GITR activation potentiates anti-tumor immunity of tumor-infiltrating lymphocytes expanded from glioblastoma by rescuing exhaustion.

Background: Adoptive cell therapy with autologous TIL has demonstrated an objective response rate (ORR) of 36% in the post-immune checkpoint inhibitor (ICI) setting in patients (pts) with advanced/unresectable melanoma (Sarnaik JCO 2021), while in ICI-naïve pts who received early-line combination of TIL and pembrolizumab, the ORR was 60%, with a 30% CR rate (O’Malley SITC 2021). Although effective, anti-PD-1 therapy is limited by poor penetration into the tumor, internalization, and endocytic clearance, in contrast with TIL, which overcome this inherent limitation. PDCD-1 gene inactivation (PD-1 KO) may enhance TIL cell therapy efficacy in the post-ICI setting and abrogate the need for systemic anti-PD-1 therapy in ICI-naïve pts. In preclinical studies, PD-1 KO TIL maintain robust effector function and phenotypic markers indicative of functional TIL (Ritthipichai ESMO 2020). We describe preclinical activity, clinical-scale manufacturing process development, and characterization of IOV-4001, an autologous PD-1 KO TIL cell product. Methods: hIL-2 NOG mice engrafted with melanoma tumor cells received adoptive transfer of autologous PD-1 KO TIL (developed with TALEN® gene editing technology in collaboration with Cellectis), mock TIL (electroporation without TALEN), mock TIL + anti-PD-1 antibody, or no adoptive transfer (n=14 each). Tumor size was measured 2×/wk for 39 days. A 22-day clinical-scale manufacturing process was established, including pre-rapid expansion protocol (pre-REP), activation, electroporation, resting, and REP, for the generation of PD-1 KO TIL. Final PD-1 KO TIL product was characterized for total viable cells (TVC), purity (% viability), identity (% CD45+CD3+), and function (PD-1 KO efficiency). Results: Day 39 mean ± SEM tumor size (mm2) for mice treated with PD-1 KO TIL (6 ± 2.8) showed superior tumor control relative to mock TIL (26 ± 8.5, P<0.05), mock TIL + anti-PD-1 (33 ± 8.8, P<0.01), and no adoptive TIL transfer (112 ± 8.4, P<0.0001). Product attributes from 6 clinical-scale manufacturing runs for PD-1 KO TIL were acceptable, with a median (range) TVC, purity, and identity of 8.3 × 109 (0.9×109-35.7×109), 94% (91%-99%), and 99% (98%-99%), respectively. Median (range) PD-1 KO efficiency was 48% (31%-84%). PD-1 KO TIL function and phenotype (differentiation, memory, activation, and exhaustion) were comparable to mock TIL. Conclusions: Anti-tumor activity of PD-1 KO TIL was superior to mock TIL suggesting that endogenous PD-1 inhibition may confer a functional advantage to the TIL over an antibody combination. PD-1 KO TIL clinical manufacturing was feasible and the TIL product quality attributes and phenotype were acceptable; importantly, lack of complete PD-1 KO may spare other PD-1-dependent in vivo cellular functions. Together, these data support clinical investigation of IOV-4001, an autologous PD-1 KO TIL cell therapy. Citation Format: Arvind Natarajan, Anand Veerapathran, Adrian Wells, Kenneth Onimus, Marcus Machin, Seth Wardell, Jamie L. Blauvelt, Madan Jagasia, Rafael Cubas. Preclinical activity and manufacturing feasibility of genetically modified PDCD-1 knockout (KO) tumor-infiltrating lymphocyte (TIL) cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2746.

