Triglyceride-mimetic approaches for targeted drug delivery to the lymphatic system: a mechanistic study
2017-01-23T21:40:54Z (GMT) by
The lymphatic system plays a number of roles in key physiological and pathological events, including propagation of the immune response, lipid transport and tumour metastases. Enhanced delivery of drugs to the lymphatic system therefore has the potential to benefit the treatment of diseases such as autoimmune disorders, immune diseases, metabolic syndrome, lymphoma and tumour metastases. Inspired by the realisation that dietary triglycerides (TG) are digested, absorbed, resynthesised and efficiently transported into and through the intestinal lymphatic system, this thesis examines TG-mimetic prodrugs as a potential conduit for drug delivery to the lymphatics. The studies reported herein investigate 1) the mechanisms underlying integration of lymph-directing prodrug strategies into lipid digestion-absorption-resynthesis-lipoprotein assembly pathways; 2) the factors that promote and that limit lymphotropic affinity and 3) potential strategies to combat such limitations. Mycophenolic acid (MPA, an immunosuppresant that acts by inhibiting lymphocyte proliferation) was chosen as a model drug for these studies as it has little intrinsic affinity for the lymph, but would benefit from targeted delivery to the lymphatic system where lymphocytes are resident at high concentrations. In the first set of studies, two classes of prodrugs of MPA (alkyl chain derivatives and TG mimetics) were examined with regard to their ability to promote lymphatic transport in rats. Three alkyl chain derivatives with varied chain lengths and linkers were examined, however, poor enzyme stability and low absorption appeared to limit lymphatic transport. In contrast, the TG mimetic, 1,3-dipalmitoyl-2-mycophenoloyl glycerol (2-MPA-TG), which was designed to promote biochemical integration into TG metabolism pathways, was significantly more effective in enhancing lymphatic transport. 2-MPA-TG resulted in increases in lymphatic transport of ~80-fold when compared to underivatised MPA. Subsequent mechanistic studies confirmed that lymphatic access of 2-MPA-TG resulted from a series of site-specific metabolic processes, including a) lipolysis in the intestinal lumen (facilitated by pancreatic lipases), b) re-esterification in enterocytes (enabled via glyceride acyltransferases), and 3) integration into intestinal LP (promoted by co-administered lipids). Each step was identified as being critical since inhibition of any of these processes resulted in significant attenuation of lymphatic transport. Importantly, following access of the re-esterification products of 2-MPA-TG into the lymphatics, conversion from pharmacologically inactive 2-MPA-TG derivatives to active free MPA was apparent and MPA concentrations in mesenteric lymph nodes were significantly higher than that obtained after administration of equimolar doses of MPA. Subsequent studies focused on the potential for structural change to the TG mimetic prodrugs to alter lymphatic transport and MPA release. The data revealed significant specificity with regards to the point of conjugation and the nature of the conjugation chemistry. MPA conjugation at the 2-position of the glycerol backbone and via an ester bond appeared to be critical requirements for optimal lymphatic transport. In addition, the insertion of straight chain alkyl linkers between MPA and the TG backbone appeared to potentiate parent drug release, although the overall extent of lymphatic transport of prodrug derivatives was not improved. Finally, methyl substitution of the alkyl chain, and β to the ester bond between the alkyl chain and the glyceryl backbone, was attempted to promote the stability of the digested monoglyceride derivative of the prodrug in the intestine. This was effective in enhancing lymphatic transport relative to the non-methylated equivalent but lymphatic transport remained below that for the directly linked 2-MPA-TG. Finally, the utility of the prodrug strategy was confirmed by studies conducted in conscious greyhound dogs where GI physiology is expected to be more representative of that in humans. Following oral administration of 2-MPA-TG to fed dogs, lymphatic transport of MPA related materials was markedly enhanced (288 fold, when compared to that after MPA dosing) and conversion to the pharmacologically active form, MPA, was evident in lymphocytes. The data demonstrate that triglyceride mimetic prodrugs provide a mechanism to increase lymphatic drug transport, and may thus ultimately provide opportunities in the treatment of lymph-resident disease.