RESUMEN
A set of GnRH analogues containing nuclear localization signal (NLS) of SV-40 virus large T-antigen have been synthesized using solid phase peptide synthesis and chemical ligation technique. Selective chemical ligation was achieved as a result of hydrazone formation in the course of interaction between NLS hydrazide and GnRH analog modified by pyruvic acid. The efficiency of synthesized peptide carriers was demonstrated in experiments with human cancer cells transfected by reporter luciferase and beta-galactosidase genes or suicide HSV-1 thymidine kinase gene. It was shown that selectivity of action on cancer cells can be achieved as a result of peptide/DNA complex penetration through the cell membrane by GnRH receptor-mediated endocytosis pathway.
Asunto(s)
Técnicas de Transferencia de Gen , Hormona Liberadora de Gonadotropina , Señales de Localización Nuclear , Antígenos Virales de Tumores/química , Antígenos Virales de Tumores/farmacología , Membrana Celular/química , Membrana Celular/metabolismo , ADN/química , ADN/farmacología , Hormona Liberadora de Gonadotropina/análogos & derivados , Hormona Liberadora de Gonadotropina/síntesis química , Hormona Liberadora de Gonadotropina/química , Hormona Liberadora de Gonadotropina/farmacología , Células Hep G2 , Humanos , Señales de Localización Nuclear/química , Señales de Localización Nuclear/farmacología , Virus 40 de los Simios/químicaRESUMEN
To improve the efficiency of anticancer drugs due to their delivery to intracellular targets a set of GnRH analogues containing nuclear localization signal (NLS) of SV-40 virus large T-antigen have been synthesized. NLS was attached to the parent molecule via ε-amino group of D-Lysine in position 1 or 6 of peptide sequence using orthogonal protection strategy. The biological activity studies revealed that incorporation of NLS moiety significantly increases cytotoxic activity of palmitoyl-containing GnRH analogues in vitro. The influence of tested peptides on tumor cells does not accompanied by the destruction of cell membrane, as confirmed in experiments with normal fibroblasts, used as a control.
Asunto(s)
Antígenos Virales de Tumores/química , Núcleo Celular/metabolismo , Portadores de Fármacos/química , Hormona Liberadora de Gonadotropina/análogos & derivados , Señales de Localización Nuclear/química , Virus 40 de los Simios/metabolismo , Secuencia de Aminoácidos , Antineoplásicos/administración & dosificación , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Portadores de Fármacos/síntesis química , Portadores de Fármacos/farmacología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Hormona Liberadora de Gonadotropina/síntesis química , Hormona Liberadora de Gonadotropina/química , Hormona Liberadora de Gonadotropina/farmacología , Humanos , Datos de Secuencia Molecular , Señales de Localización Nuclear/genética , Unión Proteica , Receptores LHRH/metabolismo , Virus 40 de los Simios/químicaRESUMEN
Luliberin analogues modified at the N-terminus were synthesized to search for drugs exerting a cytotoxic effect on cells of hormone-dependent tumors. A synthetic scheme effective in the preparation of analogues containing fatty acid residues was proposed. The cytotoxic effect of the peptides was studied on a number of cell lines of human tumors in vitro. The dependence of the antitumor effect on the length of peptide chain, amino acid sequence, and structure of the N-terminal group was demonstrated. Modification with palmitic acid was found to result in highly active compounds in the case of analogues containing more than ten aa, whereas modifications with lauric, caproic, or trimethylacetic acid led to compounds with significantly lower activities. Analogues of luliberin containing a palmitic acid residue and effectively inhibiting the growth of tumor cells in vitro were synthesized.