bims-tuinly Biomed News
on Tumor-infiltrating lymphocytes therapy
Issue of 2025–11–23
thirteen papers selected by
Pierpaolo Ginefra, Ludwig Institute for Cancer Research



  1. Front Oncol. 2025 ;15 1693912
      Adoptive cell therapy using tumor-infiltrating lymphocytes (TIL) can achieve durable responses in patients with metastatic cancers, but the long-term clonal dynamics after multiple administration and synergy with checkpoint blockade remain understudied. We present a longitudinal case study of a patient with treatment-refractory metastatic prostate cancer that achieved complete and durable tumor remission over 5-years after multiple TIL infusions and anti-PD-1 therapy. We performed longitudinal high-throughput T-cell receptor (TCR) sequencing on blood and tumor samples collected over five years to track the persistence and dynamics of TIL-derived and endogenous clonotypes. TIL-derived clonotypes exhibited sustained persistence in blood, with notable clonal expansions correlating with reduced repertoire diversity, increased clonality, and observed clinical response. Multiple TIL administration increased the patient exposure to the therapy, improving its pharmacokinetics profile over time. The third TIL infusion was followed by pembrolizumab administrations, which coincided with the re-expansion of TIL-derived clonotypes and emergence of novel clones. Serial tracking revealed clonotype stability for up to five years post-treatment. Our findings provide insights into the long-term persistence and reactivation of TIL-derived immunity and illustrate the potent synergy between adoptive transfer and PD-1 blockade by enhancing both infused and endogenous tumor-reactive T cell responses, and supporting the integration of longitudinal immunogenomic monitoring in personalized immunotherapy.
    Keywords:  TCR NGS; anti-PD-1; immune checkpoint blockade; pembrolizumab; tumor-infiltrating lymphocytes (TIL)
    DOI:  https://doi.org/10.3389/fonc.2025.1693912
  2. PeerJ. 2025 ;13 e20322
      Non-muscle-invasive bladder cancer (NMIBC) frequently recurs and progresses into an aggressive and lethal entity within five years. The clinical management of recurrent tumors remains limited. Therefore, identifying individual patients who are at a high risk of recurrence is crucial for early clinical monitoring and appropriate medical intervention, which may lead to improved outcomes. OX40 is a dual modulator that stimulates effector T cells and suppresses Tregs. It appears to be an ideal molecule for predicting survival outcomes, surpassing the predictive power of single or combined T cell signatures. It has been shown to act as an independent tumor prognostic predictor in various cancers, including non-small cell lung cancer, melanoma, and colorectal cancer. However, its potential as a prognostic tool for tumor recurrence in NMIBC has yet to be investigated. The present study aimed to investigate the potential value of OX40 as a predictor of recurrence risk in patients with NMIBC. Additionally, its downstream effectors, Foxp3 and CD8, were also evaluated. Tissue samples were collected from a cohort of 110 patients diagnosed with NMIBC. Immunohistochemistry was performed to assess the density of stromal OX40+, Foxp3+, and CD8+ tumor infiltrating lymphocytes (TILs). Following survival analysis using the Kaplan-Meier method and log-rank test, we found that tumor recurrence was associated with a decreased density of OX40+ and CD8+ TILs, an elevated density of Foxp3+ TILs, and lower ratios of OX40+/Foxp3+ and CD8+/Foxp3+ TILs. However, after adjustment, multivariate COX regression analysis indicated that only the ratio of CD8+/Foxp3+ was an independent predictor of recurrence risk. The prediction power was assessed by a receiver operating characteristic (ROC) curve analysis. The results demonstrated that the AUC value for the CD8+/Foxp3+ ratio was better than the other predictive markers. Although the expression of OX40 in TILs was associated with tumor recurrence, our results suggest that the predictive efficacy of a combination of CD8 and Foxp3 was more robust after adjustment. Future research utilizing advanced immunotyping techniques is necessary to validate these findings in larger cohorts.
