma specimens. The isolated TILs varied in CD4+ and CD8+ T-cell compositions and retained their ability to release IFNγ upon stimulation. Our results suggest that certain sarcoma subtypes have the potential to yield a sufficient number of TILs for TIL therapy.

Genetics and Therapeutic Responses to Tumor-Infiltrating Lymphocyte Therapy of Pancreatic Cancer Patient-Derived Xenograft Models.

Pancreatic cancer is the seventh leading cause of cancer-related deaths worldwide. Checkpoint immunotherapy has not yet shown encouraging results in pancreatic cancer possibly because of a poor immunogenicity and/or an immune suppressive microenvironment. The aim of this study was to develop patient-derived xenograft (PDX) models, compare their genetics to the original biopsies, and assess if autologous tumor-infiltrating lymphocytes (TILs) would have antitumoral activity in pancreatic cancer. We subcutaneously transplanted tumors from 29 patients into NOG mice to generate PDX models. We established TIL cultures and injected them into PDX mice. We analyzed histology and genetics of biopsies and PDX tumors. Tumor growths were confirmed in 11 of 29 transplantations. The PDX tumors histologically resembled their original biopsies, but because stromal cells in the PDX model tumors were from mouse, their gene expression differed from the original biopsies. Immune checkpoint ligands other than programmed death ligand-1 (PD-L1) were expressed in pancreatic cancers, but PD-L1 was rarely expressed. When it was expressed, it correlated with tumor take in PDX models. One of the 3 tumors that expressed PD-L1 was an adenosquamous cancer, and another had a mismatch repair deficiency. TILs were expanded from 6 tumors and were injected into NOG or human interleukin-2 transgenic-NOG mice carrying PDX tumors. Regression of tumors could be verified in human interleukin-2 transgenic-NOG mice in 3 of the 6 PDX models treated with autologous TILs, including the adenosquamous PDX model. PDX models of pancreatic cancer can be used to learn more about tumor characteristics and biomarkers and to evaluate responses to adoptive cell therapy and combination therapies. The major benefit of the model is that modifications of T cells can be tested in an autologous humanized mouse model to gain preclinical data to support the initiation of a clinical trial.

Futuristic approach to cancer treatment.

Cancer is becoming one of the deadliest disease in both developed as well as developing countries and continuous effort is being made to find innovative therapies for myriad types of cancers that afflict the human body. Therapeutic options for cancer have grown exponentially over the time but we are quite a way off from finding a magic bullet that can help cure cancer and based on the current evidence we may never find a catch all cure ever and it becomes crucial that we keep on innovating and find multiple ways to attack the menace of this dreaded disease. Many patients suffer recurrence of disease and require second-line or in some cases more than two lines of treatment. In this review article we have discussed the available therapies along with the newer advancements that have been made in cancer therapy. Latest developments in treatment of various cancers that have been discussed include gene editing using CRISPR/Cas9, theranostics, viral mediated therapy, artificial intelligence, tumor infiltrating lymphocyte therapy, etc.

Coupling programmed cell death 1-positive tumor-infiltrating T cells with anti-programmed cell death 1 antibody improves the efficacy of adoptive T-cell therapy

Background aimsAdoptive cell therapy (ACT) with tumor-infiltrating lymphocytes (TILs) has shown great success in clinical trials. Programmed cell death 1 (PD-1)-expressing TILs show high specificity to autologous tumor cells. However, limited therapeutic efficiency is observed as a result of the tumor immune microenvironment (TIME). MethodsCoupling PD-1+ex vivo-derived TILs with a monoclonal antibody against anti-PD-1 (aPD-1) reinvigorated the anti-tumor response of TILs against solid tumor without altering their high tumor targeting ability. ResultsUsing a melanoma-bearing mouse model, PD-1+ TILs blocked with aPD-1 (PD-1+ TILs-aPD-1) exhibited a high capability for tumor targeting as well as improved anti-tumor response in TIME. Tumor growth was substantially delayed in the mice treated with PD-1+ TILs-aPD-1. ConclusionsThe strategy utilizing TIL therapy coupled with immune checkpoint antibodies may extend to other therapeutic targets of ACT.

Adoptive cell therapy with tumor-infiltrating lymphocytes supported by checkpoint inhibition across multiple solid cancer types

BackgroundAdoptive cell therapy (ACT) with tumor-infiltrating lymphocytes (TILs) has shown remarkable results in malignant melanoma (MM), while studies on the potential in other cancer diagnoses are sparse. Further, the prospect...

