Design,Synthesis And SAR Study On Inhibitors Preventing Biosynthesis Of Fungal GPI-anchored Proteins

Fungus infections are still an important source of morbidity and mortality,they cause extravagant expenditure to the healthcare systems.Because the vast majority of life-threatening fungus infections influences people with altered immune function,the increased incidence of invasive fungus infections can be correlated with an expansion in the number of people living with conditions or treatments that effect immune function,include primary immune deficiency,HIV/AIDS,hematologic and solid organ transplantation,cancer chemotherapy,prematurity,and immune-modulatory medications.The overall mortality for invasive diseases is 30–50% that caused by Aspergillus spp.and Candida spp.,although the advent of therapeutic strategies and new diagnostic.The therapeutic method are quite limited and include only three structural classes of drugs for invasive fungus infections: polyenes,azoles,and echinocandins.As a polyene drug,Amphotericin B is a fungicidal drug and has broad spectrum activity,but it is can only use as parenteral agent and has significant toxicity.Azole antifungal drugs,such as fluconazole,voriconazole,posaconazole,and itraconazole,are now widely used for the treatment of serious fungus infections because of their more favorable safety profile and broadspectrum activity.But the isolates resistant to azoles are increasingly reported.Echinocandin derivatives are favorable safety properties with broad spectrum activity,but are exclusively parenteral agents and are has no activity to Cryptococcus neoformans.Hence,there is a critical need for new antifungus compounds that have different mechanisms of action,broad-spectrum activity,and no cross-resistance to the currently available antifungus drugs.Glycosylphosphatidylinositol(GPI)-anchored mannoproteins are one of the major cell wall components of eukaryotic microorganisms,including fungus.In fungi,more than half of GPI-anchored proteins have an unknown function,but it is noted that some of them are involved in and are likely vital to cell wall adhesion to host cells and regeneration.These findings suggest that studies on fungus GPI-anchors are a promising research area for finding new therapeutic targets for human fungus infections.Eisai Co.,Ltd.,focused on the inhibition of GPI-anchored protein biosynthesis as a therapeutic target for the treatment of fungal infections.By screening chemical libraries that inhibited fungal cell wall assembly via GPI-anchored proteins in yeast,they found an inhibitor,1-(4-butylbenzyl)isoquinoline,and its target molecule,the protein GWT1,which is required for inositol acylation at an early step of the GPI biosynthetic pathway.Although some active inhibitors of GPI-anchored proteins have been reported,the study of the structure-activity relationship is rarely reported.Combined with previous structural modification and transformation of compound 10 b,this paper focused on the optimization of our previously studied antifungal compound 41d(MIC80 = 0.0313μg / ml)to proceed compound design,synthesis and antifungal activity tests,and SAR discussion.The structure of the lead compound 41 d was divided into four parts including A ring,B ring,C ring and the linkers,which will be replaced or modified with other moieties.The optimization and SAR are discussed as follows:(1)Eight compounds of class A were designed and synthesized to simplify the structure 41 d by deleting its C ring and the linkers,and replacing B ring with phenyl ring,aliphatic cyclic rings or aliphatic chains.Compound ZCX-GPI-A2 has the highest activity(MIC80 = 1μg / ml)among class A.However,the activity of simplified structures has greatly reduced by comparing with 41 d.(2)Six compounds of class B were designed and synthesized by modifying the linkers in order to increase the water-solublity.The compounds with hydroxyl group introduced to the linker are not active.ZCX-GPI-B6,with extended linkers by one methylene group,exhibited decreased activity(MIC80 = 0.25μg / ml).Shielding the hydrogen of the amino group in the linker also reduced the activity(ZCX-GPI-B5,MIC80 = 1μg / ml).(3)The replacement of the thiophenyl ring with 1,2,3-triazole rings gave 7 compounds of class C,but all the copounds showed no activity.Thus indicated that the thiophenyl ring is an important moiety to their antifungal activity.(4)Twenty three compounds of class D were designed and synthesized by maintaining the B ring as thiophene ring and modifying the other parts of the structure,including(1)replacing amino pyridine ring of A ring with 6-quinolyl ring,3-quinolyl ring,3-isoquinolyl ring,and 7-indolyl ring;(2)replacing the substituents on the C ring,for an example,the amino moiety is converted to an oxygen atom of the connecting portion.In vitro antifungal activity test showed that ZCX-GPI-D19(MIC80 = 0.0313μg / ml)and ZCX-GPI-D21(MIC80 = 0.0313μg / ml)exhibit the same antifungal activity with the lead compound 41 d.Thus indicate that the replacement of A ring with 6-quinolyl ring,as well as the conversion of NH group of the linker into oxygen atom could increase the antifungal activity.Moreover,the C ring substituents at the ortho or para position resulted higher activity than that at meta position.(5)Seven compounds of class E designed and synthesized by replacing thiophenyl moiety of B ring with furanyl ring,and replacing 6-amino pyridine ring with quinolinyl ring.The title compound ZCX-GPI-E6 exhibited the best activity(MIC80 = 0.25μg / ml),which is lower than that of 41 d.This suggests that the activity of thiophene ring substituent is better than furan ring substituent.All the 51 compounds we design and synthesis were confirmed by H-NMR and LC-MS,and the structure-activity relationships were discussed in depth according to the antifungal activity,we provide an experimental basis for design and synthesis GPIanchored protein inhibitors for future.