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1.
Int J Nanomedicine ; 14: 5527-5540, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31413561

RESUMO

Background: Nonspecific tumor targeting, potential relapse and metastasis of tumor after treatment are the main barriers in clinical photodynamic therapy (PDT) for cancer, hence, inhibiting relapse and metastasis of tumor is significant issues in clinic. Purpose: In this work, chidamide as a histone deacetylases inhibitor (HADCi) was bound onto a pH-responsive block polymer folate polyethylene glycol-b-poly(aspartic acid) (PEG-b-PAsp) grafted folate (FA-PEG-b-PAsp) to obtain the block polymer folate polyethylene glycol-b-poly(asparaginyl-chidamide) (FA-PEG-b-PAsp-chidamide, FPPC) as multimodal tumor-targeting drug-delivery carrier to inhibiting tumor cell proliferation and tumor metastasis in mice. Methods: Model photosensitizer pyropheophorbide-a (Pha) was encapsulated by FPPC in PBS to form the polymer micelles Pha@FPPC [folate polyethylene glycol-b-poly(asparaginyl-chidamide) micelles encapsulating Pha]. Pha@FPPC was characterized by transmission electron microscope and dynamic light scattering; also, antitumor activity in vivo and in vitro were investigated by determination of cellular ROS level, detection of cell apoptosis and cell cycle arrest, PDT antitumor activity in vivo and histological analysis. Results: With favorable and stable sphere morphology under transmission electron microscope (TEM) (~93.0 nm), Pha@FPPC greatly enhanced the cellular uptake due to its folate-mediated effective endocytosis by mouse melanoma B16-F10 cells and the yield of ROS in tumor cells induced by PDT, and mainly caused necrocytosis and blocked cell growth cycle not only in G2 phase but also in G1/G0 phase after PDT. Pha@FPPC exhibited lower dark cytotoxicity in vitro and a better therapeutic index because of its higher dark cytotoxicity/photocytotoxicity ratio. Moreover, Pha@FPPC not only significantly inhibited the growth of implanted tumor and prolonged the survival time of melanoma-bearing mice due to both its folate-mediated tumor-targeting and selectively accumulation at tumor site by EPR (enhanced permeability and retention)effect as micelle nanoparticles but also remarkably prevented pulmonary metastasis of mice melanoma after PDT compared to free Pha, demonstrating its dual antitumor characteristics of PDT and HDACi. Conclusion: As a folate-mediated and acid-activated chidamide-grafted drug-delivery carrier, FPPC may have great potential to inhibit tumor metastasis in clinical photodynamic treatment for cancer because of its effective and multimodal tumor-targeting performance as photosensitizer vehicle.


Assuntos
Aminopiridinas/química , Benzamidas/química , Ácido Fólico/uso terapêutico , Micelas , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Clorofila/análogos & derivados , Clorofila/farmacologia , Sistemas de Liberação de Medicamentos , Liberação Controlada de Fármacos , Endocitose/efeitos dos fármacos , Ácido Fólico/farmacologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Nanopartículas/química , Nanopartículas/ultraestrutura , Fármacos Fotossensibilizantes/farmacologia , Polietilenoglicóis/química , Espécies Reativas de Oxigênio/metabolismo
2.
Nat Commun ; 10(1): 2678, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31213601

RESUMO

Myeloid cells contribute to tumor progression, but how the constellation of receptors they express regulates their functions within the tumor microenvironment (TME) is unclear. We demonstrate that Fcmr (Toso), the putative receptor for soluble IgM, modulates myeloid cell responses to cancer. In a syngeneic melanoma model, Fcmr ablation in myeloid cells suppressed tumor growth and extended mouse survival. Fcmr deficiency increased myeloid cell population density in this malignancy and enhanced anti-tumor immunity. Single-cell RNA sequencing of Fcmr-deficient tumor-associated mononuclear phagocytes revealed a unique subset with enhanced antigen processing/presenting properties. Conversely, Fcmr activity negatively regulated the activation and migratory capacity of myeloid cells in vivo, and T cell activation by bone marrow-derived dendritic cells in vitro. Therapeutic targeting of Fcmr during oncogenesis decreased tumor growth when used as a single agent or in combination with anti-PD-1. Thus, Fcmr regulates myeloid cell activation within the TME and may be a potential therapeutic target.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteínas de Transporte/metabolismo , Melanoma Experimental/imunologia , Proteínas de Membrana/metabolismo , Monócitos/imunologia , Neoplasias Cutâneas/imunologia , Animais , Apresentação do Antígeno/efeitos dos fármacos , Apresentação do Antígeno/imunologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinogênese/efeitos dos fármacos , Carcinogênese/imunologia , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/genética , Proteínas de Transporte/imunologia , Linhagem Celular Tumoral/transplante , Movimento Celular/efeitos dos fármacos , Movimento Celular/imunologia , Feminino , Ativação Linfocitária/imunologia , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/mortalidade , Melanoma Experimental/patologia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Análise de Sobrevida , Linfócitos T/imunologia , Microambiente Tumoral/imunologia
3.
Int J Nanomedicine ; 14: 3455-3468, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31190803

