Using multiple drugs to kill cancer cells can decrease drug resistance development. However, this approach is frequently limited by the bioavailability and toxicity of the combined agents and delivery at ratios to specific locations that synergistically kill the cancer cells. Loading the individual agents into a nanoparticle that releases the drugs at synergizing ratios at a single location is one approach to resolve this concern. Celecoxib and Plumbagin are two drugs that were identified from a screen to synergistically kill melanoma cells compared to normal cells. Combined use of these agents by traditional approaches was not possible due to poor bioavailability and toxicological concerns. This study details the development of a nanoliposomal-based agent containing Celecoxib and Plumbagin, called CelePlum-777, which is stable and releases these drugs at an optimal ratio for maximal synergistic killing efficacy. CelePlum-777 was more effective at killing melanoma than normal cells and inhibited xenograft melanoma tumor growth by up to 72% without apparent toxicity. Mechanistically, the drug combination in CelePlum-777 led to enhanced inhibition of melanoma cell proliferation mediated by decreasing levels of key cyclin important for cancer cell proliferation and survival, which was not observed with the individual agents. Thus, a novel nanoparticle based drug has been developed containing Celecoxib and Plumbagin that lacks toxicity and deliverers the agents at a synergistically killing drug ratio to kill cancer cells.

Many processes are deregulated in melanoma cells and one of those is protein production. While much is known about protein synthesis in cancer cells, effective ways of therapeutically targeting this process remains an understudied area of research. A process that is upregulated in melanoma compared to normal melanocytes is proline biosynthesis, which has been linked to both oncogene and tumor suppressor pathways, suggesting an important convergent point for therapeutic intervention. Therefore, a RNA interference (RNAi) screen of a kinase library was undertaken, identifying aldehyde dehydrogenase 18 family, member A1 (ALDH18A1) as a critically important gene in regulating melanoma cell growth through proline biosynthesis. Inhibition of ALDH18A1, the gene encoding pyrroline-5-carboxylate synthase (P5CS), significantly decreased cultured melanoma cell viability and tumor growth. Knockdown of P5CS using siRNA had no effect on apoptosis, autophagy, or the cell cycle but cell doubling time increased dramatically suggesting that there was a general slowdown in cellular metabolism. Mechanistically, targeting ALDH18A1 activated the serine/threonine protein kinase GCN2 (general control nonderepressible 2) to inhibit protein synthesis, which could be reversed with proline supplementation. Thus, targeting ALDH18A1 in melanoma can be used to disrupt proline biosynthesis to limit cell metabolism thereby increasing the cellular doubling time mediated through the GCN2 pathway.