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1.
Nat Commun ; 12(1): 5314, 2021 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-34493727

RESUMO

Adoptive T cell therapy (ACT) requires lymphodepletion preconditioning to eliminate immune-suppressive elements and enable efficient engraftment of adoptively transferred tumor-reactive T cells. As anti-CD4 monoclonal antibody depletes CD4+ immune-suppressive cells, the combination of anti-CD4 treatment and ACT has synergistic potential in cancer therapy. Here, we demonstrate a post-ACT conditioning regimen that involves transient anti-CD4 treatment (CD4post). Using murine melanoma, the combined effect of cyclophosphamide preconditioning (CTXpre), CD4post, and ex vivo primed tumor-reactive CD8+ T-cell infusion is presented. CTXpre/CD4post increases tumor suppression and host survival by accelerating the proliferation and differentiation of ex vivo primed CD8+ T cells and endogenous CD8+ T cells. Endogenous CD8+ T cells enhance effector profile and tumor-reactivity, indicating skewing of the TCR repertoire. Notably, enrichment of polyfunctional IL-18Rαhi CD8+ T cell subset is the key event in CTXpre/CD4post-induced tumor suppression. Mechanistically, the anti-tumor effect of IL-18Rαhi subset is mediated by IL-18 signaling and TCR-MHC I interaction. This study highlights the clinical relevance of CD4post in ACT and provides insights regarding the immunological nature of anti-CD4 treatment, which enhances anti-tumor response of CD8+ T cells.


Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos Alquilantes/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Ciclofosfamida/farmacologia , Subunidade alfa de Receptor de Interleucina-18/genética , Melanoma Experimental/terapia , Neoplasias Cutâneas/terapia , Linfócitos T Citotóxicos/imunologia , Transferência Adotiva , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Imunoterapia Adotiva/métodos , Interleucina-18/genética , Interleucina-18/imunologia , Subunidade alfa de Receptor de Interleucina-18/agonistas , Subunidade alfa de Receptor de Interleucina-18/imunologia , Ativação Linfocitária , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Camundongos , Camundongos Endogâmicos C57BL , Receptores CCR4/genética , Receptores CCR4/imunologia , Receptores CCR8/genética , Receptores CCR8/imunologia , Receptores Histamínicos H4/genética , Receptores Histamínicos H4/imunologia , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Análise de Sobrevida , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/transplante , Carga Tumoral/efeitos dos fármacos
2.
Nat Commun ; 12(1): 4734, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34354077

RESUMO

The tumor microenvironment (TME) is a complex amalgam of tumor cells, immune cells, endothelial cells and fibroblastic stromal cells (FSC). Cancer-associated fibroblasts are generally seen as tumor-promoting entity. However, it is conceivable that particular FSC populations within the TME contribute to immune-mediated tumor control. Here, we show that intratumoral treatment of mice with a recombinant lymphocytic choriomeningitis virus-based vaccine vector expressing a melanocyte differentiation antigen resulted in T cell-dependent long-term control of melanomas. Using single-cell RNA-seq analysis, we demonstrate that viral vector-mediated transduction reprogrammed and activated a Cxcl13-expressing FSC subset that show a pronounced immunostimulatory signature and increased expression of the inflammatory cytokine IL-33. Ablation of Il33 gene expression in Cxcl13-Cre-positive FSCs reduces the functionality of intratumoral T cells and unleashes tumor growth. Thus, reprogramming of FSCs by a self-antigen-expressing viral vector in the TME is critical for curative melanoma treatment by locally sustaining the activity of tumor-specific T cells.


Assuntos
Melanoma Experimental/terapia , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Fibroblastos Associados a Câncer/imunologia , Fibroblastos Associados a Câncer/patologia , Técnicas de Reprogramação Celular/métodos , Quimiocina CXCL13/genética , Quimiocina CXCL13/imunologia , Feminino , Vetores Genéticos , Interleucina-33/deficiência , Interleucina-33/genética , Interleucina-33/imunologia , Oxirredutases Intramoleculares/genética , Oxirredutases Intramoleculares/imunologia , Vírus da Coriomeningite Linfocítica/genética , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Células Estromais/imunologia , Células Estromais/patologia , Linfócitos T/imunologia , Linfócitos T/patologia , Microambiente Tumoral/imunologia
3.
Science ; 373(6558): 1040-1046, 2021 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-34446607

RESUMO

The antitumor efficacy of cancer immunotherapy can correlate with the presence of certain bacterial species within the gut microbiome. However, many of the molecular mechanisms that influence host response to immunotherapy remain elusive. In this study, we show that members of the bacterial genus Enterococcus improve checkpoint inhibitor immunotherapy in mouse tumor models. Active enterococci express and secrete orthologs of the NlpC/p60 peptidoglycan hydrolase SagA that generate immune-active muropeptides. Expression of SagA in nonprotective E. faecalis was sufficient to promote immunotherapy response, and its activity required the peptidoglycan sensor NOD2. Notably, SagA-engineered probiotics or synthetic muropeptides also augmented anti-PD-L1 antitumor efficacy. Taken together, our data suggest that microbiota species with specialized peptidoglycan remodeling activity and muropeptide-based therapeutics may enhance cancer immunotherapy and could be leveraged as next-generation adjuvants.