RESUMO

Background: Phospholysine phosphohistidine inorganic pyrophosphate phosphatase (LHPP) is a novel tumor suppressor. However, whether LHPP is effective to melanoma has not been investigated. Gene therapy provides a new strategy for the treatment of melanoma. Currently, it suffers from the lack of safe and effective gene delivery systems. Methods: A CRGDKGPDC peptide (iRGD) modified hybrid monomethoxy poly(ethylene glycol)-poly(D,L-lactide) nanoparticle (iDPP) was prepared and complexed with a LHPP plasmid, forming an iDPP/LHPP nanocomplex. The iDPP/LHPP nanocomplex was characterized by particle size distribution, zeta potential, morphology, cytotoxicity, and transfection efficiency. The antitumor efficacy of the nanocomplex against melanoma was studied both in vitro and in vivo. Further, the potential epigenetic changes in melanoma induced by iDPP/LHPP nanocomplex were evaluated. Results: The iDPP/LHPP nanocomplex showed high transfection efficiency and low toxicity. Moreover, the nanocomplex displayed a neutral charge that can meet the requirement of intravenous injection for targeted gene therapy. In vitro and in vivo experiments indicated that the iDPP/LHPP nanocomplex significantly inhibited the melanoma growth without causing notable adverse effects. We also found that LHPP played an important role in epigenetics. It regulated the expression of genes related to the proliferation and apoptosis chiefly at the level of transcription. Conclusion: This work demonstrates that the iDPP nanoparticle-delivered LHPP gene has a potential application in melanoma therapy through regulation of the genes associated with epigenetics.


Assuntos
Pirofosfatase Inorgânica/uso terapêutico , Melanoma Experimental/tratamento farmacológico , Nanopartículas/química , Animais , Antineoplásicos/farmacologia , Apoptose , Linhagem Celular Tumoral , Epigênese Genética/efeitos dos fármacos , Humanos , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Nanopartículas/ultraestrutura , Oligopeptídeos/química , Especificidade de Órgãos/efeitos dos fármacos , Poliésteres/química , Polietilenoglicóis/química
4.
J Photochem Photobiol B ; 197: 111533, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31254952

RESUMO

Melanoma is one of the most lethal tumors among the skin cancers, arising from complex genetic mutations in melanocyte. Melanoma microenvironment is very heterogeneous, showing complex vascular networks and immunogenicity, as well as induced acquired resistance to treatments by upregulation of multidrug resistance (MDR) mechanisms. Different studies have showed that Photodynamic Therapy (PDT) could be considered a new potential approach for melanoma treatment. PDT combines a light with a specific wavelength and a photosensitizer: when these two elements interact reactive oxygen species (ROS) are generated leading to tumor cell destruction. In this study verteporfin (Ver), a second-generation photosensitizer, has been conjugated with mesoporous silica nanoparticles (MSNs): the resulting Ver-MSNs are an efficient nanoplatforms used to enhance cargo capacity and cellular uptake. Our in vitro and in vivo studies investigated whether Ver-MSNs were able to reduce or inhibit melanoma growth. In vitro experiments performed using B16F10 mouse melanoma cells showed that Ver-MSNs stimulated by red light (693 nm) significantly decreased in vitro cells proliferation in a range of concentration between 0.1 µg/ml to 10 µg/ml. When Ver-MSNs (5 µg/ml in glycerol) were topically administrated to melanoma tumor mass developed in mice and stimulated by red light for four times in 16 days, they were able to reduce the tumor mass of 50.2 ±â€¯6,6% compared to the untreated (only glycerol) mice. In the light of this information, PDT performed using Ver-MSNs could be considered a new promising and potential approach to treat melanoma.


Assuntos
Melanoma Experimental/tratamento farmacológico , Nanopartículas Metálicas/química , Dióxido de Silício/química , Verteporfina/química , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Portadores de Fármacos/química , Feminino , Luz , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Fotoquimioterapia , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/uso terapêutico , Porosidade , Espécies Reativas de Oxigênio/metabolismo , Verteporfina/uso terapêutico
5.
Mar Drugs ; 17(5)2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31035725

RESUMO

Melanoma is one of the most malignant and aggressive types of cancer worldwide. Fibroblast growth factor 2 (FGF2) is one of the critical regulators of melanoma angiogenesis and metastasis; thus, it might be an effective anti-cancer strategy to explore FGF2-targeting drug candidates from existing drugs. In this study, we evaluate the effect of the marine drug propylene glycol alginate sodium sulfate (PSS) on FGF2-mediated angiogenesis and invasion. The data shows that FGF2 selectively bound to PSS with high affinity. PSS inhibited FGF2-mediated angiogenesis in a rat aortic ring model and suppressed FGF2-mediated invasion, but not the migration of murine melanoma B16-F10 cells. The further mechanism study indicates that PSS decreased the expression of activated matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9), and also suppressed their activity. In addition, PSS was found to decrease the level of Vimentin in B16-F10 cells, which is known to participate in the epithelial-mesenchymal transition. Notably, PSS did not elicit any changes in cancer cell viability. Based on the results above, we conclude that PSS might be a potential drug to regulate the tumor microenvironment in order to facilitate the recovery of melanoma patients.


