MULTIFUNCTIONAL HYBRID LIPID-PLGA NANOPARTICLES FOR DUAL DRUG DE-LIVERY: SURFACE FUNCTIONALIZATION WITH M12 PEPTIDE FOR MUSCLE TAR-GETING

Sarcopenia has become an increasingly significant biomedical and societal concern. This highlights the need for therapeutic strategies that can complement or replace lifestyle‑based interventions when these are insufficient or impractical. This work investigates a multimodal therapeutic strategy for sarco-penia by co‑delivering PEA and TMZ, two agents with complementary mechanisms. Previous studies showed that PEA, when delivered through hybrid lipid-PLGA nanoparticles, enhances an-ti‑inflammatory responses in C2C12 myoblasts [1]. Here, the formulation was redesigned to also en-capsulate TMZ, a metabolic modulator that inhibits mitochondrial β‑oxidation. The hybrid lipid-PLGA system enables the simultaneous incorporation of lipophilic PEA and hydrophilic TMZ within a single nanocarrier. To achieve selective muscle targeting after intravenous administration, the nanoparticles were further functionalised with the M12 peptide, a ligand known to direct nanocarriers toward muscle fibers and sarcomeric structures [2]. The optimisation process aimed to enhance drug encapsulation and release behaviour, ensuring sta-ble co‑loading and controlled systemic delivery. Substituting TMZ·dihydrochloride with TMZ‑Base signif-icantly improved nanoparticle performance, raising TMZ encapsulation from ~30% to ~74%. PEGyla-tion further increased colloidal stability and produced a slightly more sustained release profile. The re-fined co‑loaded hybrid PLGA nanoparticles also showed reduced particle size, improved reproducibil-ity, and controlled release of both TMZ and PEA. To achieve active muscle targeting, M12‑functionalised hybrid PLGA nanoparticles were then devel-oped. Functionalisation was performed by synthesising a PLGA‑PEG‑M12 conjugate through nucleo-philic substitution between NHS‑activated PLGA‑PEG and the peptide’s N‑terminal amine. FT‑IR and NMR analyses confirmed successful conjugation through the appearance of new vibrational features and peptide‑specific signals. Incorporating this conjugate directly during nanoparticle formation en-sured homogeneous peptide distribution throughout the nanosystem. The resulting M12‑functionalised nanoparticles displayed favourable physicochemical properties, with fresh particles around 146 nm and low polydispersity (PDI ~0.24). Their zeta potential was less negative than non‑functionalised ones, consistent with the peptide’s charged residues. Drug loading remained high, indicating that peptide incorporation did not compromise encapsulation. Overall, these results validate the effective M12 functionalisation and support the suitability of these nanoparticles for targeted muscle delivery.