Assuntos
Antígeno B7-H1/antagonistas & inibidores , Enterococcus/metabolismo , Inibidores de Checkpoint Imunológico/uso terapêutico , Melanoma Experimental/terapia , N-Acetil-Muramil-L-Alanina Amidase/metabolismo , Peptidoglicano/metabolismo , Animais , Carga Bacteriana , Proteínas de Bactérias/metabolismo , Enterococcus/enzimologia , Enterococcus faecalis/metabolismo , Enterococcus faecium/metabolismo , Microbioma Gastrointestinal , Imunoterapia , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína Adaptadora de Sinalização NOD2/metabolismo , Fragmentos de Peptídeos/metabolismo , Probióticos , Transdução de Sinais
4.
Int J Cancer ; 149(6): 1369-1384, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34086978

RESUMO

Oncolytic virotherapies are perceived as remarkable immunotherapies coming into view and represent highly promising cancer treatments, yet to figure out its specific immune responses and underlying barriers remains critical. Albeit recent studies have demonstrated that oncolytic viruses (OVs) could fine tune tumor microenvironment (TME) to elicit tumor suppression mainly due to effective T-cell responses, the interaction between suppressive T cells and OVs is barely undetermined. Herein, we found that regulatory T cells (Treg cells) were increased in the TME following systemic administration of oncolytic virus M1 along with the higher expression of relative cytokines and chemokines in both mouse RM-1 prostatic carcinoma model and mouse B16F10 melanoma model. Besides, Treg cells expressed high levels of CD25 post-M1 treatment, and its suppressive effect on CD8+ T cells was also elevated. Depletion of Treg cells in M1-treated groups significantly reinforced antitumor effect of M1. Specific targeting of Treg cells using cytotoxic T lymphocyte-associated protein 4 (CTLA-4) antibody (Ab) in combination with M1 treatment elicited a more profound tumor suppression and longer overall survival time than M1 alone in both tumor models. Moreover, CTLA-4 Ab further aggrandized antitumor immune response elicited by M1, including increased infiltration of CD45+ immune cells and CD8+ or CD4+ T lymphocytes, decreased ratio of Treg cells to CD4+ T lymphocytes, the intensified lymphocytotoxicity and elevated secretion of cytotoxic cytokines like interferon-γ, granzyme B and perforin. Therefore, our findings constituted a suggestive evidence that targeting Treg cells in M1-based oncolytic virotherapy may achieve a highly response in clinical cancer research.


Assuntos
Inibidores de Checkpoint Imunológico/administração & dosagem , Melanoma Experimental/terapia , Vírus Oncolíticos/fisiologia , Doenças Prostáticas/terapia , Linfócitos T Reguladores/metabolismo , Administração Intravenosa , Animais , Linfócitos T CD8-Positivos/metabolismo , Antígeno CTLA-4/antagonistas & inibidores , Linhagem Celular Tumoral , Terapia Combinada , Citocinas/metabolismo , Feminino , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Masculino , Melanoma Experimental/imunologia , Camundongos , Terapia Viral Oncolítica , Doenças Prostáticas/imunologia , Microambiente Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Theranostics ; 11(15): 7308-7321, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34158852

RESUMO

Dendritic cells (DCs) can process the antigens of cancer vaccine and thus stimulate the CD8+ T cells to recognize and kill the tumor cells that express these antigens. However, lack of promising carriers for presenting the antigens to DCs is one of the main barriers to the development of clinically effective cancer vaccines. Another limitation is the risk of inflammatory side effects induced by the adjuvants. It is still unclear how we can develop ideal adjuvant-free DC vaccine carriers without adjuvants. Methods: A 12-mer peptide carrier (CBP-12) with high affinity for Clec9a expressed on DCs was developed using an in silico rational optimization method. The therapeutic effects of the adjuvant-free vaccine comprising CBP-12 and exogenous or endogenous antigenic peptides were investigated in terms of antigen cross-presentation efficacy, specific cytotoxic T lymphocyte response, and antitumor activity. We also explored the mechanism involved in the antitumor effects of the adjuvant-free CBP-12 vaccine. Finally, we assessed the effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Results: Here, we developed CBP-12 as a vaccine carrier that enhanced the uptake and cross-presentation of the antigens, thus inducing strong CD8+ T cell responses and antitumor effects in both anti-PD-1-responsive (B16-OVA) and -resistant (B16) models, even in adjuvant-free conditions. CBP-12 bound to and activated Clec9a, thereby stimulating Clec9a+ DC to product IL-21, but not IL-12 by activating of Syk. The antitumor effects of the CBP-12 conjugated peptide vaccines could be blocked by an IL-21 neutralizing antibody. We also observed the synergistic antitumor effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Conclusions: CBP-12 could serve as an adjuvant-free peptide vaccine carrier for cancer immunotherapy.