    Keywords:  CD8; Foxp3; Non-muscle-invasive bladder cancer; OX40; Tumor infiltrating lymphocytes
    DOI:  https://doi.org/10.7717/peerj.20322
  3. Cancer Lett. 2025 Nov 13. pii: S0304-3835(25)00717-7. [Epub ahead of print] 218145
      Immune evasion in cancer is frequently mediated by checkpoint molecules such as programmed death-1 (PD-1), programmed death-ligand 1 (PD-L1) and cytotoxic T-lymphocyte-associated protein-4 (CTLA-4), which suppress T-cell recognition and perpetuate tumor progression. To counter this immunosuppressive axis, immune checkpoint inhibitors (ICIs) have been developed to reinvigorate antitumor immunity by reversing T lymphocyte dysfunction. Over 30 ICIs have been clinically approved for the treatment of malignant tumors. Despite these advances, ICI monotherapy remains limited due to the primary or acquired resistance and suboptimal efficacy in immunologically "cold" tumor microenvironments characterized by inadequate T-cell infiltration. To address these challenges, combinatorial strategies integrating ICIs with tumor-infiltrating lymphocyte (TIL) therapy are gaining momentum. This review synthesizes preclinical and clinical evidences supporting TIL-ICI synergy in solid tumors, wherein adoptively transferred tumor-reactive T cells enhance intratumoral immune activation while ICIs sustain TIL effector functions by alleviating checkpoint constraints. We further discuss emerging strategies to optimize this paradigm, including engineered TILs with enhanced persistence, biomarker-driven patient stratification, and multimodal regimens incorporating anti-angiogenics or epigenetic modulators. Despite promising early-phase trial outcomes, barriers to clinical adoption still persist, such as manufacturing complexity, immune-related toxicity management, and the paucity of Phase III validation. By delineating mechanistic rationale, current progress, and translational roadblocks, this work provides a roadmap for advancing TIL-ICI combinations toward broader therapeutic implementation.
    Keywords:  Combination therapy; Immune checkpoint inhibitors; Immunotherapy; PD-1; Tumor infiltrating lymphocytes
    DOI:  https://doi.org/10.1016/j.canlet.2025.218145
  4. Front Immunol. 2025 ;16 1651129
       Background: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality, with limited curative options for advanced disease. Natural Killer (NK) cells are critical innate immune effectors, but their anti-tumor function is severely compromised by the immunosuppressive tumor immune microenvironment (TIME), particularly through transforming growth factor-beta (TGF-β). This study investigates the pivotal role of TGF-β signaling in modulating NK cell phenotypes and functions within the HCC TIME.
    Methods: to comprehensively assess TGF-β pathway activation and its impact on NK cells, tumor-infiltrating lymphocytes (TILs) and liver-infiltrating lymphocytes (LILs) were isolated from HCC patients undergoing curative resection. Phenotypic and functional analyses were performed, along with functional restoration experiments targeting TGF-β signaling.
    Results: Tumor-infiltrating NK cells (TINKs) exhibited significant activation of both canonical (SMAD-dependent) and non-canonical (TAK1/p38 MAPK) TGF-β signaling, with a predominance of the non-canonical pathway. This activation was associated with the emergence of an ILC1-like NK subset (CD103+/CD49a+), which was nearly absent in non-tumor liver tissue. These ILC1-like cells maintained strong cytokine production and expressed high levels of inhibitory receptors (PD-1, TIM-3, TIGIT), whereas conventional NK cells (cNKs; CD103-/CD49a-/CD9-) were functionally impaired. Notably, blocking TGF-β receptor binding and SMAD3 activation restored cNK functionality.
    Discussion: our findings suggest that while non-canonical TGF-β signaling drives phenotypic reprogramming and contributes to NK cell dysfunction, canonical SMAD-dependent signaling remains a key therapeutic target for functional restoration. These results highlight the dual role of TGF-β in immune modulation and suggest that targeted pathway inhibition could enhance innate anti-tumor responses, opening new avenues for combination therapies in HCC.
