RESUMO
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.
Assuntos
Técnicas de Transferência de Genes , Hormônio Liberador de Gonadotropina , Sinais de Localização Nuclear , Antígenos Virais de Tumores/química , Antígenos Virais de Tumores/farmacologia , Membrana Celular/química , Membrana Celular/metabolismo , DNA/química , DNA/farmacologia , Hormônio Liberador de Gonadotropina/análogos & derivados , Hormônio Liberador de Gonadotropina/síntese química , Hormônio Liberador de Gonadotropina/química , Hormônio Liberador de Gonadotropina/farmacologia , Células Hep G2 , Humanos , Sinais de Localização Nuclear/química , Sinais de Localização Nuclear/farmacologia , Vírus 40 dos Símios/químicaRESUMO
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.
Assuntos
Antígenos Virais de Tumores/química , Núcleo Celular/metabolismo , Portadores de Fármacos/química , Hormônio Liberador de Gonadotropina/análogos & derivados , Sinais de Localização Nuclear/química , Vírus 40 dos Símios/metabolismo , Sequência de Aminoácidos , Antineoplásicos/administração & dosagem , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Portadores de Fármacos/síntese química , Portadores de Fármacos/farmacologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Hormônio Liberador de Gonadotropina/síntese química , Hormônio Liberador de Gonadotropina/química , Hormônio Liberador de Gonadotropina/farmacologia , Humanos , Dados de Sequência Molecular , Sinais de Localização Nuclear/genética , Ligação Proteica , Receptores LHRH/metabolismo , Vírus 40 dos Símios/químicaRESUMO
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.
