RESUMO
BACKGROUND: The pleiotropic cytokine, transforming growth factor (TGF)-ß, and CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in actively suppressing antitumor immune responses. Evidence shows that TGF-ß produced by tumor cells promotes tolerance via expansion of Tregs. Our group previously demonstrated that blockade of TGF-ß signaling with a small molecule TGF-ß receptor I antagonist (SM16) inhibited primary and metastatic tumor growth in a T cell dependent fashion. In the current study, we evaluated the effect of SM16 on Treg generation and function. METHODS: Using BALB/c, FoxP3eGFP and Rag-/- mice, we performed FACS analysis to determine if SM16 blocked de novo TGF-ß-induced Treg generation in vitro and in vivo. CD4+ T cells from lymph node and spleen were isolated from control mice or mice maintained on SM16 diet, and flow cytometry analysis was used to detect the frequency of CD4+CD25-FoxP3+ and CD4+CD25+FoxP3+ T cells. In vitro suppression assays were used to determine the ability to suppress naive T cell proliferation in vitro of both CD4+CD25+FoxP3+ and CD4+CD25-FoxP3+ T cell sub-populations. We then examined whether SM16 diet exerted an inhibitory effect on primary tumor growth and correlated with changes in FoxP3+expression. ELISA analysis was used to measure IFN-γ levels after 72 h co-culture of CD4+CD25+ T cells from tumor-bearing mice on control or SM16 diet with CD4+CD25- T cells from naive donors. RESULTS: SM16 abrogates TGF-ß-induced Treg generation in vitro but does not prevent global homeostatic expansion of CD4+ T cell sub-populations in vivo. Instead, SM16 treatment causes expansion of a population of CD4+CD25-Foxp3+ Treg-like cells without significantly altering the overall frequency of Treg in lymphoreplete naive and tumor-bearing mice. Importantly, both the CD4+CD25-Foxp3+ T cells and the CD4+CD25+Foxp3+ Tregs in mice receiving SM16 diet exhibited diminished ability to suppress naive T cell proliferation in vitro compared to Treg from mice on control diet. CONCLUSIONS: These findings suggest that blockade of TGF-ß signaling is a potentially useful strategy for blunting Treg function to enhance the anti-tumor response. Our data further suggest that the overall dampening of Treg function may involve the expansion of a quiescent Treg precursor population, which is CD4+CD25-Foxp3+.
Assuntos
Antineoplásicos/metabolismo , Antígenos CD4/metabolismo , Linfócitos T CD4-Positivos/imunologia , Fatores de Transcrição Forkhead/metabolismo , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Contagem de Células , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Homeostase , Interferon gama/metabolismo , Linfonodos/citologia , Camundongos Knockout , Neoplasias/patologia , Baço/citologia , Linfócitos T Reguladores/imunologia , Carga TumoralRESUMO
BACKGROUND: Alpha-tocopheryloxyacetic acid (α-TEA) is a semi-synthetic derivative of naturally occurring vitamin E (alpha-tocopherol) that can be delivered via an oral route. Preclinical in vitro and in vivo data demonstrated that α-TEA is a potent anti-tumor agent with a safe toxicity profile in mice. We report a comprehensive study to evaluate the toxokinetics of good manufacturing practice (GMP)-grade α-TEA in dogs after daily oral administration for 28 days, followed by a 28-day recovery period. METHODS: Male and female beagle dogs received capsules of α-TEA Lysine Salt at doses of 100, 300, 1500 mg/kg/day. α-TEA plasma levels were determined by high-performance liquid chromatography (HPLC) with mass spectrometric detection. During the treatment, animals were observe for clinical signs, food consumption, body weight, and subjected to ophthalmoscopic, and electrocardiographic assessments. At the end of the dosing period, blood was taken and toxicokinetic analyses and histopathology assessments were performed when animals were necropsied. RESULTS: Our findings showed that there was no α-TEA-related mortality or moribundity. At the highest dose, increases in white blood cells and fibrinogen levels were observed. These levels returned to normal at the end of the recovery period. Histopathological evaluation of major organs revealed no significant lesions related to α-TEA-treatment. CONCLUSION: We demonstrate that for designing clinical trials in patients, the highest non-severely toxic dose (HNSTD) of α-TEA is 1500 mg/kg/day in Beagle dogs and this data informed the design of dose-escalation studies of α-TEA in patients with advanced cancer.