Assuntos
Alginatos/farmacologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Melanoma Experimental/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Neoplasias Cutâneas/tratamento farmacológico , Alginatos/uso terapêutico , Animais , Aorta/efeitos dos fármacos , Organismos Aquáticos/química , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Embrião de Galinha , Membrana Corioalantoide , Avaliação Pré-Clínica de Medicamentos , Transição Epitelial-Mesenquimal , Humanos , Laminaria/química , Melanoma Experimental/irrigação sanguínea , Melanoma Experimental/patologia , Camundongos , Invasividade Neoplásica/patologia , Invasividade Neoplásica/prevenção & controle , Neovascularização Patológica/patologia , Neovascularização Fisiológica/efeitos dos fármacos , Técnicas de Cultura de Órgãos , Ratos , Neoplasias Cutâneas/irrigação sanguínea , Neoplasias Cutâneas/patologia , Microambiente Tumoral/efeitos dos fármacos
6.
Cancer Immunol Immunother ; 68(7): 1095-1106, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31104075

RESUMO

Checkpoint blockade immunotherapy is now a first-line treatment option for patients with melanoma. Despite achieving objective responses in about half of patients, the exact immune mechanisms elicited and those required for therapeutic success have not been clearly identified. Insight into these mechanisms is key for improving outcomes in a broader range of cancer patients. We used a murine melanoma model to track responses by different subsets of tumor-infiltrating lymphocytes (TIL) during checkpoint blockade immunotherapy. Tumors from treated mice had increased frequencies of both CD4+ and CD8+ T cells, which also showed evidence of functional reinvigoration and elevated effector cytokine production after immunotherapy. We predicted that increased T cell numbers and function within tumors reflected either infiltration by new T cells or clonal expansion by a few high-affinity tumor-reactive T cells. To address this, we compared TIL diversity before and after immunotherapy by sequencing the complementarity determining region 3 (CDR3) of all T cell receptor beta (TCRß) genes. While checkpoint blockade effectively slowed tumor progression and increased T cell frequencies, the diversity of intratumoral T cells remained stable. This was true when analyzing total T cells and when focusing on smaller subsets of effector CD4+ and CD8+ TIL as well as regulatory T cells. Our study suggests that checkpoint blockade immunotherapy does not broaden the T cell repertoire within murine melanoma tumors, but rather expands existing T cell populations and enhances effector capabilities.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Melanoma Experimental/tratamento farmacológico , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Neoplasias Cutâneas/tratamento farmacológico , Animais , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Antígeno CTLA-4/antagonistas & inibidores , Linhagem Celular Tumoral , Regiões Determinantes de Complementaridade/genética , Humanos , Imunoterapia/métodos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Resultado do Tratamento , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia
7.
Nanomedicine (Lond) ; 14(9): 1077-1093, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31050580

RESUMO

Aim: The present study was aimed at determining the antiproliferative, antioxidant, anti-inflammatory and antitumor activity of developed silymarin-nanostructured lipid carrier (NLC) gel. Materials & methods: B16 melanoma cell line and albino mice were used as ex vivo and in vivo models, respectively, to evaluate the aforementioned pharmacological activities. Results: The volume of large tumors significantly (p < 0.05) reduced from 5.02 to 3.05 mm3, levels of IL-1α and TNF-α were significantly (p < 0.001) lower and levels of superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) significantly (p < 0.0001) increased in the group treated with silymarin-NLC gel. Furthermore, in skin treated with placebo and conventional gels, a basosquamous carcinoma and squamous cell carcinoma were noticed, respectively. Conclusion: Silymarin-NLC gel presented better treatment outcomes compared with silymarin-conventional gel.


Assuntos
Antineoplásicos/uso terapêutico , Lipídeos/química , Melanoma Experimental/tratamento farmacológico , Nanocápsulas/química , Silimarina/uso terapêutico , Neoplasias Cutâneas/tratamento farmacológico , Animais , Antineoplásicos/administração & dosagem , Carcinogênese/efeitos dos fármacos , Carcinogênese/patologia , Catalase/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular/efeitos dos fármacos , Feminino , Géis , Glutationa/metabolismo , Humanos , Masculino , Malondialdeído/metabolismo , Melanoma Experimental/patologia , Camundongos , Silimarina/administração & dosagem , Neoplasias Cutâneas/patologia , Superóxido Dismutase/metabolismo , Resultado do Tratamento
8.
Medicina (Kaunas) ; 55(4)2019 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-30935124

