Role of HIVEP2 in drug-tolerant persister cancer cells and tumor recurrence

Abstract

[eng] Colorectal cancer (CRC) is a leading cause of death worldwide and tumor recurrence is a frequent complication that arises from minimal residual disease (MRD). Patients can relapse months or even years after an apparent remission of the disease when tumors regrow as metastatic and drug resistant. At these advanced stages, cancer is refractory to most available treatments, presenting the poorest prognosis and leading to high mortality rates. MRD comprises a small subpopulation of cancer cells that survive treatment and remain undetectable to the available diagnostic tools as latent seeds of the disease. These drug-tolerant persister (DTP) cells -which includes dormant and slow-cycling cancer cells (SCCCs)- are characterized by being quiescent and drug resistant due to non-genetical perturbations, since these cells can alternate between proliferating and quiescent periods. Although great efforts have been made trying to characterize and target these DTP cells, our knowledge regarding the mechanism underlying the biology of these cells is still limited. Thanks to characterization of SCCCs from different tumor origins as diverse as CRC, melanoma and glioblastoma, we identified several transcription and epigenetic factors highly expressed in these cells and other dormant models (e.g. D-HEp3), among which HIVEP2 is highly expressed. In the present work, we characterized for the first time the role of HIVEP2 in the biology of DTP cells and its implication in tumor recurrence. HIVEP2 is a transcription factor involved in the regulation of the expression of a wide range of genes, and extremely important for the regulation of inflammation and immune response and the proper development of the brain and cognitive functions. In this doctoral thesis we have uncovered the role of HIVEP2 regulating cell proliferation and self-renewal and providing intrinsic mechanisms to chemotherapy-resistance using genetically modified CRC models. Overexpression of HIVEP2 causes cell cycle arrest delaying tumor growth. However, it confers full resistance to chemotherapies such as 5-fluorouracil (5FU) in vivo. This observation could explain why CRC patients with high levels of HIVEP2 exhibit worse progression free survival after chemotherapy treatment. In silico analysis of drug sensitivity in multiple cancer cell lines depicted that those with high levels of HIVEP2 are more resistant to a wide range of chemotherapies, which we achieved to validate for 5FU in different CRC models of patient-derived xenograft organoids (PDXOs). Unfortunately, we were not able to validate the predicted sensitivity to the kinase inhibitor sapitinib. The enhance on AKT and ERK activation seems to be responsible for the increased chemoresistance, since both signaling cascades can trigger pro-survival responses. Additionally, other mechanisms identified by gene expression analysis and immunohistochemistry of tumor sections that can be involved in drug-tolerance are the upregulation of ATP-binding cassettes transporters, aberrant expression of keratins and a mucinous differentiation of the tumors. All these processes have been previously described to be involved in resistance to chemotherapies. Finally, we also provided evidence suggesting that HIVEP2 may be involved in dissemination, since we detected an increased number of dormant disseminated tumor cells in the organs of chicken embryos. Altogether, our findings highlight the importance of HIVEP2 in cancer progression and chemotherapy resistance in CRC. HIVEP2 overexpression not only confers resistance to conventional treatments, but also promotes the survival and dissemination of dormant tumor cells. These findings enhance our understanding of DTP cells and offer potential strategies for developing new targeted therapies aimed at eradicating this subpopulation of cancer cells.

[spa] El cáncer colorrectal (CRC) es una de las principales causas de muerte en todo el mundo, siendo la recurrencia tumoral, causada por la existencia de una enfermedad residual mínima (MRD), la principal complicación. Los pacientes pueden recaer meses o años después de una aparente remisión, cuando los tumores reaparecen como metastásicos y resistentes a las quimioterapias y a la mayoría de los tratamientos disponibles, lo que conlleva a altas tasas de mortalidad. La MRD se compone de una pequeña población de células cancerígenas que sobreviven al tratamiento y permanecen indetectables. Estas células persistentes tolerantes a los fármacos (DTP), que incluyen células durmientes y de ciclo lento (SCCCs), se caracterizan por un estado quiescente y una quimioresistencia intrínseca debido a alteraciones no genéticas. A pesar de los grandes esfuerzos por caracterizar y atacar estas células, nuestro conocimiento sobre su biología sigue siendo limitado. Mediante la caracterización de SCCCs de tumores de CRC, melanoma y glioblastoma, identificamos varios factores de transcripción y epigenéticos altamente expresados en esta subpoblación, entre los que destaca HIVEP2. Este trabajo caracteriza por primera vez el papel de HIVEP2 en las células DTP y su implicación en la recurrencia tumoral. HIVEP2 es un factor de transcripción que regula la expresión de muchos genes, y es crucial para la respuesta inmune y el desarrollo cerebral. En esta tesis doctoral hemos descubierto que HIVEP2 regula la proliferación celular, confiriendo mecanismos intrínsecos de quimioresistencia en CRC. La sobreexpresión de HIVEP2 detiene el ciclo celular, retrasando el crecimiento tumoral, pero a su vez, confiriendo resistencia al 5-fluorouracilo (5FU). Esto explicaría por qué los pacientes con CRC con altos niveles de HIVEP2 presentan peor supervivencia libre de enfermedad después del tratamiento con quimioterapia. El análisis in silico y la validación en organoides derivados de pacientes con CRC mostraron que las células con altos niveles de HIVEP2 son más resistentes al 5FU. Además, HIVEP2 parece estar involucrado en la diseminación tumoral, aumentando el número de células tumorales durmientes. Estos hallazgos subrayan la importancia de HIVEP2 en la progresión del cáncer y la resistencia a tratamientos, ofreciendo la posibilidad de nuevas estrategias para terapias dirigidas.