The role of Translesion Synthesis Cisplatin and TMZ resistance in glioma cells.

Author(s)

Marcela Teatin Latancia

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2022-06-09

Examining board members

Carlos Frederico Martins Menck; José Renato Rosa Cussiol; Nicolas Carlos Hoch; Elza Tiemi Sakamoto Hojo; Nadja Cristhina de Souza Pinto; Mari Cleide Sogayar

• Advisor: Carlos Frederico Martins Menck

Abstract

Glioblastoma is a cancer that presents poor prognosis, with a high ratio of recurrence. Translesion synthesis (TLS) DNA polymerases are enzymes capable of replicating unrepaired DNA damage. Tumor cells can use this process to survive lesions caused by genotoxic chemotherapeutic drugs (CMT), contributing to the mechanisms of therapy resistance. In addition, TLS is error prone and can lead to mutagenesis, increasing the resistance potential of tumor cells. Treatment with CMT, such as cisplatin or temozolomide (TMZ), induces DNA damage to promote tumor cell death. Little is known about the role of TLS polymerases in the responses to TMZ. Preliminary results obtained by Dr. Clarissa R. R. Rocha, in our group, identified a list of genes associated with the resistance to TMZ, using a library of genes activated or knocked out by the CRISPR-Cas9 system. In this list, the participation of some TLS polymerases, such as, Pol and Pol , are remarkable. Thus, our aim is to better understand the role of TLS polymerases, especially Pol , Pol and Pol , in the resistance to cisplatin and TMZ, CMT used in therapeutic glioma protocols. At first, we treated mutated POLH cells (XP-V) and complemented cells for this gene (XP-V comp) with TMZ. We observed that XP-V cells are more sensitive to TMZ, indicating that Pol must be important to overcome DNA damage promoted by TMZ. We developed U251-MG knockout cells for the POLK and POLI polymerase genes by CRISPR/Cas9. These mutated cells also had their viability impaired after treatment with TMZ, increased in genotoxic stress and cell cycle arrest, mainly in G2 phase, but not in S phase - after 48 h of treatment, in all three mutated cells, not observed in cells wild-type for these genes. To further investigate DNA replication, we performed DNA fiber assay, and, surprisingly, there was no difference between cells mutated to POLK and wild-type cells treated with TMZ. We conclude that TLS polymerases protect tumor cells from damage caused by TMZ, and, therefore, play an important role to overcome resistance to this chemotherapy. Now, we intend to understand how replication is affected in cells treated with TMZ. There is no S-phase arrest, where these enzymes generally act, and no difference in the progression fork replication indicating that the role played by these polymerases in the resistance to TMZ may be beyond TLS. Thus, we are looking for other possible functions responsible for the observed effects. In summary, we want to better understand the resistance mechanism promoted by TLS polymerases and seek new targets that collaborate with the overcome of resistance to TMZ, used in the clinic for the glioma treatment, and contribute to improve the prognosis of patients with this neoplasia. (AU)

• FAPESP's process: 18/10061-0 - Role of translesion synthesis in cisplatin and temozolomide resistance in Glioma cells
• Grantee: Marcela Teatin Latancia
• Support Opportunities: Scholarships in Brazil - Doctorate