PEGylated liposomes are attractive pharmaceutical nanocarriers; however, literature reports of ligand-targeted nanoparticles have not consistently shown successful results. Here, we employed a multifaceted synthetic strategy to prepare peptide-targeted liposomal nanoparticles with high purity, reproducibility, and precisely controlled stoichiometry of functionalities to evaluate the role of liposomal PEG coating, peptide EG-linker length, and peptide valency on cellular uptake in a systematic manner. We analyzed these parameters in two distinct disease models where the liposomes were functionalized with either HER2- or VLA-4-antagonistic peptides to target HER2-overexpressing breast cancer cells or VLA-4-overexpressing myeloma cells, respectively. When targeting peptides were tethered to nanoparticles with an EG45 (∼PEG2000) linker in a manner similar to a more traditional formulation, their cellular uptake was not enhanced compared to non-targeted versions regardless of the liposomal PEG coating used. Conversely, reduction of the liposomal PEG to PEG350 and the peptide linker to EG12 dramatically enhanced cellular uptake by ∼9 fold and ∼100 fold in the breast cancer and multiple myeloma cells, respectively. Uptake efficiency reached a maximum and a plateau with ∼2% peptide density in both disease models. Taken together, these results demonstrate the significance of using the right design elements such as the appropriate peptide EG-linker length in coordination with the appropriate liposomal PEG coating and optimal ligand density in efficient cellular uptake of liposomal nanoparticles.