Journal for Immunotherapy of Cancer最新文献

Background: Clear cell renal cell carcinoma (ccRCC) is the most common histologic type of RCC. However, the spatial and functional heterogeneity of immunosuppressive cells and the mechanisms by which their interactions promote immunosuppression in the ccRCC have not been thoroughly investigated.

Methods: To further investigate the cellular and regional heterogeneity of ccRCC, we analyzed single-cell and spatial transcriptome RNA sequencing data from four patients, which were obtained from samples from multiple regions, including the tumor core, tumor-normal interface, and distal normal tissue. On the basis, the findings were investigated in vitro using tissue and blood samples from 15 patients with ccRCC and validated in the broader samples on tissue microarrays.

Results: In this study, we revealed previously unreported subsets of both stromal and immune cells, as well as mapped their spatial location at finer resolution. In addition, we validated the clusters of tumor cells after removing batch effects according to six characterized gene sets, including epithelial-mesenchymal transition^high clusters, metastatic clusters and proximal tubule^high clusters. Importantly, we identified a special regulatory T (Treg) cell subpopulation that has the molecular characteristics of terminal effector Treg cells but expresses multiple cytokines, such as interleukin (IL)-1β and IL-18. This group of Treg cells has stronger immunosuppressive function and was associated with a worse prognosis in ccRCC cohorts. They were colocalized with MRC1 ⁺ FOLR2 ⁺ tumor-associated macrophages (TAMs) at the tumor-normal interface to form a positive feedback loop, maintaining a synergistic procarcinogenic effect. In addition, we traced the origin of IL-1β⁺ Treg cells and revealed that IL-18 can induce the expression of IL-1β in Treg cells via the ERK/NF-κB pathway.

Conclusions: We demonstrated a novel cancer-promoting Treg cell subset and its interactions with MRC1 ⁺ FOLR2 ⁺TAMs, which provides new insight into Treg cell heterogeneity and potential therapeutic targets for ccRCC.

Introduction: Neoadjuvant chemoimmunotherapy has achieved overall survival (OS) benefit for patients with resectable non-small cell lung cancer (NSCLC). Here, we present outcomes after 3 years of follow-up from the first reported study of neoadjuvant atezolizumab+chemotherapy.

Methods: This open-label, multicenter single-arm investigator-initiated phase II study conducted at three US hospitals tested up to four cycles of atezolizumab, carboplatin, and nab-paclitaxel prior to surgery. Major pathological response (MPR, primary endpoint) was previously reported; here, we report 3-year disease-free survival (DFS), OS, and clinical characteristics of patients developing brain metastases (BM) with integrated data from tumor genomics, gene expression, and quantitative immunofluorescent measurement of immune markers.

Results: Of 30 enrolled patients, 29 were taken to the operating room. 26 underwent R0 resection, with 17 experiencing MPR (10 pCR). With a median follow-up of 39.5 months, the median OS was 55.8 months, and the median DFS was 34.5 months. Landmark OS at 36 months was 77%. Among 14 patients with recurrent disease, 6 patients had BM. Patients whose tumors had mutations in STK11 and KEAP1 did not have a significantly higher incidence of BM. Reduced copy number of STK11 and KEAP1, both residing on chromosome 19p, was observed in ~1/3 of tumors. Reduced CN of STK11 was significantly associated with worse pathological response and incidence of BM.

Conclusions: Consistent with recent phase III studies, 3-year OS data with neoadjuvant atezolizumab+chemotherapy was associated with prolonged PFS and OS. Establishing associations between STK11 and KEAP1 genomic alterations and key clinical outcomes in early-stage NSCLC requires further study.

Background: Although tumor necrosis factor receptor 2 (TNFR2) has been recognized as an attractive next-generation candidate target for cancer immunotherapy, the factors that regulate the gene expression and their mechanistic effects on tumor-infiltrating regulatory T cells (Treg cells) remain poorly understood.

Methods: Single-cell RNA sequencing analysis was employed to analyze the phenotypic and functional differences between TNFR2⁺ Treg cells and TNFR2^- Treg cells. Malignant pleural effusion (MPE) from humans and mouse was used to investigate the potential mechanisms by which lactate regulates TNFR2 expression.

