RESUMO
CTL recognition of non-mutated tumor-associated antigens (TAA), present on cancer cells but also in healthy tissues, is an important element of cancer immunity, but the mechanism of its selectivity for cancer cells and opportunities for its enhancement remain elusive. In this study, we found that CTL expression of the NK receptors (NKR) DNAM-1 and NKG2D was associated with the effector status of CD8+ tumor-infiltrating lymphocytes (TIL) and long-term survival of melanoma patients. Using MART-1 and NY-ESO-1 as model TAAs, we demonstrated that DNAM-1 and NKG2D regulate T-cell receptor (TCR) functional avidity and set the threshold for TCR activation of human TAA-specific CTLs. Superior costimulatory effects of DNAM-1 over CD28 involved enhanced TCR signaling, CTL killer function and polyfunctionality. Double transduction of human CTLs with TAA-specific TCR and NKRs resulted in strongly enhanced antigen sensitivity, without a reduction in the antigen specificity and selectivity of killer function. In addition, the elevation of NKR-Ligand expression on cancer cells by chemotherapy also increased CTL recognition of cancer cells expressing low levels of TAA. Our data help to explain the ability of self-antigens to mediate tumor rejection in the absence of autoimmunity and support the development of dual-targeting adoptive T cell therapies that use NKRs to enhance the potency and selectivity of recognition of TAA-expressing cancer cells.
RESUMO
We previously reported that dendritic cell (DC)-based vaccines targeting antigens expressed by tumor-associated vascular endothelial cells (VECs) and pericytes effectively control tumor growth in translational mouse tumor models. In the current report, we examined whether the therapeutic benefits of such tumor blood vessel antigen (TBVA)-targeted vaccines could be improved by the cotargeting of tumor antigens in the s.c. B16 melanoma model. We also evaluated whether combination vaccines incorporating anti-PD-L1 checkpoint blockade and/or a chemokine-modulating (CKM; IFNα + TLR3-L [rintatolimod] + Celecoxib) regimen would improve T cell infiltration/functionality in tumors yielding enhanced treatment benefits. We report that DC-peptide or DC-tumor lysate vaccines coordinately targeting melanoma antigens and TBVAs were effective in slowing B16 growth in vivo and extending survival, with superior outcomes observed for DC-peptide-based vaccines. Peptide-based vaccines that selectively target either melanoma antigens or TBVAs elicited a CD8+ T cell repertoire recognizing both tumor cells and tumor-associated VECs and pericytes in vitro, consistent with a treatment-induced epitope spreading mechanism. Notably, combination vaccines including anti-PD-L1 + CKM yielded superior therapeutic effects on tumor growth and animal survival, in association with the potentiation of polyfunctional CD8+ T cell reactivity against both tumor cells and tumor-associated vascular cells and a pro-inflammatory TME.
RESUMO
Background: Stimulator of Interferon Genes (STING) is a dsDNA sensor that triggers type I inflammatory responses. Recent data from our group and others support the therapeutic efficacy of STING agonists applied intratumorally or systemically in a range of murine tumor models, with treatment benefits associated with tumor vascular normalization and improved immune cell recruitment and function within the tumor microenvironment (TME). However, such interventions are rarely curative and STING agonism coordinately upregulates expression of immunoregulatory interferon-stimulated genes (ISGs) including Arg2, Cox2, Isg15, Nos2, and Pdl1 that may limit treatment benefits. We hypothesized that combined treatment of melanoma-bearing mice with STING agonist ADU-S100 together with antagonists of regulatory ISGs would result in improved control of tumor growth vs. treatment with ADU-S100 alone. Methods: Mice bearing either B16 (BRAFWTPTENWT) or BPR20 (BRAFV600EPTEN-/-) melanomas were treated with STING agonist ADU-S100 plus various inhibitors of ARG2, COX2, NOS2, PD-L1, or ISG15. Tumor growth control and changes in the TME were evaluated for combination treatment vs ADU-S100 monotherapy by tumor area measurements and flow cytometry/transcriptional profiling, respectively. Results: In the B16 melanoma model, we noted improved antitumor efficacy only when ADU-S100 was combined with neutralizing/blocking antibodies against PD-L1 or ISG15, but not inhibitors of ARG2, COX2, or NOS2. Conversely, in the BPR20 melanoma model, improved tumor growth control vs. ADU-S100 monotherapy was only observed when combining ADU-S100 with ARG2i, COX2i, and NOS2i, but not anti-PD-L1 or anti-ISG15. Immune changes in the TME associated with improved treatment outcomes were subtle but included increases in proinflammatory innate immune cells and activated CD8+CD69+ T cells and varied between the two tumor models. Conclusions: These data suggest contextual differences in the relative contributions of individual regulatory ISGs that serve to operationally limit the anti-tumor efficacy of STING agonists which should be considered in future design of novel combination protocols for optimal treatment benefit.