RESUMO

Background and objectives: Cancer represents the miscommunication between and within the body cells. The mutations of the oncogenes encoding the MAPK pathways play an important role in the development of tumoral diseases. The mutations of KRAS and BRAF oncogenes are involved in colorectal cancer and melanoma, while the NRAS mutations are associated with melanoma. Thiazolidine-2,4-dione is a versatile scaffold in medicinal chemistry and a useful tool in the development of new antitumoral compounds. The aim of our study was to predict the pharmacokinetic/pharmacodynamic properties, the drug-likeness and lead-likeness of two series of synthetic 5-arylidene(chromenyl-methylene)-thiazolidinediones, the molecular docking on the oncoproteins K-Ras, N-Ras and B-Raf, and to investigate the cytotoxicity of the compounds, in order to select the best structural profile for potential anticancer agents. Materials and Methods: In our paper we studied the cytotoxicity of two series of thiazolidine-2,4-dione derivatives, their ADME-Tox properties and the molecular docking on a mutant protein of K-Ras, two isoforms of N-Ras and an isoform of B-Raf with 16 mutations. Results: The heterocyclic compounds strongly interact with K-Ras and N-Ras right after their posttranslational processing and/or compete with GDP for the nucleotide-binding site of the two GTPases. They are less active against the GDP-bound states of the two targets. All derivatives have a similar binding pattern in the active site of B-Raf. Conclusions: The data obtained encourage the further investigation of the 5-arylidene(chromenyl-methylene)-thiazolidinediones as potential new agents against the oncoproteins K-Ras, N-Ras and B-Raf.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Melanoma/tratamento farmacológico , Proteína Oncogênica p21(ras)/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Neoplasias Cutâneas/tratamento farmacológico , Tiazolidinedionas/química , Tiazolidinedionas/uso terapêutico , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Descoberta de Drogas , GTP Fosfo-Hidrolases/química , Guanosina Difosfato/química , Humanos , Melanoma Experimental/tratamento farmacológico , Camundongos , Simulação de Acoplamento Molecular/métodos , Mutação , Proteína Oncogênica p21(ras)/genética , Ligação Proteica , Estrutura Secundária de Proteína , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Tiazolidinedionas/síntese química
9.
Pharmazie ; 74(3): 163-167, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30961683

RESUMO

The natural phytoalexin resveratrol (RES) has exhibited excellent anti-tumor effects on a variety of tumors including malignant melanoma. However, its specific mechanism of anti-melanoma needs to be further explored. It has been reported, that the expression of tumor suppressor gene RUNX3 was lost or substantially decreased in melanoma. Whether RES exerts its anti-tumor effect by regulating the expression of RUNX3 gene in melanoma is worthy of study. In the present study, we found the RUNX3 promoter is hypermethylated and the expression of RUNX3 mRNA and protein are absent in melanoma cells B16F10. After intervention with RES, promoter hypermethylation of RUNX3 in B16F10 cells could be significantly decreased and mRNA and protein expression of it was upregulated in a dose-dependent manner. We further investigated the effects of RES on B16F10 xenograft models. The intervention of RES and treatment of melanoma positive drug dacarbazine (DTIC) both could significantly inhibit tumor growth in xenograft mice, but only RES could upregulate the expression of RUNX3 mRNA and protein in peripheral blood and tumor tissues. Therefore, the upregulation of mRNA and protein expression of RUNX3 resulting from promoter demethylation might be one of the mechanisms of RES inhibiting melanoma. This research has revealed a novel mechanism for RES against melanoma from the epigenetic perspective, which is helpful to improve the understanding of the anti-tumor mechanism of RES and provide new insights for the treatment of melanoma.


Assuntos
Subunidade alfa 3 de Fator de Ligação ao Core/genética , Desmetilação do DNA/efeitos dos fármacos , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/genética , Resveratrol/farmacologia , Animais , Antineoplásicos Fitogênicos/farmacologia , Linhagem Celular Tumoral , Feminino , Crescimento/efeitos dos fármacos , Xenoenxertos , Masculino , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Regulação para Cima/efeitos dos fármacos
10.
Cancer Sci ; 110(6): 1974-1986, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31012976

RESUMO

We previously found that circulating ß2 -glycoprotein I inhibits human endothelial cell migration, proliferation, and angiogenesis by diverse mechanisms. In the present study, we investigated the antitumor activities of ß2 -glycoprotein I using structure-function analysis and mapped the critical region within the ß2 -glycoprotein I peptide sequence that mediates anticancer effects. We constructed recombinant cDNA and purified different ß2 -glycoprotein I polypeptide domains using a baculovirus expression system. We found that purified ß2 -glycoprotein I, as well as recombinant ß2 -glycoprotein I full-length (D12345), polypeptide domains I-IV (D1234), and polypeptide domain I (D1) significantly inhibited melanoma cell migration, proliferation and invasion. Western blot analyses were used to determine the dysregulated expression of proteins essential for intracellular signaling pathways in B16-F10 treated with ß2 -glycoprotein I and variant recombinant polypeptides. Using a melanoma mouse model, we found that D1 polypeptide showed stronger potency in suppressing tumor growth. Structural analysis showed that fragments A and B within domain I would be the critical regions responsible for antitumor activity. Annexin A2 was identified as the counterpart molecule for ß2 -glycoprotein I by immunofluorescence and coimmunoprecipitation assays. Interaction between specific amino acids of ß2 -glycoprotein I D1 and annexin A2 was later evaluated by the molecular docking approach. Moreover, five amino acid residues were selected from fragments A and B for functional evaluation using site-directed mutagenesis, and P11A, M42A, and I55P mutations were shown to disrupt the anti-melanoma cell migration ability of ß2 -glycoprotein I. This is the first study to show the therapeutic potential of ß2 -glycoprotein I D1 in the treatment of melanoma progression.


