• Evaluation of In Vitro/In Vivo Correlations for Transdermal Delivery Systems by In Vitro Permeation Testing and Human Pharmacokinetic Studies, With and Without a Transient Heat Application

      Shin, Soo Hyeon; Stinchcomb, Audra L.; Hassan, Hazem; 0000-0001-5091-8870 (2018)
      An in vitro model that exhibits in vitro/in vivo correlations (IVIVC) is a powerful tool in biopharmaceutical drug development because it can efficiently predict drug product performance in vivo. While the concept of IVIVC has been utilized mostly for oral dosage forms, demonstrations of IVIVC with in vitro permeation testing (IVPT) for transdermal delivery systems (TDS) are emerging. The objective of this work was to evaluate IVIVC for TDS using two model drugs, nicotine and fentanyl, with different physicochemical characteristics (e.g. log P). Additionally, the effect of heat exposure (42 ± 2 °C) on the rate and extent of TDS drug delivery was evaluated. IVPT studies using excised human skin and in vivo pharmacokinetic (PK) studies in human subjects were conducted under harmonized study conditions and designs to evaluate IVIVC. The correlations were evaluated in multiple ways, including a single point comparison of parameters such as steady-state concentration and heat-induced increase in partial AUCs, as well as a point-to-point correlation (Level A IVIVC). Level A IVIVC was examined using multiple approaches. A strong IVIVC was consistently observed for nicotine TDS in presence and absence of heat, suggesting the utility of IVPT as a tool to evaluate and predict in vivo performance of nicotine TDS. The IVIVC results for fentanyl were relatively weaker, especially when IVIVC for heat effects were examined, with greater in vivo heat effects observed compared to the in vitro heat effects. A separate study evaluating IVIVC for fentanyl TDS without a heat exposure component and utilization of some PK parameters obtained directly from study subjects yielded improved IVIVC results. The findings from the present research work suggest that IVPT data generally shows good predictability of in vivo performance of TDS at normal temperature conditions. However, the usefulness of IVPT for assessing and predicting external factors such as heat, especially for lipophilic drug molecules, may have some limitations that could be further improved.
    • Evaluation of skin tape stripping in healthy human volunteers as a methodology for quantifying local drug bioavailability from dermal products

      Shukla, Sagar; Stinchcomb, Audra L.; Hassan, Hazem (2020)
      Stratum corneum (SC) tape stripping is a valuable methodology that has been used for quantifying bioavailability (BA) of topical drug products at the site of action. Although the Food and Drug Administration (FDA) tape stripping guidance was withdrawn several years ago due to variable results, with an appropriate study design, tape stripping procedures can be a reproducible BA method. Therefore, the objective of this work was to investigate the use of tape stripping to quantify BA and evaluate in vitro/in vivo correlations (IVIVC) of two model compounds (lidocaine and diclofenac). These compounds were selected for their differing physicochemical properties and skin permeation rates. Two healthy human volunteer pharmacokinetic and tape stripping studies were conducted to quantify the BA in the SC and measure the elimination rate constant through the skin (kesc). Investigator variability from the in vivo tape stripping study was also examined, and the method variability potentially induced by the investigator can be mitigated by the quality by design (QBD) approach of using transepidermal water loss (TEWL) for determining when most of the SC has been removed in each individual volunteer. TEWL readings assisted the investigator by representing the SC tape stripping endpoint, and the SC masses removed from each volunteer were similar for the two investigators. Harmonized IVPT studies were also conducted and key parameter estimates were determined (absolute bioavailability (F) and (kesc)). These parameter estimates were used to simulate in vivo SC drug concentrations. The error in the SC drug concentration predictions from both in vivo studies was usually less than 20% compared to observed values, which demonstrates the predictive power of carefully harmonized IVPT studies. IVPT studies require less time and expense than human studies, and therefore these models play an important role in the early stages of drug development to predict in vivo SC drug concentrations, and absorption of drug through the skin. In this study, in vivo kesc for a quickly permeating drug (lidocaine) appears to be well predicted by IVPT; however, further work needs to be done to predict a slowly permeating drug’s (diclofenac) SC drug concentrations and kesc.
    • The Role of the Constitutive Androstane Receptor in Cyclophosphamide-based Treatment of Lymphomas