Assuntos
Antígeno B7-H1 , Melanoma Experimental , Camundongos , Animais , Proteínas Proto-Oncogênicas B-raf , Ciclo-Oxigenase 2 , Linhagem Celular Tumoral , Interferons , Microambiente TumoralRESUMO
CD28-driven "signal 2" is critical for naïve CD8+ T cell responses to dendritic cell (DC)-presented weak antigens, including non-mutated tumor-associated antigens (TAAs). However, it is unclear how DC-primed cytotoxic T lymphocytes (CTLs) respond to the same TAAs presented by cancer cells which lack CD28 ligands. Here, we show that NK receptors (NKRs) DNAM-1 and NKG2D replace CD28 during CTL re-activation by cancer cells presenting low levels of MHC I/TAA complexes, leading to enhanced proximal TCR signaling, immune synapse formation, CTL polyfunctionality, release of cytolytic granules and antigen-specific cancer cell killing. Double-transduction of T cells with recombinant TCR and NKR constructs or upregulation of NKR-ligand expression on cancer cells by chemotherapy enabled effective recognition and killing of poorly immunogenic tumor cells by CTLs. Operational synergy between TCR and NKRs in CTL recognition explains the ability of cancer-expressed self-antigens to serve as tumor rejection antigens, helping to develop more effective therapies.
RESUMO
Background: Proinflammatory chemokines/cytokines support development and maturation of tertiary lymphoid structures (TLS) within the tumor microenvironment (TME). In the current study, we sought to investigate the prognostic value of TLS-associated chemokines/cytokines (TLS-kines) expression levels in melanoma patients by performing serum protein and tissue transcriptomic analyses, and to then correlate these data with patients clinicopathological and TME characteristics. Methods: Levels of TLS-kines in patients' sera were quantitated using a custom Luminex Multiplex Assay. The Cancer Genomic Atlas melanoma cohort (TCGA-SKCM) and a Moffitt Melanoma cohort were used for tissue transcriptomic analyses. Associations between target analytes and survival outcomes, clinicopathological variables, and correlations between TLS-kines were statistically analyzed. Results: Serum of 95 patients with melanoma were evaluated; 48 (50%) female, median age of 63, IQR 51-70 years. Serum levels of APRIL/TNFSF13 were positively correlated with levels of both CXCL10 and CXCL13. In multivariate analyses, high levels of serum APRIL/TNFSF13 were associated with improved event-free survival after adjusting for age and stage (HR = 0.64, 95% CI 0.43-0.95; p = 0.03). High expression of APRIL/TNFSF13 tumor transcripts was significantly associated with improved OS in TCGA-SKCM (HR = 0.69, 95% CI 0.52-0.93; p = 0.01) and in Moffitt Melanoma patients (HR = 0.51, 95% CI: 0.32-0.82; p = 0.006). Further incorporation of CXCL13 and CXCL10 tumor transcript levels in a 3-gene index revealed that high APRIL/CXCL10/CXCL13 expression was associated with improved OS in the TCGA SKCM cohort (HR = 0.42, 95% CI 0.19-0.94; p = 0.035). Melanoma differentially expressed genes positively associated with high APRIL/CXCL10/CXCL13 tumor expression were linked to tumor infiltration by a diverse array of proinflammatory immune cell types. Conclusion: Serum protein and tumor transcript levels of APRIL/TNFSF13 are associated with improved survival outcomes. Patients exhibiting high coordinate expression of APRIL/CXCL10/CXCL13 transcripts in their tumors displayed superior OS. Further investigation of TLS-kine expression profiles related to clinical outcomes in larger cohort studies is warranted.