Assuntos
Vacinas Anticâncer , Células Dendríticas/imunologia , Sistemas de Liberação de Medicamentos , Interleucinas/imunologia , Lectinas Tipo C/imunologia , Melanoma Experimental/imunologia , Peptídeos , Receptores Imunológicos/imunologia , Transdução de Sinais/efeitos dos fármacos , Quinase Syk/imunologia , Animais , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/farmacologia , Feminino , Interleucinas/genética , Lectinas Tipo C/genética , Melanoma Experimental/genética , Melanoma Experimental/terapia , Camundongos , Camundongos Knockout , Peptídeos/imunologia , Peptídeos/farmacologia , Receptores Imunológicos/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Quinase Syk/genética , Vacinas de Subunidades/imunologia , Vacinas de Subunidades/farmacologia
6.
ACS Appl Mater Interfaces ; 13(23): 26790-26799, 2021 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-34061496

RESUMO

For melanoma with high lethality and metastasis rate, traditional therapy has limited effects; local photothermal therapy (PTT) synergetic with immune therapy for cancer treatment can perhaps improve the situation. However, because of the natural existence of the tumor matrix barrier, the penetration depth of drugs and immune cells often dampens the efficacy of cancer treatment. Herein, we report an innovative synergetic PTT and immune therapy through dissolving microneedles for the codelivery of the hyaluronidase-modified semiconductor polymer nanoparticles containing poly(cyclopentadithiophene-alt-benzothiadiazole) and immune adjuvant polyinosinic-polycytidylic acid (PIC). Benefiting from the dissolution of an extracellular matrix of hyaluronidase, the semiconductor polymer nanoparticles and PIC penetrate the tumor deeply, under synergetic therapy with PTT, activating the immune cells and enhancing the T-cell immune response for inhibition of tumor growth and metastasis. This study provides a promising platform for effective melanoma treatment and a novel strategy to overcome the stromal barrier.


Assuntos
Matriz Extracelular/metabolismo , Hialuronoglucosaminidase/metabolismo , Melanoma Experimental/terapia , Nanopartículas/administração & dosagem , Terapia Fototérmica , Polímeros/química , Linfócitos T/imunologia , Animais , Feminino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/química , Agulhas
7.
Nat Commun ; 12(1): 3182, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34075041

RESUMO

Interleukin 9 (IL-9)-producing helper T (Th9) cells are essential for inducing anti-tumor immunity and inflammation in allergic and autoimmune diseases. Although transcription factors that are essential for Th9 cell differentiation have been identified, other signaling pathways that are required for their generation and functions are yet to be explored. Here, we identify that Epidermal Growth Factor Receptor (EGFR) is essential for IL-9 induction in helper T (Th) cells. Moreover, amphiregulin (Areg), an EGFR ligand, is critical for the amplification of Th9 cells induced by TGF-ß1 and IL-4. Furthermore, our data show that Areg-EGFR signaling induces HIF1α, which binds and transactivates IL-9 and NOS2 promoters in Th9 cells. Loss of EGFR or HIF1α abrogates Th9 cell differentiation and suppresses their anti-tumor functions. Moreover, in line with its reliance on HIF1α expression, metabolomics profiling of Th9 cells revealed that Succinate, a TCA cycle metabolite, promotes Th9 cell differentiation and Th9 cell-mediated tumor regression.


Assuntos
Receptores ErbB/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Interleucina-9/genética , Melanoma Experimental/terapia , Neoplasias Cutâneas/terapia , Linfócitos T Auxiliares-Indutores/imunologia , Anfirregulina/metabolismo , Animais , Diferenciação Celular/imunologia , Feminino , Células HEK293 , Voluntários Saudáveis , Humanos , Imunoterapia Adotiva/métodos , Melanoma Experimental/imunologia , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/genética , Cultura Primária de Células , RNA-Seq , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Neoplasias Cutâneas/imunologia , Ácido Succínico/metabolismo , Linfócitos T Auxiliares-Indutores/transplante , Ativação Transcricional/imunologia
8.
Cancer Sci ; 112(7): 2652-2663, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33934440

RESUMO

Mitochondria are key cytoplasmic organelles. Their activation is critical for the generation of T cell proliferation and cytotoxicity. Exhausted tumor-infiltrating T cells show a decreased mitochondrial function and mass. 5-Aminolevulinic acid (5-ALA), a natural amino acid that is only produced in the mitochondria, has been shown to influence metabolic functions. We hypothesized that 5-ALA with sodium ferrous citrate (SFC) might provide metabolic support for tumor-infiltrating T cells. In a mouse melanoma model, we found that 5-ALA/SFC with a programmed cell death-ligand 1 (PD-L1) blocking Ab synergized tumor regression. After treatment with 5-ALA/SFC and anti-PD-L1 Ab, tumor infiltrating lymphocytes (TILs) were not only competent for the production of cytolytic particles and cytokines (granzyme B, interleukin-2, and γ-interferon) but also showed enhanced Ki-67 activity (a proliferation marker). The number of activated T cells (PD-1+ Tim-3- ) was also significantly increased. Furthermore, we found that 5-ALA/SFC activated the mitochondrial functions, including the oxygen consumption rate, ATP level, and complex V expression. The mRNA levels of Nrf-2, HO-1, Sirt-1, and PGC-1α and the protein levels of Sirt-1 were upregulated by treatment with 5-ALA/SFC. Taken together, our findings revealed that 5-ALA/SFC could be a key metabolic regulator in exhausted T cell metabolism and suggested that 5-ALA/SFC might synergize with anti-PD-1/PD-L1 therapy to boost the intratumoral efficacy of tumor-specific T cells. Our study not only revealed a new aspect of immune metabolism, but also paved the way to develop a strategy for combined anti-PD-1/PD-L1 cancer immunotherapy.


