# Discovery of Two Novel Pyrazole Derivatives as Anticancer Agents Targeting Tubulin Polymerization and MAPK Signaling Pathways

## TL;DR

Two new pyrazole compounds were found to kill cancer cells by disrupting microtubules and affecting key signaling pathways, showing promise as potential anticancer drugs.

## Contribution

Discovery of two novel pyrazole derivatives with dual mechanisms targeting tubulin and MAPK pathways.

## Key findings

- P3C.1 and P3C.2 showed potent cytotoxicity across multiple cancer cell lines, especially in triple-negative breast cancer.
- The compounds induce apoptosis through ROS generation, mitochondrial depolarization, and cell cycle arrest.
- Transcriptome and kinase analysis revealed disruption of microtubules and altered MAPK signaling as key mechanisms.

## Abstract

Drug resistance is the major determinant of chemotherapy failure, leading to relapse and tumor progression, demonstrating the urgent need for novel antineoplastic drugs. This study aimed to evaluate the anticancer potential of two novel pyrazole derivatives, P3C.1 and P3C.2, and to elucidate their mechanism of action in cancer cells.

The cytotoxicity of the compounds was evaluated across 27 different cancer cell lines via a nuclear staining assay. Subsequent flow cytometric and biochemical analyses were performed to assess reactive oxygen species (ROS) generation, apoptosis induction, mitochondrial integrity, and cell cycle progression. Additional studies included transcriptome analyses and immunoassays to characterize the molecular mechanisms underlying drug activity.

Two novel pyrazole derivatives, P3C.1 and P3C.2, were identified with potent cytotoxicity on a variety of cancer cell lines. Among the adherent cell lines tested, the triple-negative breast cancer (TNBC) cell line MDA-MB-231 exhibited the highest sensitivity to both compounds and was therefore selected for further experimentation. In vitro assays demonstrated that both compounds induced ROS generation, mitochondrial membrane depolarization, cell cycle arrest and apoptosis. Whole-transcriptome sequencing of P3C.1 and P3C.2-treated MDA-MB-231 and two lymphoblastic leukemia cell lines revealed four genes in common associated with cell signaling and membrane dynamics. Connectivity Map (CMAP) database comparisons of shared genes for each cancer subtype revealed a strong similarity between the two compounds with tubulin inhibitors, and subsequent assays confirmed that these compounds act as microtubule-disrupting agents. Moreover, protein phosphorylation analysis indicated that both compounds induced hyperphosphorylation of JNK, and ERK1/2, along with hypophosphorylation of p38 kinases.

P3C.1 and P3C.2 emerged as promising anti-breast cancer agents with dual mechanisms of action involving microtubule disruption and altered kinase signaling, leading to induction of apoptosis.