Proteolytically Stable Cancer Targeting Peptides with High Affinity for Breast Cancer Cells

Abstract

Cancer cell targeting peptides have emerged as a highly efficient approach for selective delivery of chemotherapeutics and diagnostics to different cancer cells. However, the use of α-peptides in pharmaceutical applications is hindered by their enzymatic degradation and low bioavailability. Starting with a 10-mer α-peptide 18 that we developed previously, here we report three novel analogues of 18 that are proteolytically stable and display better (up to 3.5-fold) affinity profiles for breast cancer cells compared to 18. The design strategy involved replacement of two or three amino acids in the sequence of 18 with d-residues or β(3)-amino acids. Such replacement maintained the specificity for cancer cells (MDA-MB-435, MDA-MB-231, and MCF-7) with low affinity for control noncancerous cells (MCF-10A and HUVEC), showed an increase in secondary structure, and rendered the analogues completely stable to human serum and liver homogenate from mice. The three analogues are potentially safe with minimal cellular toxicity and are efficient targeting moieties for specific drug delivery to breast cancer cells. The strategy used here may be adapted to develop peptide analogues that will target other cancer cell types.

Novel Peptide–Doxorubucin Conjugates for Targeting Breast Cancer Cells Including the Multidrug Resistant Cells

“…Previous in vitro and in vivo studies for drug conjugates, such as DPV-maleimide, have shown that the use of TFA salts is not associated with any toxicity up to 1 mM concentrations. 28 These data reveal that both conjugates 1 and 2 are stable enough to allow sufficient time for cancer cell targeting.…”

“…To facilitate side chain attachment to Dox, previously reported peptide 18-4 was synthesized with one substitution (r8k) and a C-terminal amide group. 28 This peptide is now referred to as 18-4a. Peptide 18-4a…”

“…27 We reported a proteotytically stable decapeptide 18-4 (WxEYAAQrFL) that preferentially binds and internalizes into breast cancer cells, and minimally binds to normal mammary epithelial cells. 28 Peptide 18-4 was derived from peptide p160 (VPWMEYPAQRFL) that has specific binding to breast cancer cells. 29 Peptide 18-4 has two D-amino acid substitutions at the enzymatic labile sites which added complete proteolytic stability in human serum and liver homogenate.…”

“…In addition, 18-4 is potentially safe with minimal cellular toxicity. 28 We hypothesize that conjugation of peptide 18-4 to Dox through a hydrolysable linker will create a breast cancer specific prodrug, which can selectively target and deliver Dox to breast cancerous cells with 5 reduced delivery to normal cells. In addition, conjugation may overcome the p-glycoprotein efflux mechanism involved in multidrug resistance (MDR).…”

Design and Modification of Anticancer Peptides

“…Hilchie et al [19] designed a analog of NRC-03 by replacing all the original amino acids with the corresponding D-amino acids, named [D]-NRC-03, which exhibited higher anticancer activity than NRC-03 due to the stability against human serum or trypsin [21]. Except D-amino acid, other unnatural amino acids also have been used to modify the anticancer peptide to increase their stability or bioactivity [22][23][24]. Anticancer peptide of…”