Assuntos
Ácido Aminolevulínico/farmacologia , Antígeno B7-H1/antagonistas & inibidores , Ácido Cítrico/farmacologia , Compostos Ferrosos/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Trifosfato de Adenosina/metabolismo , Animais , Linhagem Celular Tumoral , Terapia Combinada , Feminino , Heme Oxigenase-1/metabolismo , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Antígeno Ki-67/metabolismo , Contagem de Linfócitos , Linfócitos do Interstício Tumoral/citologia , Linfócitos do Interstício Tumoral/metabolismo , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Fator 1 Nuclear Respiratório/metabolismo , Consumo de Oxigênio/efeitos dos fármacos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Sirtuína 1/metabolismo
9.
Methods Mol Biol ; 2255: 171-186, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34033103

RESUMO

Immunogenic cell death (ICD) is a form of regulated cell death that is capable of eliciting an immune response. In cancer, tumor cells undergoing ICD are known to emit damage associated molecular patterns (DAMPs) that are capable of recruiting and activating antigen presenting cells (APCs), which ultimately lead to the activation of an antitumor immune response. Surface translocation of intracellular chaperones such as calreticulin, release of TLR agonists such as high mobility box 1, and the secretion of type I IFN are some of the hallmark features seen in tumors succumbing to ICD. While detection of these molecules is suggestive of ICD induction, which alone does not certify that the treatment is an ICD inducer, an in vivo vaccination assay using injured tumor cells remains to be the gold standard method to functionally verify ICD. This chapter will discuss the necessary steps required to conduct an in vivo vaccination assay, focusing on the preparation of vaccine using treated tumor cells, and how these cells are then utilized in the animal model.


Assuntos
Antineoplásicos/farmacologia , Apoptose , Vacinas Anticâncer/administração & dosagem , Modelos Animais de Doenças , Morte Celular Imunogênica , Melanoma Experimental/terapia , Vacinação/métodos , Animais , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos
10.
J Mater Chem B ; 9(18): 3892-3899, 2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-33928989

RESUMO

Immunotherapy is revolutionizing cancer treatment. Vaccination of antigenic peptides has been identified as a promising strategy for cancer immunotherapy while insufficient immune responses were stimulated due to low antigenicity. Moreover, immune checkpoint blockade therapy is still limited by a low objective response rate. In this work, cationic polymer-lipid hybrid nanovesicle (P/LNV)-based liposomes are designed to simultaneously deliver tumor vaccines composed of anionic antigen epitopes, toll-like receptor-9 agonist (TLR9), CpG (AE/CpG), and indoleamine-2,3-dioxygenase (IDO) inhibitor, 1-methyl-tryptophan (1-MT), to increase the immunogenicity of peptide antigens and meanwhile block the immune checkpoint. P/LNV liposomes efficiently enhanced the uptake of vaccines by dendritic cells (DCs) and improved the maturation of DCs indicated by the significantly increased percentage of CD86+MHCI+ DCs, resulting in a potent cytotoxic T-lymphocyte (CTL) response against B16-OVA tumor cells in vitro. Importantly, the combination immunotherapy showed significantly higher therapeutic efficiency towards melanoma tumors in mice, compared with an untreated or individual therapy modality. Mechanistically, the co-delivery system could elicit a strong cancer-specific T-cell response, as characterized by the remarkably increased infiltration of CD8+ T cells in the tumor and draining lymph nodes. Altogether, cationic liposomes delivered with tumor vaccines and IDO inhibitor provide a promising platform for cancer immunotherapy by provoking antitumor T-cell immunity and simultaneously reversing the immunosuppressive tumor microenvironment.


Assuntos
Ilhas de CpG , Epitopos/imunologia , Imunoterapia/métodos , Lipossomos/química , Melanoma Experimental/terapia , Triptofano/análogos & derivados , Animais , Ânions/química , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , Cátions/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Epitopos/química , Lipídeos/química , Lipossomos/farmacologia , Melanoma Experimental/mortalidade , Camundongos , Camundongos Endogâmicos C57BL , Nanoestruturas/química , Polímeros/química , Taxa de Sobrevida , Triptofano/química
11.
J Med Chem ; 64(9): 5802-5815, 2021 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-33844923

RESUMO

Peptide stapling chemistry represents an attractive strategy to promote the clinical translation of protein epitope mimetics, but its use has not been applied to natural cytotoxic peptides (NCPs) to produce new oncolytic peptides. Based on a wasp venom peptide, a series of stapled anoplin peptides (StAnos) were prepared. The optimized stapled Ano-3/3s were shown to be protease-resistant and exerted superior tumor cell-selective cytotoxicity by rapid membrane disruption. In addition, Ano-3/3s induced tumor ablation in mice through the direct oncolytic effect and subsequent stimulation of immunogenic cell death. This synergistic oncolytic-immunotherapy effect is more remarkable on melanoma than on triple-negative breast cancer in vivo. The efficacies exerted by Ano-3/3s on melanoma were further characterized by CD8+ T cell infiltration, and the addition of anti-CD8 antibodies diminished the long-term antitumor effects. In summary, these results support stapled peptide chemistry as an advantageous method to enhance the NCP potency for oncolytic therapy.