Assuntos
Melanoma , Humanos , Feminino , Pessoa de Meia-Idade , Idoso , Masculino , Prognóstico , Melanoma/genética , Citocinas , Perfilação da Expressão Gênica , Genômica , Microambiente Tumoral/genéticaRESUMO
We sought to develop a sentinel lymph node gene expression signature score predictive of disease recurrence in patients with cutaneous melanoma. Gene expression profiling was performed on SLN biopsies using U133A 2.0 Affymetrix gene chips. The top 25 genes associated with recurrence-free survival (RFS) were selected and a penalized regression function was used to select 12 genes with a non-zero coefficient. A proportional hazards regression model was used to evaluate the association between clinical covariates, gene signature score, and RFS. Among the 45 patients evaluated, 23 (51%) had a positive SLN. Twenty-one (46.7%) patients developed disease recurrence. For the top 25 differentially expressed genes (DEG), 12 non-zero penalized coefficients were estimated (CLGN, C1QTNF3, ADORA3, ARHGAP8, DCTN1, ASPSCR1, CHRFAM7A, ZNF223, PDE6G, CXCL3, HEXIM1, HLA-DRB). This 12-gene signature score was significantly associated with RFS (p < 0.0001) and produced a bootstrap C index of 0.888. In univariate analysis, Breslow thickness, presence of primary tumor ulceration, SLN positivity were each significantly associated with RFS. After simultaneously adjusting for these prognostic factors in relation to the gene signature, the 12-gene score remained a significant independent predictor for RFS (p < 0.0001). This SLN 12-gene signature risk score is associated with melanoma recurrence regardless of SLN status and may be used as a prognostic factor for RFS.
RESUMO
This collection contains 10 reports published In Frontiers in Oncology between August 2020 and March 2022 broadly focused on the immunobiology of renal cell carcinoma (RCC), the impact of immunotherapy in the setting of RCC, and the identification of biomarkers that are prognostic of RCC patient outcomes and response to immunotherapy.
RESUMO
Peripheral neurons comprise a critical component of the tumor microenvironment (TME). The role of the autonomic innervation in cancer has been firmly established. However, the effect of the afferent (sensory) neurons on tumor progression remains unclear. Utilizing surgical and chemical skin sensory denervation methods, we showed that afferent neurons supported the growth of melanoma tumors in vivo and demonstrated that sensory innervation limited the activation of effective antitumor immune responses. Specifically, sensory ablation led to improved leukocyte recruitment into tumors, with decreased presence of lymphoid and myeloid immunosuppressive cells and increased activation of T-effector cells within the TME. Cutaneous sensory nerves hindered the maturation of intratumoral high endothelial venules and limited the formation of mature tertiary lymphoid-like structures containing organized clusters of CD4+ T cells and B cells. Denervation further increased T-cell clonality and expanded the B-cell repertoire in the TME. Importantly, CD8a depletion prevented denervation-dependent antitumor effects. Finally, we observed that gene signatures of inflammation and the content of neuron-associated transcripts inversely correlated in human primary cutaneous melanomas, with the latter representing a negative prognostic marker of patient overall survival. Our results suggest that tumor-associated sensory neurons negatively regulate the development of protective antitumor immune responses within the TME, thereby defining a novel target for therapeutic intervention in the melanoma setting.
Assuntos
Melanoma , Neoplasias Cutâneas , Estruturas Linfoides Terciárias , Humanos , Imunidade , Microambiente TumoralRESUMO
Tumor-infiltrating lymphocytes (TILs) contain substantial numbers of CD4+ T cells mediating pro- and antitumor functions. While CD4+ Tregs are well characterized and known to promote tumor immune evasion, the fingerprint of CD4+ Th cells that recognizes tumor antigens and serves to restrict disease progression has remained poorly discriminated. In this issue of the JCI, Duhen et al. analyzed tumors from patients with head and neck squamous cell carcinoma or colon carcinoma and identified a unique programmed cell death 1-positive, ICOS1-positive (PD-1+ICOS1+) subpopulation of CD4+ TILs highly enriched for the ability to recognize tumor-associated antigens. These cells localized proximally to MHC II+ antigen-presenting cells and CD8+ T cells within tumors, where they appeared to proliferate and function almost exclusively as Th cells. These potentially therapeutic Th cells can be monitored for patient prognosis and are expected to have substantial utility in developing personalized adoptive cell- and vaccine-based immunotherapeutic approaches for improving patient outcomes.
Assuntos
Neoplasias de Cabeça e Pescoço , Microambiente Tumoral , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Linfócitos do Interstício Tumoral , PrognósticoRESUMO
BACKGROUND: Given equivocal results related to overall survival (OS) for patients with multiple primary melanomas (MPMs) compared with those with single primary melanomas (SPMs) in previous reports, the authors sought to determine whether OS differs between these 2 cohorts in their center using their UPCI-96-99 database. Secondary aims were to assess the differences in recurrence-free survival (RFS). In a subset of patients, transcriptomic profiling of peripheral blood mononuclear cells (PBMCs) was performed to assess disease-associated genes of interest. METHODS: This retrospective case-controlled study included patients with MPMs and age-, sex-, and stage-matched controls with SPMs at a 1:1 ratio. Cox regression models were used to evaluate the effect of the presence of MPMs on death and recurrence. NanoString PanCancer Immune Profiling was used to assess peripheral blood immune status in patients. RESULTS: In total, 320 patients were evaluated. The mean patient age was 47 years; 43.8% were male. Patients with MPMs had worse RFS and OS (P = .023 and P = .0019, respectively). The presence of MPMs was associated with an increased risk of death (hazard ratio [HR], 4.52, P = .0006), and increased risk of disease recurrence (HR, 2.17; P = .004) after adjusting for age, sex, and stage. The degree of tumor-infiltrating lymphocytes (TILs) was different between the first melanoma of MPMs and SPMs. Expression of CXCL6 and FOXJ1 was increased in PBMCs isolated from patients with MPMs. CONCLUSIONS: Patients with MPMs had worse RFS and OS compared with patients with SPMs. Immunologic differences were also observed, including TIL content and expression of CXCL6/FOXJ1 in PBMCs of patients with MPMs, which warrant further investigation.
