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The physiological pH changes and peristalsis activities in gastrointestinal (GI) tract have big impact on the dissolution of oral drug products, when those oral drug products include APIs with pH-dependent solubility. It is well documented that predicting the bioperformance of those oral drug products can be challenging using compendial methods. To overcome this limitation, in vivo predictive dissolution apparatuses, such as the transfer model, have been developed to predict bioperformance of oral formulation candidates and drug products. In this manuscript we utilize a new transfer-model dissolution apparatus, the gastrointestinal simulator-α (GIS-α), to characterize its behavior in terms of transfer kinetics and pH, assess its reproducibility and adaptability to mimic different transfer conditions, as well as study dissolution of ketoconazole and dipyridamole as model BCS class IIb compounds. Availability of commercially available dissolution transfer systems with similar configuration to compendial dissolution apparatus, may be helpful to simplify and standardize in vivo predictive dissolution methodologies for BCS class IIb compounds in the future.

Exploring the intraluminal behavior of an oral drug product in the human gastrointestinal (GI) tract remains challenging. Many in vivo techniques are available to investigate the impact of GI physiology on oral drug behavior in fasting state conditions. However, little is known about the intraluminal behavior of a drug in postprandial conditions. In a previous report, we described the mean solution and total concentrations of ibuprofen after oral administration of an immediate-release (IR) tablet in fed state conditions. In parallel, blood samples were taken to assess systemic concentrations. The purpose of this work was to statistically evaluate the impact of GI physiology (e.g., pH, contractile events) within and between individuals (intra and intersubject variability) for a total of 17 healthy subjects. In addition, a pharmacokinetic (PK) analysis was performed by noncompartmental analysis, and PK parameters were correlated with underlying physiological factors (pH, time to phase III contractions postdose) and study parameters (e.g., ingested amount of calories, coadministered water). Moreover, individual plasma profiles were deconvoluted to assess the fraction absorbed as a function of time, demonstrating the link between intraluminal and systemic behavior of the drug. The results demonstrated that the in vivo dissolution of ibuprofen depends on the present gastric pH and motility events at the time of administration. Both intraluminal factors were responsible for explaining 63% of plasma Cmax variability among all individuals. For the first time, an in-depth analysis was performed on a large data set derived from an aspiration/motility study, quantifying the impact of physiology on systemic behavior of an orally administered drug product in fed state conditions. The data obtained from this study will help us to develop an in vitro biorelevant dissolution approach and optimize in silico tools in order to predict the in vivo performance of orally administered drug products, especially in fed state conditions. ispartof: MOLECULAR PHARMACEUTICS vol:15 issue:12 pages:5468-5478 ispartof: location:United States status: published

Summary of the In Vivo Predictive Dissolution (iPD) - Oral Drug Delivery (ODD) Conference 2018

The formulation developments and the in vivo assessment of Biopharmaceutical Classification System (BCS) class II drugs are challenging due to their low solubility and high permeability in the human gastrointestinal (GI) tract. Since the GI environment influences the drug dissolution of BCS class II drugs, the human GI characteristics should be incorporated into the in vitro dissolution system to predict bioperformance of BCS class II drugs. An absorptive compartment may be important in dissolution apparatus for BCS class II drugs, especially for bases (BCS IIb) because of high permeability, precipitation, and supersaturation. Thus, the in vitro dissolution system with an absorptive compartment may help predicting the in vivo phenomena of BCS class II drugs better than compendial dissolution apparatuses. In this study, an absorptive compartment (a biphasic device) was introduced to a gastrointestinal simulator. This addition was evaluated if this in vitro system could improve the prediction of in vivo dissolution for BCS class IIb drugs, ketoconazole and raloxifene, and subsequent absorption. The gastrointestinal simulator is a practical in vivo predictive tool and exhibited an improved in vivo prediction utilizing the biphasic format and thus a better tool for evaluating the bioperformance of BCS class IIb drugs than compendial apparatuses.

