Journal for Immunotherapy of Cancer最新文献

Background: Colorectal cancer (CRC) progression is shaped by the tumor microenvironment, particularly tumor-associated macrophages (TAMs), which often adopt immunosuppressive functions. CD300e, a myeloid receptor involved in immune regulation, has an uncharacterized role in CRC.

Methods: Functional studies were conducted in azoxymethane/dextran sodium sulfate and MC38 murine CRC models using CD300e systemic and myeloid-specific CD300e knockout mice, and adoptive transfer experiments assessed macrophage-intrinsic effects. Human studies included analysis of CD300e expression in matched tumor and normal tissue from patients with CRC and in vitro co-culture of patient-derived colon tumor organoids with monocytes to study CD300e induction and TAM polarization.

Results: In vivo, CD300e deficiency led to reduced tumor burden, enhanced major histocompatibility complex expression on TAMs, and improved T-cell responses. CD300e-deficient macrophages exhibited increased phagocytic activity, antigen presentation, and support for T-cell proliferation and cytotoxicity. Adoptive transfer confirmed that macrophage-intrinsic CD300e expression is sufficient to suppress T-cell function and promote tumor growth. In patients with CRC, CD300e is selectively upregulated in tumor-infiltrating monocytes and macrophages, driving a suppressive phenotype marked by impaired antigen presentation. Tumor-derived signals in vitro induce CD300e expression and promote a protumorigenic macrophage profile.

Conclusions: Our findings identify CD300e as a critical regulator of macrophage-mediated immune suppression in CRC and a potential target for reprogramming TAMs to enhance immunotherapy.

Background: Neutralization of interferon (IFN)-γ abrogates the efficacy of anti-programmed death-ligand 1 (PD-(L)1) checkpoint inhibitors. Most epithelial cells do not constitutively express major histocompatibility complex (MHC) class II but can be induced to do so by IFN-γ. Inducible tumor-specific MHC class II (tsMHC-II) underlies responsiveness to anti-PD-(L)1. Retrospective studies show that tsMHC-II positivity associates with improved outcomes in patients treated with anti-PD-(L)1. The ANICCA-Class II single-arm Bayesian phase II trial prospectively explored whether positive tsMHC-II status could be a useful selection marker for anti-programmed cell death protein-1 (PD-1) in proficient mismatch repair colorectal cancer (pMMR CRC). In parallel, we retrospectively evaluated the potential predictive power of immunoscore-immune checkpoint (IS-IC) for outcome with single-agent immune checkpoint blockade.

Methods: Patients with histologically confirmed locally advanced/metastatic pMMR CRC with >1% MHC class II expression, Eastern Cooperative Oncology Group performance status 0-2, aged ≥18 years were eligible. Participants received 480 mg nivolumab every 28 days for up to 24 cycles. The primary outcome was durable clinical benefit (DCB) defined as participants remaining progression-free at their third trial-specific scan since treatment start (ie, at approximately 27 weeks). Secondary outcomes included progression-free survival time (PFS) and overall survival time (OS).

Results: 35 participants were treated: 65.7% of participants' cancers were tsMHC-II ≥5%. 3/35 patients achieved DCB (8.6%), estimating the true DCB rate (R) of 11% (95% credible interval 3% to 22%) with 0.002 probability that the true DCBR was >30%, below the required 0.5 to warrant further research. The higher tsMHC-II cut-point ≥5% was not more useful in predicting duration of disease stabilization. All three participants who achieved DCB had no evidence of liver metastases (LM); DCBR 23.1% in those without versus 0% in those with LM. PFS and OS were significantly greater in those without LM. There was no evidence that IS-IC high predicted for prolonged time on treatment or improved tumor growth inhibition.

Conclusions: In pMMR CRC, tsMHC-II positivity fails to identify a subset of patients with metastatic pMMR CRC obtaining potentially meaningful benefit from single-agent anti-PD-1. Although numbers are limited, there is no clear evidence that IS-IC is predictive of outcome with single-agent anti-PD-1. The poor outcome in those with LM underscores the need for therapies that overcome the systemic immunosuppression driven by LM.