Assuntos
Tocoferóis/farmacocinética , Tocoferóis/toxicidade , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Antineoplásicos/farmacocinética , Antineoplásicos/toxicidade , Biópsia , Contagem de Células Sanguíneas , Análise Química do Sangue , Coagulação Sanguínea/efeitos dos fármacos , Testes de Coagulação Sanguínea , Cães , Feminino , Lisina , Masculino , Sais , Fatores de Tempo , Tocoferóis/administração & dosagem , Tocoferóis/química , Testes de Toxicidade , Toxicocinética , UrináliseRESUMO
The effects of α-tocopheryl succinate (α-TOS), α-tocopheryl acetyl ether (α-TEA) and triphenylphosphonium-tagged vitamin E succinate (mitochondrially targeted vitamin E succinate; MitoVES) on energy-related mitochondrial functions were determined in mitochondria isolated from AS-30D hepatoma and rat liver, bovine heart sub-mitochondrial particles (SMPs), and in rodent and human carcinoma cell lines and rat hepatocytes. In isolated mitochondria, MitoVES stimulated basal respiration and ATP hydrolysis, but inhibited net state 3 (ADP-stimulated) respiration and Ca(2+) uptake, by collapsing the membrane potential at low doses (1-10µM). Uncoupled mitochondrial respiration and basal respiration of SMPs were inhibited by the three drugs at concentrations at least one order of magnitude higher and with different efficacy: MitoVES>α-TEA>α-TOS. At high doses (>10µM), the respiratory complex II (CII) was the most sensitive MitoVES target. Acting as an uncoupler at low doses, this agent stimulated total O(2) uptake, collapsed ∆ψ(m), inhibited oxidative phosphorylation and induced ATP depletion in rodent and human cancer cells more potently than in normal rat hepatocytes. These findings revealed that in situ tumor mitochondria are preferred targets of the drug, indicating its clinical relevance.
Assuntos
Mitocôndrias/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Vitamina E/análogos & derivados , Trifosfato de Adenosina/química , Animais , Cálcio/metabolismo , Bovinos , Linhagem Celular Tumoral , Respiração Celular/efeitos dos fármacos , Complexo I de Transporte de Elétrons/metabolismo , Complexo II de Transporte de Elétrons/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/fisiologia , RatosRESUMO
Cancer immunotherapy such as anti-PD-1/anti-PD-L1 immune checkpoint blockade (ICB) can provide significant clinical benefit in patients with advanced malignancies. However, most patients eventually develop progressive disease, thus necessitating additional therapeutic options. We have developed a novel agent, a-TEA-LS, that selectively induces tumor cell death while sparing healthy tissues, leading to increased activation of tumor-reactive T cells and tumor regression. In the current study, we explored the impact of combined a-TEA-LS + ICB in orthotopic and spontaneously arising murine models of mammary carcinoma. We found that a-TEA-LS + ICB led to increased production of pro-inflammatory cytokines that were associated with a reduction in tumor growth and prolonged survival. Together, these data demonstrate the potential utility of a-TEA-LS + ICB for the treatment of breast cancer and provide the rationale for clinical translation of this novel approach.
