RESUMO
It is becoming increasingly clear that processive DNA replication is threatened not only by DNA damage but also by secondary structures that can form in the DNA template. Failure to resolve these structures promptly leads to both genetic instability, for instance DNA breaks and rearrangements, and to epigenetic instability, in which inaccurate propagation of the parental chromatin state leads to unscheduled changes in gene expression. Multiple overlapping mechanisms are needed to deal with the wide range of potential DNA structural challenges to replication. This review focuses on the emerging mechanisms by which specialised DNA polymerases, best known for their role in the replication of damaged DNA, contribute to the replication of undamaged but structured DNA, particularly G quadruplexes.
Assuntos
Replicação do DNA , DNA Polimerase Dirigida por DNA/metabolismo , DNA/química , Quadruplex G , DNA/metabolismoRESUMO
BACKGROUND & AIMS: Patients with colorectal tumors with microsatellite instability (MSI) have better prognoses than patients with tumors without MSI, but have a poor response to 5-fluorouracilbased chemotherapy. A dominant-negative form of heat shock protein (HSP)110 (HSP110DE9) expressed by cancer cells with MSI, via exon skipping caused by somatic deletions in the T(17) intron repeat, sensitizes the cells to 5-fluorouracil and oxaliplatin.We investigated whether HSP110 T(17) could be used to identify patients with colorectal cancer who would benefit from adjuvant chemotherapy with 5-fluorouracil and oxaliplatin. METHODS: We characterized the interaction between HSP110 and HSP110DE9 using surface plasmon resonance. By using polymerase chain reaction and fragment analysis, we examined how the size of somatic allelic deletions in HSP110 T(17) affected the HSP110 protein expressed by tumor cells. We screened 329 consecutive patients with stage IIIII colorectal tumors with MSI who underwent surgical resection at tertiary medical centers for HSP110 T(17). RESULTS: HSP110 and HSP110DE9 interacted in a1:1 ratio. Tumor cells with large deletions in T(17) had increased ratios of HSP110DE9:HSP110, owing to the loss of expression of full-length HSP110. Deletions in HSP110 T(17) were mostly biallelic in primary tumor samples with MSI. Patients with stage IIIII cancer who received chemotherapy and had large HSP110 T(17) deletions (≥5 bp; 18 of 77 patients, 23.4%) had longer times of relapse-free survival than patients with small or no deletions (≤4 bp; 59 of 77 patients, 76.6%) in multivariate analysis (hazard ratio, 0.16; 95% confidence interval, 0.0120.8; P = .03). We found a significant interaction between chemotherapy and T17 deletion (P =.009). CONCLUSIONS: About 25% of patients with stages IIIII colorectal tumors with MSI have an excellent response to chemotherapy, due to large, biallelic deletions in the T(17) intron repeat of HSP110 in tumor DNA.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Sequência de Bases , Biomarcadores Tumorais/genética , Neoplasias Colorretais/tratamento farmacológico , Proteínas de Choque Térmico HSP110/genética , Instabilidade de Microssatélites , Deleção de Sequência , Idoso , Antineoplásicos/administração & dosagem , Biomarcadores Tumorais/química , Biomarcadores Tumorais/metabolismo , Western Blotting , Linhagem Celular Tumoral , Quimioterapia Adjuvante , Colectomia , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/cirurgia , Feminino , Fluoruracila/administração & dosagem , Seguimentos , Proteínas de Choque Térmico HSP110/química , Proteínas de Choque Térmico HSP110/metabolismo , Humanos , Íntrons , Leucovorina/administração & dosagem , Masculino , Modelos Moleculares , Compostos Organoplatínicos/administração & dosagem , Oxaliplatina , Estudos Retrospectivos , Ressonância de Plasmônio de Superfície , Análise de Sobrevida , Resultado do TratamentoRESUMO
Recently we have shown that the mitogen-activated protein kinase (MAPK) MAPK14/p38α is involved in resistance of colon cancer cells to camptothecin-related drugs. Here we further investigated the cellular mechanisms involved in such drug resistance and showed that, in HCT116 human colorectal adenocarcinoma cells in which TP53 was genetically ablated (HCT116-TP53KO), overexpression of constitutively active MAPK14/p38α decreases cell sensitivity to SN-38 (the active metabolite of irinotecan), inhibits cell proliferation and induces survival-autophagy. Since autophagy is known to facilitate cancer cell resistance to chemotherapy and radiation treatment, we then investigated the relationship between MAPK14/p38α, autophagy and resistance to irinotecan. We demonstrated that induction of autophagy by SN38 is dependent on MAPK14/p38α activation. Finally, we showed that inhibition of MAPK14/p38α or autophagy both sensitizes HCT116-TP53KO cells to drug therapy. Our data proved that the two effects are interrelated, since the role of autophagy in drug resistance required the MAPK14/p38α. Our results highlight the existence of a new mechanism of resistance to camptothecin-related drugs: upon SN38 induction, MAPK14/p38α is activated and triggers survival-promoting autophagy to protect tumor cells against the cytotoxic effects of the drug. Colon cancer cells could thus be sensitized to drug therapy by inhibiting either MAPK14/p38 or autophagy.
Assuntos
Autofagia/efeitos dos fármacos , Camptotecina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteína Quinase 14 Ativada por Mitógeno/metabolismo , Proteína Supressora de Tumor p53/deficiência , Camptotecina/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Ativação Enzimática/efeitos dos fármacos , Técnicas de Inativação de Genes , Células HCT116 , Humanos , Irinotecano , Proteína Supressora de Tumor p53/metabolismo , Vacúolos/efeitos dos fármacos , Vacúolos/metabolismo , Vacúolos/ultraestruturaRESUMO
Secreted Frizzled-related proteins (Sfrps) are extracellular regulators of Wnt signalling and play important roles in developmental and oncogenic processes. They are known to be upregulated in regenerating muscle and in myoblast cultures but their function is unknown. Here, we show that the addition of recombinant Sfrp1 or Sfrp2 to C2C12 cell line cultures or to primary cultures of satellite cells results in the inhibition of myotube formation with no significant effect on the cell cycle or apoptosis. Even though at confluence, treated and untreated cultures are identical in appearance, analyses have shown that, for maximum effect, the cells have to be treated while they are proliferating. Furthermore, removal of Sfrp from the culture medium during differentiation restores normal myotube formation. We conclude that Sfrp1 and Sfrp2 act to prevent myoblasts from entering the terminal differentiation process.