The patient receives a pharmaceutical product, not a drug. The pharmaceutical products are formulated with a drug, an active ingredient to produce the maximum therapeutic effect after oral absorption. Therefore, it is the product we must optimize for the patients. In order to assure the safety and efficacy of pharmaceutical products, we need an in vivo predictive tool for oral product performance in patients. Currently, we are a surprisingly long way from accomplishing that objective. If the 20th century was the ‘age of the drug’, i.e., the ‘magic bullet’, the 21st century must become the ‘age of the guided missile’, i.e., the delivery system, including the form of the active pharmaceutical ingredient (API) (‘drug’). The physical form of the drug and the delivery system must be optimized to maximize the therapeutic benefits of pharmaceutical products for humans. Oral immediate release (IR) dosage forms cannot be optimal for all drugs or likely even any drugs (APIs). Still, the formulation of pharmaceutical products has to be optimized for patients. But how do we optimize oral delivery of drugs? It is usually through ‘trial and error’, in humans! We need a better way to optimize the oral dosage forms. We have suggested to select different dissolution methodologies for this optimization based on BCS Subclasses. In this article, we present the predicted in vivo drug dissolution profile of ketoconazole as a model drug from our laboratory utilizing a gastrointestinal simulator (GIS), which is an adaptation of the ASD system. GIS consists of three chambers representing stomach, duodenum, and jejunum, to create the human gastrointestinal tract-like environment and enable the control the gastric emptying rate. This dissolution system allows the monitoring of the drug dissolution phenomena and the observation of the supersaturation and the precipitation of pharmaceutical products, which is useful information to predict in vivo dissolution of pharmaceutical products. This system can provide the actual input needed to accurately predict the input into the systemic circulation required by many of the absorption prediction packages available today.

The development of formulations and the assessment of oral drug absorption for Biopharmaceutical Classification System (BCS) class IIb drugs is often a difficult issue due to the potential for supersaturation and precipitation in the gastrointestinal (GI) tract. The physiological environment in the GI tract largely influences in vivo drug dissolution rates of those drugs. Thus, those physiological factors should be incorporated into the in vitro system to better assess in vivo performance of BCS class IIb drugs. In order to predict oral bioperformance, an in vitro dissolution system with multiple compartments incorporating physiologically relevant factors would be expected to more accurately predict in vivo phenomena than a one-compartment dissolution system like USP Apparatus 2 because, for example, the pH change occurring in the human GI tract can be better replicated in a multi-compartmental platform. The Gastrointestinal Simulator (GIS) consists of three compartments, the gastric, duodenal and jejunal chambers, and is a practical in vitro dissolution apparatus to predict in vivo dissolution for oral dosage forms. This system can demonstrate supersaturation and precipitation and, therefore, has the potential to predict in vivo bioperformance of oral dosage forms where this phenomenon may occur. In this report, in vitro studies were performed with dipyridamole and ketoconazole to evaluate the precipitation rates and the relationship between the supersaturation levels and oral absorption of BCS class II weak base drugs. To evaluate the impact of observed supersaturation levels on oral absorption, a study utilizing the GIS in combination with mouse intestinal infusion was conducted. Supersaturation levels observed in the GIS enhanced dipyridamole and ketoconazole absorption in mouse, and a good correlation between their supersaturation levels and their concentration in plasma was observed. The GIS, therefore, appears to represent in vivo dissolution phenomena and demonstrate supersaturation and precipitation of dipyridamole and ketoconazole. We therefore conclude that the GIS has been shown to be a good biopredictive tool to predict in vivo bioperformance of BCS class IIb drugs that can be used to optimize oral formulations.