TPS9608 Background: Metastatic melanoma (MM) patients who have progressed following immune checkpoint inhibitors (ICI; anti–PD-1/CTLA-4 therapy) and targeted agents (BRAF/MEK inhibitors) have limited treatment options and dismal prognosis. An autologous tumor-infiltrating lymphocytes (TIL) therapy (LN-144, lifileucel) has demonstrated efficacy and durable responses in heavily pretreated advanced melanoma. In a phase II study of TIL therapy (lifileucel) with traditional lymphodepletion (LD), using fludarabine 25 mg/m2 for 5 days and cyclophosphamide (Cy) 60 mg/kg for 2 days, in 153 MM patients previously treated with ICI and BRAF/MEK inhibitors, an overall response rate (ORR) of 31% was noted, and the median duration of response (DOR) was not reached after 27.6 month follow up. We hypothesize that a reduced dose of Flu/Cy LD for TIL therapy will have similar TIL expansion and persistence post-infusion, resulting in similar efficacy with a reduced toxicity profile. Methods: Study NCT06151847 is a single-center, open-label phase II pilot trial evaluating the efficacy, in vivo persistence, and safety of TIL therapy after reduced dose Flu/Cy LD. Key inclusion criteria include unresectable or metastatic melanoma (stage IIIC-IV) with disease progression after one or more lines of therapy. Four of the 12 planned patients have been enrolled as of February 2, 2024. Central TIL manufacturing from at least a 1.5 cm tumor specimen involves ex vivo expansion through cell culture in the presence of the interleukin (IL)-2 and an anti-CD3 monoclonal antibody (1-150 x109 viable cells). Patients will receive an outpatient reduced-dose Flu/Cy LD regimen of Flu (30 mg/m2 on days -4, -3, -2, -1) and Cy (750 mg/m2 on days -4, -3, -2). Infusion of TIL will be performed on day 0 followed by high-dose IL-2 (600,000 IU/kg up to 6 doses). The primary objective will be to ascertain TIL persistence using T-cell receptor (TCR) sequencing at day 42 which will be compared to historical data already generated utilizing traditional Flu/Cy LD. The secondary objectives will be to evaluate the efficacy parameters including ORR, DOR, progression-free survival (PFS), and overall survival (OS), and the safety profile of TIL in combination with a reduced dose Flu/Cy LD regimen. Exploratory objectives will involvecorrelative analyses to characterize the immunome and microenvironment. If TIL persistence is roughly equivocal between the reduced and high-dose Flu/Cy LD regimens, the design of a larger-scale trial using reduced-dose Flu/Cy LD for TIL therapy may be considered. Study follow-up will continue for one year. Clinical trial information: NCT06151847 .

NK ACTIVITY OF TUMOR INFILTRATING AND LYMPH NODE LYMPHOCYTES IN HUMAN PULMONARY TUMORS

INTRODUCTION: TILs (Tumor infiltrating lymphocytes) therapy has been shown to be highly effective in melanoma and holds great potential for improving cancer immunotherapy in other cancers. The key limitation for TIL therapy is the complex process to activate and expand TILs. Thus, there is a strong need for refining strategies to enable TIL immunotherapy. To address this challenge, we hypothesized that we can directly activate and expand TILs in vivo, thereby eliminating the complex production process. We took advantage of a multifunctional nanoparticle (NP) platform to engineer NPs with multiple co-stimulatory molecules. We aimed to show that our in vivo approach can effectively stimulate TILs. METHODS: Co-stimulatory molecules functionalized NPs (TIL-NPs) were synthesized through reacting Azide-functionalized NP cores with DBCO-functionalized molecules. TIL-NPs were injected intratumorally to promote the expansion and activation of TILs in vivo. The MC38 colorectal tumor model was used for evaluating the effects of TIL-NPs on TILs activation. For the in vitro experiment, TILs from tumors of MC38-bearing mice were incubated with MC38 cells following various stimulation strategies. For the in vivo experiment, a bilateral tumor model was established, with only the right flank receiving TIL-NP injection. RESULTS: We successfully engineered NPs containing anti-CD3, anti-41BB and IL-2. These NPs were 126.2 nm in size. Per miligram NP conjugated with approximately 50μg immune activating molecules. TILs from MC38 tumors were incubated with TIL-NPs. Free antibodies were used as control. We found TILs stimulated by TIL-NPs demonstrated greater cytotoxicity against MC38 cells compared to the groups treated with free antibodies or unstimulated TILs. IFN-γ levels were elevated in all treatment groups, with the most significant increase observed in the TIL-NPs group. To determine if TIL-NPs increased T-cell activation in vivo, we evaluate the antitumor effects of TIL-NPs in a bilateral tumor model. One tumor received TIL-NPs while the other tumor was used to assess systemic treatment responses. Intratumorally injected TIL-NPs were highly effective in eradicating the injected tumor when compared to that of control and free antibodies groups. All of the tumors on the right flank/injected completely regressed, achieving a 100% complete response (CR) rate. Importantly, the TIL-NPs also provided a robust systemic response, providing 60% CRs in the contralateral flank tumors. Overall, TIL-NPs significantly prolonged survival of mice. Functionalizing these molecules on NPs also induced clustering of their respective receptors and improved signaling. CONCLUSION: TIL-NPs engineered with multiple co-stimulatory molecules can directly stimulate TILs in vivo, providing a systemic immunotherapy response against cancer. Citation Format: Xin Wen, Lifu Ruan, Longzhen Zhang, Tian Zhang, Andrew Wang. Engineered nanoparticles promote direct in vivo expansion and activation of tumor infiltrating lymphocytes for cancer growth inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3477.

