Toward the development of a novel non-RGD cyclic peptide drug conjugate for treatment of human metastatic melanoma

Abstract

The newly discovered short (9 amino acid) non-RGD S-S bridged cyclic peptide ALOS-4 (H-cycl(Cys-Ser-Ser-Ala-Gly-Ser-Leu-Phe-Cys)-OH), which binds to integrin αvβ3 is investigated as peptide carrier for targeted drug delivery against human metastatic melanoma. ALOS4 binds specifically the αvβ3 overexpressing human metastatic melanoma WM-266-4 cell line both in vitro and in ex vivo assays. Coupling ALOS4 to the topoisomerase I inhibitor Camptothecin (ALOS4-CPT) increases the cytotoxicity of CPT against human metastatic melanoma cells while reduces dramatically the cytotoxicity against non-cancerous cells as measured by the levels of γH2A.X, active caspase 3 and cell viability. Moreover, conjugating ALOS4 to CPT even increases the chemo-stability of CPT under physiological pH. Bioinformatic analysis using Rosetta platform revealed potential docking sites of ALOS4 on the αvβ3 integrin which are distinct from the RGD binding sites. We propose to use this specific non-RGD cyclic peptide as the therapeutic carrier for conjugation of drugs in order to improve efficacy and reduce toxicity of currently available treatments of human malignant melanoma.

Conjugates of Cell Adhesion Peptides for Therapeutics and Diagnostics Against Cancer and Autoimmune Diseases

“…Recently, a new non-RGD cyclic peptide called ALOS4 (cyclo-(CSSAGSLFC)) was found to bind to overexpressed integrin α V β 3 in malignant melanoma cells (Figure 1; Table 1) [37]. ALOS4 peptide has been similarly explored as a tumor targeting peptide due to its binding affinity for α V β 3 integrin.…”

“…ALOS4 peptide has been similarly explored as a tumor targeting peptide due to its binding affinity for α V β 3 integrin. The binding site for ALOS4 on α V β 3 integrin is different from the binding site of RGD peptide [37]. However, this peptide can be used as an alternative method of delivering cytotoxic drug to cancer cells overexpressing α V β 3 integrin.…”

“…ALOS4 was conjugated to fluorescein isothiocyanate (FITC) via a γ-aminobutyric acid (GABA) linker, to make FITC-GABA-ALOS4 [37]. This molecule was engulfed by WM-266-4 melanoma cells via binding to upregulated α V β 3 integrins, as detected by FACScan [37]. The high deposition of FITC-GABA-ALOS4 was found at the tumor xenograft of WM-266-4 melanoma cells in nude mice, indicating tumor targeting by ALOS4 peptide.…”

“…The results showed that CPT-GABA-ALOS4 killed WM-266-4 cells in a dose-dependent manner to as much as 70% at 10 μM. In contrast, it only killed 30% of non-malignant HEK-293 with the same dose [37]. Because non-malignant HEK-293 do not overexpress α V β 3 integrin on their cell surfaces, the results suggest that selectivity of ALOS constructs for WM-266-4 over HEK-293 cells are due to the overexpression of α V β 3 integrins on WM-266-4 cells.…”

“…In addition, the free CPT and two other cytotoxic drugs consistently killed considerable percentages of cell-populations in both WM-266-4 and non-malignant HEK-293. This study presents a promising non-RGD targeting peptide for the enhancement of chemo-stability and effective tumor-specific delivery of cytotoxic drugs [37]. …”