# Anti-Biofilm Agents to Overcome _Pseudomonas aeruginosa Antibiotic Resistance_
## 3 Anti-Biofilm Agents to Tackle _P. aeruginosa_
### Analogs of Native Autoinducers  
(1)  AHL autoinducer analogs targeting LasR and RhlR  
A first strategy for the development of odDHL and BHL analogs has been the modification of the long acyl chain by the introduction of diverse aromatic rings. The development by [PERSON] et al. of novel AFLL analogs by solid phase synthesis gave them easy access to a wide variety of derivatives [51]. Notably, the introduction of bromophenyl and indole groups led to **ABA-5** and **ABA-6**, which inhibit LasR with IC\({}_{50}\) of 16.1 and 14.8 \(\upmu\)M, respectively. Thanks to their anti-LasR activity, these two compounds inhibit 100% of biofilm formation at 50 \(\upmu\)M (Figure 6). In 2024, the \(N\)-acyl chain of homoserine lactone (HSL) was substituted by [PERSON] et al. by a cinnamyl group [52]. Interestingly, the compound **ABA-7** bearing this conjugated carbonyl system and methoxy electron-donating group in _para_ position of the phenyl ring reduced biofilm formation by 36% at 100 \(\upmu\)M. Molecular docking studies revealed the formation of key hydrogen bonds between the HSL nucleus and Trp60, Asp73, Ser129 and Tyr56 residues in the autoinduction site of LasR, similar to those formed by the native autoinducer odDHL. Other modifications of the acyl chain of native AI consisted in introducing various spacers between the HSL core and an aromatic fragment, as described by [PERSON] et al. with the analog **ABA-8**[53]. The presence of a dithiocarbamate group enables the molecule to form two additional H-bonds between the phenylurea and the key residue Arg61 (Figure 6). In fact, **ABA-8**, which is the most active compound of the series, is the only molecule able to establish these essential interactions for potent anti-LasR activity. With a potent efficacy on all three QS systems (68%, 59% and 73% inhibition of LasR, RhlR and PqsR at 3 \(\upmu\)M, respectively), **ABA-8** inhibits biofilm formation by 36% at 15 \(\upmu\)M.  
Finally, [PERSON] et al. reported the replacement of the HSL core with a 3-amino-2-oxazolinone nucleus, well-known for its antimicrobial properties. [54] Pharmacomodulations on this scaffold brought out the compound **ABA-9** bearing a three-carbon acyl chain ended by a phenoxy group substituted by a bromine in its _para_ position (Figure 6). **ABA-9** revealed a good anti-biofilm activity with 30% inhibition of biofilm formation at 20 \(\upmu\)M.  
Figure 5: Essential interactions between natural autoinducers odDHL, BHL and PQS and their corresponding receptors LasR, RhlR and PqsR. No cytotoxicity data have been reported for those molecules. 2. AQ autoinducer analogs targeting PqsR The design of AQ analogs is mainly based on the use of quinolone biososters (Figure 7). Notably, [PERSON] et al. designed four ABAs based on pyranone and pyridinone patterns. First, the compound **ABA-10** was isolated from a series of 3-hydroxy-6-methyl-4_H_-pyran-4-ones substituted by a benzyl group in position 2 [55]. In fact, the _para_ substitution with a nitro group of this aromatic ring gave the best anti-biofilm activity with an IC\({}_{50}\) of 21.3 uM against the PAO1 strain. Interestingly, **ABA-10** showed no cytotoxicity below 50 uM in an MTT assay on murine macrophage RAW264.7 cells. Then, the authors demonstrated the relevance of the pyridinone scaffold, often found in iron chelators, for combining biofilm inhibition and iron trapping [56]. Their study explored various side alkyl chain lengths and different linkers between aromatic rings. The best compound, **ABA-11**, potently reduced biofilm formation (IC\({}_{50}\) of 6.6 uM) with an innovative mode of action. Indeed, the inhibition of QS systems is associated with a vectorization allowed via recognition by the pyoverdine receptor Fpva. The cytotoxicity of this siderophore analog appeared very weak on Vero monkey kidney cells (15% cell viability inhibition at 25 uM). From 2022, this research team combined the amide spacer of **ABA-11** with the suitably substituted aromatic ring of **ABA-10**[57; 58].