Cardiac Toxicity Associated with Cancer Immunotherapy and Biological Drugs.

Simple SummaryImmunotherapy is increasingly being used to treat solid tumors and lymphoproliferative diseases. The main classes of drugs are: HER-2-targeted therapies, CTLA-blockers, PD/PDL-1 inhibitors, CAR-T therapy. All these drugs are associated with meaningful cardiac toxicity, ranging from a transient decline of left ventricular function with complete reversibility to myocarditis with a high fatality rate.Cancer immunotherapy significantly contributed to an improvement in the prognosis of cancer patients. Immunotherapy, including human epidermal growth factor receptor 2 (HER2)-targeted therapies, immune checkpoint inhibitors (ICI), and chimeric antigen receptor-modified T (CAR-T), share the characteristic to exploit the capabilities of the immune system to kill cancerous cells. Trastuzumab is a monoclonal antibody against HER2 that prevents HER2-mediated signaling; it is administered mainly in HER2-positive cancers, such as breast, colorectal, biliary tract, and non-small-cell lung cancers. Immune checkpoint inhibitors (ICI) inhibit the binding of CTLA-4 or PD-1 to PDL-1, allowing T cells to kill cancerous cells. ICI can be used in melanomas, non-small-cell lung cancer, urothelial, and head and neck cancer. There are two main types of T-cell transfer therapy: tumor-infiltrating lymphocytes (or TIL) therapy and chimeric antigen receptor-modified T (CAR-T) cell therapy, mainly applied for B-cell lymphoma and leukemia and mantle-cell lymphoma. HER2-targeted therapies, mainly trastuzumab, are associated with left ventricular dysfunction, usually reversible and rarely life-threatening. PD/PDL-1 inhibitors can cause myocarditis, rare but potentially fulminant and associated with a high fatality rate. CAR-T therapy is associated with several cardiac toxic effects, mainly in the context of a systemic adverse effect, the cytokines release syndrome.

Adoptive NK Cell Therapy: A Promising Treatment Prospect for Metastatic Melanoma.

Simple SummaryThe incidence of metastatic melanoma has been increasing over the past years with current therapies showing limited efficacy to cure the disease. Therefore, other options are being investigated, such as adoptive cell therapy (ACT) where activated immune cells are infused into a patient to attack melanoma. Natural killer (NK) cells are part of the innate immune system and extremely suitable for this kind of therapy since they show minimal toxicities in the clinical setting. In this review, we focus on current strategies for NK cell therapy and the development of new approaches that hold great promise for the treatment of advanced melanoma. Adoptive cell therapy (ACT) represents a promising alternative approach for patients with treatment-resistant metastatic melanoma. Lately, tumor infiltrating lymphocyte (TIL) therapy and chimeric antigen receptor (CAR)-T cell therapy have shown improved clinical outcome, compared to conventional chemotherapy or immunotherapy. Nevertheless, they are limited by immune escape of the tumor, cytokine release syndrome, and manufacturing challenges of autologous therapies. Conversely, the clinical use of Natural Killer (NK) cells has demonstrated a favorable clinical safety profile with minimal toxicities, providing an encouraging treatment alternative. Unlike T cells, NK cells are activated, amongst other mechanisms, by the downregulation of HLA class I molecules, thereby overcoming the hurdle of tumor immune escape. However, impairment of NK cell function has been observed in melanoma patients, resulting in deteriorated natural defense. To overcome this limitation, “activated” autologous or allogeneic NK cells have been infused into melanoma patients in early clinical trials, showing encouraging clinical benefit. Furthermore, as several NK cell-based therapeutics are being developed for different cancers, an emerging variety of approaches to increase migration and infiltration of adoptively transferred NK cells towards solid tumors is under preclinical investigation. These developments point to adoptive NK cell therapy as a highly promising treatment for metastatic melanoma in the future.

1008P cytoTIL15: A novel TIL therapy for melanoma with superior potency and enhanced persistence without IL2 to improve safety & efficacy and expand patient eligibility

Tumor-infiltrating lymphocyte (TIL) therapy is at the cusp of approval for heavily pretreated patients with solid tumor malignancies. TIL therapy currently requires IL2 for in vivo maintenance of TILs, significantly limiting its application due to patient safety and eligibility hurdles. cytoTIL15 is a TIL product engineered with regulatable membrane bound IL15 (mbIL15) designed via our cytoDRiVE® platform. Armoring TILs with endogenous mbIL15 has several advantages over systemic IL2. Unlike IL2, IL15 does not increase immunosuppressive regulatory T-cells and drives T-cell differentiation towards a stem-cell memory phenotype associated with long-term persistence.