Assuntos
Neoplasias da Mama , Inibidores de Checkpoint Imunológico , Humanos , Animais , Camundongos , Feminino , Imunoterapia , CitocinasRESUMO
Effective tumor immunotherapy may require not only activation of anti-tumor effector cells, but also abrogation of tumor-mediated immunosuppression. The cytokine TGF-ß, is frequently elevated in the tumor microenvironment and is a potent immunosuppressive agent and promoter of tumor metastasis. OX40 (CD134) is a member of the TNF-α receptor superfamily and ligation by agonistic antibody (anti-OX40) enhances effector function, expansion, and survival of activated T cells. In this study, we examined the therapeutic efficacy and anti-tumor immune response induced by the combination of a small molecule TGF-ß signaling inhibitor, SM16, plus anti-OX40 in the poorly immunogenic, highly metastatic, TGF-ß-secreting 4T1 mammary tumor model. Our data show that SM16 and anti-OX40 mutually enhanced each other to elicit a potent anti-tumor effect against established primary tumors, with a 79% reduction in tumor size, a 95% reduction in the number of metastatic lung nodules, and a cure rate of 38%. This positive treatment outcome was associated with a 3.2-fold increase of tumor-infiltrating, activated CD8+ T cells, an overall accumulation of CD4+ and CD8+ T cells, and an increased tumor-specific effector T cell response. Complete abrogation of the therapeutic effect in vivo following depletion of CD4+ and CD8+ T cells suggests that the anti-tumor efficacy of SM16+ anti-OX40 therapy is T cell dependent. Mice that were cured of their tumors were able to reject tumor re-challenge and manifested a significant tumor-specific peripheral memory IFN-γ response. Taken together, these data suggest that combining a TGF-ß signaling inhibitor with anti-OX40 is a viable approach for treating metastatic breast cancer.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Compostos Azabicíclicos/administração & dosagem , Carcinoma/tratamento farmacológico , Imunoterapia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Fator de Crescimento Transformador beta/antagonistas & inibidores , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Compostos Azabicíclicos/efeitos adversos , Carcinoma/patologia , Progressão da Doença , Sinergismo Farmacológico , Feminino , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Metástase Neoplásica , Transplante de Neoplasias , Receptores OX40/agonistas , Receptores OX40/imunologia , Transdução de Sinais/efeitos dos fármacos , Carga TumoralRESUMO
Alpha-tocopheryloxy acetic acid (α-TEA) is an ether derivative of vitamin E and has been shown to suppress tumor growth in various murine and human xenograft tumor models, including melanoma, breast, lung, prostate, and ovarian cancers. The purpose of this study was to assess its safety and pharmacokinetics after repeat dosing in a preclinical murine model. Male and female mice received α-TEA doses of 100, 300, or 1500 mg/kg/day by daily oral gavage for 28 days. α-TEA serum levels were determined weekly by high-performance liquid chromatography with mass spectrometric detection. After 28 days of dosing, complete blood counts were taken, blood chemistry was analyzed, and histology was performed. Pharmacokinetic parameters were determined after single dosing. There was no mortality, and we found no clinical signs of toxicity in any of the α-TEA doses tested. Histopathological evaluation of major organs (heart, lung, kidney, liver, spleen, jejunum, ileum, and cecum) revealed no significant α-TEA treatment-related lesions. Blood counts revealed low-grade anemia but no other significant differences between treatment and control groups. Blood chemistry revealed moderate liver toxicity that was dose dependent and was absent in the lowest dose group. There were no significant sex-specific differences in the toxicity profile. The half-life of orally administered α-TEA was determined to be 52 h. This is the first report comprehensively evaluating the toxicity profile of this novel anticancer drug and will facilitate the design of clinical trials to evaluate the safety and antitumor efficacy of α-TEA in patients with cancer.
Assuntos
Antineoplásicos/farmacocinética , Tocoferóis/farmacocinética , Anemia/induzido quimicamente , Animais , Antineoplásicos/toxicidade , Doença Hepática Induzida por Substâncias e Drogas , Avaliação Pré-Clínica de Medicamentos , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Tocoferóis/toxicidadeRESUMO
INTRODUCTION: α-Tocopheryloxyacetic acid (α-TEA) is a novel ether derivative of α-tocopherol that has generated interest as a chemotherapeutic agent because of its selective toxicity toward tumor cells and its ability to suppress tumor growth in various rodent and human xenograft models. We previously reported that oral α-TEA inhibited the growth of both a transplanted (4T1) and a spontaneous MMTV-PyMT mouse model of breast cancer. METHODS: Because little is known about the possible immunological mechanisms underlying the in vivo α-TEA effects, we evaluated the impact of α-TEA therapy on the immune response by characterizing immune cell populations infiltrating the tumor site. RESULTS: α-TEA treatment resulted in higher frequencies of activated T cells in the tumor microenvironment and twofold and sixfold higher ratios of CD4⺠and CD8âºT cells to regulatory T cells, respectively. This finding was correlated with an increased ability of tumor-draining lymph node cells and splenocytes from α-TEA-treated mice to secrete interferon (IFN)-γ in response to CD3 or to mediate a cytolytic response in a tumor-specific fashion, respectively. That the α-TEA-mediated antitumor effect had a T cell-dependent component was demonstrated by the partial abrogation of tumor suppression when CD4⺠and CD8⺠T cells were depleted. We also determined the intratumoral cytokine and chemokine profile and found that α-TEA treatment increased intratumoral IFN-γ levels but decreased interleukin (IL)-4 levels, suggesting a shift toward a TH1 response. In addition, α-TEA induced higher levels of the inflammatory cytokine IL-6 and the chemokine CCL5. CONCLUSIONS: Taken together, these data suggest that α-TEA treatment, in addition to its direct cytotoxic effects, enhanced the anti-tumor immune response. This study provides a better understanding of the mechanisms of action of α-TEA and its effect on the immune system and may prove useful in designing immune-stimulating strategies to boost the antitumor effects of α-TEA in breast cancer patients.
