Poster M4 - ESTIMATING TENOFOVIR and ATAZANAVIR DIFFUSION LENGTH IN THE LOBULE FROM IR-MALDESI IMAGE DATA

Introduction: Combination antiretroviral therapy (cART) has been transformative for treating HIV, enabling durable viral suppression in plasma for patients [1-3]. However, recent studies in humans and non-human primates have revealed that antiretroviral drugs do not distribute uniformly across tissues, and in some instances, are considerably excluded from lymph nodes (LNs) [4-7]. Imaging techniques like IR-MALDESI have visualized up to 30-fold variations in antiretroviral levels within the same LN, with the highest concentrations in sinuses and lowest in the lobules [7].
The LN’s unique structure and microenvironment may underlie this heterogeneous drug distribution. The lobules house the densely cellular regions critical for antigen presentation and lymphocyte interactions yet are semi-isolated from lymphatic sinuses and blood vessels. The controlled transport into the lobule through these two pathways potentially limits the ARVs’ capacity for free diffusion into the lobules, which could be the underlying reason for the observed heterogeneous distribution of these drugs within the LNs. Additionally, many HIV drugs are irreversibly phosphorylated following uptake into the cell, blocking exit of the drugs from the cell. Combined with the fact that the lobules are extremely cell-dense regions, this means that free drug in the extracellular space will be exceedingly sparse. Based on this understanding, we hypothesize that the transport of certain HIV antiretroviral drugs, such as Atazanavir (ATZ) and Tenofovir (TFV), may be dominated by cell-mediated transport via lymphocytes within the lobule.

Materials and

Methods: In this study, a steady-state reaction-diffusion model is used to describe the radial diffusion of ARVs from LN vasculature based on IR-MALDESI measurements of ARV distributions in non-human primate tissue for drugs Efavirenz (EFV), TFV, and ATZ representing multiple classes. As it shown in figure 1, each blood vessel is considered as an independent major source of drug. If we assume that the drug, whether intracellular or extracellular, transports within the lobule in a diffusion-like manner, and that the drug also decays within the lobule, then the steady-state concentration from a single point source follows a Bessel-K function of the distance from the center of the source. Bessel-K function depends only on the diffusion length (DL) of the ARVs. Then the confidence intervals for ARV’s DL were calculated using MCMC method, by a binomial distribution function to predict presence of drug in each pixel. Then, the results were compared with known values for DL of the free drugs and the effective DL of the drugs inside the lymphocytes in the LN [3]. Our analysis was done based on nine MALDI images for distribution of ARVs in LN. Nine LNs for TFV, six LNs for ATZ, three LNs for EFV.

Results, Conclusions, and Discussions: The DL of drugs within lymphocytes was calculated using this formula: √(D/δ), where D is the diffusion rate for lymphocytes, and δ is the decay rate for drugs inside the lymphocytes. The half-life of TFV within lymphocytes ranges from 15.5 to 60 hours [8], while the half-life of ATZ within lymphocytes is 12.1 hours [9]. The DL for free drug was calculated using the same formula, where D represents the effective diffusion rate for drugs within the LN lobules, and δ is the decay rate for the drugs in the tissue.
Figure 2-3 compares the 95% confidence intervals of the DLs for TFV and ATZ in each LN against the DLs of free drug and cell-mediated drug transport. Our results demonstrate that the DLs of TFV and ATZ fall within the range expected for cell-mediated drug transport mechanisms. This suggests that lymphocyte-facilitated transport plays a significant role in the distribution of these antiretroviral agents within the LN lobules.
For EFV, our model failed to reject the null hypothesis, which indicates a uniform distribution of EFV inside the LNs. This result is consistent with our observations from the EFV data gathered from LNs.

The findings of this study provide valuable insights into the mechanisms governing the distribution of antiretroviral drugs within LNs. The calculated DLs of TFV and ATZ fell within the range expected for cell-mediated drug transport, suggesting that lymphocyte-facilitated transport plays a crucial role in the dissemination of these antiretroviral agents within the LN lobules. This observation aligns with the understanding that the lobules are densely populated with cells and semi-isolated from the lymphatic sinuses and blood vessels, hindering free diffusion of drugs.
Overall, these results highlight the significance of considering cell-mediated transport mechanisms in the context of LN drug distribution, particularly for antiretroviral agents like TFV and ATZ. Understanding these mechanisms can inform the development of more effective strategies to enhance drug delivery to lymphoid tissues, which are critical reservoirs for HIV persistence.

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