Assuntos
Membrana Celular/metabolismo , Melanoma Experimental/terapia , Peptídeos/uso terapêutico , Venenos de Vespas/metabolismo , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Desenho de Fármacos , Feminino , Interações Hidrofóbicas e Hidrofílicas , Morte Celular Imunogênica/efeitos dos fármacos , Imunoterapia , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Peptídeos/química , Peptídeos/farmacologia , Análise de Sobrevida , Transplante Homólogo , Neoplasias de Mama Triplo Negativas/imunologia , Neoplasias de Mama Triplo Negativas/terapia , Venenos de Vespas/química
12.
J Hematol Oncol ; 14(1): 43, 2021 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-33731208

RESUMO

BACKGROUND: Transgelin-2 is a 22 kDa actin-binding protein that has been proposed to act as an oncogenic factor, capable of contributing to tumorigenesis in a wide range of human malignancies. However, little is known whether this tiny protein also plays an important role in immunity, thereby keeping body from the cancer development and metastasis. Here, we investigated the functions of transgelin-2 in dendritic cell (DC) immunity. Further, we investigated whether the non-viral transduction of cell-permeable transgelin-2 peptide potentially enhance DC-based cancer immunotherapy. METHODS: To understand the functions of transgelin-2 in DCs, we utilized bone marrow-derived DCs (BMDCs) purified from transgelin-2 knockout (Tagln2-/-) mice. To observe the dynamic cellular mechanism of transgelin-2, we utilized confocal microscopy and flow cytometry. To monitor DC migration and cognate T-DC interaction in vivo, we used intravital two-photon microscopy. For the solid and metastasis tumor models, OVA+ B16F10 melanoma were inoculated into the C57BL/6 mice via intravenously (i.v.) and subcutaneously (s.c.), respectively. OTI TCR T cells were used for the adoptive transfer experiments. Cell-permeable, de-ubiquitinated recombinant transgelin-2 was purified from Escherichia coli and applied for DC-based adoptive immunotherapy. RESULTS: We found that transgelin-2 is remarkably expressed in BMDCs during maturation and lipopolysaccharide activation, suggesting that this protein plays a role in DC-based immunity. Although Tagln2-/- BMDCs exhibited no changes in maturation, they showed significant defects in their abilities to home to draining lymph nodes (LNs) and prime T cells to produce antigen-specific T cell clones, and these changes were associated with a failure to suppress tumor growth and metastasis of OVA+ B16F10 melanoma cells in mice. Tagln2-/- BMDCs had defects in filopodia-like membrane protrusion and podosome formation due to the attenuation of the signals that modulate actin remodeling in vitro and formed short, unstable contacts with cognate CD4+ T cells in vivo. Strikingly, non-viral transduction of cell-permeable, de-ubiquitinated recombinant transgelin-2 potentiated DC functions to suppress tumor growth and metastasis. CONCLUSION: This work demonstrates that transgelin-2 is an essential protein for both cancer and immunity. Therefore, transgelin-2 can act as a double-edged sword depending on how we apply this protein to cancer therapy. Engineering and clinical application of this protein may unveil a new era in DC-based cancer immunotherapy. Our findings indicate that cell-permeable transgelin-2 have a potential clinical value as a cancer immunotherapy based on DCs.


Assuntos
Transferência Adotiva , Células Dendríticas/imunologia , Melanoma Experimental/terapia , Proteínas dos Microfilamentos/imunologia , Proteínas Musculares/imunologia , Animais , Movimento Celular , Células Cultivadas , Células Dendríticas/citologia , Feminino , Imunidade , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Proteínas Musculares/genética
13.
J Immunol ; 206(8): 1966-1975, 2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33722878

RESUMO

Inflammation has long been associated with cancer initiation and progression; however, how inflammation causes immune suppression in the tumor microenvironment and resistance to immunotherapy is not well understood. In this study, we show that both innate proinflammatory cytokine IL-1α and immunotherapy-induced IL-1α make melanoma resistant to immunotherapy. In a mouse melanoma model, we found that tumor size was inversely correlated with response to immunotherapy. Large tumors had higher levels of IL-1α, Th2 cytokines, polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs), and regulatory T cells but lower levels of IL-12, Th1 cytokines, and activated T cells. We found that therapy with adenovirus-encoded CD40L (rAd.CD40L) increased tumor levels of IL-1α and PMN-MDSCs. Blocking the IL-1 signaling pathway significantly decreased rAd.CD40L-induced PMN-MDSCs and their associated PD-L1 expression in the tumor microenvironment and enhanced tumor-specific immunity. Similarly, blocking the IL-1 signaling pathway improved the antimelanoma activity of anti-PD-L1 Ab therapy. Our study suggests that blocking the IL-1α signaling pathway may increase the efficacy of immunotherapies against melanoma.