BackgroundAdoptive Cell Transfer (ACT) with tumor infiltrating lymphocytes (TIL) is a novel immunotherapeutic strategy that has shown encouraging results in metastatic melanoma, cervical cancer, and ovarian cancer.1–5 The immunogenic tumor...

Background: T cell exhaustion is driven by persistence of antigen and inflammation, common features of cancer. The success of a checkpoint inhibitor blockade may depend upon reactivation of pre-existing tumor-specific CD8+ T cells in the tumor microenvironment. Using an in vitro model and tumor infiltrating lymphocytes (TILs) isolated from multiple tumors, we explored microRNAs (miRNA, miR) involved in T cell dysfunction in order to identify pathways that may be important for altering this phenotype following immunotherapy. Methods: We used an established in vitro model of T cell exhaustion: healthy donor CD8+ T cells were stimulated with anti-CD3/CD28 for six days. We used RNASeq and quantitative PCR to evaluate genomic (mRNA and microRNA (miR)) changes associated with T cell function and paired differentially expressed microRNAs with predicted target genes. Exhaustion of CD8+ TILs isolated from melanoma (n = 2), NSCLC (n = 3), renal (n = 3), bladder (n = 10), and colorectal tumors (n = 2) was measured by flow cytometry analysis of PD-1/TIM-3. miRNA and mRNA relationships identified by the in vitro model were evaluated in a subset of TILs from NSCLC, renal, and bladder tumors (n = 6). Results: Results showed expected phenotypic and functional changes across 6 donors stimulated chronically with anti-CD3/CD28: 2-4-fold increased PD-1 and TIM-3 surface expression with a 3-5-fold loss of intracellular IFNg production (p < 0.05). Comparing exhausted T cells with unstimulated T cells revealed ~1300 differentially expressed genes and ~100 differentially expressed miRs (p < 0.05). A set of differentially expressed genes/miRs from the in vitro exhaustion model was then confirmed in TILs from NSCLC, bladder, and renal tumors. Differentially expressed miRs included microRNAs-155 and 181a, previously implicated in regulating IFNg signaling and T cell proliferation, respectively. Focusing on miRs that were altered between activation and exhaustion revealed a novel miR exhaustion signature, which was increased ≥4-fold relative to activated T cells. Evaluating this miR exhaustion signature in TCGA demonstrated a 16-fold range in expression across indications, with melanoma, head and neck, and NSCLC, among the highest median expression, while renal, HCC and rectal cancers were among the lowest. In HCC, GITR and OX40 pathway activation signatures were increased >2-fold in tumors with high miR signature, while in renal cancer, B cell and CD40 pathway signatures were enriched. In melanoma, high miR signature showed a trend toward increased myeloid-derived suppressor cell (MDSC) signature expression (>1.5-fold change), while the opposite trend was observed in head and neck cancer. Conclusions: We identified a novel miRNA exhaustion signature associated with immune-related pathways in multiple tumor types. This signature may help generate hypotheses to guide prioritization of specific IO combination treatments. Citation Format: Lydia M. Greenlees, Michael Kuziora, Yinong Sebastian, Todd Creasy, Young S. Lee, Fernanda Pilataxi, Nick Holoweckyj, Li Cheng, Brandon W. Higgs, Koustubh Ranade, Katie Streicher. microRNA signature of T cell exhaustion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5390.