Assuntos
Melanoma , Neoplasias Cutâneas , Feminino , Humanos , Linfócitos do Interstício Tumoral , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Prognóstico , Estudos RetrospectivosRESUMO
BACKGROUND: Melanoma of unknown primary (MUP) represents a poorly understood group of patients both clinically and immunologically. We investigated differences in prognosis and candidate immune biomarkers in patients with unknown compared with those with known primary melanoma enrolled in the E1609 adjuvant trial that tested ipilimumab at 3 and 10 mg/kg vs high-dose interferon-alfa (HDI). PATIENTS AND METHODS: MUP status was defined as initial presentation with cutaneous, nodal or distant metastasis without a known primary. Relapse-free survival (RFS) and overall survival (OS) rates were estimated by the Kaplan-Meier method. Stratified (by stage) log-rank test was used to compare RFS and OS by primary tumor status. Gene expression profiling (GEP) was performed on the tumor biopsies of a subset of patients. Similarly, peripheral blood samples were tested for candidate soluble and cellular immune biomarkers. RESULTS: MUP cases represented 12.8% of the total population (N=1699) including 11.7% on the ipilimumab arms and 14.7% on the HDI arm. Stratifying by stage, RFS (p=0.001) and overall survival (OS) (p=0.009) showed outcomes significantly better for patients with unknown primary. The primary tumor status remained prognostically significant after adjusting for treatment and stage in multivariate Cox proportional hazards models. Including only ipilimumab-treated patients, RFS (p=0.005) and OS (p=0.023) were significantly better in favor of those with unknown primary. Among patients with GEP data (n=718; 102 MUP, 616 known), GEP identified pathways and genes related to autoimmunity, inflammation, immune cell infiltration and immune activation that were significantly enriched in the MUP tumors compared with known primaries. Further investigation into infiltrating immune cell types estimated significant enrichment with CD8 +and CD4+T cells, B cells and NK cells as well as significantly higher major histocompatibility complex (MHC)-I and MHC-II scores in MUP compared with known primary. Among patients tested for circulating biomarkers (n=321; 66 unknown and 255 known), patients with MUP had significantly higher circulating levels of IL-2R (p=0.04). CONCLUSION: Patients with MUP and high-risk melanoma had significantly better prognosis and evidence of significantly enhanced immune activation within the TME and the circulation, supporting the designation of MUP as a distinct prognostic marker in patients with high-risk melanoma.
Assuntos
Melanoma/mortalidade , Neoplasias Primárias Desconhecidas/mortalidade , Neoplasias Cutâneas/mortalidade , Adolescente , Adulto , Criança , Perfilação da Expressão Gênica , Humanos , Melanoma/imunologia , Melanoma/patologia , Neoplasias Primárias Desconhecidas/imunologia , Neoplasias Primárias Desconhecidas/patologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Adulto JovemRESUMO
PURPOSE: Despite unprecedented responses to immune checkpoint inhibitors and targeted therapy in melanoma, a major subset of patients progresses and have few effective salvage options. We have previously demonstrated robust, selective uptake of the peptidomimetic LLP2A labeled with Cu-64 ([64Cu]-LLP2A) for positron emission tomography (PET) imaging in subcutaneous and metastatic models of B16F10 murine melanoma. LLP2A binds with high affinity to very late antigen-4 (VLA-4, integrin α4ß1), a transmembrane protein overexpressed in melanoma and other cancers that facilitates tumor growth and metastasis. Yet B16F10 fails to faithfully reflect human melanoma biology, as it lacks certain oncogenic driver mutations, including BRAF mutations found in ≥ 50 % of clinical specimens. Here, we evaluated the PET tracer [64Cu]-CB-TE1A1P-PEG4-LLP2A ([64Cu]-LLP2A) in novel, translational BRAFV600E mutant melanoma models differing in VLA-4 expression-BPR (VLA-4-) and BPRα (VLA-4+). PROCEDURES: BPR cells were transduced with α4 (CD49d) to overexpress intact cell surface VLA-4 (BPRα). The binding affinity of [64Cu]-LLP2A to BPR and BPRα cells was determined by saturation binding assays. [64Cu]-LLP2A internalization into B16F10, BPR, and BPRα cells was quantified via a plate-based assay. Tracer biodistribution and PET/CT imaging were evaluated in mice bearing subcutaneous BPR and BPRα tumors. RESULTS: [64Cu]-LLP2A demonstrated high binding affinity to BPRα (Kd = 1.4 nM) but indeterminate binding to BPR cells. VLA-4+ BPRα and B16F10 displayed comparable time-dependent [64Cu]-LLP2A internalization, whereas BPR internalization was undetectable. PET/CT showed increased tracer uptake in BPRα tumors vs. BPR tumors in vivo, which was validated by significantly greater (p < 0.0001) BPRα tumor uptake in biodistribution analyses. CONCLUSIONS: [64Cu]-LLP2A discriminates BPRα (VLA-4+) vs. BPR (VLA-4-) melanomas in vivo, supporting translation of these BRAF-mutated melanoma models via prospective imaging and theranostic studies. These results extend the utility of LLP2A to selectively target clinically relevant and therapy-resistant tumor variants toward its use for therapeutic patient care.