Assuntos
Movimento Celular/efeitos dos fármacos , Melanoma Experimental/tratamento farmacológico , Peptídeos/farmacologia , beta 2-Glicoproteína I/química , Sequência de Aminoácidos , Animais , Sítios de Ligação/genética , Linhagem Celular Tumoral , Masculino , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Camundongos Endogâmicos C57BL , Simulação de Acoplamento Molecular , Mutagênese Sítio-Dirigida , Peptídeos/química , Peptídeos/metabolismo , Domínios Proteicos , Homologia de Sequência de Aminoácidos , beta 2-Glicoproteína I/genética , beta 2-Glicoproteína I/metabolismo
11.
Nanoscale ; 11(16): 7931-7943, 2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-30964937

RESUMO

Since mannose receptors (MRs) are expressed on the surfaces of dendritic cells (DCs), the most professional antigen presenting cells in our body, DNA vaccine carriers containing either covalently grafted mannosyl- or mannose-mimicking shikimoyl-ligands are being increasingly used in ex vivo DC-transfection based DNA vaccination. To this end, we have recently demonstrated that ex vivo immunization of mice with liposomes of shikimoylated cationic amphiphiles containing a 6-amino hexanoic acid spacer group in the head-group region in complexation with melanoma antigen (MART1) encoded DNA vaccine (pCMV-MART1) induces long lasting anti-melanoma immune responses (C. Voshavar, et al., J. Med. Chem., 2017, 60, 1605-1610). This finding prompted us to examine, in the present investigation, the efficacies of gold nanoparticles conjugated to the mannose-mimicking shikimoyl ligand (SL) via a 6-amino hexane thiol spacer (AuNPs-SL) for use in ex vivo DC-transfection based genetic immunization. Herein, we report on the design, synthesis, physico-chemical characterization and bioactivities of AuNPs-SL. Dynamic light scattering and transmission electron microscopy studies revealed the hydrodynamic diameters of theAuNPs-SL nanoconjugates to be within the range of 23-44 nm and their surface potentials within the range of 9-28 mV. MTT-assay showed the non-cytotoxic nature of AuNPs-SL and the findings in the electrophoretic gel retardation assays revealed strong DNA binding properties of the AuNPs-SL. Importantly, subcutaneous immunization of C57BL/6J mice with DCs ex vivo transfected with an electrostatic complex of AuNPs-SL & melanoma antigen (MART1) encoded DNA vaccine (p-CMV-MART1) induced a long lasting (100 days) anti-tumor immune response in immunized mice upon subsequent challenge with a lethal dose of melanoma. Notably, mice immunized with either autologous mbmDCs ex vivo pre-transfected with nanoplexes of shikimoylated AuNPs-SL & an irrelevant pCMV-SPORT-ß-gal plasmid (without having encoded melanoma antigen) or untransfected DCs showed no lasting protection against subsequent tumor challenge. The presently described shikimoyl-decorated gold nanoparticles (AuNPs-SL) are expected to find future use in ex vivo DC-transfection based genetic immunization against cancer and other infectious diseases.


Assuntos
Vacinas Anticâncer/imunologia , Ouro/química , Nanopartículas Metálicas/química , Nanoestruturas/química , Safrol/química , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Dendríticas/citologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Feminino , Imunidade Celular , Interferon gama/metabolismo , Ligantes , Antígeno MART-1/genética , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/mortalidade , Camundongos , Camundongos Endogâmicos C57BL , Nanoestruturas/uso terapêutico , Nanoestruturas/toxicidade , Plasmídeos/genética , Plasmídeos/metabolismo , Taxa de Sobrevida , Transplante Homólogo
12.
Int J Immunopathol Pharmacol ; 33: 2058738419843366, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30968715

RESUMO

Poor prognosis is associated with melanoma due to immunosuppression profiles, suggesting that immune alterations have an important role in the occurrence, growth, and metastasis of melanoma. Here, we found that PCC0208018, a small-molecule compound, enhanced T cell proliferation and activation to release interferon gamma (IFN-γ) and interleukin-2 (IL-2) without blocking the programmed cell death 1 (PD-1)/programmed cell death-ligand 1 (PD-L1) binding and did not directly affect tumor cell viability in vitro. Furthermore, PCC0208018 increased the phosphorylation of protein kinase B (PKB/AKT) as well as extracellular regulated protein kinases (ERK) in human peripheral blood mononuclear cells (PBMCs) in vitro. The secretion of cytokines induced by PCC0208018 was significantly suppressed by the PI3K inhibitor GDC-0941. In B16-F10 melanoma-harboring mice, PCC0208018 significantly inhibited tumor growth as well as increasing CD3+, CD3+CD4+, and CD3+CD8+ T cell abundance in tumors without affecting PD-L1 expression. This study showed that PCC0208018 potentially increased PBMCs proliferation and function by activating the phosphatidylinositol 3 kinase (PI3K)/AKT and mitogen-activated protein kinase (MEK)/ERK pathways to exert antitumor effects.