Results: Treg cells with high TNFR2 expression exhibited elevated levels of immune checkpoint molecules. Additionally, the high expression of TNFR2 on Treg cells was positively correlated with a poor prognosis in MPE patients. Moreover, we revealed that lactate upregulated TNFR2 expression on Treg cells, thereby enhancing their immunosuppressive function in MPE. Mechanistically, lactate modulated the gene transcription of transcription factor nuclear factor-κB p65 (NF-κB p65) through histone H3K18 lactylation (H3K18la), subsequently upregulating the gene expression of TNFR2 and expediting the progression of MPE. Notably, lactate metabolism blockade combined with immune checkpoint blockade (ICB) therapy effectively enhanced the efficacy of ICB therapy, prolonged the survival time of MPE mice, and improved immunosuppression in the microenvironment of MPE.

Conclusions: The study explains the mechanism that regulates TNFR2 expression on Treg cells and its function in MPE progression, providing novel insights into the epigenetic regulation of tumor development and metabolic strategies for MPE treatment by targeting lactate metabolism in Treg cells.

Introduction: Despite significant successes, immune checkpoint blockade fails to achieve clinical responses in a significant proportion of patients, predictive markers for responses are imperfect and immune-related adverse events (irAEs) are unpredictable. We used T-cell receptor (TCR) sequencing to systematically analyze prospectively collected patient blood samples from a randomized clinical trial of dual immune checkpoint inhibitor therapy to evaluate changes in the T-cell repertoire and their association with response and irAEs.

Methods: Patients with immunotherapy-naïve metastatic non-small cell lung cancer (NSCLC) were treated with ipilimumab and nivolumab according to trial protocol (LONESTAR, NCT03391869). Blood samples were systematically obtained at baseline (n=107), after 12 weeks of ipilimumab and nivolumab (n=91), and at the time of grade ≥2 irAEs (n=77). For analysis of T-cell repertoire, we performed immunoSEQ to assess the complementary determining region 3β region of the TCR involved in antigen binding.

Results: A total of 250 samples from 119 patients were analyzed. Patients who had a response to therapy exhibited greater T-cell diversity at baseline. Interestingly, patients with irAEs demonstrated lower T-cell richness at the time of toxicity compared with those without irAEs.

Conclusion: Our study highlights the potential impact of peripheral blood T-cell repertoire on clinical response and toxicities from the combination of ipilimumab and nivolumab in patients with metastatic NSCLC. These findings suggest that analysis of peripheral blood T-cell repertoire may help to guide patient selection for treatment with ipilimumab and nivolumab.

Trial registration number: NCT03391869.

Background: The use of immune checkpoint inhibitors (CPIs) has become a dominant regimen in modern cancer therapy, however immune resistance induced by tumor-associated macrophages (TAMs) with immune suppressive and evasion properties limits responses. Therefore, the rational design of immune modulators that can control the immune suppressive properties of TAMs and polarize them, as well as dendritic cells (DCs), toward a more proinflammatory phenotype is a principal objective in cancer immunotherapy.

Methods: Here, using a protein engineering approach to enhance cytokine residence in the tumor microenvironment, we examined combined stimulation of the myeloid compartment via tumor stroma-binding granulocyte-macrophage colony-stimulating factor (GM-CSF) to enhance responses in both DCs and T cells via stroma-binding interleukin-12 (IL-12). We evaluated tumor responses at the levels of outcome, cellular responses, and cytokine responses in both the tumors and the tumor-draining lymph nodes. We further investigated the potentiation of DC response to IL-12 by GM-CSF stimulation ex vivo.

Results: Engineered GM-CSF restored an antitumorigenic tumor myeloid microenvironment otherwise suppressed by TAMs, while engineered IL-12 provided effector signals to T cells, thereby boosting both tumor-resident antitumor macrophage and CD8⁺ T cell populations. Furthermore, engineered GM-CSF potentiated DC response to IL-12, upregulating DC expression of IL-12 receptor and enhancing their expression of proinflammatory cytokines and chemokines on IL-12 exposure. This resulted in remarkable synergistic efficacy in multiple solid tumor models treated with the dual cytokine combination. The combination therapy also improved the efficacy of CPI in a CPI-resistant genetically-engineered melanoma model and exhibited synergistic antitumor efficacy in a pulmonary metastasis model.

Conclusion: Our strategy provides a rational design for combination immunotherapy targeting both myeloid and lymphoid compartments through complementary mechanisms.

Background: The immune-related adverse event (irAE), pneumonitis, is a potentially fatal complication of immune checkpoint inhibitors (ICIs). Preventing its progression is crucial, emphasizing the need for effective screening tests. We evaluated the feasibility of using Krebs von den Lungen-6 (KL-6), a marker for interstitial pneumonitis, as a screening tool for pneumonitis.

