Discussion
Patient noncompliance is a significant barrier to successful TB management. According to a recent meta-analysis, patients who missed 10% or more of their doses contributed to poor treatment outcomes (113). The risk of mortality, morbidity, and drug resistance increases dramatically in those who do not show adherence to the TB treatment15. Noncompliance with TB treatment is multifaceted, but pill burden and dosing frequency play a significant role (114). As a result, there is an urgent need for new formulations and fixed dose combinations (FDC) that can provide all drugs in one pill with reduced dosing frequency.
Our center has worked on PLGA nano-formulation of anti-tubercular drugs for the treatment of pulmonary tuberculosis10. The nanoformulation has been evaluated for oral and inhalation administration in animals and a phase one ascending dose study is ongoing 14. A sustained release profile and better therapeutic effect were been seen 14. However, there is not a single human study of this or similar formulation for the evaluation of pharmacokinetics, pharmacodynamics and biodistribution profile. Before conducting Pharmacodynamics studies in humans, we require complete evaluation of safety, pharmacokinetics and biodistribution profile in humans16. Therefore, we have elucidated the biodistribution pattern of PLGA loaded rifampicin nanoparticles in humans under gamma camera by using radiolabeled rifampicin. We exclusively tested rifampicin in this investigation because we wanted to get accurate results free from drug interference. Another reason for using only rifampicin was that it is used to treatMeningococcal meningitis, Methicillin-Resistant Staphylococcus Aureus (MRSA), Neisseria gonorrhoeae, Haemophilus influenzae, and Legionella pneumonia infections in addition to tuberculosis.
Radio-pharmaceuticals have an emerging role in diagnosis, therapeutic, or in both (theranostics). They have also been used in biodistribution and mass-balance studies to see the fate of a drug in a living system non-invasively. In this study, we labeled rifampicin by indirect method with 99mTc to track the distribution of nanoparticles in healthy human volunteers. The various parameters have been standardized to obtain the highest radiochemical purity. When 500µg of rifampicin, 15mCi activity, pH ranges between 6.5-7, was incubated at 80oC for 30 minutes in step one and at 100oC for 40 minutes in step two, more than 97 percent radio-labeling yield was obtained. Animals were used to evaluate the labeled rifampicin for polarity and distribution pattern changes. The labeled rifampicin was found in every organ, with the highest concentration in the stomach, followed by the lungs, liver, and intestines, which is consistent with previous findings17.
In our study, we have evaluated rifampicin PLGA nanoparticles because we wanted to study how the drug will behave when given as a nano-formulation. This information would be applicable to other anti-tubercular drugs too. A significant difference in plasma level of rifampicin between the 5 to 15 hours of oral administration in nanoformulation of rifampicin group compared to conventional group (Fig.7). In nanoformulation group, the plasma levels of rifampicin were detectable over five days but were below the limit of quantification (BLQ) of 250 ng/ml. We have done the confirmatory animal PK and biodistribution studies, in which, again, we got the plasma level of the drug in BLQ. The reason for limited sustained release could be dissipation of nanoparticles with feces after 24 hours which is average gut transit time18. In addition to this, significant drug concentration was observed in lymph nodes and spleen on the day sixth after administration. This presence of drug could be attributed to the smaller sized (100 nm or less) nanoparticles, absorbed through the para-cellular spaces from GI track1920. Further in lymph nodes the drug concentrations detected was also high i.e. up to 12 µg/ml on 6th-day. The possible explanation could be the denominator effect as the lymph nodes harvested from each animal were 200-300 µgs of weight. A significant difference in the PK parameters in the two groups have been found, as elimination constant in nanoformation has been decreased, which could have increased the other parameters like t1/2, AUC, AUMC, and MRT21. The significant difference in AUC/MIC and percentage T>MIC could be because of sustained release behavior of nanoformulation.
The free movement of nanoparticles throughout the GIT indicates major proportion of nanoparticles remained un-absorbed. Nanoparticles were started to get accumulated in the distal parts of the GI tract from the 5th hour of oral administration, which is in concordance with gastro-illic transit time. Lucia et al. 2019 have done an ex-vivo study of PLGA-PEG nanoparticles loaded with a fluorescent dye, rhodamine B, and the anti-cancerous drug paclitaxel. In that study, they saw the nanoparticles were in the gut lumen and started accumulating in the large intestine as time increased after oral administration.
They have also explored the absorption pattern of nanoparticles in the GI track, persistent luminal localization of nanoparticles has been seen at different time points 22. Same pattern has been seen in our study, we found nanoparticles were get accumulated in large intestines gamma camera and showed the release of drugs from there. The higher counts in the intestines and lower counts in blood and urine over the 24 hours in the nano-formulation group as compared to the conventional group confirm the accumulation of nanoparticles in the large intestines.
In conclusion, our study has shown the bio-distribution and pharmacokinetic profile of PLGA nanoparticles in humans. Nanoparticles get accumulated into distal parts of the gut from where the encapsulated drug is released in a sustained release manner. Although the sustained release of seven days was not observed in healthy human volunteers, an increase in pharmacokinetic parameters in the nanoformulation group could be utilized to decrease the dose of ATT for the treatment of pulmonary tuberculosis.
Acknowledgment:
This work was funded by ICMR, New Delhi, India.
Conflict of interest
We don’t have any conflict of interest.
Data availability and statement
Research data are not shared
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