Assuntos
Antineoplásicos/farmacologia , Neoplasias Mamárias Experimentais/imunologia , Tocoferóis/farmacologia , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citocinas/imunologia , Citotoxicidade Imunológica/efeitos dos fármacos , Feminino , Estimativa de Kaplan-Meier , Linfonodos/efeitos dos fármacos , Linfonodos/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Tocoferóis/administração & dosagem , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologiaRESUMO
BACKGROUND: HER2/neu is an oncogene that facilitates neoplastic transformation due to its ability to transduce growth signals in a ligand-independent manner, is over-expressed in 20-30% of human breast cancers correlating with aggressive disease and has been successfully targeted with trastuzumab (Herceptin®). Because trastuzumab alone achieves only a 15-30% response rate, it is now commonly combined with conventional chemotherapeutic drugs. While the combination of trastuzumab plus chemotherapy has greatly improved response rates and increased survival, these conventional chemotherapy drugs are frequently associated with gastrointestinal and cardiac toxicity, bone marrow and immune suppression. These drawbacks necessitate the development of new, less toxic drugs that can be combined with trastuzumab. Recently, we reported that orally administered alpha-tocopheryloxyacetic acid (α-TEA), a novel ether derivative of alpha-tocopherol, dramatically suppressed primary tumor growth and reduced the incidence of lung metastases both in a transplanted and a spontaneous mouse model of breast cancer without discernable toxicity. METHODS: In this study we examined the effect of α-TEA plus HER2/neu-specific antibody treatment on HER2/neu-expressing breast cancer cells in vitro and in a HER2/neu positive human xenograft tumor model in vivo. RESULTS: We show in vitro that α-TEA plus anti-HER2/neu antibody has an increased cytotoxic effect against murine mammary tumor cells and human breast cancer cells and that the anti-tumor effect of α-TEA is independent of HER2/neu status. More importantly, in a human breast cancer xenograft model, the combination of α-TEA plus trastuzumab resulted in faster tumor regression and more tumor-free animals than trastuzumab alone. CONCLUSION: Due to the cancer cell selectivity of α-TEA, and because α-TEA kills both HER2/neu positive and HER2/neu negative breast cancer cells, it has the potential to be effective and less toxic than existing chemotherapeutic drugs when used in combination with HER2/neu antibody.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/uso terapêutico , Antioxidantes/farmacologia , Neoplasias da Mama/tratamento farmacológico , Tocoferóis/farmacologia , Animais , Western Blotting , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Feminino , Citometria de Fluxo , Humanos , Camundongos , Modelos Biológicos , Fragmentos de Peptídeos/metabolismo , TrastuzumabRESUMO
BACKGROUND: Transforming growth factor beta (TGF-beta) plays a complex role in breast carcinogenesis. Initially functioning as a tumor suppressor, this cytokine later contributes to the progression of malignant cells by enhancing their invasive and metastatic potential as well as suppressing antitumor immunity. The purpose of this study was to investigate the efficacy of SM16, a novel small molecule ALK5 kinase inhibitor, to treat a highly metastatic, TGF-beta-producing murine mammary carcinoma (4T1). MATERIALS AND METHODS: Mice bearing established 4T1 tumors were treated with SM16 intraperitoneally (i.p.) or orally, and primary and metastatic tumor growth was assessed. RESULTS: SM16 inhibited Smad2 phosphorylation in cultured 4T1 tumor cells as well as primary and metastatic 4T1 tumor tissue. Blockade of TGF-beta signal transduction in 4T1 tumor cells by SM16 prevented TGF-beta-induced morphological changes and inhibited TGF-beta-induced invasion in vitro. When delivered via daily i.p. injection or orally through mouse chow, SM16 inhibited the growth of primary and metastatic 4T1 tumors. Splenocytes isolated from mice on the SM16 diet displayed enhanced IFN-gamma production and antitumor CTL activity. Furthermore, SM16 failed to inhibit the growth and metastasis of established 4T1 tumors in immunodeficient SCID mice. CONCLUSION: Taken together, the data indicate that the antitumor efficacy of SM16 is dependent on an immune-mediated mechanism and that SM16 may represent a safe and effective treatment for metastatic breast cancer.