Assuntos
Integrina alfa4beta1 , Melanoma , Animais , Linhagem Celular Tumoral , Radioisótopos de Cobre , Modelos Animais de Doenças , Humanos , Integrina alfa4beta1/metabolismo , Melanoma/diagnóstico por imagem , Melanoma/genética , Camundongos , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Tomografia por Emissão de Pósitrons/métodos , Estudos Prospectivos , Proteínas Proto-Oncogênicas B-raf/genética , Distribuição TecidualRESUMO
Epigenetic mechanisms play vital roles not only in cancer initiation and progression, but also in the activation, differentiation and effector function(s) of immune cells. In this review, we summarize current literature related to epigenomic dynamics in immune cells impacting immune cell fate and functionality, and the immunogenicity of cancer cells. Some important immune-associated genes, such as granzyme B, IFN-γ, IL-2, IL-12, FoxP3 and STING, are regulated via epigenetic mechanisms in immune or/and cancer cells, as are immune checkpoint molecules (PD-1, CTLA-4, TIM-3, LAG-3, TIGIT) expressed by immune cells and tumor-associated stromal cells. Thus, therapeutic strategies implementing epigenetic modulating drugs are expected to significantly impact the tumor microenvironment (TME) by promoting transcriptional and metabolic reprogramming in local immune cell populations, resulting in inhibition of immunosuppressive cells (MDSCs and Treg) and the activation of anti-tumor T effector cells, professional antigen presenting cells (APC), as well as cancer cells which can serve as non-professional APC. In the latter instance, epigenetic modulating agents may coordinately promote tumor immunogenicity by inducing de novo expression of transcriptionally repressed tumor-associated antigens, increasing expression of neoantigens and MHC processing/presentation machinery, and activating tumor immunogenic cell death (ICD). ICD provides a rich source of immunogens for anti-tumor T cell cross-priming and sensitizing cancer cells to interventional immunotherapy. In this way, epigenetic modulators may be envisioned as effective components in combination immunotherapy approaches capable of mediating superior therapeutic efficacy.