Assuntos
Resistencia a Medicamentos Antineoplásicos/imunologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Imunoterapia/métodos , Interleucina-1alfa/imunologia , Melanoma Experimental/terapia , Animais , Linhagem Celular Tumoral , Citocinas/imunologia , Citocinas/metabolismo , Humanos , Inibidores de Checkpoint Imunológico/imunologia , Interleucina-1alfa/metabolismo , Estimativa de Kaplan-Meier , Melanoma Experimental/imunologia , Camundongos Endogâmicos C57BL , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/metabolismo , Neutrófilos/imunologia , Neutrófilos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia
14.
J Am Chem Soc ; 143(13): 5127-5140, 2021 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-33764762

RESUMO

Immunotherapy has provided a promising strategy for the treatment of cancers. However, even in tumors with high antigen burdens, the systemic inhibition of the antigen presentation still greatly restricts the application of immunotherapy. Here, we construct a tumor protein-engineering system based on the functional tripeptide, Asp-Phe-Tyr (DFY), which can automatically collect and deliver immunogenetic tumor proteins from targeted cells to immune cells. Through a tyrosinase-catalyzed polymerization, the DFY tripeptide selectively accumulates in tyrosinase high-expressed melanoma cells. Then quinone-rich intermediates are covalently linked with tumor-specific proteins by Michael addition and form tumor protein-carried microfibers that could be engulfed by antigen-presenting cells and exhibited tumor antigenic properties for boosting immune effect. In melanoma cells with deficient antigen presentation, this system can successfully enrich and transport tumor antigen-containing proteins to immune cells. Furthermore, in the in vivo study on murine melanoma, the transdermal delivery of the DFY tripeptide suppressed the tumor growth and the postsurgery recurrence. Our findings provide an avenue for the regulation of the immune system on an organism by taking advantage of certain polymerization reactions by virtue of chemical biology.


Assuntos
Imunoterapia/métodos , Melanoma Experimental/terapia , Monofenol Mono-Oxigenase/metabolismo , Oligopeptídeos/uso terapêutico , Administração Cutânea , Animais , Células Apresentadoras de Antígenos/imunologia , Catálise , Melanoma Experimental/imunologia , Camundongos , Oligopeptídeos/administração & dosagem , Oligopeptídeos/metabolismo , Polimerização
15.
J Clin Invest ; 131(8)2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33690225

RESUMO

Melanomas commonly undergo a phenotype switch, from a proliferative to an invasive state. Such tumor cell plasticity contributes to immunotherapy resistance; however, the mechanisms are not completely understood and thus are therapeutically unexploited. Using melanoma mouse models, we demonstrated that blocking the MNK1/2-eIF4E axis inhibited melanoma phenotype switching and sensitized melanoma to anti-PD-1 immunotherapy. We showed that phospho-eIF4E-deficient murine melanomas expressed high levels of melanocytic antigens, with similar results verified in patient melanomas. Mechanistically, we identified phospho-eIF4E-mediated translational control of NGFR, a critical effector of phenotype switching. Genetic ablation of phospho-eIF4E reprogrammed the immunosuppressive microenvironment, exemplified by lowered production of inflammatory factors, decreased PD-L1 expression on dendritic cells and myeloid-derived suppressor cells, and increased CD8+ T cell infiltrates. Finally, dual blockade of the MNK1/2-eIF4E axis and the PD-1/PD-L1 immune checkpoint demonstrated efficacy in multiple melanoma models regardless of their genomic classification. An increase in the presence of intratumoral stem-like TCF1+PD-1+CD8+ T cells, a characteristic essential for durable antitumor immunity, was detected in mice given a MNK1/2 inhibitor and anti-PD-1 therapy. Using MNK1/2 inhibitors to repress phospho-eIF4E thus offers a strategy to inhibit melanoma plasticity and improve response to anti-PD-1 immunotherapy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Fator de Iniciação 4E em Eucariotos/imunologia , Imunidade Celular , Sistema de Sinalização das MAP Quinases/imunologia , Melanoma Experimental/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Linhagem Celular Tumoral , Fator de Iniciação 4E em Eucariotos/genética , Imunoterapia , Sistema de Sinalização das MAP Quinases/genética , Melanoma Experimental/genética , Melanoma Experimental/terapia , Camundongos , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Proteínas Serina-Treonina Quinases/genética , Receptor de Fator de Crescimento Neural/genética , Receptor de Fator de Crescimento Neural/imunologia
16.
Cancer Res ; 81(8): 1965-1976, 2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33589517