Assuntos
Antineoplásicos/farmacologia , Melanoma Experimental/metabolismo , Neoplasias Cutâneas/metabolismo , Linfócitos T/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Células CHO , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Células Cultivadas , Cricetinae , Cricetulus , Humanos , Masculino , Melanoma Experimental/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Cutâneas/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Pharm Res ; 36(4): 65, 2019 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-30859335

RESUMO

PURPOSE: Tumor-associated macrophages (TAMs) with immune-suppressive M2-like phenotype constitute a significant part of tumor and support its growth, thus making an attractive therapeutic target for cancer therapy. To improve the delivery of drugs that control the survival and/or functions of TAMs, we developed nanoparticulate drug carriers with high affinity for TAMs. METHODS: Poly(lactic-co-glycolic acid) nanoparticles were coated with M2pep, a peptide ligand selectively binding to M2-polarized macrophages, via a simple surface modification method based on tannic acid-iron complex. The interactions of M2pep-coated nanoparticles with macrophages of different phenotypes were tested in vitro and in vivo. PLX3397, an inhibitor of the colony stimulating factor-1 (CSF-1)/CSF-1 receptor (CSF-1R) pathway and macrophage survival, was delivered to B16F10 tumors via M2pep-modified PLGA nanoparticles. RESULTS: In bone marrow-derived macrophages polarized to M2 phenotype, M2pep-coated nanoparticles showed greater cellular uptake than those without M2pep. Consistently, M2pep-coated nanoparticles showed relatively high localization of CD206+ macrophages in B16F10 tumors. PLX3397 encapsulated in M2pep-coated nanoparticles attenuated tumor growth better than the free drug counterpart. CONCLUSION: These results support that M2pep-coating can help nanoparticles to interact with M2-like TAMs and facilitate the delivery of drugs that control the tumor-supportive functions of TAMs.


Assuntos
Macrófagos/efeitos dos fármacos , Nanopartículas/química , Peptídeos/química , Polímeros/química , Aminopiridinas/química , Aminopiridinas/farmacologia , Animais , Linhagem Celular , Linhagem Celular Tumoral , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Humanos , Masculino , Melanoma Experimental/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Pirróis/química , Pirróis/farmacologia , Células RAW 264.7
14.
Drug Deliv ; 26(1): 376-383, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30905216

RESUMO

Metformin hydrochloride (Met) is commonly used for antidiabetic therapy though its antimelanoma action is also reported. Conventional oral administration method of Met is not appropriate for therapy of melanoma because of large dose, adverse reactions, and low efficiency. Here, a transdermal Met-loaded cubic phase was developed for local treatment of melanoma. In silico formulation optimization of the cubic phases was done, and the corresponding formulations were prepared and characterized. The optimized formulations were screened based on the stable microstructure and proper fluidity. Highly efficient mouse skin permeability of Met was found with the cubic phases compared to Met solutions. High antimelanoma effect of transdermal Met-loaded cubic phases also was shown by the significant decrease of tumor volume and the improvement of melanoma cell apoptosis on the B16 melanoma mice. Met-loaded cubic phases are a promising topically applied medication for local therapies of melanoma.


Assuntos
Antineoplásicos/administração & dosagem , Melanoma Experimental/tratamento farmacológico , Metformina/administração & dosagem , Neoplasias Cutâneas/tratamento farmacológico , Administração Cutânea , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Simulação por Computador , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/farmacocinética , Hipoglicemiantes/farmacologia , Masculino , Melanoma Experimental/patologia , Metformina/farmacocinética , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Permeabilidade , Absorção Cutânea , Neoplasias Cutâneas/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Artif Cells Nanomed Biotechnol ; 47(1): 904-916, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30856018

RESUMO

The aim of present study was to develop the efficient targeting of Concanavalin-A conjugated nanotransfersomal gel to bind directly to melanocytes gel layer against UVB induced skin carcinoma. Carbopol loaded nanotransfersomal gel have prepared by modified rotary evaporation sonication technique & conjugated synthesized by carbodiimide method and they were characterized the morphology, zeta potential, penetration and cell viability. In vitro release studies & skin permeation have determined using Franz diffusion cell and confocal laser scanning microscope (CLSM). The conjugated formulation showed vesicles size, polydispersity index, zeta potential and % conjugation efficiency of 179.0 ± 0.32 nm, 0.197 ± 0.07, 35.1 ± 0.21 mV and 89.73 ± 1.29% respectively. The surface morphology was confirmed by transmission electron microscopy (TEM) and FTIR to make sure the compatibility among its ingredients. Con-A conjugated nanotransfersomal gel showed toxicity on melanoma (A375) in a concentration range of 0.4-2.0 mg/mL, but less toxicity toward HaCaT cells. The MTT assay has analyzed against two different cell lines, to determine their anti-cancer potentials and their targeting ability. Conjugated formulation were found to decrease the cell viability, higher skin targeting efficacy in in-vitro & in-vivo. Concanavalin conjugated nanotransfersomal gel of apigenin promise an efficient and economic approach for the skin cancer.


