Design and synthesis of 4′-O-alkyl-chitobiosyl-4-methylumbelliferone as human chitinase fluorogenic substrates

• New probes are reported that report faithfully on chitotriosidase activity.
• Transglycosylation is effectively prevented by sterically bulky appendages.
• Central glucosazido building block to act as an effective acceptor and donor synthon.
• trans-Selective glycosylations without the use of a neighbouring group.

The synthesis of three fluorogenic chitobiosyl derivatives, modified at the non-reducing 4′-OH with, either a methyl, an isopropyl or a cyclohexylmethyl substituent, is described. The 4′-capped 4-methylumbelliferyl chitobiosides are hydrolysed by the human chitinase CHIT1 following Michaelis–Menten kinetics and in contrast to unmodified chitobiosyl-4-methylumbelliferone do not undergo transglycosylation. The compounds are also relatively poor hexosaminidase substrates and thus provide useful alternatives to 4′-deoxychitobiosyl-4-methylumbelliferone, previously reported by us as fluorogenic substrate to monitor CHIT1 activity as a marker for Gaucher disease state.

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