Background: The effectiveness of immunotherapies against glioblastoma (GB) remains limited. A major obstacle in advancing new strategies is the reliance on non-autologous systems, which do not accurately mimic the true extent of inter-patient heterogeneity in both immune responses and tumor susceptibility. This often leads to misleading conclusions about therapeutic efficacy and targetability.

Methods: In this study, we addressed this critical gap by employing a fully autologous model. We phenotypically characterized primary αβ and γδT cells from the peripheral blood and tumors of 40 brain tumor patients, including 36 with confirmed GB, and expanded and functionally assessed the autologous anti-GB reactivity in a subset of patients.

Results: Notably, only Vδ2⁺ and Vδ2^- γδT cells, but not αβT cells, recognized autologous tumors. While Vδ2^- γδT cells showed activity in a subset of patients, Vδ2⁺ γδTILs from all patients responded to autologous GB cells in the presence of pamidronate. In patients, a higher percentage of Vδ2⁺ γδTILs was associated with longer overall survival. However, the potency of Vδ2⁺ γδTILs varied markedly between individuals, highlighting substantial inter-patient heterogeneity in γδT cell-mediated tumor recognition. This variability was driven by differences in both immune cell-intrinsic features and tumor-intrinsic factors, including expression of BTN2A1 and especially BTN3A, the ligands of the Vδ2⁺ γδTCR. Functional assays revealed that anti-GB reactivity was further modulated by stimulatory and inhibitory co-receptors such as NKG2D, CD94, and TIGIT. Transcriptomic analysis linked Vδ2⁺ γδT cell reactivity to extracellular matrix (ECM) pathways and disrupting ECM components such as LAMA5 and TGFB1 enhanced T cell responses. Knockout of ITGA3, a LAMA5 receptor, increased BTN2A1 and BTN3A expression on GB cells, improving immune recognition.

Conclusions: This study demonstrates that inter-patient heterogeneity in Vδ2⁺ γδTIL responses to GB is driven by the extracellular matrix-BTN3A axis. Autologous systems effectively capture this heterogeneity, offering a reliable platform to identify determinants of both immune function and tumor vulnerability, insights that are essential for the rational design of γδTIL-based immunotherapies.

Background: Adoptive transfer of T-cell receptor-engineered T cells (TCR-T cells) has shown promising efficacy in solid tumor treatment, but achieving clinical benefit typically requires infusion of tens of billions of cells. The commonly used rapid expansion protocol (REP), based on the CD3-agonistic OKT3 antibody and irradiated allogeneic feeder cells, exponentially expands tumor-infiltrating lymphocytes (TILs). However, the effect of REP on TCR-T cell frequency and phenotype remains unclear. This study aimed to evaluate the impact of REP on TCR-T cells and to assess the potential of a neoantigen-specific stimulation platform, NeoExpand, as a strategy to selectively expand TCR-T cells with favorable phenotypes.

Methods: We compared the effects of REP and NeoExpand on the frequency, yield, and phenotype of TCR-T cells engineered against shared neoantigens. Various autologous antigen-presenting cell (APC) types, including dendritic cells and bulk peripheral blood mononuclear cells (PBMCs), were tested as stimulators in NeoExpand. Additionally, we combined NeoExpand with CD3 activation, with or without allogeneic feeders, to improve scalability. The role of exogenous interleukin (IL)-21 in shaping TCR-T cell phenotypes was also investigated.

Results: REP impaired the frequency and phenotype of TCR-T cells, particularly CD4⁺ and CD8⁺ subsets matched to their engineered TCRs. In contrast, NeoExpand selectively increased TCR-T cell frequency and improved their phenotypes but did not consistently achieve higher total yields compared with REP. Among the tested autologous APCs, PBMCs effectively supported selective expansion. Further optimization using PBMCs as APCs revealed that combining NeoExpand with CD3 activation enabled exponential expansion without compromising selectivity. Inclusion of IL-21 during NeoExpand promoted a naïve/stem-like phenotype in CD8⁺ TCR-T cells, with minimal effect on CD4⁺ TCR-T cells.