Abstract LB150: Clinical Feasibility and Treatment Outcomes with Unselected Autologous Tumor Infiltrating Lymphocyte Therapy in Patients with Advanced Cutaneous Melanoma

Abstract Introduction: A minority of patients (pts) with advanced melanoma achieve long-term survival with immunotherapy and those who relapse following immune checkpoint inhibition (ICI) or BRAFi have limited treatment options. The intrinsic anti-tumor activity and broad neoantigen-specific TCR repertoire of unselected autologous TIL may provide advantages over other treatments in solid tumors, including ICI-refractory melanoma. Methods: This is a retrospective analysis of a single-center experience of TIL for treatment of advanced cutaneous melanoma. Unselected autologous young TIL derived from digested tumor tissue were manufactured under a MHRA Manufacturing Specials license. Pts with advanced cutaneous melanoma and no other treatment options received non-myeloablative lymphodepleting chemotherapy (cyclophosphamide 60 mg/kg/d x 2d, fludarabine 25 mg/m2/d x 5 d [Cy/Flu]), followed by TIL infusion and post-TIL high-dose (HD) IL-2 (600,000-720,000 IU/Kg) on a compassionate use basis. Efficacy for 15 imaging-evaluable pts was reported by investigator assessment of CT/MRI per RECIST 1.1; 6 additional pts were followed using non-RECIST 1.1 imaging (PET) and clinical monitoring. Clinically significant adverse events (AEs) with onset post-TIL infusion were reported for all treated pts. Data cutoff date: 31-DEC-2019. Results: Between OCT-2011 and AUG-2019, 21 pts with advanced cutaneous melanoma were treated with Cy/Flu, TIL (median 31.9 x109 cells infused), and HD IL-2 (median 8 doses). All had high-risk metastatic disease (median 4 sites, stage IV M1d [38%]), an average of 3 prior therapies (any ICI [91%], PD-1 [57%], BRAFi [52%], and MEKi [24%]). With a median follow up of 52.2 mos, the response rate in imaging-evaluable pts (n=15) was 53% with a CR rate of 13%; DCR was 73% including 3 (20%) pts with stable disease. Additional durable responses were observed in the 6 pts followed by PET and clinical monitoring. Responses were generally consistent across subgroups including age, number of disease sites, tumor burden, brain metastases, number of prior lines of therapies, prior PD-1 blockade, prior BRAFi, and prior MEKi. For all treated pts, the median survival was 21.3 mos. Toxicity was generally self-limited and consistent with Cy/Flu and HD IL-2. Common AEs (≥20%, any grade) were thrombocytopenia (62%), pyrexia (57%), rigors (43%), neutropenia (29%), tachycardia (29%), pulmonary edema (24%), and vascular leak (24%); no treatment-related deaths were observed. Conclusion: The high response rate observed in this series exceeds the 41% ORR estimated for TIL in advanced cutaneous melanoma (meta-analysis by Dafni et al. Ann Oncol., 2019) and highlights the successful bench-to-bedside application of unselected autologous TIL to address unmet medical need in advanced melanoma. Use of tumor digests as starting material for manufacturing of TIL demonstrates feasibility of this approach. A multicenter Phase 2 trial of this therapy in advanced melanoma is planned for 2021. Citation Format: Robert E. Hawkins, Yizhou Jiang, Paul C. Lorigan, Fiona C. Thistlethwaite, Manon Pillai, Martine Thomas, Natalia Kirilova, John S. Bridgeman, Gray Kueberuwa, Ryan S. Guest, Zachary J. Roberts. Clinical Feasibility and Treatment Outcomes with Unselected Autologous Tumor Infiltrating Lymphocyte Therapy in Patients with Advanced Cutaneous Melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB150.

Clinical efficacy of T-cell therapy after short-term BRAF-inhibitor priming in patients with checkpoint inhibitor-resistant metastatic melanoma

PurposeDespite impressive response rates following adoptive transfer of autologous tumor-infiltrating lymphocytes (TILs) in patients with metastatic melanoma, improvement is needed to increase the efficacy and broaden the applicability of this...

