RESUMO
Internal human xenografts provide valuable animal models to study the microenvironments and metastatic processes occurring in human cancers. However, the use of such models is hampered by the logistical difficulties of reproducibly and simply assessing tumor burden. We developed a high-sensitivity assay for quantifying human DNA in small volumes of mouse plasma, enabling in-life monitoring of systemic tumor burden. Growth kinetics analyses of various xenograft models showed the utility of circulating human DNA as a biomarker. We found that human DNA concentration reproducibly increased with disease progression and decreased after successful therapeutic intervention. A marked, transient spike in circulating human tumor DNA occurred immediately after cytotoxic therapy or surgery. This simple assay may find broad utility in target validation studies and preclinical drug development programs.
Assuntos
Biomarcadores Tumorais/sangue , Neoplasias do Colo/sangue , DNA de Neoplasias/sangue , Animais , Biomarcadores Tumorais/genética , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Primers do DNA , Modelos Animais de Doenças , Feminino , Células HCT116 , Humanos , Elementos Nucleotídeos Longos e Dispersos/genética , Camundongos , Camundongos Nus , Transplante de Neoplasias , Transplante HeterólogoRESUMO
For the production of DNA microarrays from PCR products, purification of the the DNA fragments prior to spotting is a major expense in cost and time. Also, a considerable amount of material is lost during this process and contamination might occur. Here, a protocol is presented that permits the manufacture of microarrays from unpurified PCR products on aminated surfaces such as glass slides coated with the widely used poly(L-lysine) or aminosilane. The presence of primer molecules in the PCR sample does not increase the non-specific signal upon hybridisation. Overall, signal intensity on arrays made of unpurified PCR products is 94% of the intensity obtained with the respective purified molecules. This slight loss in signal, however, is offset by a reduced variation in the amount of DNA present at the individual spot positions across an array, apart from the considerable savings in time and cost. In addition, a larger number of arrays can be made from one batch of amplification products.