Side chain effect in the modulation of αvβ3/α5β1 integrin activity via clickable isoxazoline-RGD-mimetics: development of molecular delivery systems

Abstract

Construction of small molecule ligand (SML) based delivery systems has been performed starting from a polyfunctionalized isoxazoline scaffold, whose α v β 3 and α 5 β 1 integrins' potency has been already established. The synthesis of this novel class of ligands was obtained by conjugation of linkers to the heterocyclic core via Huisgen-click reaction, with the aim to use them as "shuttles" for selective delivery of diagnostic agents to cancer cells, exploring the effects of the side chains in the interaction with the target. Compounds 17b and 24 showed excellent potency towards α 5 β 1 integrin acting as selective antagonist and agonist respectively. Further investigations confirmed their effects on target receptor through the analysis of fibronectin-induced ERK1/2 phosphorylation. In addition, confocal microscopy analysis allowed us to follow the fate of EGFP conjugated α 5 β 1 integrin and 17b FITCconjugated (compound 31) inside the cells. Moreover, the stability in water solution at different values of pH and in bovine serum confirmed the possible exploitation of these peptidomimetic molecules for pharmaceutical application.

Selective Integrin Ligands Promote Cell Internalization of the Antineoplastic Agent Fluorouracil

“…Although these alterations impair crucial components of apoptotic process, it has been shown that 5-FU is able to trigger apoptosis in a time- and concentration-dependent manner in Jurkat cells . HEK293 cells were employed as noncancerous control cells; moreover, this cell line does not express α 4 nor α 5 integrin subunit, whereas it endogenously expresses the integrin subunit β 1 . ,, …”

“…53 HEK293 cells were employed as noncancerous control cells; moreover, this cell line does not express α 4 nor α 5 integrin subunit, whereas it endogenously expresses the integrin subunit β 1 . 14,32,54 Jurkat, K562, and HEK293 cell lines were exposed to 5-FU; reference compounds A, 13, 15. and the three 5-FU-conjugates D, E, and F (10−50−100 μM) for 72 h. Apoptosis was evaluated by Annexin V assay as described in the Materials and Methods. As shown in Figure 4, 5-FU was able to induce apoptosis in all three cell lines considered; interestingly, the proapoptotic effect of 5-FU was concentration-dependent.…”

“…53 HEK293 cells were employed as noncancerous control cells; moreover, this cell line does not express α 4 nor α 5 integrin subunit, whereas it endogenously expresses the integrin subunit β 1 . 14 , 32 , 54…”

“…In addition, integrins can be internalized upon specific ligand binding − and therefore they may be used as shuttles to selectively release the antineoplastic drug only inside integrin expressing cancer cells . Chemotherapy is considered the standard of care for several locally advanced cancers.…”

Fast MacMillan’s Imidazolidinone-Catalyzed Enantioselective Synthesis of Polyfunctionalized 4-Isoxazoline Scaffolds

“…Our synthetic ligands contained 4-isoxazolines as the core of the peptidomimetics, which are further decorated with suitable branches for binding therapeutics and diagnostics. 8 …”