Biomaterials folded into nanometer scale sized particles (NPs) can be utilized as targeted drug delivery systems for cancer therapy. Lonidamine (LND), a small anticancer drug which inhibits glycolysis was previously examined as a treatment for benign prostate hypertrophy, however it shows severe liver toxicity. NPs composed of the negatively charged Polypeptide, poly gamma glutamic acid (g-PGA) and a designed amphiphilic and cationic β-sheet Peptide (denoted PoP-NPs) loaded with LND, denoted LND-PoP-NPs were previously used in our lab to successfully target the mitochondria when coated with peptide (LND-mPoP-NPs).[1,2] In this study, we found a new procedure to increase LND-mPoP-NPs efficacy by concentrating the NP solution using lyophilization and re-dissolution of the obtained powder in a tenth of the original volume (designated as C-LND-mPoP-NPs). The C-LND-mPoP-NPs were found to enhance drug efficacy without showing any toxicity in MDA-MB-231 breast cancer cells and in xenograft mice model.