Novel targeted lipid prodrugs to improve cellular absorption of acyclovir and topical delivery to the eye via nanomicellar approach
Abstract
Biotinylated lipid prodrugs of acyclovir (ACV) were designed to target the
sodium dependent multivitamin transporter (SMVT) on the intestine and cornea to
facilitate ACV cellular transport. All the prodrugs were screened for their uptake across
MDCK-MDR1, Caco-2, human corneal epithelial cells (HCEC) and interaction with
SMVT on freshly excised rabbit corneas. Further confirmation of interaction of
biotinylated lipid prodrugs with SMVT was evaluated by docking analysis. The
cytotoxicity of the prodrugs was evaluated on MDCK-MDR1, Caco-2, HCEC and rabbit
primary corneal epithelial cells (rPCEC). The enzymatic stability of all the prodrugs was
studied in various ocular tissue homogenates. Finally, the in vitro antiviral activity of the
prodrugs was studied against HSV-1, HSV-2, HCMV and EBV viruses. Uptake of
biotinylated lipid prodrugs (B-R-ACV and B-12HS-ACV) was significantly higher than
B-ACV, R-ACV, 12HS-ACV and ACV in all cell lines studied. Transepithelial transport
studies across rabbit cornea showed lower [3H] biotin permeability in the presence of
biotin and biotinylated prodrugs, indicating recognition of the prodrugs by SMVT on
cornea. Average Vina scores obtained from docking studies further confirm that
biotinylated lipid prodrugs of ACV possess higher affinity to SMVT over B-ACV. None of the prodrugs exhibited any cytotoxicity on MDCK-MDR1, Caco-2, HCEC and rPCEC cells which suggest that these compounds were safe and non-toxic. B-R-ACV and B-
12HS-ACV were found to be relatively more stable in all ocular tissue homogenates in
comparison to non-lipidated but biotinylated prodrug, B-ACV. B-R-ACV and B-12HSACV
were found to possess excellent antiviral activity against HSV-1 and HSV-2.
Therefore, biotinylated lipid prodrugs of ACV appeared to possess enhanced affinity
towards SMVT. Synergistic improvement in cellular uptake is probably due to
recognition of the prodrugs by SMVT on the intestine/cornea and lipid mediated
transcellular diffusion. These compounds displayed excellent tissue stability and
minimal/no cytotoxicity. Also, these novel compounds exhibited excellent antiviral
activity against HSV-1, HSV-2 and EBV. Micellar carriers composed of two non-ionic surfactants, Vitamin E TPGS
stabilized with octoxynol-40 in a defined ratio were investigated for the delivery of
biotinylated lipid prodrug of ACV to the eye. Mixed nanomicelles were characterized by
their hydrodynamic diameter, zeta-potential, polydispersity index and surface
morphology. Furthermore, in vitro biocompatibility studies such as cytotoxicity
assessment, pro-inflammatory gene and cytokine expression analyses were carried out by
MTT assay and real time PCR analysis, respectively. The average micelle size was 10.46
nm with a PDI of 0.086 for unloaded micelles, and 10.78 nm with a PDI of 0.075 for
prodrug loaded micelles. TEM analysis confirmed that the micelles were spherical and
homogenous, and devoid of aggregation. Unloaded and loaded micelles did not differ in
terms of morphology. The particle sizes visualized by TEM were very similar to the size
obtained by DLS. The formulations did not exhibit any cytotoxic or inflammatory effects
on HCEC cells.
Table of Contents
Literature review -- Sodium dependent multivitamin transporter (SMVT): a potential target for drug delivery -- Rationale for investigation -- Targeted lipid based drug conjugates: a novel strategy for drug delivery -- Functional and molecular aspects of biotin uptake via SMVT in human corneal epithelial (HCEC) and retinal pigment epithelial (D407) cells -- Novel biotinylated lipid prodrugs of acylovir for the treatment of herpetic keratitis (HK): transporter recognition, tissue stability and antiviral activity -- Synthesis and antiviral activities of biotinylates lipid prodrug of acyclovir (B-10HD-ACV): interaction with SMVT -- Nanomicelles: an emerging platform for drug delivery to the eye -- Aqueous nanomicellar formulation for topical delivery of biotinylated lipid prodrug of acyclovir: formulation development and ocular biocompatibility -- Summary and recommendations -- Appendix
Degree
Ph.D.