BackgroundTumor-associated macrophage (TAM) infiltration has been shown to modulate response to immune checkpoint inhibitors in various cancers, but its role in metastatic clear cell renal cell carcinoma (mccRCC) remains unclear. Here, we investigated the role of CD163+ TAMs as a potential determinant of clinical outcomes to first-line anti-PD-1 therapy (nivolumab) in patients with mccRCC enrolled in the HCRN GU16-260 trial. Moreover, as recent data suggest that the interaction between TAMs and tumor infiltrating lymphocytes (TILs) promotes T cell exhaustion, we explored the spatial relationship between CD163+ TAMs and CD8+ TILs in different states of exhaustion (ie, terminally exhausted (TE) and non-terminally exhausted (NTE) CD8+ TILs).MethodsPre-treatment tumor samples from 67 patients were analyzed by multiplex immunofluorescence to identify CD163+ TAMs, CD8+PD-1+TIM-3+ and/or LAG-3 + (TE CD8+), and CD8+PD-1+TIM-3−LAG-3− (NTE CD8+) TILs. Associations between the natural log of density of CD163+ TAMs with progression-free survival (PFS) and objective response rate (ORR) were assessed using univariable Cox and logistic regression models, respectively. An optimized cutoff was determined using minimum p value for ORR. For each tumor, the density of TE CD8+ TILs and the density of NTE CD8+ TILs were calculated within a 30 µm radius area centered on CD163+ TAMs (proximal area) and outside of this area (non-proximal area), using the ‘sf’ package within R software. The densities of TE and NTE CD8+ TILs were compared in proximal versus non-proximal areas across all tumor samples using the Wilcoxon signed-rank test. For each CD8+ TIL population (TE and NTE), the enrichment in proximity of CD163+ TAMs was assessed by calculating the difference in densities in proximal and non-proximal areas normalized by the density in the overall tumor area. The level of enrichment in TE CD8+ TILs versus NTE CD8+ TILs in proximity of CD163+ TAMs was compared using the Wilcoxon signed-rank test.ResultsThe density of CD163+ TAMs, analyzed as a continuous variable was positively associated with ORR (OR: 2.21, 95% CI: 1.33 to 3.69, P = .002) and PFS (HR: 0.77, 95% CI: 0.61 to 0.97, P = .028). At an optimized cutoff, patients with high density of CD163+ TAMs (n = 34, 50.7%) had higher ORR (65% vs. 15%, P < .001) and longer median PFS (16.6 months, 95% CI: 5.5-32.9 vs. 5.5 months, 95% CI: 4.1-10.6, P = .009) compared to patients with low density of CD163+ TAMs (n = 33, 49.3%). The density of CD163+ TAMs was moderately correlated with the density of TE CD8+ TILs (Spearman correlation, r = 0.55) and weakly correlated with the density of NTE CD8+ TILs (r = 0.32). Proximity analysis showed that the density of TE CD8+ TILs was significantly higher in the area proximal to the CD163+ TAMs compared to the non-proximal area (median density: 123.3/mm2 vs. 37.2/mm2; P < .001). Similarly, the density of NTE CD8+ TILs was significantly higher in the area proximal to the CD163+ TAMs compared to the non-proximal area (median density: 127.2/mm2 vs. 66.8/mm2; P < .001). The level of enrichment in proximity of CD163+ TAMs was higher for TE CD8+ TILs compared to NTE CD8+ TILs (0.77 vs. 0.58; p = 0.0011).ConclusionsHigh levels of CD163+ TAMs are associated with improved outcomes to anti-PD-1 therapy in mccRCC. In addition, exhausted CD8+ TILs preferentially localize in proximity of CD163+ TAMs in ccRCC tissues, supporting that TAM-T cell interactions are critical for driving T cell dysfunction. Taken together, our data are consistent with the hypothesis that the efficacy of PD-1 blockade may be in part mediated by reprogramming TAMs from a pro-tumorigenic to an anti-tumorigenic state.