Assuntos
Apigenina/química , Concanavalina A/química , Sistemas de Liberação de Medicamentos/métodos , Melanoma Experimental/tratamento farmacológico , Preparações Farmacêuticas/administração & dosagem , Neoplasias Cutâneas/tratamento farmacológico , Administração Tópica , Animais , Linhagem Celular Tumoral , Sobrevivência Celular , Liberação Controlada de Fármacos , Estabilidade de Medicamentos , Cabras , Humanos , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Tamanho da Partícula , Preparações Farmacêuticas/química , Absorção Cutânea , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Propriedades de Superfície , Raios Ultravioleta
16.
Nat Commun ; 10(1): 1147, 2019 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-30850596

RESUMO

DNA nanostructures are promising drug carriers with their intrinsic biocompatibility, uniformity and versatility. However, rapid serum disintegration leads to low bioavailability at targeted sites following systemic administration, hindering their biomedical applications. Here we demonstrate transdermal delivery of framework nucleic acids (FNAs) through topical applications. By designing FNAs with distinct shapes and sizes, we interrogate their penetration on mice and human skin explant. Skin histology reveals size-dependent penetration, with FNAs ≤75 nm effectively reaching dermis layer. 17 nm-tetrahedral FNAs show greatest penetration to 350 µm from skin periphery. Importantly, structural integrity is maintained during the skin penetration. Employing a mouse melanoma model, topical application of doxorubicin-loaded FNAs accommodates ≥2-fold improvement in drug accumulation and tumor inhibition relative to topically-applied free doxorubicin, or doxorubicin loaded in liposomes and polymeric nanoparticles. Programmable penetration with minimal systemic biodistribution underlines FNA potential as localized transdermal drug delivery carriers.


Assuntos
Antibióticos Antineoplásicos/farmacologia , Preparações de Ação Retardada/farmacocinética , Doxorrubicina/farmacologia , Sistemas de Liberação de Medicamentos/métodos , Melanoma Experimental/tratamento farmacológico , Ácidos Nucleicos/química , Neoplasias Cutâneas/tratamento farmacológico , Administração Cutânea , Animais , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Preparações de Ação Retardada/química , Doxorrubicina/química , Doxorrubicina/farmacocinética , Humanos , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Nus , Ácidos Nucleicos/farmacocinética , Permeabilidade , Pele/efeitos dos fármacos , Pele/metabolismo , Pele/patologia , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Suínos
17.
Mar Drugs ; 17(2)2019 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-30821275

RESUMO

Lj-RGD3, which contains three Arg⁻Gly⁻Asp (RGD) motifs, was first identified from the buccal glands of Lampetra japonica and has been shown to suppress the tumor progression in the previous studies. Apart from the three RGD motifs, Lj-RGD3 is also characterized by its high content of histidine in its amino acid sequence. In order to clarify whether the histidine-rich characterization of Lj-RGD3 is also associated with its anti-tumor activity, mutants were designed in which the three RGD motifs (Lj-112), or all histidines (Lj-27) or both (Lj-26) were deleted. Furthermore, a mutant (Lj-42) in which all histidines and three RGD motifs were respectively substituted with alanines and three Ala⁻Gly⁻Asp (AGD) motifs, as well as a mutant (Lj-41) in which all histidines were substituted with alanines was synthesized to avoid alterations in structure which might further cause changes in the peptides' functions. After recombination and purification, recombinant Lj-112 (rLj-112), recombinant Lj-27 (rLj-27), recombinant Lj-41 (rLj-41), and recombinant Lj-RGD3 (rLj-RGD3) exhibited anti-proliferative activity in B16 cells, respectively; while recombinant Lj-26 (rLj-26) and recombinant Lj-42 (rLj-42) did not affect the proliferation of B16 cells significantly. In addition, the anti-proliferative activity of rLj-112 in B16 cells was due to apoptosis. Typical apoptosis features were observed, including chromatin condensation, fragmented DNA, and increased levels of cleaved caspase 3/caspase 7/nuclear enzyme poly (ADP-ribose) polymerase (PARP) in B16 cells. Similar to rLj-RGD3, rLj-112 was also capable of suppressing the migration and invasion of B16 cells by disturbing the F-actin arrangement. After labeling with FITC, rLj-112 was found localized in the cytoplasm of B16 cells, which induced the internalization of epidermal growth factor receptor (EGFR), suggesting that rLj-112 might block the EGFR mediated signaling pathway. Actually, the phosphorylation level of EGFR and its downstream signal molecules including Akt, PI3K, p38, and ERK1/2 was reduced in the rLj-112 treated B16 cells. In vivo, rLj-112 also inhibited the growth, weight, and volume of the tumors in B16 xenografted C57BL/6 mice without reducing their body weight, indicating that rLj-112 might be safe and might be used as an effective anti-tumor drug in the near future.


Assuntos
Receptores ErbB/metabolismo , Venenos de Peixe/genética , Venenos de Peixe/farmacologia , Oligopeptídeos/genética , Oligopeptídeos/farmacologia , Sequência de Aminoácidos , Animais , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 7/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/antagonistas & inibidores , Feminino , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/metabolismo , Poli(ADP-Ribose) Polimerase-1/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Distribuição Aleatória , Transdução de Sinais/efeitos dos fármacos , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Colloids Surf B Biointerfaces ; 177: 356-361, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30772670