Stem cell properties of GT101, a tumor infiltrating lymphocyte (TIL) product with multiple indications.

e14506 Background: Tumor infiltrating lymphocyte (TIL) is a kind of adoptive T cell therapy that has demonstrated clinical benefits in several types of solid tumors. GT101 is an autologous TIL therapy for the treatment of multiple solid tumors. Previous studies showed that in vitro expansion resulted in over 80% terminally differentiated effector T cells in the final TIL products, which may show good cytotoxicity but weak capability of persistence and proliferation in vivo. Herein we present phenotype and function characterization of our TIL product GT101 with a focus on its stemness attributes. Methods: We have produced more than 30 batches of GT101 TIL samples from 10 different cancer indications and analyzed their proliferation, cytokine production and phenotype at preREP and/or REP stage. Results: GT101 products keep a consistent T cell phenotype and good proliferation capacity, and are capable of secreting large amounts of cytokines after stimulation by anti-CD3/CD28 antibody, reflecting its good functionality. The residues in the final products, such as feeder cells and tumor cells, are below the detection limit. In contrast to equivalent TIL products previously reported across the world, we find that GT101 features a manufacturing process that preserves the stem cell characteristics of T cells. Analysis of all the 30 GT101 TIL samples demonstrates that the average percentage of central memory T cells (Tcm) and naïve T cells is 40-50% and 10-20% respectively in both CD8+ and CD4+ populations at the end of preREP stage. Meanwhile, CD8+TCF1+ cells are close to 50%, mainly in the form of PD-1-. In the phase of REP, the stemness of T cell is still well maintained—the average proportion of both Tcm and naïve T cells only decreases by about 5%, and that of CD8+TCF1+ cells down by about 10%. Conclusions: In general, GT101 demonstrates good stemness with a higher percentage of Tcm and TCF1+ than equivalent products previously reported across the world, suggesting better performance in persistence and proliferation in clinic.

In-depth immune and molecular profiling of melanoma patients receiving adoptive T-cell therapy reveals biomarkers of efficacy in ATATIL study.

2533 Background: Adoptive cell therapy (ACT) using tumor-infiltrating lymphocytes (TIL) has demonstrated a curative potential for patients with metastatic melanoma (MM). Nevertheless, activity remains unsatisfactory in many patients, requiring development of biomarkers that predict therapeutic efficacy. We report results of a single-center phase I study to assess feasibility, safety and efficacy of TIL-ACT in MM patients (NCT03475134). Methods: Patients with MM refractory to at least one prior line of therapy received TIL therapy with lymphodepleting chemotherapy before T-cell infusion, followed by high-dose interleukin-2. RDG- and FDG-PET imaging was performed before and after TIL infusion. Multispectral immuno-fluorescence (mIF) imaging and bulk-RNA sequencing (Seq) were performed on tumor samples pre-ACT and post-ACT (day+30 and upon progression). Single-cell RNA-Seq and TCR-Seq were performed on pre-ACT tumor and ACT product, as well as on tumor-reactive and neoantigen-specific TILs and on longitudinal blood samples. Results: As of 02/02/2021, thirteen patients (enrolled between March 2018 and December 2020) have successfully completed TIL-ACT therapy, with a median follow-up of 9.5 months (IQR 3.0 -24.6). Median age was 53 years (range 20-69) and all were previously treated with PD-1 based blockade. Median number of TILs infused was 55.0 x109 cells (range 12.8-84.7). The best overall response rate by RECIST 1.1 and disease control rate in evaluable patients was 41.7% (5/12) and 50% (6/12) respectively at 3 months. Two patients have an ongoing near-complete response at 3 years. Up to data cut-off, 10 patients have progressed by RECIST v1.1, with median PFS of 4.8 months (95% CI 1.5 - 9.6), while median OS is not reached. mIF revealed biomarkers of response, which may allow proper identification of patients in subsequent studies. In addition, deep sequencing of bulk and neoepitope-specific TIL clonotypes highlighted transcriptomic signatures revealing cell programs regulating in vitro expansion, in vivo blood persistence as well as tumor infiltration post-ACT. RGD-PET data will also be presented. Conclusions: We demonstrate reproducibility of TIL-ACT in our center, consistently with previous reports. Comprehensive translational studies reveal immune correlates of clinical responses that contribute to the understanding of mechanisms of TIL potency and will guide the development of next-generation cell products. Clinical trial information: NCT03475134.