Weakly basic drugs exhibit a pH-dependent dissolution profile in the gastrointestinal (GI) tract, which makes it difficult to predict their oral absorption profile. The aim of this study was to investigate the utility of the gastrointestinal simulator (GIS), a novel in vivo predictive dissolution (iPD) methodology, in predicting the in vivo behavior of the weakly basic drug dipyridamole when coupled with in silico analysis. The GIS is a multicompartmental dissolution apparatus, which represents physiological gastric emptying in the fasted state. Kinetic parameters for drug dissolution and precipitation were optimized by fitting a curve to the dissolved drug amount-time profiles in the United States Pharmacopeia apparatus II and GIS. Optimized parameters were incorporated into mathematical equations to describe the mass transport kinetics of dipyridamole in the GI tract. By using this in silico model, intraluminal drug concentration-time profile was simulated. The predicted profile of dipyridamole in the duodenal compartment adequately captured observed data. In addition, the plasma concentration-time profile was also predicted using pharmacokinetic parameters following intravenous administration. On the basis of the comparison with observed data, the in silico approach coupled with the GIS successfully predicted in vivo pharmacokinetic profiles. Although further investigations are still required to generalize, these results indicated that incorporating GIS data into mathematical equations improves the predictability of in vivo behavior of weakly basic drugs like dipyridamole.

The goal of this project was to explore and to statistically evaluate the responsible gastrointestinal (GI) factors that are significant factors in explaining the systemic exposure of ibuprofen, between and within human subjects. In a previous study, we determined the solution and total concentrations of ibuprofen as a function of time in aspirated GI fluids, after oral administration of an 800 mg IR tablet (reference standard) of ibuprofen to 20 healthy volunteers in fasted state conditions. In addition, we determined luminal pH and motility pressure recordings that were simultaneously monitored along the GI tract. Blood samples were taken to determine ibuprofen plasma levels. In this work, an in-depth statistical and pharmacokinetic analysis was performed to explain which underlying GI variables are determining the systemic concentrations of ibuprofen between (inter-) and within (intra-) subjects. In addition, the obtained plasma profiles were deconvoluted to link the fraction absorbed with the fraction dissolved. Multiple linear regressions were performed to explain and quantitatively express the impact of underlying GI physiology on systemic exposure of the drug (in terms of plasma Cmax/AUC and plasma Tmax). The exploratory analysis of the correlation between plasma Cmax/AUC and the time to the first phase III contractions postdose (TMMC-III) explains ∼40% of the variability in plasma Cmax for all fasted state subjects. We have experimentally shown that the in vivo intestinal dissolution of ibuprofen is dependent upon physiological variables like, in this case, pH and postdose phase III contractions. For the first time, this work presents a thorough statistical analysis explaining how the GI behavior of an ionized drug can explain the systemic exposure of the drug based on the individual profiles of participating subjects. This creates a scientifically based and rational framework that emphasizes the importance of including pH and motility in a predictive in vivo dissolution methodology to forecast the in vivo performance of a drug product. Moreover, as no extensive first-pass metabolism is considered for ibuprofen, this study demonstrates how intraluminal drug behavior is reflecting the systemic exposure of a drug. ispartof: MOLECULAR PHARMACEUTICS vol:15 issue:12 pages:5454-5467 ispartof: location:United States status: published

The purpose of this study was to assess the feasibility of a multicompartmental in vitro dissolution apparatus, gastrointestinal simulator (GIS), in assessing the drug dissolution of 2 commercially available oral dosage forms for itraconazole (ICZ). The GIS consists of 3 chambers, mimicking the upper gastrointestinal tract. In vitro dissolution of ICZ capsule or oral solution was evaluated in United States Pharmacopeia apparatus II and GIS. To investigate the suitability of fasted state simulated intestinal fluid (FaSSIF) to predict better in vivo, FaSSIF as well as phosphate buffer were used as dissolution media. Area under the dissolved drug amount-time curve (AUDC) was calculated for each dosage form in each apparatus, and the ratios of AUDCoral solution to AUDCcapsule were compared with human pharmacokinetic data. Based on this comparison, GIS with FaSSIF can adequately distinguish the pharmacokinetic profiles of 2 oral dosage forms for ICZ. Additionally, Caco-2 cell transepithelial transport study in combination with GIS revealed that improved drug dissolution by formulations resulted in enhanced permeation of ICZ through cell monolayer, suggesting the observed ICZ concentration in the GIS will directly reflect systemic exposure. These results indicate GIS would be a powerful tool to assess the formulations of ICZ as well as other Biopharmaceutics Classification System class II drug formulations.

