The pH-(Low) Insertion Peptide (pHLIP) can target cancer cells based on their low extracellular pH (pHe) in vivo. Under slightly acidic conditions, pHLIP inserts into membrane, forming a transmembrane helix. Thus, pHLIP has the potential to selectively deliver drugs directly into the cytoplasm of cancer cells. Herein, using structure-activity-relationship (SAR) studies (by changing pHLIP sequence or side-chain structures), we searched and found new pHLIP variants that would, as compared to the original 'WT' pHLIP, (a) insert more readily at the tumor pHe of 6.5-7.0, (b) with insertion occurring over a narrower pH range (i.e. sharper pH response), and (c) have increased solubility. Furthermore, to study 'WT' pHLIP insertion in cellular membranes in more detail, as well as to evaluate the promising variants (from liposomal SAR studies) in cancer cells, we developed fluorescently self-quenched pHLIP probes that become more fluorescent upon interactions with cells. We learned that (a) compared to 'WT' pHLIP, D14E, D25E and Asp25Aad variants all insert more readily at the plasmamembrane, (b) a large portion of WT pHLIP insertion occur in endosomal membranes, (c) negatively charged polar N-terminus improved the pH-response of pHLIP insertion in cells, and (d) pHLIP insertion is cell-line dependent.

Towards drug delivery via pHLIP insertion in the plasmamembrane of cancer cells at tumor extracellular pH

248th National Meeting (2014)

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