Assuntos
Compostos Azabicíclicos/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Administração Oral , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Imunofluorescência , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Nus , Proteínas Serina-Treonina Quinases/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Tumors employ strategies to escape immune control. The principle aim of most cancer immunotherapies is to restore effective immune surveillance. Among the different processes regulating immune escape, tumor microenvironment-associated soluble factors, and/or cell surface-bound molecules are mostly responsible for dysfunctional activity of tumor-specific CD8+T cells. These dynamic immunosuppressive networks prevent tumor rejection at several levels, limiting also the success of immunotherapies. Nevertheless, the recent clinical development of immune checkpoint inhibitors or of molecules modulating cellular targets and immunosuppressive enzymes highlights the great potential of approaches based on the selective disruption of immunosuppressive networks. Currently, the administration of different categories of immunotherapy in combination regimens is the ultimate modality for impacting the survival of cancer patients. With the advent of immune checkpoint inhibitors, designed to mount an effective antitumor immune response, profound changes occurred in cancer immunotherapy: from a global stimulation of the immune system to a specific targeting of an immune component. This review will specifically highlight the players, the mechanisms limiting an efficient antitumor response and the current immunotherapy modalities tailored to target immune suppressive pathways. We also discuss the ongoing challenges encountered by these strategies and provide suggestions for circumventing hurdles to new immunotherapeutic approaches, including the use of relevant biomarkers in the optimization of immunotherapy regimens and the identification of patients who can benefit from defined immune-based approaches.
RESUMO
Mitochondria have recently emerged as new and promising targets for cancer prevention and therapy. One of the reasons for this is that mitochondria are instrumental to many types of cell death and often lie downstream from the initial actions of anti-cancer drugs. Unlike the tumour suppressor gene encoding p53 that is notoriously prone to inactivating mutations but whose function is essential for induction of apoptosis by DNA-targeting agents (such as doxorubicin or 5-fluorouracil), mitochondria present targets that are not so compromised by genetic mutation and whose targeting overcomes problems with mutations of upstream targets such as p53. We have recently proposed a novel class of anti-cancer agents, mitocans that exert their anti-cancer activity by destabilising mitochondria, promoting the selective induction of apoptotic death in tumour cells. In this communication, we review recent findings on mitocans and propose a common basis for their mode of action in inducing apoptosis of cancer cells. We use as an example the analogues of vitamin E that are proving to be cancer cell-specific and may soon be developed into efficient anti-cancer drugs.
