FOLFOX is a combination of folinic acid (FnA), 5-fluorouracil (5-Fu) and oxaliplatin (OxP). It has been used as the standard treatment for colorectal cancer (CRC) and hepatocellular carcinoma (HCC). This treatment is effective, but its high toxicity is dose limiting, and the drugs need to be taken for a long time. To lower the toxicity so that higher doses can be administered with minimal side effects, two lipid-based membrane–core (MC) nanoformulations, Nano-Folox and Nano-FdUMP, have recently been developed by using the nanoprecipitation technique. The combination of Nano-Folox (containing platinum drug and FnA) and Nano-FdUMP (containing fluorine drug) significantly improves the antitumor effect against CRC and HCC relative to FOLFOX (the combination of free drugs), resulting in long-term survival of animals without significant toxic signs. Here, we describe two formulation protocols. First, for Nano-Folox, a Pt(DACH)•FnA nanoprecipitate is formed by [Pt(DACH)(H2O)2]2+ (the active form of OxP) and FnA2−, and the resultant nanoprecipitate is encapsulated inside the lipid nanoparticles (NPs) modified with the PEGylated aminoethyl anisamide (AEAA, a targeting ligand for sigma-1 receptor overexpressing on CRC and HCC). Second, for Nano-FdUMP, FdUMP (the active metabolite of 5-Fu) is entrapped inside the amorphous Ca3(PO4)2 nanoprecipitate, and the resultant Ca3(PO4)2•FdUMP nanoprecipitate is encapsulated into the AEAA-targeted PEGylated lipid NPs. The procedures for Nano-Folox and Nano-FdUMP take ~17 h and ~4 h, respectively (~17 h if they are prepared simultaneously). Procedures for the physicochemical (~30 h) and cytotoxic (~54 h) characterization are also described.
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This study was supported by an NIH grant (CA198999, to L.H.). We thank O. Gololobova at the University of North Carolina at Chapel Hill for assistance with ICP-MS and thank Z. Cong at Jilin University for assistance with HPLC.
L.H. is a consultant for PDS Biotechnology, Samyang Biopharmaceutical Co. and Stemirna. J.G. declares no competing interests.
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Key references using this protocol
Guo, J. et al. ACS Nano 14, 5075–5089 (2020): https://doi.org/10.1021/acsnano.0c01676
Guo, J. et al. Mol. Cancer 20, 10 (2021): https://doi.org/10.1186/s12943-020-01297-0
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Guo, J., Huang, L. Formulation of two lipid-based membrane–core nanoparticles for FOLFOX combination therapy. Nat Protoc (2022). https://doi.org/10.1038/s41596-022-00698-3