Structure- and ligand-based optimization of fragment binding tyrosinase from Agaricus bisporus to develop anti-melanogenic agents

Tyrosinases (TYR, EC 1.14.18.1) are metalloenzymes, occurring in a broad range of organisms. The catalytic domain is highly conserved and encloses a pair of copper atoms and six histidine residues coordinating them. The human isozyme, catalysing the conversion of L-tyrosine in L-dopaquinone, is involved in the physiological biosynthesis of melanin. However, its unregulated production could be responsible for hyperpigmentation and related skin diseases. Therefore, TYR inhibition was investigated as a viable strategy for tackling these diseases. Efficient and selective tyrosinase inhibitors are mainly identified by two approches: i) extraction and isolation from natural sources; ii) organic synthesis and structural optimization of lead compounds. Even if a lot of natural and synthetic inhibitors have so far been reported, many of them suffered from side effects in humans. In this scenario, my PhD research project was directed at the design, synthesis and in vitro evaluation of a large collection of TYR inhibitors with potentially better pharmacological profile. The 4-fluorobenzyl and 4-(1-piperazinyl)phenol moieties have been elected as key portions, given their ability to bind the bi-copper active site of TYR. Previous in silico and crystallographic studies on tyrosinases suggested that the 4-fluorobenzyl motif exhibited a prominent role. Following this assumption, the 4-fluorobenzylpiperazine fragment was explored by developing three series of derivatives; moreover, biososteric replacements on the obtained hit compound (IC50= 0.18 µM) resulted in (4-pyridinyl)-4H-1,2,4-triazole-based analogs. The structural optimization by means of different types of aromatic "tails" led to AbTYR inhibitors exhibiting higher efficacy than the reference compound kojic acid (IC50= 17.76 µM) and revealing antimelanogenic effects in absence of cytotoxicity. In parallel, it was investigated the 4-(1-piperazinyl)phenol bulding block, which similarly to other natural phenols exibits per se TYR inhibitory properties (IC50= 28.87 µM). The introduction of a variety of substituted benzoyl rings led to an improvement of the inhibition for several derivatives. Finally, hybrids between the two well-investigated fragments were designed and prepared. In addition to a relevant inhibitory activity, the best active compound (IC50= 3.80 µM) showed antioxidant effects.Altogether, the collected full SAR considerations for the newer classes of inhibitors as well as the reported data about their biological properties could provide the basis for future insights in the TYRIs field.