Assuntos
Antineoplásicos/farmacologia , Mitocôndrias/efeitos dos fármacos , Vitamina E/análogos & derivados , Animais , Apoptose/efeitos dos fármacos , Humanos , Transdução de Sinais/efeitos dos fármacosRESUMO
Dendritic cell (DC)-based vaccines have exhibited minimal effectiveness in treating established tumors, likely because of factors present in the tumor microenvironment. One such factor is transforming growth factor beta (TGF-beta), a cytokine that is produced by numerous tumor types and has been demonstrated to impair DC functions in vitro. We have evaluated the effect of TGF-beta on the immunostimulatory activities of DCs. We demonstrate that TGF-beta exposure inhibits the ability of DCs to present antigen, stimulate tumor-sensitized T lymphocytes, and migrate to draining lymph nodes. Neutralization of TGF-beta using the TGF-beta-neutralizing monoclonal antibody 2G7 enhanced the ability of DC vaccines to inhibit the growth of established 4T1 murine mammary tumors. Treatment of 4T1 tumors transduced with the antisense TGF-beta transgene (4T1-asT) with the combination of DC and 2G7 monoclonal antibody inhibited tumor growth and resulted in complete regression of tumors in 40% of the mice. These results demonstrate that neutralization of TGF-beta in tumor-bearing mice enhances the efficacy of DC-based vaccines.
Assuntos
Anticorpos Monoclonais/farmacologia , Apresentação de Antígeno/efeitos dos fármacos , Vacinas Anticâncer/imunologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Fator de Crescimento Transformador beta/farmacologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Apresentação de Antígeno/imunologia , Antígenos CD40/metabolismo , Movimento Celular/efeitos dos fármacos , Movimento Celular/imunologia , DNA Antissenso/genética , DNA Antissenso/farmacologia , Feminino , Imunoterapia Adotiva , Linfonodos/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Neoplasias Mamárias Experimentais/imunologia , Neoplasias Mamárias Experimentais/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Linfócitos T/imunologia , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/imunologiaRESUMO
In preclinical tumor models, αOX40 therapy is often successful at treating small tumors, but is less effective once the tumors become large. For a tumor immunotherapy to be successful to cure large tumors, it will most likely require not only an agonist to boost effector T-cell function but also inhibitors of T-cell suppression. In this study, we show that combining αOX40 antibodies with an inhibitor of the TGFß receptor (SM16) synergizes to elicit complete regression of large established MCA205 and CT26 tumors. Evaluation of tumor-infiltrating T cells showed that SM16/αOX40 dual therapy resulted in an increase in proliferating granzyme B(+) CD8 T cells, which produced higher levels of IFNγ, compared with treatment with either agent alone. We also found that the dual treatment increased pSTAT3 expression in both CD4 and CD8 T cells isolated from tumors. Because others have published that STAT3 signaling is detrimental to T-cell function within the tumor microenvironment, we explored whether deletion of STAT3 in OX40-expressing cells would affect this potent combination therapy. Surprisingly, we found that deletion of STAT3 in OX40-expressing cells decreased the efficacy of this combination therapy, showing that the full therapeutic potential of this treatment depends on STAT3 signaling, most likely in the T cells of tumor-bearing mice.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias/metabolismo , Receptores OX40/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Fator de Transcrição STAT3/metabolismo , Animais , Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Compostos Azabicíclicos/administração & dosagem , Linhagem Celular Tumoral , Feminino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Receptores OX40/imunologia , Transdução de SinaisRESUMO
Stimulation of apoptosis has been reported as the primary mechanism of tumor cell death induced by alpha-tocopheryloxyacetic acid (α-TEA), an esterase-resistant, semi-synthetic derivative of vitamin E (R-R-R-α-tocopherol). New information now shows that α-TEA also triggers tumor cell autophagy and promotes antigen cross-presentation. Autophagosome-enriched fractions of α-TEA-treated tumor cells (α-TAGS) efficiently cross-primed antigen-specific CD8 (+) T cells and vaccination with dendritic cells (DC) pulsed with α-TAGS reduced lung metastases and increased survival of tumor-bearing mice. Taken together, these observations suggest that both autophagy and apoptosis signaling programs are activated in tumor cells by α-TEA treatment and may contribute to tumor cell death. We propose that autophagy-dependent enhancement of cross-presentation is a novel mechanism of α-TEA-mediated tumor immunity and that α-TEA can be exploited as an adjuvant to enhance the antitumor response.
