Research On The Design,synthesis And Biological Activity Of Derivatives Of α-aminophosphonate

Since the discovery of the biological activity of amino phosphonic acid structure and its existence in organisms half a century ago,the chemical and biological research on these compounds has gradually developed into a unique branch of phosphorus chemistry.As natural amino acid analogues,especially,research ofα-aminophosphonate and its derivatives is the most extensive,current biological activity related has been extended to the antibacterial,antifungal,antitumor,antiviral,anti-inflammatory,parasitic resistance,oxidation resistance and even diabetes,lipid-decreasing fields,shows theα-aminophosphonate and its derivatives as advantage structure with wide space of exploration.Inspired by the concept of multi-target for drug development and in combination with the previous experience of finding many highly active molecules and new activities by coupling two pharmacophore groups in our laboratory,this research intends to carry out based on the novel conjugates ofα-aminophosphate and explore its design,synthesis,biological activity and structure-activity relationship;New compounds with dual mechanisms of action or new mechanisms of action are expected to be obtained and expand the research of organophosphorus compounds in pharmaceutical chemistry.1.Design and synthesis of target compoundsThe design of multi-target drugs will combine pharmacophore with same or different action mechanism through molecular coupling to obtain new compound with double action mechanism or new action mechanism,which is a powerful weapon against drug-resistant bacteria infection and complex and comprehensive diseases.As the analogues of natural amino acids,the biological activity ofα-aminophosphate mainly comes from the N-C-P characteristic structure in their molecules,which has a very large research space for derivatives.Therefore,this study draws on the concept of multi-target drug design and adopts the principle of drug combination to design derivatives in three directions while keeping the molecular framework unchanged.It is hoped that the active molecules with better structural activity than the single parent can be obtained by coupling pharmacophore with different mechanisms of action.Referring to the existing design ideas of heterozygous molecules,this study introduced a variety of pharmacophore based on the parent structure ofα-aminophosphonate and the target molecular model of"A-Linker-B".We designed a total of about 3,000 target molecules through the modification ofα-aminophosphonate substituents,different linkers,and pharmacophore.With the help of Plexus,a physical-chemical property computing platform of Eli Lilly,we optimized compounds and selected drug-like molecules,and finally obtained seven series（TM1-TM8）and a total of 117 target compounds.In view of the structure of the target compound,the synthesis of the target molecule was carried out using the synthesis route starting from the preparation ofα-aminophosphonate.Through experimental exploration,combined with column chromatography and thin plate chromatography,10 intermediates and 117 target compounds were synthesized.The structures of the compounds were confirmed by ¹H NMR,¹³C NMR and HR MS.2.The bioactivity of the target compoundsThe derivatives ofα-aminophosphonate have wide activity,among which the active molecules of antibacterial,antifungal and antitumor are common.In order to control the possible activity of the molecule,the authors introduced sulfonamide through sulfonyl reaction and introduced azole ring and fluoroquinolone structure in the mode of"A-Linker-B".Therefore,6 sensitive pathogenic strains and Pichia pastorius were selected to test the antibacterial and antifungal activity of the target molecules.Their inhibitory activities against Mycobacterium smegmatis and three types of tumor cells were then measured to study the anti-tuberculosis and anticancer activity of the target molecules.Anti-sensitive strain activity:Almost all molecules containing sulfonamide structure derivatives showed weak inhibitory activity against Acinetobacter baumannii,but the MIC value was far lower than that of the positive control drug.Other compounds showed weak inhibitory activity against Micrococcus gambogans,but none of the other four bacteria showed inhibition in the test concentration range.Almost all structural derivatives containing azole showed weak inhibitory activity against Acinetobacter baumannii,only a few target molecules showed MIC value of 128μg/m L against Escherichia coli,Salmonella and Pseudomonas aeruginosa,and all molecules showed no inhibitory ability against Staphylococcus aureus and Micrococcus gambogoma within the tested concentration range.Sulfonamides and azole derivatives showed selective inhibition against gram-negative bacteria.In contrast,most fluoroquinolone-containing derivatives showed high inhibitory activity against Staphylococcus aureus,with MIC values of 0.8～25.6μg/m L,and TM7-7 and TM7-8MIC values of 1.6μg/m L and 0.8μg/m L,respectively,which were equivalent to gatifloxacin,a corresponding fluoroquinolone drug.For the other strains,only a few molecules showed weak inhibitory activity,reflecting the selective inhibitory effect of fluoroquinolone conjugated derivatives on gram-positive bacteria.Antifungal activity:The target molecules containing sulfonamide fragments and azole fragments showed antifungal activity,with MIC values ranging from 128 to 256μg/m L in most cases,and only MIC values of the compound TM1-21 reached 64μg/m L.n the whole,the inhibitive activities of aromatic sulfonyl phenyl ring para and ortho-substituted compounds were weaker than those of meso-substituted compounds.Among the 56 azole derivatives,34 showed significant inhibitory effects on the growth of Pichia coli with MIC values ranging from 32 to 128μg/m L.Among them,the MIC value of the six compounds was 64μg/m L（TM2-3,TM2-11,TM3-2,TM3-8,TM3-9and TM4-3）,and the inhibitory effect of TM2-6 was the strongest,and the MIC value reached 32μg/m L,which was stronger than the positive control antibacterial drugs xacin,cefthiophene,vancomycin and ampicillin.The efficacy intensity was the same as that of fluconazole,which was worthy of further study.Overall,the effect of the active fragment on the activity of anti-picea was the azole fragment>sulfonamide fragment>fluoroquinolone fragment.Aromatic sulfonyl benzene ring para substitution and ortho substitution compounds in sulfonamide derivatives were weaker than meso-substituted compounds in inhibiting activity.Among azole derivatives,TM3 series compounds performed better when considering the the total number of active molecules or the number of highly active molecules,which may be related to the unique properties of electron-absorbing substituted fluorine atoms.In terms of the number of ester carbon atoms,methyl derivatives were slightly more dominant than ethyl derivatives in azole derivatives.When n-butyl derivatives of TM4series are added for comparison,it could be seen that with the increase of phosphate alkyl carbon chain,the inhibitory activity of derivatives was lower.It also shows that the N-C-P structure which is most affected by the steric hindrance of phosphate ester alkyl should be the active structure which plays an inhibitory role.Anti-mycobacterium smegmatis activity:In anti-TB activity tests,fluoroquinolone-containing derivatives exhibited inhibitory activity comparable to that of the positive control drug isoniazid,with MIC values of 12.5μg/m L or below,among which the MIC values of TM7-7 were 3.125μg/m L,and the MIC values of the remaining compounds were all greater than 100μg/m L.Anti-cancer activity:In the anti-tumor activity test,only HCT116 cells of three types of tumor cells showed weak inhibitory activity at a concentration of 5μM,among which only IM1-2 and TM2-4 showed an inhibitory rate of 25%.Some compounds are still being tested,and the activity against citrus canker disease is in a state of waiting to be measured.In this study,10 intermediates and 117 target compounds were synthesized.At test concentrations,most of the molecules were less resistant to bacteria,and some were barely as active as the positive control drugs.Most of the molecules showed good in vitro antibacterial activity against pichia yeast,some of which were as good or better than vancomycin.Only a few molecules have weak antitumor activity;The activity of pathogenic bacteria in citrus remains to be observed.