    Keywords:  ILC1-like cells; NK cells; TGF-β; functional restoration; hepatocellular carcinoma; immune function
    DOI:  https://doi.org/10.3389/fimmu.2025.1651129
  5. Nature. 2025 Nov 19.
      Emerging evidence suggests a correlation between CD8+ T cell-tumour cell proximity and anti-tumour immune response1,2. However, it remains unclear whether these cells exist as functional clusters that can be isolated from clinical samples. Here, using conventional and imaging flow cytometry, we show that from 21 out of 21 human melanoma metastases, we could isolate heterotypic clusters, comprising CD8+ T cells interacting with one or more tumour cells and/or antigen-presenting cells (APCs). Single-cell RNA-sequencing analysis revealed that T cells from clusters were enriched for gene signatures associated with tumour reactivity and exhaustion. Clustered T cells exhibited increased TCR clonality indicative of expansion, whereas TCR-matched T cells showed more exhaustion and co-modulation when conjugated to APCs than when conjugated to tumour cells. T cells that were expanded from clusters ex vivo exerted on average ninefold increased killing activity towards autologous melanomas, which was accompanied by enhanced cytokine production. After adoptive cell transfer into mice, T cells from clusters showed improved patient-derived melanoma control, which was associated with increased T cell infiltration and activation. Together, these results demonstrate that tumour-reactive CD8+ T cells are enriched in functional clusters with tumour cells and/or APCs and that they can be isolated and expanded from clinical samples. Typically excluded by single-cell gating in flow cytometry, these distinct heterotypic T cell clusters are a valuable source to decipher functional tumour-immune cell interactions and may also be therapeutically explored.
    DOI:  https://doi.org/10.1038/s41586-025-09754-w
  6. Adv Sci (Weinh). 2025 Nov 18. e12793
      Regulatory T cells (Tregs) are pivotal mediators of immunosuppression in hepatocellular carcinoma, but strategies for selectively disrupting their function remain underdeveloped. Here, puerarin, a natural isoflavone is identifed as a selective immunometabolic modulator. It impairs mitochondrial metabolism in tumor-infiltrating Tregs (Ti-Tregs) without affecting conventional T cells. Mechanistically, puerarin directly binds to MIC19-a core subunit of the mitochondrial contact site and cristae organizing system-leading to its degradation and disruption of the MIC19-MIC60 complex. This disruption causes cristae disorganization, reduces oxidative phosphorylation, and weakens the immunosuppressive function of Ti-Tregs. In vivo, puerarin decreases Ti-Treg infiltration, thereby enhancing antitumor immunity without causing systemic toxicity. Furthermore, MIC19 knockdown and site-directed mutagenesis studies validate the role of critical MIC19 residues (His180, Gln187, and Tyr211) in puerarin's activity. These results reveal a mechanism by which puerarin suppresses mitochondrial metabolism of Ti-Tregs and emphasize the therapeutic potential of natural compounds in metabolic targeting for cancer immunotherapy.
    Keywords:  Regulator T cells; hepatocellular carcinoma; immunotherapy; mitochondrial metabolism; puerarin
    DOI:  https://doi.org/10.1002/advs.202512793
  7. Front Immunol. 2025 ;16 1691746
      Effective anti-tumor immunity critically depends on functional CD8+ T cells, yet in almost all solid tumors, these cells become dysfunctional, exhausted, or spatially excluded. This breakdown of immune surveillance arises not only from cell-intrinsic T cell exhaustion but also from multimodal communication among tumor, stromal, and immune cells within the tumor microenvironment (TME). This communication is mediated not only through direct receptor-ligand interactions but also through a suite of indirect mechanisms, such as metabolic competition, secretion of immunosuppressive metabolites and cytokines, extracellular vesicle exchange, and even mitochondrial transfer via tunneling nanotubes or membrane transfer through T cell trogocytosis. Together, these suppressive interactions impair CD8+ T cell metabolism, effector function, and persistence, thereby enabling tumor immune evasion. In this review, we summarize current understanding of how multimodal cell-cell communication, including immune checkpoints, metabolic reprogramming, and stromal crosstalk, cooperatively drive CD8+ T cell dysfunction. We also highlight emerging therapeutic strategies aimed at rewiring these suppressive networks, with emphasis on translational potential. A deeper understanding of the spatial, molecular, and metabolic context of CD8+ T cell suppression offers new avenues to enhance the efficacy of cancer immunotherapies.