RESUMO

Inactivation of tumor-infiltrating lymphocytes (TIL) is one of the mechanisms mitigating antitumor immunity during tumor onset and progression. Epigenetic abnormalities are regarded as a major culprit contributing to the dysfunction of TILs within tumor microenvironments. In this study, we used a murine model of melanoma to discover that Tet2 inactivation significantly enhances the antitumor activity of TILs with an efficacy comparable to immune checkpoint inhibition imposed by anti-PD-L1 treatment. Single-cell RNA-sequencing analysis suggested that Tet2-deficient TILs exhibit effector-like features. Transcriptomic and ATAC-sequencing analysis showed that Tet2 ablation reshapes chromatin accessibility and favors binding of transcription factors geared toward CD8+ T-cell activation. Furthermore, the ETS family of transcription factors contributed to augmented CD8+ T-cell function following Tet2 depletion. Overall, our study establishes that Tet2 constitutes one of the epigenetic barriers that account for dysfunction of TILs and that Tet2 inactivation could promote antitumor immunity to suppress tumor growth. SIGNIFICANCE: This study suggests that ablation of TET2+ from TILs could promote their antitumor function by reshaping chromatin accessibility for key transcription factors and enhancing the transcription of genes essential for antitumor activity.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Proteínas de Ligação a DNA/deficiência , Ativação Linfocitária/imunologia , Linfócitos do Interstício Tumoral/imunologia , Melanoma Experimental/imunologia , Proteínas Proto-Oncogênicas/deficiência , Transferência Adotiva/métodos , Animais , Cromatina/metabolismo , Desmetilação do DNA , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Epigênese Genética , Deleção de Genes , Inativação Gênica , Inibidores de Checkpoint Imunológico/uso terapêutico , MAP Quinase Quinase Quinases , Melanoma Experimental/metabolismo , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Perforina/metabolismo , Proteínas Proto-Oncogênicas/genética , Análise de Sequência de RNA , Fatores de Transcrição/metabolismo , Microambiente Tumoral/imunologia , Fator de Necrose Tumoral alfa/metabolismo
17.
Hum Vaccin Immunother ; 17(7): 1910-1922, 2021 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-33522416

RESUMO

Anti-programmed death ligand 1 (PD-L1) therapy has been beneficial in treating patients with certain cancers. Here, we tested whether anti-PD-L1 therapy is effective for controlling different types of tumors using animal models of TC-1, MC38 and B16. We found that, despite PD-L1 expression, anti-PD-L1 therapy showed little and some antitumor activity in the TC-1 and MC38 models. However, anti-PD-L1 therapy exhibited a more dramatic antitumor effect in the B16 model. This difference in antitumor responses was likely associated with the CD8 + T cell infiltration status of tumor tissues. In the B16 model, CD8 + T cells and to a lesser degree NK cells were found to be responsible for the antitumor response of anti-PD-L1 therapy, as determined by immune cell subset depletion. In particular, CD8 + T cells from B16-bearing mice produced an IFN-γ in response to B16 cells and citrate phosphate buffer-treated B16 cell peptide elutes but not to an immunodominant class I epitope, Trp2180-188, suggesting that CD8 + T cells that recognize neoantigens were induced in B16 tumor-bearing mice and then reactivated by anti-PD-L1 for tumor control. When B16 tumor-bearing mice were treated with anti-PD-L1 in combination with Trp2180-188 peptide vaccines, they displayed significantly more tumor control than either single therapy. Taken together, these studies show that B16 melanomas are more effectively controlled through reactivation of tumor-infiltrating lymphocytes by anti-PD-L1 therapy. Moreover, combined therapy using anti-PD-L1 and Trp2 peptide vaccines is more beneficial for controlling B16 melanomas through reactivation of neoantigen-specific CD8 + T cells and induction of Trp2-specific CD8 + T cells.


Assuntos
Vacinas Anticâncer , Melanoma Experimental , Animais , Antígeno B7-H1 , Linfócitos T CD8-Positivos , Linhagem Celular Tumoral , Humanos , Células Matadoras Naturais , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos
18.
Nat Commun ; 12(1): 832, 2021 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-33547304

RESUMO

The two T cell inhibitory receptors PD-1 and TIM-3 are co-expressed during exhausted T cell differentiation, and recent evidence suggests that their crosstalk regulates T cell exhaustion and immunotherapy efficacy; however, the molecular mechanism is unclear. Here we show that PD-1 contributes to the persistence of PD-1+TIM-3+ T cells by binding to the TIM-3 ligand galectin-9 (Gal-9) and attenuates Gal-9/TIM-3-induced cell death. Anti-Gal-9 therapy selectively expands intratumoral TIM-3+ cytotoxic CD8 T cells and immunosuppressive regulatory T cells (Treg cells). The combination of anti-Gal-9 and an agonistic antibody to the co-stimulatory receptor GITR (glucocorticoid-induced tumor necrosis factor receptor-related protein) that depletes Treg cells induces synergistic antitumor activity. Gal-9 expression and secretion are promoted by interferon ß and γ, and high Gal-9 expression correlates with poor prognosis in multiple human cancers. Our work uncovers a function for PD-1 in exhausted T cell survival and suggests Gal-9 as a promising target for immunotherapy.