BACKGROUND Tumor-infiltrating lymphocyte (TIL) therapy is an emerging treatment option for treatment-refractory melanoma, FDA-approved in 2024. TIL therapy involves tumor harvest, isolation and ex vivo expansion of TIL, followed by lymphodepletion prior to TIL infusion. Studies for TIL therapy approval excluded patients with uncontrolled melanoma brain metastases (MBMs). Given the timeline from TIL harvest to infusion and use of lymphodepletion there are concerns about intralesional hemorrhage, intracranial progression and radiation therapy (RT) toxicity. While RT remains a cornerstone in management of MBMs, there are limited data on management during the peri-TIL therapy window and the safety/efficacy of integrating RT. Herein we review our institutional experience on clinical outcomes and toxicity among patients undergoing RT for MBMs that underwent TIL harvest. METHODS We performed a retrospective analysis of patients with advanced melanoma that underwent TIL harvest at a single institution from May to December 2024. Patients with MBMs were stratified by receipt of RT and timing of development of MBMs in relation to date of TIL harvest. Medical records were reviewed with attention to clinical outcomes, RT parameters and adverse events (CTCAEv5.0) related to TIL therapy or RT. RESULTS A total of 9 patients with MBMs and undergoing TIL harvest were identified. Median age was 63.2 years, and 55% of patients harbored a BRAF V600E variant. Median time from TIL harvest to infusion was 2.1 months and median follow-up post-harvest was 4.4 (2.1-8.3) months. One patient did not undergo infusion due to TIL product not meeting specification. Two patients had known MBMs and underwent RT &amp;gt;3mo prior to TIL harvest. 55% (5 of 9) of patients developed new MBMs during the window from TIL harvest to infusion. 67% (6 of 9 patients) underwent brain-directed RT. 83% (5 of 6) were treated with focal stereotactic RT with 10 MBMs treated over 6 treatment courses. One patient underwent hippocampal-avoidant WBRT. Of note, 50% of patients that underwent RT had evidence of possible intralesional bleed noted on brain MRI prior to RT. RT-related toxicity was minimal with grade 1-2 fatigue, grade 1 headaches and no radiographic or symptomatic radiation necrosis. There were no intracranial bleeding events after TIL harvest. Neurotoxicity related to TIL therapy was observed including grade 2-3 encephalopathy, grade 3 fatigue, and grade 2 headaches. At last follow up, regional brain control was 67% and irradiated local control was 83%. CONCLUSION Our institutional experience suggests that treatment of MBMs with RT in patients undergoing TIL therapy is feasible and safe. Given the risk of intracranial progression or development of new MBMs between TIL harvest and infusion, RT is an option as a consolidative ‘bridging’ therapy for intracranial disease control.

Introduction: Autologous tumor-infiltrating lymphocytes (TIL) engineered to secrete IL-12 have shown robust clinical efficacy in metastatic melanoma. However, systemic toxicity from elevated circulating IL-12 has limited its application. IOV-5001 is a next-generation TIL therapy engineered to inducibly express membrane-tethered IL-12 (TeIL12) upon antigen engagement, restricting IL-12 activity to the tumor microenvironment. This study evaluated the antitumor efficacy and molecular phenotype of IOV-5001. Methods: TIL were derived from solid tumor tissues, cultured, and transduced with a lentiviral vector encoding inducible TeIL12 bicistronically with a growth factor. Antitumor activity of IOV-5001 was tested in vitro via a tumor antigen-directed TCR killing assay or against autologous patient-derived endometrial cancer organoid (PDO), and/or tumor digests. Transcriptional profiling (bulk and single-cell) was used to characterize the molecular features of IOV-5001. Results: IOV-5001 TIL were successfully generated and effectively expressed TeIL12 upon T cell receptor (TCR) engagement. The expression of TeIL12, soluble IL-12, and IFN-gamma was lacking in the absence of stimulation, indicating that the activity of IOV-5001 TIL is dependent on TCR engagement. IOV-5001 demonstrated superior killing and effector cytokine production compared with unmodified TIL in a tumor antigen-directed TCR killing assay and against autologous tumor, irrespective of aldesleukin support. The antitumor activity was further enhanced when IOV-5001 was expanded in an optimized media. Analysis of bulk gene expression profiling revealed transcriptional profiles for IOV-5001 characterized by upregulation of genes involved in cell cycle, telomere maintenance, mitochondrial metabolism, IL-12, and TCR signaling. Downregulated genes exhibited an enrichment for IL-10 and IL-5 signaling, and prostanoid/eicosanoid ligand binding receptors. Single-cell analysis confirmed these patterns were enriched within tethered cytokine-positive TIL. Furthermore, single-cell analysis of preREP cells demonstrated that proportions of putative tumor-reactive CD8 T cells were similar in IOV-5001 TIL among tethered cytokine-positive and -negative TIL, indicating unbiased transduction. Conclusions: These findings demonstrate that IOV-5001 with inducible and membrane-tethered IL-12 confers superior antitumor activity compared with unmodified TIL. The enhanced molecular and functional characteristics highlight IOV-5001 as a promising product for enhancing TIL efficacy in solid tumors. Citation Format: Patrick Innamarato, Joe Dean, Mohammed Alkhouli, Marcus Machin, Nathan Gilbert, Sadie Johnson, Gerard Sapena, Nino Mtchedlidze, Behzad Damirchi, Robyn Oldham, Andrew Yuhas, Judy Fang, Jamie Blauvelt, Shari Pilon-Thomas, Rongsu Qi, Hequn Yin, Sean Hall. IOV-5001, autologous tumor-infiltrating lymphocytes (TIL) armored with inducible membrane-tethered IL-12, shows enhanced antitumor efficacy with an improved cellular state [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4863.

