Sequence modification of heptapeptide selected by phage display as homing device for HT-29 colon cancer cells to improve the anti-tumour activity of drug delivery systems

Abstract

Development of peptide-based conjugates for targeted tumour therapy is a current research topic providing new possibilities in cancer treatment. In this study, VHLGYAT heptapeptide selected by phage display technique for HT-29 human colon cancer was investigated as homing peptide for drug delivery. Daunomycin was conjugated to the N-terminus of the peptide directly or through Cathepsin B cleavable spacers. Conjugates showed moderate in vitro cytostatic effect. Therefore, sequence modifications were performed by Ala-scan and positional scanning resulting in conjugates with much higher bioactivity. Conjugates in which Gly was replaced by amino acids with bulky apolaric side chains provided the best efficacy. The influence of the cellular uptake, stability and drug release on the anti-tumour activity was investigated. It was found that mainly the difference in the cellular uptake of the conjugates generated the distinct effect on cell viability. One of the most efficient conjugate Dau=Aoa-LRRY-VHLFYAT-NH 2 showed tumour growth inhibition on orthotopically developed HT-29 colon cancer in mice with negligible toxic side effect compared to the free drug. We also indicate that this sequence is not specific to HT-29 cells, but it has a remarkable effect on many other cancer cells. Nevertheless, the Phe-containing conjugate was more active in all cases compared to the conjugate with the parent sequence. The literature data suggested that this sequence is highly overlapped with peptides that recognize Hsp70 membrane bound protein overexpressed in many types of tumours.

Improved In Vivo Anti-Tumor and Anti-Metastatic Effect of GnRH-III-Daunorubicin Analogs on Colorectal and Breast Carcinoma Bearing Mice

“…On the contrary, we obtained relatively high IC 50 and relative potency values in case of MRC-5 cells, which suggested that the conjugates are highly tumor selective, except for PANC-1 cells, where the IC 50 value of the free Dau was also substantially higher than for the other cancer cell lines. Higher values of relative potency indicate a loss of potency of the conjugates with respect to free Dau [42,43].…”

In Vivo Tumor Growth Inhibition and Antiangiogenic Effect of Cyclic NGR Peptide-Daunorubicin Conjugates Developed for Targeted Drug Delivery

“…A variety of synthetic therapeutics have been used which target receptors, and hence revealed a significant tumor growth inhibition in vitro and in vivo. Thus, HT-29 human colon tumors were implanted to the intestine of immunodeficient SCID mice [75][76][77].…”

Energy‐resolved HCD fragmentation of daunorubicin‐peptide conjugates

“…Daunorubicin was used for the development of a large number of peptide-drug conjugates in our laboratory that showed efficient antitumor activity not only in vitro, but also in vivo. 4,5 Daunorubicin consists of an anthraquinone aglycon part and a daunosamine sugar moiety linked to the tetracycline by a glycosidic bond. Due to the complex structure of PDCs, tandem mass spectrometry is an indispensable tool for verification of the structure and purity of synthetic products.…”

“…Mass spectrometry is widely used to determine the release of drugs from PDCs, and to identify their metabolites. [4][5][6][7][8] In the case of anthracycline containing PDCs, however, mass spectrometric analysis is hindered by the degradation of the compounds during electrospray ionization (ESI). ESI is a soft ionization technique, producing intact, singly or multiply protonated molecules from peptides.…”