Pentoxifylline and Norcantharidin Synergistically Suppress Melanoma Growth in Mice: A Multi-Modal In Vivo and In
Israel Lara-Vega1, Minerva Nájera-Martínez1, Armando Vega-López1
1Laboratorio de Toxicología Ambiental, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Av. Wilfrido Massieu s/n, Unidad Profesional Zacatenco, Mexico City 07738, Mexico.
Abstract:
Melanoma is a highly aggressive skin cancer with limited therapeutic response. Targeting intracellular signaling pathways and promoting tumor cell differentiation are promising therapeutic strategies. Pentoxifylline (PTX) and norcantharidin (NCTD) have demonstrated antitumor properties, but their combined mechanisms of action in melanoma remain poorly understood. The effects of PTX (30 and 60 mg/kg) and NCTD (0.75 and 3 mg/kg), administered alone or in combination, in a DBA/2J murine B16-F1 melanoma model via intraperitoneal and intratumoral (IT) routes were evaluated. Tumor growth was monitored, and molecular analyses included RNA sequencing and immunofluorescence quantification of PI3K, AKT1, mTOR, ERBB2, BRAF, and MITF protein levels, and molecular docking simulations were performed. In the final stage of the experiment, combination therapy significantly reduced tumor volume compared to monotherapies, with the relative tumor volume decreasing from 18.1 ± 1.2 (SD) in the IT Control group to 0.6 ± 0.1 (SD) in the IT combination-treated group (n = 6 per group; p < 0.001). RNA-seq revealed over 3000 differentially expressed genes in intratumoral treatments, with enrichment in pathways related to oxidative stress, immune response, and translation regulation (KEGG and Reactome analyses). Minimal transcript-level changes were observed for BRAF and PI3K/AKT/mTOR genes; however, immunofluorescence showed reduced total and phosphorylated levels of PI3K, AKT1, mTOR, BRAF, and ERBB2. MITF protein levels and pigmentation increased, especially in PTX-treated groups, indicating enhanced melanocytic differentiation. Docking analyses predicted direct binding of both drugs to PI3K, AKT1, mTOR, and BRAF, with affinities ranging from -5.7 to -7.4 kcal/mol. The combination of PTX and NCTD suppresses melanoma progression through dual mechanisms: inhibition of PI3K/AKT/mTOR signaling and promotion of tumor cell differentiation.
Insights
Pentoxifylline (PTX) and norcantharidin (NCTD) combination therapy significantly reduced melanoma tumor volume by inhibiting PI3K/AKT/mTOR signaling and promoting melanocytic differentiation.
Area of Science:
- Oncology
- Molecular Biology
- Pharmacology
Background:
- Melanoma is an aggressive skin cancer with limited treatment options.
- Targeting intracellular pathways and promoting tumor cell differentiation are key therapeutic strategies.
- The combined effects of Pentoxifylline (PTX) and norcantharidin (NCTD) on melanoma are not well understood.
Purpose of the Study:
- To investigate the combined therapeutic effects of PTX and NCTD on melanoma.
- To elucidate the molecular mechanisms underlying their combined action.
- To evaluate their impact on intracellular signaling pathways and tumor cell differentiation.
Main Methods:
- Utilized a DBA/2J murine B16-F1 melanoma model.
- Administered PTX and NCTD via intraperitoneal and intratumoral routes, alone and in combination.
- Performed RNA sequencing, immunofluorescence, and molecular docking simulations.
Main Results:
- Combination therapy significantly reduced tumor volume compared to monotherapies (p < 0.001).
- RNA-seq identified over 3000 differentially expressed genes, enriched in oxidative stress and immune response pathways.
- Immunofluorescence revealed decreased PI3K, AKT1, mTOR, BRAF, and ERBB2 levels, alongside increased MITF and pigmentation, indicating differentiation.
Conclusions:
- The combination of PTX and NCTD suppresses melanoma progression.
- Dual mechanisms involve inhibition of the PI3K/AKT/mTOR signaling pathway.
- Promotion of tumor cell differentiation is another key mechanism of action.
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