Gly-Phe-Gly-OH

We synthesized four novel lipophilic derivatives of phenylalanyl-glycine (Phe-Gly), C4-Phe-Gly, Phe-Gly-C4, C6-Phe-Gly and C8-Phe-Gly by chemical modification with butyric acid (C4), caproic acid (C6) and octanoic acid (C8). The effect of the acylation on the stability, permeability and accumulation of Phe-Gly in the intestine was investigated by in vitro studies. The stability of Phe-Gly in homogenates of duodenal and colonic membranes was low, but was significantly improved by the acylation except for Phe-Gly-C4. In the transport studies, a modified Ussing chamber was used for the intestinal permeability experiments with Phe-Gly and its acyl derivatives. The permeability of native Phe-Gly and Phe-Gly-C4 across the intestinal membrane was not observed during the transport studies. However, the permeability of Phe-Gly was much improved by chemical modification with various fatty acids to its N-terminal portion. The permeability of acyl-Phe-Gly derivatives across the intestinal membrane decreased with increasing the chain length of fatty acids. In addition, the intestinal tissue accumulation of acyl-Phe-Gly derivatives at the end of the transport studies was much higher than that of native Phe-Gly. The intestinal tissue accumulation of acyl-Phe-Gly in the duodenum increased as the chain length of fatty acids increased. Furthermore, intestinal permeability of C4-Phe-Gly was slightly inhibited in the presence of 5 mM ceftibuten and was significantly reduced under low temperature condition. We observed a directional difference in the transport of C4-Phe-Gly (the mucosal to serosal transport of C4-Phe-Gly was higher than its serosal to mucosal transport) suggesting that C4-Phe-Gly might be transported by a carrier-mediated process as well as other dipeptides. These findings indicate that acylation might be useful approach to enhance the transport of Phe-Gly, a model dipeptide, transported by a carrier-mediated process.

Trastuzumab conjugates consisting of exatecan derivatives were prepared and their biological activities and physicochemical properties were evaluated. The ADCs showed strong efficacy and a low aggregation rate. The exatecan derivatives were covalently connected via a peptidyl spacer (Gly-Gly-Phe-Gly), which is assumed to be stable in circulation, and were cleaved by lysosomal enzymes following ADC internalization into tumor tissue. These anti-HER2 ADCs exhibited a high potency, specifically against HER2-positive cancer cell lines in vitro. The ADCs, bearing exatecan derivatives which have more than two methylene chains, exhibited superior cytotoxicity. It was speculated that steric hindrance of the cleavable amide moiety could be involved in the drug release. The adequate alkyl lengths of exatecan derivatives (13, 14, 15) were from two to four in terms of aggregation rate. The ADC having a hydrophilic moiety showed good efficacy in a HER2-positive and Trastuzumab-resistant breast carcinoma cell model in mice.

Protein-mimics are of great interest for their structure, stability, and properties. We are interested in the synthesis of protein-mimics containing triazole linkages as peptide-bond surrogate by topochemical azide-alkyne cycloaddition (TAAC) polymerization of azide- and alkyne-modified peptides. The rationally designed dipeptide N3 -CH2 CO-Phe-NHCH2 CCH (1) crystallized in a parallel β-sheet arrangement and are head-to-tail aligned in a direction perpendicular to the β-sheet-direction. Upon heating, crystals of 1 underwent single-crystal-to-single-crystal polymerization forming a triazole-linked pseudoprotein with Gly-Phe-Gly repeats. During TAAC polymerization, the pseudoprotein evolved as helical chains. These helical chains are laterally assembled by backbone hydrogen bonding in a direction perpendicular to the helical axis to form helical sheets. This interesting helical-sheet orientation in the crystal resembles the cross-α-amyloids, where α-helices are arranged laterally as sheets.