Assuntos
Apresentação de Antígeno , Apoptose , Autofagia/fisiologia , Regulação Neoplásica da Expressão Gênica , Tocoferóis/farmacologia , Animais , Autofagia/efeitos dos fármacos , Linfócitos T CD8-Positivos/citologia , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , Neoplasias/imunologia , Neoplasias/patologia , Tocoferóis/química , Vitamina E/análogos & derivadosRESUMO
The semisynthetic vitamin E derivative alpha-tocopheryloxyacetic acid (α-TEA) induces tumor cell apoptosis and may offer a simple adjuvant supplement for cancer therapy if its mechanisms can be better understood. Here we report that α-TEA also triggers tumor cell autophagy and that it improves cross-presentation of tumor antigens to the immune system. α-TEA stimulated both apoptosis and autophagy in murine mammary and lung cancer cells and inhibition of caspase-dependent apoptosis enhanced α-TEA-induced autophagy. Cell exposure to α-TEA generated double-membrane-bound vesicles indicative of autophagosomes, which efficiently cross-primed antigen-specific CD8(+) T cells. Notably, vaccination with dendritic cells pulsed with α-TEA-generated autophagosomes reduced lung metastases and increased the survival of tumor-bearing mice. Taken together, our findings suggest that both autophagy and apoptosis signaling programs are activated during α-TEA-induced tumor cell killing. We suggest that the ability of α-TEA to stimulate autophagy and enhance cross-priming of CD8(+) T cells might be exploited as an adjuvant strategy to improve stimulation of antitumor immune responses.
Assuntos
Adenocarcinoma/tratamento farmacológico , Apresentação de Antígeno/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Apresentação Cruzada/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Mamárias Animais/tratamento farmacológico , Tocoferóis/farmacologia , Adenocarcinoma/imunologia , Adenocarcinoma de Pulmão , Animais , Apoptose/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Sobrevivência Celular , Células Dendríticas/imunologia , Feminino , Neoplasias Pulmonares/imunologia , Ativação Linfocitária , Neoplasias Mamárias Animais/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , VacinaçãoAssuntos
Autofagia , Bioensaio/normas , Animais , Autofagia/fisiologia , Bioensaio/métodos , Simulação por Computador , HumanosRESUMO
We recently demonstrated the antitumor efficacy of orally administered alpha-tocopheryloxyacetic acid (alpha-TEA), a redox silent and nonhydrolyzable derivative of naturally occurring vitamin E. In order to move alpha-TEA closer to the clinic to benefit patients with breast cancer, the present study had two goals. First, to determine the minimal effective treatment dose; and second, to test the efficacy of dietary administration of alpha-TEA in the clinically relevant MMTV-PyMT mouse model of spontaneous breast cancer that more closely resembles human disease. The minimal effective dose of alpha-TEA was evaluated in the transplantable 4T1 tumor model and we show a dose-dependent decrease of primary tumor growth and reduction of metastatic spread to the lung. Six-week-old MMTV-PyMT mice were treated with oral alpha-TEA for 9 weeks, with no apparent signs of drug toxicity. The alpha-TEA treatment delayed tumor development and significantly slowed tumor progression, resulting in a 6-fold reduction of the average cumulative tumor size. In addition, oral alpha-TEA caused an 80% reduction in spontaneous metastases. In situ analysis of tumor tissue identified apoptosis as an important mechanism of alpha-TEA-mediated tumor suppression in addition to inhibition of tumor cell proliferation. This study shows, for the first time, the ability of orally administered alpha-TEA to delay tumor onset and to inhibit the progression and metastatic spread of a clinically relevant model of spontaneous breast cancer. Our finding of the high efficacy in this tumor model highlights the translational potential of oral alpha-TEA therapy.