Assuntos
Epigênese Genética , Imunidade , Imunomodulação/genética , Imunoterapia , Neoplasias/etiologia , Neoplasias/terapia , Animais , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Biomarcadores Tumorais , Ensaios Clínicos como Assunto , Terapia Combinada , Gerenciamento Clínico , Suscetibilidade a Doenças , Desenvolvimento de Medicamentos , Metabolismo Energético , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoterapia/métodos , Neoplasias/metabolismo , Transdução de Sinais/efeitos dos fármacos , Resultado do Tratamento , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
BACKGROUND: A first-in-human, randomized pilot phase II clinical trial combining vaccines targeting overexpressed, non-mutated tumor blood vessel antigens (TBVA) and tyrosine kinase inhibitor dasatinib was conducted in human leukocyte antigen (HLA)-A2+ patients with advanced melanoma. METHODS: Patient monocyte-derived type-1-polarized dendritic cells were loaded with HLA-A2-presented peptides derived from TBVA (DLK1, EphA2, HBB, NRP1, RGS5, TEM1) and injected intradermally as a vaccine into the upper extremities every other week. Patients were randomized into one of two treatment arms receiving oral dasatinib (70 mg two times per day) beginning in week 5 (Arm A) or in week 1 (Arm B). Trial endpoints included T cell response to vaccine peptides (interferon-γ enzyme-linked immunosorbent spot), objective clinical response (Response Evaluation Criteria in Solid Tumors V.1.1) and exploratory tumor, blood and serum profiling of immune-associated genes/proteins. RESULTS: Sixteen patients with advanced-stage cutaneous (n=10), mucosal (n=1) or uveal (n=5) melanoma were accrued, 15 of whom had previously progressed on programmed cell death protein 1 (PD-1) blockade. Of 13 evaluable patients, 6 patients developed specific peripheral blood T cell responses against ≥3 vaccine-associated peptides, with further evidence of epitope spreading. All six patients with specific CD8+ T cell response to vaccine-targeted antigens exhibited evidence of T cell receptor (TCR) convergence in association with preferred clinical outcomes (four partial response and two stabilization of disease (SD)). Seven patients failed to respond to vaccination (one SD and six progressive disease). Patients in Arm B (immediate dasatinib) outperformed those in Arm A (delayed dasatinib) for immune response rate (IRR; 66.7% vs 28.6%), objective response rate (ORR) (66.7% vs 0%), overall survival (median 15.45 vs 3.47 months; p=0.0086) and progression-free survival (median 7.87 vs 1.97 months; p=0.063). IRR (80% vs 25%) and ORR (60% vs 12.5%) was greater for females versus male patients. Tumors in patients exhibiting response to treatment displayed (1) evidence of innate and adaptive immune-mediated inflammation and TCR convergence at baseline, (2) on-treatment transcriptional changes associated with reduced hypoxia/acidosis/glycolysis, and (3) increased inflammatory immune cell infiltration and tertiary lymphoid structure neogenesis. CONCLUSIONS: Combined vaccination against TBVA plus dasatinib was safe and resulted in coordinating immunologic and/or objective clinical responses in 6/13 (46%) evaluable patients with melanoma, particularly those initiating treatment with both agents. TRIAL REGISTRATION NUMBER: NCT01876212.
Assuntos
Antígenos de Neoplasias/uso terapêutico , Antineoplásicos/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Dasatinibe/uso terapêutico , Células Dendríticas/metabolismo , Melanoma/tratamento farmacológico , Antineoplásicos/farmacologia , Vacinas Anticâncer/farmacologia , Dasatinibe/farmacologia , Feminino , Humanos , Masculino , Melanoma/patologia , Projetos Piloto , Estudos ProspectivosRESUMO
CD40 crosslinking plays an important role in regulating cell migration, adhesion and proliferation in renal cell carcinoma (RCC). CD40/CD40L interaction on RCC cells activates different intracellular pathways but the molecular mechanisms leading to cell scattering are not yet clearly defined. Aim of our study was to investigate the main intracellular pathways activated by CD40 ligation and their specific involvement in RCC cell migration. CD40 ligation increased the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun NH (2)-terminal kinase (JNK) and p38 MAPK. Furthermore, CD40 crosslinking activated different transcriptional factors on RCC cell lines: AP-1, NFkB and some members of the Nuclear Factor of Activated T cells (NFAT) family. Interestingly, the specific inhibition of NFAT factors by cyclosporine A, completely blocked RCC cell motility induced by CD40 ligation. In tumor tissue, we observed a higher expression of NFAT factors and in particular an increased activation and nuclear migration of NFATc4 on RCC tumor tissues belonging to patients that developed metastases when compared to those who did not. Moreover, CD40-CD40L interaction induced a cytoskeleton reorganization and increased the expression of integrin ß1 on RCC cell lines, and this effect was reversed by cyclosporine A and NFAT inhibition. These data suggest that CD40 ligation induces the activation of different intracellular signaling pathways, in particular the NFATs factors, that could represent a potential therapeutic target in the setting of patients with metastatic RCC.