Conclusions: These findings demonstrate that NeoExpand enables selective expansion of TCR-T cells while preserving favorable phenotypes and scalability. The approach supports further clinical evaluation of NeoExpand as a strategy to generate high-quality TCR-T cell products for adoptive cell therapy.

Background: High-risk neuroblastoma (HR-NBL) is an aggressive tumor of the sympathetic nervous system with high risk of relapse and poor overall survival. Allogeneic hematopoietic cell transplant (allo-HCT) has been used previously in patients with HR-NBL; however, graft-versus-host disease (GVHD) and disease progression have limited clinical application. Ex vivo stimulated allogeneic natural killer (NK) cells represent an approach to enhance the graft-versus-tumor (GVT) effect without exacerbation of GVHD but have not shown efficacy in NBL.

Methods: Ex vivo stimulated NK cells from C57BL/6NCr (B6) mice were expanded with soluble interleukin-15 (IL-15) and IL-15 receptor alpha (IL-15Rα) alone or with irradiated CD137L/CD54⁺ aggressive variant of the Neuro-2a murine neuroblastoma cell line (15-4P) at a 1:1 ratio for 10-12 days. Allogeneic NK cells were then analyzed for activation, proliferation, cytokine production, and cytotoxicity against two murine NBL cell lines, Neuro2a and NXS2, in the absence or presence of anti-T-cell immunoglobulin and mucin-domain containing-3 (TIM-3). Lethally irradiated B6AJF1 mice received allo-HCT from B6 donors followed by NBL challenge after 7 days to mimic tumor relapse. Select groups received anti-TIM-3 starting on day 9 for every 4 days with/without infusions of 15-4P B6 NK cells on days 14, 21, and 28. In select experiments, T cell and NK cells were selectively depleted to establish contribution to the GVT effect. All groups were analyzed for tumor growth, GVHD and survival.

Results: Co-culturing NK cells with 15-4P results in 78-fold expansion with increased expression of Kiel-67 (Ki-67) and Natural Killer Group 2, Member D (NKG2D), NKp46, TNF-Related Apoptosis-Inducing Ligand (TRAIL) and TIM-3. 15-4P stimulated allogeneic NK cells showed enhanced cytotoxicity against NBL compared with IL-15 NK cells alone but was limited in part due to high expression of TIM-3 ligands on Neuro-2a compared with NXS2. The addition of TIM-3 blockade further enhanced NK cytotoxicity versus Neuro-2a, with enhanced 15-4P NK cell degranulation, Eomesodermin, TRAIL and Fas Ligand expression observed. In vivo, the combination of 15-4P stimulated allogeneic NK cells and TIM-3 blockade after allo-HCT resulted in prolonged survival against NBL with decreased tumor burden compared with NK cells or anti-TIM-3 alone. Depletion of NK cells, but not T cells, abrogated the GVT effect.

Conclusion: Allo-HCT can be a platform for treating NBL using combination ex vivo stimulated allogeneic NK cell therapy with TIM-3 blockade to enhance the GVT effect without inducing GVHD.

Background: Bone metastasis (BM) drives therapeutic resistance and mortality in nasopharyngeal carcinoma (NPC). Tumor metabolites are crucial for NPC metastasis; however, the mechanisms by which these metabolites synergistically alter the immune microenvironment to promote BM remain unclear.

Methods: In this study, limited immune infiltration was observed in the NPC BM tumor microenvironment. Multiomics analysis has identified sphingosine kinase 1 (SPHK1) as a pivotal mediator driving BM and immune evasion in NPC, orchestrating the production of 1-phosphorylated sphingosine (S1P), which is critical for NPC pathogenesis.

