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1.
J Invest Dermatol ; 126(4): 882-9, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16439964

RESUMO

Selectin-mediated leukocyte adhesion to endothelia, the crucial first step initiating the pathogenic cascade of inflammation, is an attractive target for specific therapies. The small-molecule macrolide, efomycine M, inhibits selectin-mediated leukocyte adhesion in vitro and in vivo, and effectively alleviates inflammatory disorders in vivo. To define the molecular basis of the therapeutically relevant antiadhesive properties of efomycines, several new species of this family were purified and/or synthesized. Efomycines E and G were isolated from Steptomyces BS1261. Efomycine O was synthesized by Lewis acid-catalyzed acetalization and efomycine M was generated by base-catalyzed deglycosylation. Efomycine S resulted from ester cleavage of the macrolide ring system, and efomycine T represents the peracetylated form of efomycine M. When the functional activity of efomycines on adhesion of leukocytes to vascular endothelium was studied, some remarkable differences between the compounds became apparent, inasmuch as efomycines E, G, M, and O significantly inhibited adhesion of both human and porcine leukocytes to the vascular endothelium, whereas efomycines S and T did not show any biological activity. A novel docking engine (ProPose), generating an improved, fully configurable protein-ligand interaction model, demonstrated that biological activities of efomycines can be predicted in silico, thus highlighting the utility of such combinatorial approaches.


Assuntos
Adesão Celular/efeitos dos fármacos , Macrolídeos/química , Macrolídeos/farmacologia , Selectinas/efeitos dos fármacos , Animais , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/imunologia , Leucócitos/efeitos dos fármacos , Leucócitos/imunologia , Macrolídeos/isolamento & purificação , Estrutura Molecular , Streptomyces/metabolismo , Relação Estrutura-Atividade , Suínos
4.
J Invest Dermatol ; 122(5): 1266-76, 2004 May.
Artigo em Inglês | MEDLINE | ID: mdl-15140231

RESUMO

Bypassing molecular mechanisms of apoptosis deficiency may be of great utility for the successful treatment of malignant tumors. We have discovered that imiquimod, a small-molecule immunomodulator, exerts rather tumor-selective direct pro-apoptotic activity in vivo and in vitro towards cutaneous metastases of malignant melanoma, an aggressive skin tumor. This pro-apoptotic activity was not detectable with resiquimod, a closely related structural analogue whose pro-inflammatory activity is even greater than that of imiquimod. Unresponsiveness of some melanoma metastases to imiquimod in vivo corresponded to resistance towards imiquimod-induced apoptosis in vivo and in vitro. At the molecular level, the pro-apoptotic activity of imiquimod was independent of membrane-bound death receptors, but depended on Bcl-2 expression as demonstrated by overexpression of Bcl-2 in melanoma cells. Imiquimod is the first topical compound with the potential to bypass molecular mechanisms of apoptosis deficiency, a concept that may be relevant for other tumors as well.


Assuntos
Aminoquinolinas/administração & dosagem , Antineoplásicos/administração & dosagem , Apoptose/efeitos dos fármacos , Melanoma/tratamento farmacológico , Neoplasias Cutâneas/tratamento farmacológico , Administração Tópica , Aminoquinolinas/farmacologia , Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose , Biópsia , Linhagem Celular Tumoral/citologia , Linhagem Celular Tumoral/efeitos dos fármacos , Humanos , Imidazóis/farmacologia , Imiquimode , Melanócitos/citologia , Melanócitos/efeitos dos fármacos , Melanoma/imunologia , Melanoma/secundário , Glicoproteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Receptores do Fator de Necrose Tumoral/metabolismo , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Ligante Indutor de Apoptose Relacionado a TNF , Fator de Necrose Tumoral alfa/metabolismo , Receptor fas/metabolismo
5.
Arch Dermatol Res ; 296(2): 59-66, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15278364

RESUMO

Molecular events underlying the progression of malignant tumors through the surrounding tissue are largely mediated by membrane-bound adhesion molecules. Basal-cell adhesion molecule (B-CAM), a 90-kDa laminin receptor of the immunoglobulin superfamily, is induced in some epithelial malignancies. Its function in these tumors, however, still remains obscure. We demonstrated that expression of B-CAM is very weak, if detectable at all, in normal epidermis but is strongly induced in both basal cell carcinomas and squamous cell carcinomas of the skin, and most pronounced at the basal surface of the tumor nests. Interestingly, the only known B-CAM ligand, laminin, was markedly upregulated within corresponding microanatomical sites surrounding the tumor nests, suggesting that both molecules may interact there. Consistent with this hypothesis, we were able to directly demonstrate binding of a B-CAM/Fc chimeric molecule to the peritumoral stroma in situ. Finally, in proof-of-principle experiments, human B-CAM was overexpressed both in murine and in human fibroblasts. The haptotactic migration of these novel B-CAM+ cell populations on a laminin matrix was significantly increased (P = 0.02) as compared to mock-transfected cells when integrin-mediated adhesion was blocked by chelation of divalent cations. Thus, our findings provide the first direct experimental evidence that interactions of B-CAM and laminin may be involved in progression of epithelial skin tumors.