RESUMO

Phosphoester compounds are promising materials with expected biocompatibility; however, little has been reported on the use of phosphoester compounds for micelle formulations. In this study, paclitaxel (PTX)-encapsulated micelles were prepared using four kinds of alkyl di(MePEG-lactate) phosphates. From the results of the determination of critical micelle concentrations and an in vitro stability test, it was shown that a compound to which 1-eicosanol was introduced as a side chain was desirable in the preparation of PTX-encapsulated micelles (PTX-micelles). The mean volume diameter and PTX content of the micelles were 135.7 ± 52.2 nm and 3.9% ± 0.2%, respectively. in vitro release tests of the micelles were performed at different pH levels. Twenty-four hours after the start of the release test, the cumulative PTX release rate of PTX-micelles at pH 5.0 reached 96.2%, which was three times higher than that at pH 7.4. As a result of the degradation test of the compound used for the micelle, it was confirmed that this compound degraded faster at pH 5.0 than at pH 7.4. The hemolysis rate of drug-free micelles was 0.8%-1.4%, and the biocompatibility of this micelle as a drug carrier was suggested. In addition, the effectiveness of PTX-micelles in cancer treatment was evaluated via biodistribution study. PTX concentration in the tumor was significantly increased in the group administered PTX-micelles as compared with the group administered PTX solution. These results suggest that phosphoester compounds are useful in preparing biocompatible pH-responsive carriers.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Materiais Biocompatíveis/farmacologia , Micelas , Organofosfatos/química , Paclitaxel/farmacologia , Animais , Antineoplásicos Fitogênicos/química , Materiais Biocompatíveis/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Portadores de Fármacos/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Masculino , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Camundongos , Estrutura Molecular , Paclitaxel/química , Tamanho da Partícula , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Propriedades de Superfície
19.
Int J Mol Sci ; 20(3)2019 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-30708951

RESUMO

Malignant melanoma is a highly aggressive form of skin cancer which has a propensity for metastasis. Epithelial mesenchymal transition (EMT) plays a primordial role in the progression of metastatic disease. Metastatic melanoma is resistant to conventional therapies. Hence, researchers have been exploring alternative approaches, including the utility of bioactive phytochemicals to manage metastatic disease. In the present study, we investigated the potential of cirsiliol, a flavonoid isolated from Centaurea jacea L., in modulating the aggressive behavior of B16F10 metastatic melanoma cells, including EMT, and associated molecular mechanisms of action. Cirsiliol was found to be effective in restraining the colony formation and migration of fibronectin-induced B16F10 metastatic melanoma cells. Cirsiliol inhibited the activity and expression of matrix metalloproteinase-9 (MMP-9). Cirsiliol also suppressed the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (also known as Akt)/nuclear factor-κB (NF-κB) signaling pathway which, in turn, caused upregulation of E-cadherin and downregulation of N-cadherin, Snail and Twist. Based on these results, cirsiliol may be considered a promising compound against EMT in the therapeutic management of malignant melanoma.


Assuntos
Transição Epitelial-Mesenquimal/efeitos dos fármacos , Flavonas/administração & dosagem , Melanoma Experimental/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Neoplasias Cutâneas/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Flavonas/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , Melanoma Experimental/metabolismo , Camundongos , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Cutâneas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Proc Natl Acad Sci U S A ; 116(8): 3100-3105, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30718426

RESUMO

Denileukin diftitox (DAB-IL-2, Ontak) is a diphtheria-toxin-based fusion protein that depletes CD25-positive cells including regulatory T cells and has been approved for the treatment of persistent or recurrent cutaneous T cell lymphoma. However, the clinical use of denileukin diftitox was limited by vascular leak toxicity and production issues related to drug aggregation and purity. We found that a single amino acid substitution (V6A) in a motif associated with vascular leak induction yields a fully active, second-generation biologic, s-DAB-IL-2(V6A), which elicits 50-fold less human umbilical vein endothelial cell monolayer permeation and is 3.7-fold less lethal to mice by LD50 analysis than s-DAB-IL-2. Additionally, to overcome aggregation problems, we developed a production method for the fusion toxin using Corynebacterium diphtheriae that secretes fully folded, biologically active, monomeric s-DAB-IL-2 into the culture medium. Using the poorly immunogenic mouse B16F10 melanoma model, we initiated treatment 7 days after tumor challenge and observed that, while both s-DAB-IL-2(V6A) and s-DAB-IL-2 are inhibitors of tumor growth, the capacity to treat with higher doses of s-DAB-IL-2(V6A) could provide a superior activity window. In a sequential dual-therapy study in tumors that have progressed for 10 days, both s-DAB-IL-2(V6A) and s-DAB-IL-2 given before checkpoint inhibition with anti-programmed cell death-1 (anti-PD-1) antibodies inhibited tumor growth, while either drug given as monotherapy had less effect. s-DAB-IL-2(V6A), a fully monomeric protein with reduced vascular leak, is a second-generation diphtheria-toxin-based fusion protein with promise as a cancer immunotherapeutic both alone and in conjunction with PD-1 blockade.


Assuntos
Toxina Diftérica/administração & dosagem , Interleucina-2/administração & dosagem , Melanoma Experimental/tratamento farmacológico , Receptor de Morte Celular Programada 1/genética , Substituição de Aminoácidos/genética , Anticorpos/administração & dosagem , Proliferação de Células/efeitos dos fármacos , Corynebacterium diphtheriae/química , Corynebacterium diphtheriae/patogenicidade , Toxina Diftérica/química , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Imunossupressores/administração & dosagem , Imunotoxinas/administração & dosagem , Interleucina-2/química , Subunidade alfa de Receptor de Interleucina-2/efeitos dos fármacos , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Receptores de Interleucina-2/genética , Receptores de Interleucina-2/imunologia , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/química , Linfócitos T Reguladores/efeitos dos fármacos
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