## 6 PLGA Nanoparticles Functionalization
### BBB Transcytosis
#### 6.2.1 Cell Penetrating Peptides
CRT is used to improve BBB penetration of drugs, since it is an iron-mimic peptide that targets the protein complex of transferrin and the transferrin receptor (TfR). The average NP size was around 130 nm, which was acceptable for BBB penetration. Consistent with previous reports [152, 153], the S1 and curcumin loaded NPs crossed the BBB with low efficiency, whereas the CRT-NP-S1+Cur NPs had remarkably improved cellular uptake and BBB penetration ability in vitro, which suggests that the CRT peptide increases permeation of the BBB to PLGA NPs. According to these results, the higher number of CRT-NPs in the brain, the more they penetrate, indicting the contribution of CRT to increasing the penetration of PLGA NPs. The results of the behavioural tests demonstrated that these NPs significantly improved the spatial memory and recognition in AD mice, in addition to remarkably decreasing the A\(\beta\) deposit burden. Therefore,  
Figure 8: Conjugation of peptides to the surface of PLGA NPs through the maleimide-thiol reaction (based on [134]). compared to other PLGA NPs, CRT peptide-modified PLGA NPs codelivering S1 and curcumin showed potential advances for the treatment of AD mice [107]. Following the same line, B6 peptide (CGHKAKGRPK) is also considered as a promising candidate for enhancing drug delivery into the CNS due to its ability to target TftR as a substitute for the transferrin protein [154; 155; 156]. In fact, it is derived from a phage display library, and has previously shown high permeability across the BBB [157] and higher accumulation in brain capillary endothelial cells [158]. Thus, it is suggested that it would also improve curcumin bioavailability in the brain. Therefore, [PERSON] et al. [106] prepared PLGA-PEG NPs loaded with curcumin and conjugated with the B6 peptide, obtaining NPs with a size less than 150 nm. These PLGA-PEG-B6/Cur NPs significantly increased curcumin cellular uptake in vitro and could remarkably improve cognitive impairment in APP/PS1 mice, indicating that these NPs can profoundly improve curcumin delivery efficiency to the brain. Moreover, ex vivo studies demonstrated that they could reduce hippocampal \(\beta\)-amyloid formation and deposits, as well as tau hyperphosphorylation. Thus, these NPs may serve as an interesting strategy for the treatment of AD [106]. Another strategy to overcome the hydrophobic nature and nonsolubility in water of curcumin is conjugating PLGA NPs with a targeting Tet-1 peptide moiety. Tet-1 is a 12- amino acid peptide (HLNILSTLWKYR) identified through phage display, which has the binding features of tetanus toxin [159], thus presenting affinity to neurons, and could target cargos to the brain, bypassing the BBB [160]. For this reason, [PERSON] et al. [104] synthesized water-soluble PLGA coated-curcumin NPs and coupled these NPs with Tet-1, resulting in NPs with a size between 150-200 nm. Surface modification with Tet-1 peptide increased neuronal targeting efficiency in vitro. Indeed, enhanced neuronal uptake of curcumin-PLGA NPs was observed when compared to nontargeted curcumin-PLGA NPs. Therefore, these results indicate that Tet-1-targeted PLGA-coated curcumin NPs can be additional tools for the treatment of neurological diseases, in particular AD, with respect to its antiamyloid and antioxidant activities [104]. However, this is a preliminary in vitro study that needs more detailed in vivo investigations to draw further conclusions. [PERSON] et al. [101] employed another phage display peptide library with the purpose of isolating peptides that may be exploited to target delivery systems to the BBB and finally selected a 12-amino-acid-peptide (denoted as Pep TGN). They covalently conjugated Pep TGN (TGNYKALHPHNG) onto the surface of PEG-PLGA based NPs. The surface-modified PLGA NPs with Pep TGN had sizes about 100 nm and resulted in an improved delivery of NPs across the BBB and brain targeting.