    Keywords:  CD8+ T cell; dysfunction; multimodal cell-cell communication; suppression; tumor microenvironment
    DOI:  https://doi.org/10.3389/fimmu.2025.1691746
  8. Sci Rep. 2025 Nov 20. 15(1): 40914
      Immunotherapy has opened new avenues of treatment for patients with advanced non-small cell lung cancer (NSCLC) without previous hope of survival. Unfortunately, only a small percentage of patients benefit from it and there is not an effective biomarker to predict patients' response. Since T cells are key effectors of antitumor immunity, T cell receptor (TCR) has emerged as a potential predictive biomarker. Here, we evaluated the potential of baseline TCR repertoire, as a predictive biomarker in advanced NSCLC patients treated with pembrolizumab at first-line. After obtaining peripheral blood and tissue samples at baseline, next-generation sequencing targeting TCRβ/γ was performed. We found an uneven tumor-infiltrating TCRβ repertoire, and the use of various tumor-infiltrating and circulating TRBV/J genes were able of predicting the immunotherapy response. Our results support the potential of evaluating tissue and circulating TCRβ repertoire prior to pembrolizumab, revealing it as a promising immunotherapy response biomarker in NSCLC patients.
    Keywords:  Immunotherapy response; Non-small cell lung cancer; Pembrolizumab; Predictive biomarker; T cell receptor; TCR repertoire
    DOI:  https://doi.org/10.1038/s41598-025-21612-3
  9. Clin Cancer Res. 2025 Nov 21.
       PURPOSE: Exhaustion represents a collection of programmed T cell differentiation states and an important mode of T cell dysfunction. T cell progression from progenitor to terminal exhaustion is associated with upregulation of the transcription factor TOX. Our understanding of factors regulating TOX expression and the transition from progenitor to terminal exhaustion, however, remains incomplete.
    EXPERIMENTAL DESIGN: Single-cell RNA sequencing was used to evaluate expression of TNF receptors on human and murine tumor-infiltrating CD8+ T cells. Flow cytometry was utilized to assess exhaustion markers and TNF receptors on CD8+ T cells. Bulk RNA sequencing was used to demonstrate the role of TNFR2 on the overall exhaustion profile. Finally, the effect of TNFR2 on the overall anti-tumor response was established using TNFR2 KO mice and an antagonist.
    RESULTS: We reveal that upregulation of TNFR2 coincides with the gain of phenotypic markers and functions reflective of terminal exhaustion. Loss of TNFR2 affords a novel population of T cells that express TIM3 but possess diminished TOX levels and contain functional characteristics of both progenitor and terminally exhausted cells. TIM3+ TNFR2 KO T cells exhibit reduced exhaustion transcriptional programs and enhanced AP1 pathway signatures. Finally, TNFR2 KO mice demonstrate improved T cell-dependent control of tumor and chronic lymphocytic choriomeningitis (cLCMV) infection, while pharmacologic antagonism of TNFR2 licenses responses to checkpoint blockade in multiple subcutaneous and intracranial tumor models.
    CONCLUSIONS: Our data place TNFR2 signaling as a potential upstream regulator of TOX expression in T cells and propose TNFR2 antagonism as a novel immunotherapeutic strategy.
    DOI:  https://doi.org/10.1158/1078-0432.CCR-24-3455
  10. bioRxiv. 2025 Oct 05. pii: 2025.10.03.680201. [Epub ahead of print]
      Methionine is an essential amino acid critical for T cell activation. While methionine restriction (MR) combined with immune checkpoint blockade has been shown to enhance T cell function, the impact of methionine on adoptive T cell therapies is largely unexplored. Here, we examined the functionality of T cells under MR and pharmaceutical inhibition of the methionine cycle (MAT2Ai), using primary T cells and a murine adoptive T cell therapy model. In vitro , transient MR or MAT2Ai treatment increased interferon gamma (IFNγ) expression in CD8 + T cells, whereas sustained MR led to the upregulation of T cell exhaustion-associated markers. Mechanistically, transient MR suppressed the polyamine synthesis pathway, and supplementation with polyamines reversed MR-induced IFNγ expression. Genetic ablation of s-adenosylmethionine decarboxylase, an enzyme in the polyamine synthesis pathway, recapitulated the effect of MR, indicating that transient MR enhances T cell function by inhibiting polyamine synthesis. Despite this, transient MR treatment of ovalbumin (OVA)-specific (OT-I) CD8 + T cells prior to adoptive transfer did not improve antitumor efficacy against EG7-OVA tumors in vivo . In contrast, sustained dietary MR accelerated EG7-OVA tumor growth in mice treated with OT-I T cells, demonstrating that methionine availability is essential for the activity of adoptively transferred T cells. These findings suggest that enhancing methionine availability in the tumor microenvironment may improve the efficacy of adoptive T cell therapies.
