[en] In normal epithelial cells, p38α has a well-established role as a tumor suppressor. However, recent reports have illustrated pro-tumorigenic functions for the p38α pathway by promoting tumor cell survival and proliferation. Given that the role of p38 MAPKs have been shown to be cell and context dependent, we focused our study on the function of p38α in breast cancer progression. To investigate the role of p38α in breast tumor progression in vivo, we combined the polyoma middle T (PyMT) mouse model with floxed conditional alleles of p38α. This way, we analyzed the role of p38α in already developed tumors, where we observed a progressive tumor regression following p38α deletion (p38αΔ). This tumor reduction correlated with decreased cell proliferation, increased cell death and elevated levels of DNA damage. To investigate the molecular mechanisms underlying mammary tumor regression, we established epithelial cell cultures from PyMT-induced tumors. As increased levels of γ-H2AX were observed in tumors with reduced p38α, we examined DNA damage signaling and genome integrity in the PyMT-expressing epithelial cells. These cultures reproduced the increased levels of γ-H2AX observed in vivo and interestingly, the localization of γ-H2AX in these cells was not restricted to the nuclei, but also appeared in DNA bridges and micronuclei, potentially reflecting DNA double strand breaks resulting from chromosome segregation errors during cytokinesis. Increased levels of DNA damage are known to impact on DNA replication and hamper its progression. We directly assessed the effect of p38α deletion on DNA replication by analyzing stretched DNA fibers. We found a lower replication rate in p38αΔ cells and the analysis of fibers with bidirectional tracks revealed an increased rate of asymmetric forks, indicative of a high frequency of fork stalling. The defects in replication fork progression and the increased DNA damage led us to investigate the status of the DNA damage response, an essential network that monitors DNA replication and coordinates cellular responses. We observed defects in ssDNA generation, RPA accumulation, RPA and CHK1 phosphorylation and RAD51 recruitment after treatment with campothecin, a topoisomerase I inhibitor that induces DNA damage specifically in S-phase, indicating that ATR signaling, DNA-end resection and homologous recombination DNA repair were defective in p38α deficient cells. Moreover, we identified CtIP, a key factor that promotes DNA-end resection in mammalian cells, as a p38α substrate. De-regulation of CtIP due to lack of p38α could impact on the DNA damage response and explain many of the described phenotypes. Replication stress and DNA damage have been proposed to be the major cause of chromosome instability (CIN) in cancer, leading to both numerical and structural chromosome alterations. Using time- lapse analysis, we observed that some cells showed cytokinesis failure, reflected by the presence of DNA bridges between the daughter cells, and gave rise to multinucleated cells. However, even in the presence of DNA bridges, most of the p38αΔ cells were able to eventually divide, but exhibited micronuclei after mitosis. This indicated that p38α deletion induced missegregation, ultimately leading to aneuploidy. Our results suggested that targeting p38α would increase tumor cell sensitivity to CIN-inducing agents. We tested this hypothesis both in vitro and in vivo, and found that pharmacological inhibition of p38α increased the effect of both paclitaxel and docetaxel in the PyMT model. These results were further confirmed in patient-derived xenografts, where p38α inhibition enhanced, accelerated or prolonged the anti-tumoral response observed with the taxanes alone. These results were consistent with the idea that p38α contributes to the DNA damage response and facilitates the survival of cancer cells with high levels of CIN. Moreover, they confirmed the hypothesis that targeting p38α sensitizes cancer cells to CIN-inducing agents such as taxanes, raising the possibility of translating p38α inhibitors into the clinic.

[es] La proteína quinasa p38α está considerada como un supresor tumoral. Sin embargo, estudios recientes sugieren que puede promover el crecimiento y la supervivencia en algunos tipos de células tumorales, así como que su inhibición coopera con determinados agentes quimioterapéuticos. Así pues, la función de p38α parece depender tanto del tipo de célula como del contexto. Por ello, este trabajo abordó el papel de p38α en el contexto concreto de la progresión del cáncer de mama. Usando el modelo "Polyoma middle T" (PyMT), hemos observado que la expresión de p38α en las células epiteliales que conforman el tumor es esencial para el crecimiento del mismo. Para analizar los mecanismos moleculares implicados establecimos un sistema ex vivo consistente en líneas celulares derivadas de tumores inducidos por PyMT. Observamos que en ausencia de p38α estas células mostraban estrés replicativo, un mayor daño en el ADN y un incremento en los errores de segregación que correlacionaban con una menor viabilidad celular. Los defectos durante la replicación y el aumento en el daño en el ADN nos llevaron a investigar el estado de la ruta de respuesta al daño en el ADN. Así pues, encontramos que las células deficientes en p38α mostraban defectos en la generación de ADN monocatenario, en la activación de ATR y en el reclutamiento de RAD51 a los focos de daño, lo que sugería que la reparación de ADN por recombinación homóloga estaba afectada. Además identificamos CtIP, un factor esencial para la resección del ADN, como sustrato directo de p38α. La desregulación de CtIP influiría negativamente en la respuesta al daño en el ADN y explicaría en gran medida los fenotipos descritos anteriormente. En conjunto, nuestros datos sugieren que p38α es necesaria para una eficiente reparación y replicación del ADN y por tanto para el mantenimiento de la estabilidad cromosómica en las células epiteliales de cáncer de mama. Los resultados anteriores sugerían que la intervención farmacológica de p38α podría incrementar la sensibilidad a agentes inductores de inestabilidad cromosómica como los taxanos. Confirmamos esta teoría usando el modelo PyMT y xenografías derivadas de pacientes, comprobando que en ambos casos la inhibición de p38α potenció, aceleró o prolongó el efecto anti-tumoral de los taxanos paclitaxel y docetaxel.