## 2 Chemistry and Structure-Activity Relationship of Styrylquinolines _in vitro_
### Modulation of the spacer  
In a search for new structural features able to improve the biological profile of this class of inhibitors, the linker between the two aromatic units was modified. First of all the double bond was reduced to increase the conformational liberty between the two aromatic subunits. In an attempt to optimize the binding within the active site of the enzyme, polar functions able to interact through hydrogen bond with near amino acids were intercalated between the two rings, instead of the initial olefinic linker. Thus, compounds **79-82** possessing an amide, a hydrazide or a urea spacer were prepared, keeping constant the 7-CO\({}_{2}\)H, 8-OH pattern on the quinoline ring (Figure 3) [18]. Synthetically, the reduced quinoline **78** was easily available through catalytic hydrogenation of the corresponding SQL **35**[26]. The others derivatives were prepared using the pivotal acid **84** available in 5 steps from 8-hydroxy-2-methyl-quinoline-7-carboxylic acid (**3b**). Activation of carboxylic group of **84** using N-hydroxysuccinimide, followed by condensation with the requisite amines **87** provided amides **79** in 15-90% overall yield, after TFA deprotection using guaiacol as _tert_-butyl cation scavenger.  
Similarly condensation of **85** with hydrazines **88** afforded hydrazides **81** in 55-65% overall yield. On the other hand, [PERSON] rearrangement of acyl azide **86** and condensation of the intermediate isocyanate with aromatic amines gave the expected urea **82** which were deprotected as previously upon treatment with TFA. Curtius rearrangement in presence of benzyl alcohol, followed by hydrogenolysis of the resulting benzyl carbamate gave the C-2 amino-derivative which was converted to retro-amide **80** by treatment with benzoyl chloride and TFA deprotection (Scheme 9) [18].  
**Scheme 9.** Synthetic scheme of amides **79**, **80**, hydrazide **81** and urea **82**.  
The results of the biological evaluation of these linker-modified compounds keeping constant the 3',4'-dihydroxyphenyl subunit are depicted in Table 8. The reduction of the central double bond did not affect the integrase inhibition potency (compound **78**). In first place, this result demonstrated that the electron circulation from the catechol system towards the quinoline nitrogen, according to a \"pull-push effect\" was not required for inhibition. Furthermore an increased conformational liberty appeared not to be detrimental to the inhibition potency. Quinoline-2-carboxamide **92**, which possessed a two-carbon spacer displayed a slightly improved activity compared to the corresponding styrylquinoline **35**, whereas higher the length of the arm lower the activity. In this respect, both ureas **94** and **95** were found inactive (Table 8).  
Figure 3: Structure of inhibitors exhibiting a nitrogen containing spacer.  
To further study the influence of the ancillary ring on spacer-modified compounds, quinolines **96-106** were prepared according to Scheme 9 using various amine and hydrazine derivatives [18]. Dimeric hydrazide **106** was prepared upon treatment of _N_-hydroxysuccinimide **85** with hydrazine followed by TFA deprotection [19]. The biological evaluation of these new compounds is shown in Table 9. However in no case, were compounds with nitrogen modified spacers found to be more active than the corresponding styrylquinolines. Interestingly enough, the 3,5-dinitrophenylhydrazone derivative **104** was found to me unexpectedly active, although devoid of a hydroxyl group on the right hand. This result confirmed that the catechol moiety is not required for _in vitro_ IN inhibition.