Assuntos
Proliferação de Células/efeitos dos fármacos , Neoplasias Mamárias Experimentais/tratamento farmacológico , Tocoferóis/farmacologia , Carga Tumoral/efeitos dos fármacos , Administração Oral , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Feminino , Marcação In Situ das Extremidades Cortadas , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Tocoferóis/administração & dosagem , Tocoferóis/sangue , Vitaminas/administração & dosagem , Vitaminas/farmacologiaRESUMO
A bigenic MUC1.Tg/MIN mouse model was developed by crossing Apc/(MIN/+) (MIN) mice with human MUC1 transgenic mice to evaluate MUC1 antigen-specific immunotherapy of intestinal adenomas. The MUC1.Tg/MIN mice developed adenomas at a rate comparable to that of MIN mice and had similar levels of serum MUC1 antigen. A MUC1-based vaccine consisting of MHC class I-restricted MUC1 peptides, a MHC class II-restricted pan-helper peptide, unmethylated CpG oligodeoxynucleotide and GM-CSF caused flattening of adenomas and significantly reduced the number of large adenomas. Immunization was successful in generating a MUC1-directed immune response evidenced by increased MUC1 peptide-specific anti-tumor cytotoxicity and IFN-gamma secretion by lymphocytes.
Assuntos
Adenoma/terapia , Vacinas Anticâncer , Modelos Animais de Doenças , Neoplasias Intestinais/terapia , Mucina-1 , Fragmentos de Peptídeos , Adenoma/imunologia , Adenoma/patologia , Adjuvantes Imunológicos , Sequência de Aminoácidos , Animais , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Humanos , Imunoterapia , Neoplasias Intestinais/imunologia , Neoplasias Intestinais/patologia , Masculino , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Mucina-1/administração & dosagem , Mucina-1/química , Mucina-1/genética , Mucina-1/imunologia , Oligodesoxirribonucleotídeos/administração & dosagem , Oligodesoxirribonucleotídeos/imunologia , Fragmentos de Peptídeos/administração & dosagem , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , VacinaçãoRESUMO
"Mitocans" from the vitamin E group of selective anticancer drugs, alpha-tocopheryl succinate (alpha-TOS) and its ether analogue alpha-TEA, triggered apoptosis in proliferating but not arrested endothelial cells. Angiogenic endothelial cells exposed to the vitamin E analogues, unlike their arrested counterparts, readily accumulated reactive oxygen species (ROS) by interfering with the mitochondrial redox chain and activating the intrinsic apoptotic pathway. The vitamin E analogues inhibited angiogenesis in vitro as assessed using the "wound-healing" and "tube-forming" models. Endothelial cells deficient in mitochondrial DNA (mtDNA) were resistant to the vitamin E analogues, both in ROS accumulation and apoptosis induction, maintaining their angiogenic potential. alpha-TOS inhibited angiogenesis in a mouse cancer model, as documented by ultrasound imaging. We conclude that vitamin E analogues selectively kill angiogenic endothelial cells, suppressing tumor growth, which has intriguing clinical implications.
Assuntos
Antineoplásicos/uso terapêutico , Mitocôndrias/fisiologia , Neovascularização Patológica/prevenção & controle , Estresse Oxidativo/fisiologia , Vitamina E/análogos & derivados , Vitamina E/uso terapêutico , Inibidores da Angiogênese/uso terapêutico , Apoptose/efeitos dos fármacos , DNA Mitocondrial/genética , Resistência a Medicamentos , Endotélio Vascular , Humanos , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacosRESUMO
Alpha tocopheryl succinate (alpha-TOS) is a non-toxic vitamin E analog under study for its anti-cancer properties. In an earlier study, we showed that alpha-TOS, when used in combination with non-matured dendritic cells (nmDC) to treat pre-established tumors, acts as an effective adjuvant. In this study, we have used vesiculated alpha-TOS (Valpha-TOS), a more soluble form of alpha-TOS that is relevant for clinical use, in combination with dendritic cells to treat pre-established murine tumors. We demonstrate that Valpha-TOS kills tumor cells in vitro and inhibits the growth of pre-established murine lung carcinoma (3LLD122) as effectively as alpha-TOS. The combination of Valpha-TOS plus non-matured or TNF-alpha-matured DC is more effective at inhibiting the growth of established tumors than Valpha-TOS alone. We also observed that Valpha-TOS induces expression of heat shock proteins in tumor cells and that co-incubation of non-matured DC with lysate derived from Valpha-TOS-treated tumor cells leads to DC maturation evidenced by up-regulation of co-stimulatory molecules and secretion of IL-12p70. This study therefore demonstrates the immunomodulatory properties of Valpha-TOS that may account for its adjuvant effect when combined with DC vaccines to treat established tumors.