For over two decades, tumor infiltrating lymphocytes (TILs) have been utilized as an adoptive cell transfer therapy to treat cancer in clinical trials. In combination with added immunotherapy treatments, such as IL-2 or anti-CTLA4, TIL therapy has further improved clinic response rate. Following isolation from a patient’s own tumor specimen, TILs that recognize tumor-specific known or unknown antigens can be expanded. These cells are then infused back into tumor-bearing patients to attack the autologous cancer cell population. Despite progress in clinical research, preclinical and translational models of autologous tumor:TIL coculture have been difficult to establish. Our present study established autologous tumor:TIL coculture methods directly from patient tumor specimens. Models were established from BioDuro’s viably frozen tumor bank, containing more than 100,000 patient biospecimens. Each patient specimen contains a heterogeneous population of cell types, including tumor cells, TILs and TAFs (tumor associated fibroblasts). To establish our patient-derived coculture system, these cell populations were expanded under specific growth conditions to then serve as components of the coculture system. In this study, TILs and monocytes from a cohort of patient tumor specimens were analyzed by FACS to evaluate surface receptor expression. Leukocytes, T cells, monocytes and B cells were marked with CD45, CD3, CD14, CD20, respectively. PD-1 expression levels were also evaluated, with the results showing significant individual variation of expression on cell subpopulations. Established methods were used for the expansion of TILS, supporting either CD4 or CD8 T cell phenotypes. Autologous tumor cells were also expanded, concurrently. Afterward, TILs and tumor cells were cocultured and tumor cell killing was monitored using high content fluorescent image cytometry. We found that autologous patient-matched TILs showed preferential tumor cell killing as compared to coculture with patient-unmatched TILs. This response was observed in a dose-dependent manner. In conclusion, BioDuro’s biobank of viably frozen tumor and TIL patient specimens serves as an expansive and powerful immunooncology resource to establish patient-derived autologous tumor:TIL coculture assays. These assays, along with the ability to readily study larger populations of cancer patients, provide a powerful in vitro platform to test immunooncology drug candidates. Citation Format: Kaede Hinata, Patrice Cuff, Wei Liang, Thomas Broudy. Development of autologous tumor: TIL coculture from patient-derived samples for in vitro preclinical immuno-oncology studies [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C117. doi:10.1158/1535-7163.TARG-19-C117

BackgroundPatients with RCC may achieve remission with immune-checkpoint inhibitors (ICI); however, most patients will progress. Adoptive cell therapy with autologous TIL allows for expansion of T-cells from tumor tissue leading...

