Cellular Activity of CQWW Nullomer-Derived Peptides.

Analysis of observed protein sequences across all species within the UniProtKB/Swiss-Prot data set reveals CQWW as the shortest absent stretch of amino acids. While DNA can be found encoding the CQWW sequence, it has never been observed to be translated or included in manually curated sets of proteins, existing only in predicted, tentative sequences and in a single mature antibody sequence. We have synthesized this "nullomer" peptide, along with 13 derivatives, reversed, truncated, stereoisomers, and alanine-scanning peptides, conjugated to polyarginine stretches to increase cellular uptake. We observed their impact against a healthy neuronal line and six patient-derived glioblastoma cell lines spanning three clinical subtypes. Results reveal IC values averaging 4.9 μM for inhibition of cell survival across tested oncogenic cell lines. High-content phenotypic analysis of cellular features and reverse-phase protein arrays failed to discern a clear mode of action for the nullomer peptide but suggests mitochondrial impairment through the inhibition of GSK3 and isoforms, supported by observations of reduced mitochondrial stain intensities. With a recent increase in interest in nullomer peptides, we see the results in this study as a starting point for further investigation into this potentially therapeutic peptide class.