Assuntos
Biomarcadores Tumorais/metabolismo , Antígenos CD40/metabolismo , Ligante de CD40/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Renais/patologia , Fatores de Transcrição NFATC/metabolismo , Idoso , Apoptose , Biomarcadores Tumorais/genética , Antígenos CD40/genética , Ligante de CD40/genética , Movimento Celular , Proliferação de Células , Reagentes de Ligações Cruzadas , Feminino , Humanos , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Masculino , Pessoa de Meia-Idade , Fatores de Transcrição NFATC/genética , Metástase Neoplásica , Prognóstico , Células Tumorais CultivadasRESUMO
The interrogation of intrinsic and adaptive resistance to cancer immunotherapy has identified lack of antigen presentation and type I interferon signaling as biomarkers of non-T-cell-inflamed tumors and clinical progression. A myriad of pre-clinical studies have implicated the cGAS/stimulator of interferon genes (STING) pathway, a cytosolic DNA-sensing pathway that drives activation of type I interferons and other inflammatory cytokines, in the host immune response against tumors. The STING pathway is also increasingly understood to have other anti-tumor functions such as modulation of the vasculature and augmentation of adaptive immunity via the support of tertiary lymphoid structure development. Many natural and synthetic STING agonists have entered clinical development with the first generation of intra-tumor delivered cyclic dinucleotides demonstrating safety but only modest systemic activity. The development of more potent and selective STING agonists as well as novel delivery systems that would allow for sustained inflammation in the tumor microenvironment could potentially augment response rates to current immunotherapy approaches and overcome acquired resistance. In this review, we will focus on the latest developments in STING-targeted therapies and provide an update on the clinical development and application of STING agonists administered alone, or in combination with immune checkpoint blockade or other approaches.
RESUMO
Regulator-of-G-protein-signaling-5 (RGS5), a pro-apoptotic/anti-proliferative protein, is a signature molecule of tumor-associated pericytes, highly expressed in several cancers, and is associated with tumor growth and poor prognosis. Surprisingly, despite the negative influence of intrinsic RGS5 expression on pericyte survival, RGS5highpericytes accumulate in progressively growing tumors. However, responsible factor(s) and altered-pathway(s) are yet to report. RGS5 binds with Gαi/q and promotes pericyte apoptosis in vitro, subsequently blocking GPCR-downstream PI3K-AKT signaling leading to Bcl2 downregulation and promotion of PUMA-p53-Bax-mediated mitochondrial damage. However, within tumor microenvironment (TME), TGFß appeared to limit the cytocidal action of RGS5 in tumor-residing RGS5highpericytes. We observed that in the presence of high RGS5 concentrations, TGFß-TGFßR interactions in the tumor-associated pericytes lead to the promotion of pSmad2-RGS5 binding and nuclear trafficking of RGS5, which coordinately suppressed RGS5-Gαi/q and pSmad2/3-Smad4 pairing. The RGS5-TGFß-pSmad2 axis thus mitigates both RGS5- and TGFß-dependent cellular apoptosis, resulting in sustained pericyte survival/expansion within the TME by rescuing PI3K-AKT signaling and preventing mitochondrial damage and caspase activation. This study reports a novel mechanism by which TGFß fortifies and promotes survival of tumor pericytes by switching pro- to anti-apoptotic RGS5 signaling in TME. Understanding this altered RGS5 signaling might prove beneficial in designing future cancer therapy.
Assuntos
Neoplasias/genética , Pericitos/metabolismo , Proteínas RGS/metabolismo , Proteína Smad2/metabolismo , Animais , Feminino , Humanos , Camundongos , Transdução de Sinais , TransfecçãoRESUMO
Tertiary lymphoid structures (TLS), also known as ectopic lymphoid structures (ELS) or tertiary lymphoid organs (TLO), represent a unique subset of lymphoid tissues noted for their architectural similarity to lymph nodes, but which conditionally form in peripheral tissues in a milieu of sustained inflammation. TLS serve as regional sites for induction and expansion of the host B and T cell repertoires via an operational paradigm involving mature dendritic cells (DC) and specialized endothelial cells (i.e. high endothelial venules; HEV) in a process directed by TLS-associated cytokines and chemokines. Recent clinical correlations have been reported for the presence of TLS within tumor biopsies with overall patient survival and responsiveness to interventional immunotherapy. Hence, therapeutic strategies to conditionally reinforce TLS formation within the tumor microenvironment (TME) via the targeting of DC, vascular endothelial cells (VEC) and local cytokine/chemokine profiles are actively being developed and tested in translational tumor models and early phase clinical trials. In this regard, a subset of agents that promote tumor vascular normalization (VN) have been observed to coordinately support the development of a pro-inflammatory TME, maturation of DC and VEC, local production of TLS-inducing cytokines and chemokines, and therapeutic TLS formation. This mini-review will focus on STING agonists, which were originally developed as anti-angiogenic agents, but which have recently been shown to be effective in promoting VN and TLS formation within the therapeutic TME. Future application of these drugs in combination immunotherapy approaches for greater therapeutic efficacy is further discussed.