Assuntos
Adenocarcinoma/terapia , Neoplasias do Colo/terapia , Galectinas/imunologia , Regulação Neoplásica da Expressão Gênica/imunologia , Proteína Relacionada a TNFR Induzida por Glucocorticoide/imunologia , Receptor Celular 2 do Vírus da Hepatite A/imunologia , Receptor de Morte Celular Programada 1/imunologia , Adenocarcinoma/genética , Adenocarcinoma/imunologia , Adenocarcinoma/mortalidade , Animais , Anticorpos/farmacologia , Antineoplásicos Imunológicos/farmacologia , Neoplasias do Colo/genética , Neoplasias do Colo/imunologia , Neoplasias do Colo/mortalidade , Galectinas/antagonistas & inibidores , Galectinas/genética , Proteína Relacionada a TNFR Induzida por Glucocorticoide/agonistas , Proteína Relacionada a TNFR Induzida por Glucocorticoide/genética , Receptor Celular 2 do Vírus da Hepatite A/genética , Humanos , Imunoterapia/métodos , Células Jurkat , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos BALB C , Receptor de Morte Celular Programada 1/genética , Ligação Proteica , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/terapia , Análise de Sobrevida , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
19.
J Clin Invest ; 131(6)2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33561014

RESUMO

Most clinically used anticancer mAbs are of the IgG isotype, which can eliminate tumor cells through NK cell-mediated antibody-dependent cellular cytotoxicity and macrophage-mediated antibody-dependent phagocytosis. IgG, however, ineffectively recruits neutrophils as effector cells. IgA mAbs induce migration and activation of neutrophils through the IgA Fc receptor (FcαRI) but are unable to activate NK cells and have poorer half-life. Here, we combined the agonistic activity of IgG mAbs and FcαRI targeting in a therapeutic bispecific antibody format. The resulting TrisomAb molecules recruited NK cells, macrophages, and neutrophils as effector cells for eradication of tumor cells in vitro and in vivo. Moreover, TrisomAb had long in vivo half-life and strongly decreased B16F10gp75 tumor outgrowth in mice. Importantly, neutrophils of colorectal cancer patients effectively eliminated tumor cells in the presence of anti-EGFR TrisomAb but were less efficient in mediating killing in the presence of IgG anti-EGFR mAb (cetuximab). The clinical application of TrisomAb may provide potential alternatives for cancer patients who do not benefit from current IgG mAb therapy.


Assuntos
Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antineoplásicos Imunológicos/farmacologia , Neutrófilos/imunologia , Animais , Anticorpos Biespecíficos/farmacologia , Anticorpos Monoclonais/farmacologia , Antígenos CD/imunologia , Linhagem Celular Tumoral , Cetuximab/farmacologia , Feminino , Células HCT116 , Humanos , Imunoglobulina G/imunologia , Células Matadoras Naturais/imunologia , Macrófagos/imunologia , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Imunológicos , Receptores Fc/imunologia
20.
Theranostics ; 11(2): 540-554, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33391491

RESUMO

Background: Advanced stage cancers with a suppressive tumor microenvironment (TME) are often refractory to immune checkpoint inhibitor (ICI) therapy. Recent studies have shown that focused ultrasound (FUS) TME-modulation can synergize ICI therapy, but enhancing survival outcomes in poorly immunogenic tumors remains challenging. Here, we investigated the role of focused ultrasound based boiling histotripsy (HT) and in-situ anti-CD40 agonist antibody (αCD40) combinatorial therapy in enhancing therapeutic efficacy against ICI refractory murine melanoma. Methods: Unilateral and bilateral large (~330-400 mm3) poorly immunogenic B16F10 melanoma tumors were established in the flank regions of mice. Tumors were exposed to single local HT followed by an in-situ administration of αCD40 (HT+ αCD40: HT40). Inflammatory signatures post treatment were assessed using pan-cancer immune profiling and flow cytometry. The ability of HT40 ± ICI to enhance local and systemic effects was determined by immunological characterization of the harvested tissues, and by tumor growth delay of local and distant untreated tumors 4-6 weeks post treatment. Results: Immune profiling revealed that HT40 upregulated a variety of inflammatory markers in the tumors. Immunologically, HT40 treated tumors showed an increased population of granzyme B+ expressing functional CD8+ T cells (~4-fold) as well as an increased M1 to M2 macrophage ratio (~2-3-fold) and CD8+ T: regulatory T cell ratio (~5-fold) compared to the untreated control. Systemically, the proliferation rates of the melanoma-specific memory T cell population were significantly enhanced by HT40 treatment. Finally, the combination of HT40 and ICI therapy (anti-CTLA-4 and anti-PD-L1) caused superior inhibition of distant untreated tumors, and prolonged survival rates compared to the control. Conclusions: Data suggest that HT40 reprograms immunologically cold tumors and sensitizes them to ICI therapy. This approach may be clinically useful for treating advanced stage melanoma cancers.


Assuntos
Antígenos CD40/imunologia , Linfócitos T CD8-Positivos/imunologia , Ablação por Ultrassom Focalizado de Alta Intensidade/métodos , Inibidores de Checkpoint Imunológico/uso terapêutico , Melanoma Experimental/terapia , Linfócitos T Reguladores/imunologia , Microambiente Tumoral/imunologia , Animais , Terapia Combinada , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL
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