      Hedrich, William Dominic; Wang, Hongbing, Ph.D.; Hassan, Hazem (2018)
      Cyclophosphamide (CPA) is an alkylating prodrug which has been utilized extensively in combination chemotherapies for the treatment of cancers and autoimmune disorders since its introduction to the market in the late 1950s. The metabolic conversion of CPA to its pharmacologically active metabolite 4-OH-CPA is catalyzed primarily by cytochrome P450 (CYP) 2B6. CPA is also subject to metabolism by CYP3A4 to an inactive metabolite, N-dechoroethyl-CPA (N-DCE-CPA), and a neurotoxic byproduct, chloroacetaldehyde. CPA is the backbone of the frontline chemotherapeutic regimen used for the treatment of non-Hodgkin lymphoma which combines the CHOP (CPA-doxorubicin-vincristine-prednisone) regimen with rituximab, a monoclonal CD20 antibody. The constitutive androstane receptor (CAR, NR1I3), an orphan nuclear receptor, is recognized as the key mediator of xenobiotic-induced expression of CYP2B6, Importantly, mounting evidence suggests that activation of hCAR leads to preferential induction of CYP2B6 over CYP3A4 which suggests that selective hCAR activation may enhance CPA bioactivation and enhance the efficacy:toxicity ratio of CHOP chemotherapy for NHL. CHOP chemotherapy has been associated with severe and cumulative cardiotoxicity arising from the doxorubicin component of the regimen and it is recommended that lymphoma patients with existing heart conditions avoid treatment with the full CHOP combination. Recently, it has been demonstrated that Nrf2 (nuclear factor (erythroid-derived 2))-like 2, NFE2L2) plays a key role in governing doxorubicin-induced cardiotoxicity. Nrf2 regulates the expression of important antioxidant genes and proteins which protect tissues from damage due to oxidative stress and inflammation. It has been shown both that insufficient Nrf2 expression results in hypersensitivity to doxorubicin cardiotoxicity and that stimulation of Nrf2 by small molecule activators can provide protection from doxorubicin-mediated toxicity. Our hypothesis was that hCAR activation will increase hepatic expression of CYP2B6 while having a negligible impact on other genes responsible for the disposition of CHOP drugs. Further, activation of Nrf2 in cardiac tissue may provide protection against cardiotoxicity induced by the doxorubicin component of CHOP. Together, these gene expression alterations will lead to augmented antineoplastic activity of CHOP in target lymphoma cells while alleviating the untoward cardiotoxicity associated with this regimen. This hypothesis was tested with a variety of methods including a novel hepatocyte-lymphoma-cardiomyocyte cell co-culture system as an in vitro model for studying the biotransformation of CPA and therapeutic effects of CHOP as well as the off-target toxicity in healthy tissues in an environment that closely resembles the in vivo condition. Using this system we successfully demonstrated that activation of hCAR with small molecule activators can significantly increase the anticancer activity of the CHOP regimen in lymphoma cells. Additionally, activation of Nrf2 in cardiomyocytes in co-culture significantly reduced the doxorubicin-induced cardiotoxicity of CHOP. Utilizing a hCAR-transgenic mouse model, we were able to show in vivo that the combination of a selective hCAR activator alongside CHOP significantly increases the anticancer activity of CHOP in a lymphoma tumor xenograft study. Taken together, these results implicate hCAR and Nrf2 as drug targets for facilitating CHOP-based treatment of lymphomas. We were able to identify several compounds from the NIH Chemical Genomics Center Pharmaceutical Collection which activate both hCAR and Nrf2 and have provided preliminary evidence for their utility in CHOP-based lymphoma treatment.