Assuntos
Antineoplásicos/uso terapêutico , Proteínas de Membrana/agonistas , Neoplasias/tratamento farmacológico , Estruturas Linfoides Terciárias/imunologia , Microambiente Tumoral/imunologia , Animais , Citocinas/metabolismo , Humanos , Imunoterapia , Mediadores da Inflamação/metabolismo , Proteínas de Membrana/metabolismo , Neoplasias/imunologia , Neoplasias/metabolismo , Neoplasias/patologia , Transdução de Sinais , Estruturas Linfoides Terciárias/metabolismo , Estruturas Linfoides Terciárias/patologiaRESUMO
Recent advances in immunotherapy have enabled rapid evolution of novel interventional approaches designed to reinvigorate and expand patient immune responses against cancer. An emerging approach in cancer immunology involves the conditional induction of tertiary lymphoid structures (TLS), which are non-encapsulated ectopic lymphoid structures forming at sites of chronic, pathologic inflammation. Cutaneous melanoma (CM), a highly-immunogenic form of solid cancer, continues to rise in both incidence and mortality rate, with recent reports supporting a positive correlation between the presence of TLS in melanoma and beneficial treatment outcomes amongst advanced-stage patients. In this context, TLS in CM are postulated to serve as dynamic centers for the initiation of robust anti-tumor responses within affected regions of active disease. Given their potential importance to patient outcome, significant effort has been recently devoted to gaining a better understanding of TLS neogenesis and the influence these lymphoid organs exert within the tumor microenvironment. Here, we briefly review TLS structure, function, and response to treatment in the setting of CM. To uncover potential tumor-intrinsic mechanisms that regulate TLS formation, we have taken the novel perspective of evaluating TLS induction in melanomas impacted by common driver mutations in BRAF, PTEN, NRAS, KIT, PRDM1, and MITF. Through analysis of The Cancer Genome Atlas (TCGA), we show expression of DNA repair proteins (DRPs) including BRCA1, PAXIP, ERCC1, ERCC2, ERCC3, MSH2, and PMS2 to be negatively correlated with expression of pro-TLS genes, suggesting DRP loss may favor TLS development in support of improved patient outcome and patient response to interventional immunotherapy.
Assuntos
Biomarcadores Tumorais/genética , Melanoma/genética , Mutação , Neoplasias Cutâneas/genética , Estruturas Linfoides Terciárias/imunologia , Animais , Humanos , Imunoterapia , Melanoma/imunologia , Melanoma/patologia , Melanoma/terapia , Prognóstico , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/terapia , Estruturas Linfoides Terciárias/patologia , Microambiente TumoralRESUMO
BACKGROUND: The degree of immune infiltration in tumors, especially CD8+ T cells, greatly impacts patient disease course and response to interventional immunotherapy. Enhancement of tumor infiltrating lymphocyte (TIL) is a critical element of efficacious therapy and one that may be achieved via administration of agents that promote tumor vascular normalization (VN) and/or induce the development of tertiary lymphoid structures (TLS) within the tumor microenvironment (TME). METHODS: Low-dose stimulator of interferon genes (STING) agonist ADU S-100 (5 µg/mouse) was delivered intratumorally to established subcutaneous B16.F10 melanomas on days 10, 14 and 17 post-tumor inoculation. Treated and control tumors were isolated at various time points to assess transcriptional changes associated with VN and TLS formation via quantitative PCR (qPCR), with corollary immune cell composition changes in isolated tissues determined using flow cytometry and immunofluorescence microscopy. In vitro assays were performed on CD11c+ BMDCs treated with 2.5 µg/mL ADU S-100 or CD11c+ DCs isolated from tumor digests and associated transcriptional changes analyzed via qPCR or profiled using DNA microarrays. For T cell repertoireß-CDR3 analyses, T cell CDR3 was sequenced from gDNA isolated from splenocytes and enzymatically digested tumors. RESULTS: We report that activation of STING within the TME leads to slowed melanoma growth in association with increased production of antiangiogenic factors including Tnfsf15 (Vegi) and Cxcl10, and TLS-inducing factors including Ccl19, Ccl21, Lta, Ltb and Light. Therapeutic responses resulting from intratumoral STING activation were characterized by improved VN, enhanced tumor infiltration by CD8+ T cells and CD11c+ DCs and local TLS neogenesis, all of which were dependent on host expression of STING. Consistent with a central role for DC in TLS formation, ADU S-100-activated mCD11c+ DCs also exhibited upregulated expression of TLS promoting factors including lymphotoxin-α (LTA), interleukin (IL)-36, inflammatory chemokines and type I interferons in vitro and in vivo. TLS formation in ADU S-100-treated mice was associated with the development of a highly oligoclonal TIL repertoire enriched in expanded T cell clonotypes unique to the TME and not detected in the periphery. CONCLUSIONS: Our data support the premise that i.t. delivery of low-dose STING agonist promotes VN and a proinflammatory TME supportive of TLS formation, enrichment in the TIL repertoire and tumor growth control.