Page 122 - SRPSKO DRUŠTVO ISTRAŽIVAČA RAKA

Serbian Association for Cancer Research SDIRSACR

SESSION 5

EXPLORING THE ROLE OF MDR MECHANISMS AND STEM CELL BIOLOGY IN CANCER AND CANCER
TREATMENT STRATEGIES

P25

Expression profiling of p53 family isoforms in targeted therapy-resistant primary melanoma cell lines

Janja Josić, Anđela Horvat, Christine Supina Pavić, Ignacija Vlašić, Neda Slade

Laboratory for Protein Dynamics, Division of Molecular Medicine, Ruđer Bošković Institute, Zagreb, Croatia

Keywords: alternative splicing, drug resistance, melanoma, MITF transcription factor, tumor suppressor protein p53

Background: Metastatic melanoma is the most aggressive form of skin cancer that is responsible for more than 80% of
cutaneous cancer-related deaths. This cancer is characterized by a high frequency of somatic mutations, predominantly
in the BRAF gene. Patients harboring the BRAF V600E mutation are treated with targeted therapies, including BRAF
kinase inhibitors (BRAFi) alone or combined with MEK inhibitors (MEKi). Despite initial responses, resistance to these
therapies develops rapidly, often driven by molecular alterations. One such alteration is the changed expression of p53
family isoforms, which regulate the activity of the tumor suppressor p53 and may influence resistance mechanisms.
Material and Methods: To better understand these molecular changes, we established and validated primary
melanoma cell lines derived from metastatic melanoma patients who received combined BRAFi/MEKi therapy. We
aimed to characterize the expression profiles of p53 family members in these cells. We assessed the expression of p53,
p63, and p73 isoforms at both the mRNA and protein levels using quantitative PCR (qPCR) and Western blot analysis
across various melanoma sublines. Additionally, we measured the expression of MITF, which encodes the central
transcription factor that regulates melanogenesis, via qPCR. Expression of MITF in melanoma sublines was compared
to that in non-melanoma cancer cell lines, including prostate, ovarian, and neuroblastoma lines.
Results: We observed that primary cell sublines differ in morphology and consist of three distinct subtypes: melanocytic
cells, mesenchymal cells and a mixed population. In all cell sublines the expression of MITF gene was determined and
compared with MITF expression in different non-melanocytic cancer cell lines. All melanoma sublines demonstrated
higher MITF expression than the non-melanoma cancer lines. However, levels of MITF varied significantly among
the melanoma sublines. The results of p53 isoform expression revealed the differences in both mRNA and protein
expression levels between the individual cell sublines.
Conclusions: These preliminary findings suggest that specific p53 family isoforms may be associated with the
development of resistance to combined BRAFi/MEKi targeted therapy in melanoma. Further research is required to
confirm whether these isoforms can serve as reliable biomarkers for acquired therapy resistance and to explore their
potential roles in treatment outcomes.

Acknowledgments and funding: This project is funded by The Croatian Science Foundation grants DOK-2023-10 and
IP-2022-10-1375.

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