    DOI:  https://doi.org/10.1101/2025.10.03.680201
  11. Int J Surg Pathol. 2025 Nov 17. 10668969251382887
      The microsatellite stable with epithelial to mesenchymal transition (MSS/EMT) subtype includes some of the most aggressive gastric tumors with the worst prognosis. The purpose of this study is to characterize the MSS/EMT subtype by investigating potential associations with clinicopathologic features and specific elements of the tumor microenvironment: tumor-infiltrating lymphocytes and tumor budding. From a retrospectively collected bi-centric cohort, we first selected microsatellite stable samples and then assessed the immunohistochemical expression of E-cadherin, β-catenin, and zinc finger E-box binding homeobox 1 to identify the epithelial to mesenchymal transition. Our findings revealed that the MSS/EMT subtype represented 43% of our series, which was a high prevalence compared to previous studies. This subtype included patients who were significantly younger than non-MSS/EMT patients (p = .017). It was associated with diffuse and mixed-type histology (p = .047) and with nonconventional carcinomas (p = .023). The MSS/EMT did not correlate with the advanced stage. Regarding the tumor microenvironment elements, the MSS/EMT subtype was associated with low density of tumor-infiltrating lymphocytes (p < 10-3), while no association was found with tumor budding score. In conclusion, the highly prevalent and distinct MSS/EMT gastric cancer subtype features low tumor-infiltrating lymphocyte density, suggesting an immunosuppressive tumor microenvironment and therapy resistance.
    Keywords:  epithelial to mesenchymal transition; immunohistochemistry; molecular classification; stomach neoplasm
    DOI:  https://doi.org/10.1177/10668969251382887
  12. Cancer Immunol Immunother. 2025 Nov 18. 74(12): 378
       BACKGROUND: Melanoma is one of the deadliest forms of skin cancer. Irreversible electroporation (IRE) is an innovative, non-thermal ablation technology for treating irresectable solid cancers. However, most IRE treatments are incapable of cancer eradication and only temporarily prolong patient survival.
    METHODS: In this study, we developed a novel IRE + Combo treatment regimen that combines IRE-ablation with Combo-adjuvant [CpG, anti-PD-L1 antibody (PD-L1-Ab) and CD40-agonist] and investigated its anti-tumor immunity in a mouse BL6-10OVA (BLOVA) melanoma model.
    RESULTS: We demonstrated that inclusion of the CD40-agonist in the IRE + Combo treatment regimen promoted a more robust CD8+ T cell response (6.89%) when compared with IRE + CpG/PD-L1-Ab (2.67%) or IRE alone (0.21%) treatments, leading to eradication of subcutaneous BLOVA melanoma in 5/8 of BLOVA-bearing mice and simultaneous elimination of lung melanoma metastases. Addition of CD40-agonist to the IRE + Combo treatment regimen also induced a higher frequency (17.1%) of CD8+CD103+ conventional type-1 dendritic cells (cDC1s) with up-regulated expression of CD54, CD80, MHC II, Bcl-xL and 41BBL in tumor-drainage lymph nodes (TDLNs) relative to the control IRE + CpG/PD-L1-Ab (12.1%) and IRE alone (9.0%) treatment groups. We also show that CD40-agonist stimulated a higher frequency of CD103+TCF1+ tissue-resident memory T (TRM) cells (32.1%) in TDLNs when compared with the two control (15.3% and 6.7%) treatment groups, and that these TRM cells exhibited enhanced mitochondrial content and greater relative expression of the effector cytokines IFN-γ and TNF-α and the transcriptional regulators TRAF1, p38-MAPK and PGC-1α.
    CONCLUSION: Taken together, this study establishes that the CD40-agonist greatly potentiates the efficacy of IRE-ablation for metastatic melanoma by promoting unexpected CD8+CD103+ cDC1 and CD103+TCF1+ TRM cell responses and suggests the importance of targeting CD40-signaling to improve the efficacy of cancer IRE-ablation therapy.
    Keywords:  CD103+ TRM cell; CD40-agonist; CD8+CD103+ cDC1; IRE-ablation; Metastatic melanoma; PD-1 blockade
    DOI:  https://doi.org/10.1007/s00262-025-04217-7