Design,Synthesis And Biological Activities Of Amide And Pyridazine Derivatives As Novel PDE4 Inhibitors

Phosphodiesterase-4(PDE-4)is bimetallic hydrolase that specifically catalyze the hydrolysis of second messenger cAMP,and is highly expressed in inflammatory and immune cell type’s relevant tissues,central nervous system(CNS)and smooth muscles of the lung.The involvement of PDE4 in pathological processes associated with these tissues suggests that it is a great potential drug target.The PDE4 family is comprised of 4 primary gene products(PDE4A,PDE4B,PDE4C and PDE4D),which have three highly homologous regions in amino acid sequence,including one catalytic domain in a central portion of amino acid sequence and two upstream conserved regions(UCR1 and 2)in the N terminus.In spite of safety-related issues,some PDE4 inhibitors have been approved,such as roflumilast for the treatment of COPD and apremilast for psoriatic arthritis.However,many other PDE4 inhibitors with good activities were discontinued from development because of the potential overlap of efficacy and undesired side effects,such as nausea and emesis.It prompted us to discover novel PDE4 inhibitors.In this paper,we describe the design and synthesis of(1)fourty-eight catechol-based amides derivatives and(2)twenty-two pyridazine derivatives as PDE4 inhibitors,using roflupramand chlorbipram as lead compounds.Part I.Design and synthesis of catechol-based amides derivatives as novel PDE4D inhibitorsWithin the last decade,considerable efforts for the biological role of PDE4D have implicated that the PDE4D gene was associated with inflammation,depression,cognition-enhancing effects,brain cancer and ischemic stroke.On the other hand,PDE4D gene might be associated with the CNS-related emesis and diarrhea side effects.It is not entirely clear whether PDE4D gene is the sole cause of the said adverse events,but its potential prompted us to design and synthesize selective PDE4D inhibitors.Recently,our research group has been interested in the synthesis and biological evaluation of PDE4D inhibitors,and found roflupram showed antidepressant effects and memory-enhancing effects without emetic potency in animal models.Thus,we design and synthesize a series of novel catechol-based amidesderivatives as PDE4D inhibitors,using roflupram and roflumilast as leading compounds.In particular,amides of varioussubstituted amine were extensively explored,and structural modifications of the catechol moieties were performed to maximize the inhibitory potency.And these catechol-based amide derivatives were synthesized via seven or four steps with good yields from 3,4-dihydroxybenzaldehyde and methyl 3-hydroxy-4-methoxybenzoate respectively.Since recent studies have indicated that the catalytic domains of all subtypes PDE4 own a single active-site amino acid determining nucleotide selectivity,and have the high degree of sequence conservation.Therefore,preliminary tests on the catalytic domains of PDE 4(PDE4 CAT)in vitro activities were performed,using rolipram as positive control.The results showed that most of compounds CB-B01～04 and CB-C01～10 exhibited less PDE4 inhibitory activities than rolipram,while most of compounds CB-A01～18 and CB-D01～16 showed potent PDE4 inhibitory activities.More interestingly,compounds CB-A03,CB-A07,CB-A09,CB-A10,CB-A11,CB-A14,CB-A15,CB-A16,CB-D10 and CB-D15 showed most PDE4 inhibitory activity,and up to 27.5-fold higher activity than rolipram.Our structure-activity relationship studies of compounds CB-A01～18,CB-B01～A4,CB-CO1～10 and CB-D01-16 revealed that the effect of the substitutions on N atome toward PDE4 are as follows:1)when the substitutions(R3)on N atome is alkyl group,i-propyl>n-propyl>n-butyl>4-MeOPhCH2>PhCH2>diethyl>2-MeOPhOCH2CH2>n-Octyl>>3,4-MeOPhCH2;2)when the substitutions(R3)on N atome is aryl group,2-ClPh≈Py-3->Ph>N-Me-N-Py-2->>3,4-diMeOPh.And then,compounds CB-A03,CB-A07,CB-A09,CB-A10,CB-A11,CB-A14,CB-A15,CB-A16,CB-D10 and CB-D15 have been further tested on PDE4A4,and PDE4D7.Most of these compounds showed higher activity than rolipram.Except compound CB-A07,the PDE4D7 inhibitory activities of these compounds are higher than their inhibitory activities for PDE4A4.Moreover,CB-A16 with isopropyl group showed an increased inhibitory activity for PDE4D with 2.4-fold selectivity.To understand their nature of interactions with the PDE4D(PDB code:1XOQ),molecular docking studies were performed using CB-A11,CB-C08 and CB-D15.As anticipated,3-(cyclpentyloxy)-4-(difluoromethoxy)-phenyl moiety in these compounds occupied the pocket of it in the native ligand(roflumilast)with a small change.And the cyclpentyloxy moiety and difluoromethoxy moiety formed two hydrogen bonds with NH2 group of Gln369.Part Ⅱ.Design and synthesis of pyridazine derivatives as PDE4 inhibitorsIn our recent study,we have reported a novel PDE4 inhibitor chlorbipram without emetic effects in beagle dogs.Thus,twenty-two pyridazine derivatives(MBPD-A01～13 and MBPD-B01-09)as novel PDE4D inhibitors were designed and synthesized by using chlorbipram and roflumilast as leading compounds.Compounds MBPD-A01y13 were prepared by the reaction between substituted benzylbromide and corresponding pyridazinones.Compounds MBPD-B01～03 were synthesized by using substituted phenylacetonitrile and 3,6-dichloropyridazine as starting materials with good yields.Compounds MBPD-B04～08 were prepared by the reactions between MBPD-B02 and MBPD-B03 with alkyl amines.And compound MBPD-B09 was obtained by the hydrolytic reaction of compound MBPD-B01.The preliminary tests of these compounds on the catalytic domains of PDE 4(PDE 4 CAT)in vitro activity were performed using as rolipram a positive control.The results showed that most of compounds MBPD-A01-13 exhibited very weaker inhibitory activities for PDE4CAT,while MBPD-B01～09 showed higher inhibitory activities than MBPD-A01-13.Compared with rolipram,MBPD-B01 showed comparable inhibitory activities.Meanwhile,we docked chlorbipram into PDE4D(PDB code:3G4K)to understand the interactions of it with the PDE4D,and found two reasonable binding conformations.One of twoconformationsis similar to D159687).Both poses have predicted Ki at micromolar level(1.1 and 5.9μM,respectively),which below the predicted Ki(36.6 nM)of D159687.We guess the carbonyl oxygen on the benzyl ring causes the problem.So,we simply modify the third benzyl ring of D159687 which is replaced by pyridazine ring,and design biphenyl-pyridazine 81.Docking of compound 81 shows a great predicted Ki(93.5 nM),which is comparable with predicted Ki of D159687(36.6 nM).This result supports our conjecture,and can explain why MBPD-B01-09 showed higher PDE4 inhibitory activities than MBPD-A01～13.