127O Response to TIL therapy depends on a tumor-intrinsic IL-2 signaling axis that drives expansion of CD25+ tumor-reactive T cells

The recent U.S. FDA approval of lifileucel, a non-engineered, autologous tumor-infiltrating lymphocyte (TIL) therapy, for unresectable or metastatic melanoma represents a major milestone for cellular therapies in solid tumors. This review examines the clinical foundation, regulatory development, limitations, and evolution of TIL therapy in metastatic melanoma. Randomized academic data from the phase III M14TIL trial established the efficacy of TIL therapy. The C-144-01 study leading to lifileucel approval demonstrated median duration of response of 36.5 months, median overall survival (OS) of 13.9 months, and estimated 5-year OS rate of 19.7%, a major advance in this anti-PD-1/PD-L1 resistant cohort without effective treatment options. Despite durable responses, classical TIL therapy requires intensive nonmyeloablative lymphodepletion and high-dose interleukin-2 (IL-2), contributing to substantial toxicity and treatment-related mortality that remain barriers to broader implementation. We discuss safety-driven trial terminations related to cytokine augmentation and feasibility or strategic factors underlying discontinuation of programs, underscoring translational challenges beyond biologic efficacy. Engineered TIL platforms aim to improve persistence and reduce systemic cytokine dependence. OBX-115, designed with regulatable membrane-bound IL-15 expression, eliminates the need for IL-2 infusion and has shown early clinical activity. KSQ-001EX uses CRISPR/Cas9 to inactivate SOCS1, while KSQ-004EX additionally targets Regnase-1 to enhance TIL function. Emerging strategies including IL-2-independent expansion platforms, PD-1-edited TILs, and neoantigen-enriched products illustrate ongoing innovation. TIL therapy remains among the most promising strategies in melanoma and solid tumors after immunotherapy failure. Ongoing research aims to optimize cell dose, phenotype, tumor procurement, treatment sequencing, and rational combinations to improve durable benefit.

Simple SummaryTreatment options for colorectal cancer (CRC) patients with liver metastases are often limited to liver surgery with or without chemotherapy. However, not all patients present operable colorectal liver metastases (CRLMs). Thus, alternative therapies that exploit the anti-tumor potential of tumor-infiltrating lymphocytes (TILs) are being evaluated. The establishment of markers connecting the phenotype to the function of tumor-reactive CD8+ TILs could aid diagnostic and therapeutic advances. In this regard, tissue-resident memory T cells (TRM cells) could be a potential candidate for therapies targeting TILs. Putative tumor-reactive TRM cells among CD8+ TILs likely co-express CD103 and CD39, since these markers indicate stable tumor residency and repeated response to antigens from the tumor environment, respectively. Our phenotypic and functional analyses of TILs in CRLM, with a specific focus on CD103+CD8+ TRM cells, may guide the improvement of TIL-mediated CRC treatments.The diversity of T cells in the human liver may reflect the composition of TILs in CRLM. Our ex vivo characterization of CRLM vs. adjacent liver tissue detected CD103+CD39+CD8+ TRM cells predominantly in CRLM, which prompted further assessments. These TRM cells responded to cognate antigens in vitro. As functional activities of autologous TILs are central to the implementation of personalized cancer treatments, we applied a patient-derived xenograft (PDX) model to monitor TILs’ capacity to control CRLM-derived tumors in vivo. We established PDX mice with CRLMs from two patients, and in vitro expansion of their respective TILs resulted in opposing CD4+ vs. CD8+ TIL ratios. These CRLMs also displayed mutated KRAS, which enabled trametinib-mediated inhibition of MEK. Regardless of the TIL subset ratio, persistent or transient control of CRLM-derived tumors of limited size by the transferred TILs was observed only after trametinib treatment. Of note, a portion of transferred TILs was observed as CD103+CD8+ TRM cells that strictly accumulated within the autologous CRLM-derived tumor rather than in the spleen or blood. Thus, the predominance of CD103+CD39+CD8+ TRM cells in CRLM relative to the adjacent liver and the propensity of CD103+CD8+ TRM cells to repopulate the autologous tumor may identify these TILs as strategic targets for therapies against advanced CRC.

128O Evolution of transcriptomic and epigenomic intra-tumor heterogeneity in high-grade serous ovarian cancer with chemotherapy