RESUMO
It is commonly accepted that cancer cell progression is accompanied by accumulation of genetic changes. Here we searched for copy number variations in melanoma and asked whether homozygous losses always cumulate during tumor cell progression. Therefore we investigated either melanoma cell lines or tissue derived from the primary lesion and from the lymph node metastasis of the same individual patient. In vitro studies of melanoma cell lines revealed high migratory and anchorage independent growth of metastasis-derived cells. Surprisingly, whole genome DNA analysis of a primum-derived cell line revealed a total of 10 homozygous losses, whereas the matched metastasis-derived cell line only shared five of those losses. We further tested these cells in a mouse model for intradermal melanoma growth and detected fast growth of the metastasis-derived cell line and no growth of primum-derived cells. Additionally, we screened matched pairs of patient-derived melanoma primum and metastasis samples and we could also identify a case with homozygous deletions exclusively present in the primary lesion. Therefore, we suggest that tumor cell progression at the metastatic niche can occur parallel and independently from the primary tumor. We propose that for mutation-targeted therapy genotyping should be performed not only from primary, but also from metastatic melanoma.
Assuntos
Deleção Cromossômica , Melanoma/patologia , Neoplasias Cutâneas/patologia , Adulto , Animais , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células , Forma Celular , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 10/genética , Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 4/genética , Variações do Número de Cópias de DNA , Feminino , Estudo de Associação Genômica Ampla , Humanos , Metástase Linfática , Masculino , Melanoma/genética , Melanoma/metabolismo , Camundongos , Camundongos SCID , Invasividade Neoplásica , Transplante de Neoplasias , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Cromossomo X/genéticaRESUMO
Sphingosine kinase 1 (Sphk1), a lipid kinase implicated in cell transformation and tumor growth, is overexpressed in gastric cancer and is linked with a poor prognosis. The biological relevance of Sphk1 expression in gastric cancer is unclear. Here, we studied the functional significance of Sphk1 as a novel molecular target for gastric cancer by using an antisense oligonucleotide approach in vitro and in vivo. Gastric cancer cell lines (MKN28 and N87) were treated with Sphk1 with locked nucleic acid-antisense oligonucleotides (LNA-ASO). Sphk1 target regulation, cell growth, and apoptosis were assessed for single-agent Sphk1 LNA-ASO and for combinations with doxorubicin. Athymic nude mice xenografted with gastric cancer cells were treated with Sphk1 LNA and assessed for tumor growth and Sphk1 target regulation, in vivo. In vitro, nanomolar concentrations of Sphk1 LNA-ASO induced an approximately two-fold reduction in Sphk1 mRNA in both the cell lines. This resulted in a 1.6-fold increase in apoptosis and inhibited the growth of gastric cancer cells by more than 50% (P < 0.05). The combination of Sphk1 LNA-ASO with doxorubicin resulted in significant chemosensitization. In vivo, Sphk1 LNA-ASO displayed neither mRNA target regulation in xenografts nor antitumor activity in two independent nude mouse xenograft models. In conclusion, the potent single-agent activity and the synergistic effect of Sphk1 LNA-ASO in combination with chemotherapy in vitro highlight Sphk1 as a biologically relevant molecular target for gastric cancer. Further studies are warranted to overcome the challenge of delivering Sphk1-targeting RNA-therapeutics to solid tumors in vivo.