Methods: We examined 500 patients with cancer divided into two groups: those with cancer other than non-small cell lung cancer (NSCLC) (Group 1, n=382) and those with NSCLC (Group 2, n=118). KL-6 levels were monitored before and during ICI treatment and analyzed for their correlation with pneumonitis.

Results: In Group 1, 37 patients (9.7%) developed pneumonitis. KL-6 levels were significantly elevated at irAE onset (pre: 222.0 U/mL, post: 743.0 U/mL, p<0.0001). Receiver operating characteristic curve analysis showed an area under the curve (AUC) of 0.903 (sensitivity 81.1%, specificity 91.6%) with a cut-off value 1.52 times pre-KL-6 levels, indicating that KL-6 is a reliable biomarker for pneumonitis. In these patients, the KL-6 level increased regardless of pneumonitis severity and was significantly elevated in patients with both symptomatic (pre: 205.0 U/mL, post: 674.5 U/mL, p<0.0001) and asymptomatic pneumonitis (pre: 314.0 U/mL, post: 743.0 U/mL, p<0.0001) at irAE onset. After irAE treatment, KL-6 levels in steroid-responsive patients remained unchanged; however, steroid-unresponsive patients had a significant increase in KL-6 levels at 1 month (1078 U/mL, p=0.031) compared with at irAE onset (678.0 U/mL). In Group 2, 24 patients (20.3%) developed irAE pneumonitis, with KL-6 levels elevated (pre: 360.5 U/mL, post: 506.5 U/mL, p=0.029) and an AUC of 0.683, indicating that KL-6 was less reliable in patients with NSCLC.

Conclusions: KL-6 is a viable screening biomarker in ICI-induced pneumonitis, particularly in patients without NSCLC. In patients with NSCLC, the significance of KL-6 monitoring is limited as it is not effective for detecting ICI-induced pneumonitis; their treatment is typically managed by pulmonary specialists. Early detection through KL-6 monitoring facilitates timely intervention for ICI-induced pneumonitis, potentially preventing treatment interruptions and reducing the need for immunosuppressants.

Background: Granzyme B (GrB) is a key effector molecule, delivered by cytotoxic T lymphocytes and natural killer cells during immune surveillance to induce cell death. Fusion proteins and immunoconjugates represent an innovative therapeutic approach to specifically deliver a deadly payload to target cells. Epithelial membrane protein-2 (EMP2) is highly expressed in invasive breast cancer (BC), including triple-negative BC (TNBC), and represents an attractive therapeutic target.

Methods: We designed a novel fusion protein (GrB-Fc-KS49) composed of an active GrB fused to an anti-EMP2 single-chain antibody tethered through the immunoglobulin G heavy chain (Fc) domain. We assessed the construct's GrB enzymatic activity, anti-EMP2 binding affinity, and cytotoxicity against a panel of BC cells. The construct's pharmacokinetics (PK), toxicity profile, and in vivo efficacy were also evaluated.

Results: GrB-Fc-KS49 exhibited comparable GrB enzymatic activity to commercial GrB, as well as high affinity to an EMP2 peptide, with the dissociation constant in the picomolar range. The fusion protein rapidly internalized into EMP2+cancer cells and showed in vitro cytotoxicity to cell lines expressing surface EMP2, with half-maximal cytotoxicity (IC₅₀) values below 100 nM for most positive lines. Ex vivo stability at 37°C indicated a half-life exceeding 96 hours while in vivo PK indicated a biexponential plasma clearance, with a moderate initial clearance (t_1/2α=18.4 hours) and a much slower terminal clearance rate (t_1/2β=73.1 hours). No toxicity was measured in a Chem16 panel between the control and the GrB-Fc-KS49. In vivo, the GrB-Fc-KS49 showed efficacy against a TNBC syngeneic (4T1/_FLuc) mouse model, reducing tumor volume and cell proliferation and increasing cell death compared with controls. Treatment using an EMT6 mouse model confirmed these results. In addition to a significant impact on cell proliferation, GrB-Fc-KS49 treatment also resulted in a dramatic increase of tumor-infiltrating CD45+ cells and redistribution of tumor-associated macrophages. Transcriptomic analysis of tumors post-treatment confirmed the remodeling of the immune tumor microenvironment by the GrB-Fc-KS49 immunotoxin.