Results: The aberrant buildup of lipid metabolites, along with immune microenvironment shifts, serves as a critical driver of NPC BM. Mechanistically, S1P enhanced osteoclast recruitment via S1PR3 binding and activated the Hippo pathway, worsening bone colonization and facilitating immune evasion by expanding the exhausted CD8⁺ T cell population.

Conclusions: The synergy between the SPHK1 inhibitor PF543 and anti-programmed cell death protein 1 therapy amplified treatment effectiveness beyond standalone approaches. Overall, the SPHK1/S1P pathway advances NPC growth and aids in suppressing immune defense. Regulation of lipid metabolism may be a therapeutic target against BM in NPC and may improve the effectiveness of immunotherapy.

Background: Intratumoral vaccines offer a promising avenue in cancer immunotherapy by harnessing the tumor microenvironment to stimulate immune responses. However, challenges persist in maximizing their effectiveness and addressing immune suppression within tumors.

Methods: Conventional in situ vaccine (CisVac) and α-CD137 antibody were administered in implanted mouse models of lung and colon cancer to evaluate therapeutic efficacy. The initiation mechanism of CD8⁺ T cells was determined using antibody blockade and transgenic mouse models. Single-cell RNA sequencing was used to further characterize the activation mechanism of T cells.

Results: Our findings indicate that this synergistic approach markedly inhibits tumor growth while eliciting a robust antitumor immune response, characterized by heightened activation and cytotoxic differentiation of CD8⁺ T cells, as well as the polarization of conventional dendritic cells (cDC1). Mechanistically, it promotes the cDC1-dependent proportion and cytokine production of tumor antigen-specific CD8⁺ T cells, mobilizes and establishes enduring systemic immune memory. Our single-cell transcriptome analyses revealed that the therapy facilitates a functional remodeling of regulatory T cells (Tregs) with upregulated inflammatory genes, potentially attenuating immune suppression. Cell-cell communication analyses highlighted interactions between CD4⁺ Th1-like/Th17 cells and monocytes/DCs through the CD40L-CD40 pathway, indicating potential intercellular regulatory mechanisms.

Conclusions: Our results highlight the transformative potential of the CisVac/α-CD137 combination in cancer immunotherapy, paving the way for further exploration of its clinical utility and long-term efficacy.

Background: Stimulator of interferon genes (STING) agonist drugs can induce expression of interferon stimulated genes (ISGs) and proinflammatory cytokine production aimed to enhance antitumor immunity. The purpose of the current study was to determine the safety, pharmacokinetic, and systemic and intratumoral pharmacodynamic properties of a novel, intravenously delivered STING agonist in client-owned dogs with cancer.

Methods: GSK856, a small-molecule dimeric amidobenzimidazole STING agonist, was administered intravenously to dogs with naturally developing tumors. Patients received two doses of GSK856 1 week apart, followed by definitive-intent surgical tumor removal.

Results: 19 dogs diagnosed with various solid tumor types, including malignant melanoma (oral mucosa, n=9; digit, n=1; conjunctiva, n=1), soft tissue sarcoma (5), rhabdomyosarcoma (1), oral fibrosarcoma (1), and mammary squamous cell carcinoma (1), were enrolled. Systemic pharmacokinetic analysis revealed rapid plasma clearance of GSK856 within 30 min of bolus administration. Clinical adverse events of fever, lethargy, and nausea were transient. Concurrent elevation in serum cytokines, including interleukin-6, was consistent with cytokine release syndrome following activation of the STING pathway. Transcriptional analyses of pretreatment and post-treatment blood and tumor tissue revealed robust induction of ISGs.

Conclusions: These data identify tolerated dose levels for a novel, intravenously delivered STING agonist compound that results in on-target effects in systemic and intratumoral immune responses in dogs with solid tumors.