Assuntos
Carcinoma Basocelular/metabolismo , Carcinoma de Células Escamosas/metabolismo , Moléculas de Adesão Celular/metabolismo , Laminina/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias Cutâneas/metabolismo , Animais , Carcinoma Basocelular/etiologia , Carcinoma de Células Escamosas/etiologia , Adesão Celular , Moléculas de Adesão Celular/genética , Movimento Celular , Células Cultivadas , Humanos , Técnicas In Vitro , Ligantes , Sistema do Grupo Sanguíneo Lutheran , Camundongos , Células NIH 3T3 , Proteínas de Neoplasias/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Neoplasias Cutâneas/etiologia , Transfecção , Regulação para Cima
6.
J Natl Cancer Inst ; 100(12): 862-75, 2008 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-18544741

RESUMO

BACKGROUND: Increasing the efficacy of chemotherapeutics by reducing chemoresistance may be a useful strategy in cancer therapy. Constitutive activation of nuclear factor-kappa B (NF-kappaB) is a hallmark of various cancers, including melanoma, which is almost universally resistant to chemotherapy. NF-kappaB is regulated by inhibitory kappaB (IkappaB) proteins, which are in turn phosphorylated by the IkappaB kinase (IKK) complex. METHODS: The effect on NF-kappaB activity of a novel small-molecule inhibitor of the beta subunit of IKK (KINK-1; kinase inhibitor of nuclear factor-kappaB-1) was assessed by measuring phosphorylation of the alpha subunit of IkappaB by immunoblotting, DNA binding by electrophoretic mobility shift assays, and nuclear translocation of NF-kappaB using immunofluorescence. Regulation of NF-kappaB-dependent gene expression was determined by microarray analysis, real-time and semiquantitative reverse transcription polymerase chain reaction (RT-PCR), and Western blot analyses. The effects of KINK-1 (alone and in combination with cytostatic agents) on melanoma cells were characterized by assessing proliferation, soft agar colony formation, and markers of apoptosis. The antitumoral efficacy of KINK-1 in combination with the cytostatic agents doxorubicin or camptothecin (all injected intraperitoneally) was tested in vivo by measuring lung weight and counting metastases in C57BL6 mice (groups of six) bearing metastases of melanoma cells. All statistical tests were two-sided. Results KINK-1 strongly suppressed both constitutive and induced NF-kappaB activity in melanoma cells. It reduced the expression of NF-kappaB-dependent gene products that regulate proliferation, cytokine production, and antiapoptotic responses but exhibited little antiproliferative or proapoptotic activity at the cellular level. However, KINK-1 markedly increased the activities of some cytostatic agents in vitro and abrogated doxorubicin-induced NF-kappaB activation. Combined treatment of C57BL6 mice that had been injected with melanoma cells with KINK-1 and doxorubicin or camptothecin reduced metastases and pulmonary tumor mass compared with either treatment alone (mean lung weight 19 days after injection of melanoma cells of mice treated with 3 mg/kg KINK-1 alone, 1 mg/kg doxorubicin alone, and 1 mg/kg doxorubicin plus 3 mg/kg KINK-1 = 260 mg, 95% confidence interval (CI) = 216 to 305 mg; 268 mg, 95% CI = 224 to 313 mg; and 181 mg, 95% CI = 171 to 192 mg, respectively, P < .001 from t tests comparing mean lung weight of double-treated mice to that in mice treated with either compound alone). CONCLUSION: Inhibition of constitutive and induced IKKbeta-activity through treatment with KINK-1 might increase tumor susceptibility to chemotherapy.


Assuntos
Antineoplásicos/farmacologia , Quinase I-kappa B/antagonistas & inibidores , Melanoma/tratamento farmacológico , Melanoma/metabolismo , NF-kappa B/antagonistas & inibidores , Oxazinas/farmacologia , Piridinas/farmacologia , Animais , Antineoplásicos/efeitos adversos , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Ensaio de Desvio de Mobilidade Eletroforética , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Melanoma/enzimologia , Melanoma/secundário , Camundongos , Mitose/efeitos dos fármacos , Oxazinas/efeitos adversos , Piridinas/efeitos adversos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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