Conclusions: GrB-Fc-KS49 showed high specificity and cytotoxicity towards EMP2-positive cells. In vivo, it reduced tumor burden and increased the recruitment of immune cells into the tumor, suggesting that GrB-Fc-KS49 is a promising therapeutic candidate against BC.

Background: Chimeric antigen receptor (CAR)-T cell therapy has emerged as a transformative modality in the treatment of patients with cancer. However, it is increasingly evident that this therapeutic approach is not without its challenges. The unique nature of CAR-T cells as living drugs introduces a distinct set of side effects. As the application of CAR-T cell therapy expands to treat a broader range of diseases, it becomes increasingly important to devise effective strategies for handling the associated toxicities. Challenges in treating patients with CAR-T cells include addressing complications such as cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, and cytopenias. This comprehensive review seeks to systematically identify, categorize and elucidate all previously described neurological and psychological side effects associated with CAR-T cell therapy, shedding light on the pertinent laboratory findings that underscore these phenomena.

Methods: PubMed, Springer Link, and ScienceDirect were systematically searched for empirical studies on adult patients with cancer receiving CAR-T cell therapy for hemato-oncological malignancies. Quality assessment was conducted using Version 2 of the Cochrane risk-of-bias tool (RoB 2) for randomized trials and adherence to the STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) checklist for observational studies. The synthesis of findings was conducted via a narrative approach, consolidating the diverse array of data into a coherent framework.

Results: From an initial pool of 2,276 citations, 546 studies met the inclusion criteria, exhibiting a rich tapestry of heterogeneity in terms of study characteristics and patient samples. The incidence of neuropsychological symptoms varied notably across different CAR-T cell products and hematological malignancies. Among the most frequently reported neuropsychological symptoms were aphasia, attention deficits, impaired consciousness, and disorientation, alongside a constellation of other symptoms including confusion, cognitive impairment, memory loss, writing difficulties, fatigue, headache, agitation, tremor, seizures, and psychomotor retardation. Early intervention strategies, including corticosteroids and tocilizumab, have shown the potential to reduce the intensity of neuropsychological symptoms and prevent their progression to critical complications.

Conclusion: These insights underscore the imperative of extending neuropsychological assessments beyond the conventional Immune Effector Cell-Associated Encephalopathy score framework.

Background: The adoptive cell transfer (ACT) of T cell receptor (TCR)-engineered T cells targeting the HLA-A2-restricted epitope NY-ESO-1_157-165 (A2/NY) has yielded important clinical responses against several cancers. A variety of approaches are being taken to augment tumor control by ACT including TCR affinity-optimization and T-cell coengineering strategies to address the suppressive tumor microenvironment (TME). Most TCRs of clinical interest are evaluated in immunocompromised mice to enable human T-cell engraftment and do not recapitulate the dynamic interplay that occurs with endogenous immunity in a treated patient. A variety of humanized mouse models have been described but they have limitations in immune reconstitution and are technically challenging to implement. Here, we have developed a chimeric syngeneic tumor model in which A2Kb transgenic C57BL/6 mice are engrafted with B16 expressing A2Kb:NY as a single chain trimer (SCT) and treated by ACT with murine T cells expressing A2/NY TCRs comprising human variable fused to mouse constant regions.

Methods: We compared the function of a supraphysiological affinity A2/NY TCR (wtc51m), a computationally designed TCR in an optimal affinity range (DMβ), and a near non-binding TCR (V49I), engineered in both primary human and murine T cells by lentiviral and retroviral transduction, respectively. We evaluated a variety of strategies to stably express A2Kb:NY on the surface of mouse tumor cell lines including B16 melanoma, ultimately achieving success with an SCT comprising human β2m fused by GS linkers to both the NY-peptide and to α1 of the HLA complex. ACT studies were performed in B16-A2Kb:NY tumor-bearing, non-preconditioned immune-competent HLA-A*0201/H-2Kb (A2Kb) transgenic C57BL/6 mice and tumors characterized post-transfer.

Results: We observed significantly improved function of DMβ-T cells as well as superior infiltration and tumor control upon ACT as compared to the control TCR-T cells. Moreover, with our chimeric syngeneic tumor model, we were able to track dynamic and favorable changes in the TME upon DMβ-T cell transfer.

Conclusions: We have developed a robust, simple, and inexpensive preclinical strategy for evaluating human TCRs in the context of a fully competent murine immune system that can aid in the development of coengineered TCR-T cells and combination treatments translated to the clinic.