Background: Tumor-associated macrophages (TAMs) abundantly infiltrate tumors and possess potent antitumor capabilities. "Don't eat me" signals like CD47 allow tumors to evade macrophages and proliferate unchecked. CD47 is upregulated in many tumors and interacts with the SIRPα expressed on macrophages to restrict effector function. Similarly, CD24 interacts with the Siglec-10 on TAMs to inhibit engulfment. Despite their potential, there is still a lack of effective therapeutics targeting macrophages. Recent clinical trials targeting CD47 have demonstrated limited efficacy and significant side effects in solid tumors, primarily due to the expression of CD47 on healthy cells such as red blood cells (RBCs). We therefore developed novel anti-CD47 variable domain of heavy chain of heavy-chain antibodies (vHHs) with strong ligand-blocking activity while demonstrating minimal binding to RBCs and incorporated these vHHs to generate an anti-CD47/CD24 bispecific antibody that preserves Fc-effector function and achieves improved tumor targeting while maintaining the blockade of antiphagocytic signals elicited by both CD47 and CD24.

Methods: Yeast display was employed to generate vHHs targeting CD47 and fully human monoclonal antibodies against CD24, respectively. The antigen binding epitopes of the vHHs to CD47 were predicted using AlphaFold3. Bispecific antibodies were designed, constructed, and characterized in vitro. Antitumor efficacy was evaluated in a human immune cell reconstitution mouse model, while safety was evaluated using a humanized syngeneic mouse model. Furthermore, the underlying mechanisms and alterations in tumor microenvironment were explored ex vivo.

Results: VHHs targeting CD47 and a fully human antibody against CD24 were identified, all exhibiting potent ligand-blocking activity. The bispecific antibody BiAb-103C, engineered on a human IgG1 scaffold, had strong binding to CD47⁺CD24⁺ tumor cells and could effectively inhibit the CD47-SIRPα interaction. Fc-effector activity was observed towards CD24 (but not CD47) single-positive cells to promote phagocytosis and antibody-dependent cellular cytotoxicity of CD47⁺CD24⁺ tumor cells. In mice, antibody candidates demonstrated notable antitumor activity alongside favorable safety observations.

Conclusions: Our study presents the discovery of an anti-CD47/CD24 bispecific antibody that offers a promising therapeutic strategy to address the challenges associated with both the efficacy and safety of CD47-targeting agents, offers insight into macrophage-driven cancer immunotherapy, and could potentially provide a therapeutic option for patients non-responsive to immunotherapy.

Background: Immune checkpoint inhibitors (ICI) have improved survival in various cancers, but their success in breast cancer, specifically triple-negative breast cancer, remains limited, benefiting less than 10% of patients.

Methods: We modeled ICI response in vivo to unravel the mechanisms underlying immunotherapy efficacy, identify mechanisms of resistance in non-responsive tumors, and ascertain the therapeutic benefits of different chemotherapeutic combinations with ICI in breast cancer. We investigated the impact of ICI as monotherapy and in combination with other therapeutics in mouse models of mammary cancer, which we found robustly suppressed primary tumor growth and extended survival.

Results: Interestingly, even within a single model, responses to ICI were highly variable. Resistance was not reliably retained by transplantation into syngeneic hosts, suggesting a role for systemic host immunity rather than tumor-autonomous mechanisms. Transcriptomic analysis of the primary tumor landscape by fine-needle aspiration revealed that upregulated cytotoxic T-cell response and inflammatory interferon signaling (both at baseline and post anti-programmed death-ligand 1 administration) corresponded to favorable response to ICI. Longitudinal analysis of the peripheral blood uncovered enhanced myeloid cell recruitment in resistant mice, prior to therapy initiation. Similar effects were observed through longitudinal assessment of peripheral blood in patients with ICI-treated human breast cancer. Blocking myeloid cell recruitment with navaraxin (CXCR1/2 inhibitor) improved ICI responses, further suppressing tumor growth and improving survival.

Conclusions: These findings provide insight into resistance mechanisms and suggest the potential for minimally invasive strategies (sampling of systemic immune cells from peripheral blood) to identify patients likely to respond to ICI. This approach may help inform de-escalation strategies to mitigate therapeutic toxicities and limit unnecessary treatments.