The formulation developments and the in vivo assessment of Biopharmaceutical Classification System (BCS) class II drugs are challenging due to their low solubility and high permeability in the human gastrointestinal (GI) tract. Since the pH change in the GI environment influences the drug dissolution of BCS class IIa and IIb drugs but not for IIc drugs, the incorporation of the human GI characteristics into the in vitro dissolution system may not be crucial for BCS class IIc drugs to predict their bioperformance. An absorptive compartment in dissolution apparatus would be important to predict bioperformance of BCS class IIc drugs due to their physicochemical characteristics, low solubility and high permeability. In this study, an absorptive compartment (biphasic system) was introduced to a gastrointestinal simulator (GIS) and USP dissolution apparatus II for the evaluation of in vivo prediction for BCS class IIc drugs. Those in vitro systems with the biphasic phase were evaluated for the prediction of in vivo dissolution of BCS class IIc drugs, donepezil and danazol, and subsequent absorption. For the evaluation of in vivo dissolution of BCS class IIc drugs, simpler dissolution apparatus, USP II, with the absorptive phase might be adequate a dissolution method to predict in vivo.

The “3rd International Symposium on BA/BE of Oral Drug Products – Biopharmaceutics meets Galenics (BA/BE 2018)” was held during 11–12th of October 2018 at INFARMED, the Portuguese regulatory medicines agency, in Lisbon, Portugal. Summary of this meeting and perspective/directions of biopharmaceutical researches are discussed in this opinion paper.

The discovery pipelines in pharmaceutical companies are highly populated with poorly soluble compounds, demanding for an appropriate solution to overcome this problem. Generating supersaturated concentrations for these drugs in the gastrointestinal (GI) tract has been put forward as a promising strategy to assure sufficient systemic exposure after oral intake of the drug. Numerous publications have shown to which extent supersaturating dosage forms can create supersaturated concentrations by testing these formulations in in vitro dissolution models. The obtained dissolution profiles can serve as a valuable input for computational software programs to simulate the systemic outcome of the drug. To improve our fundamental knowledge and understanding of the impact of supersaturation/precipitation on oral absorption, attempts have been made to develop biopredictive in vitro dissolution systems such as the Gastrointestinal Simulator (GIS), the dissolution/permeation system, and the biphasic test system. Moreover, aspiration studies in humans have been performed throughout the years to measure simultaneously GI and systemic concentrations of the drug in order to directly envisage the impact of supersaturation on systemic exposure. This manuscript aims to provide a comprehensive review with respect to the predictive power of in vitro tools towards the in vivo performance of supersaturating dosage forms. Different case studies will be thoroughly presented where a plethora of supersaturating formulations was tested applying a biopredictive setting. Recent investigations on liquid-liquid phase separation of supersaturated solutions, which makes comprehension of the dissolution process of supersaturating dosage forms extremely challenging, are also discussed.

Over the past decade, formulation predictive dissolution (fPD) testing has gained increasing attention. Another mindset is pushed forward where scientists in our field are more confident to explore the in vivo behavior of an oral drug product by performing predictive in vitro dissolution studies. Similarly, there is an increasing interest in the application of modern computational fluid dynamics (CFD) frameworks and high-performance computing platforms to study the local processes underlying absorption within the gastrointestinal (GI) tract. In that way, CFD and computing platforms both can inform future PBPK-based in silico frameworks and determine the GI-motility-driven hydrodynamic impacts that should be incorporated into in vitro dissolution methods for in vivo relevance. Current compendial dissolution methods are not always reliable to predict the in vivo behavior, especially not for biopharmaceutics classification system (BCS) class 2/4 compounds suffering from a low aqueous solubility. Developing a predictive dissolution test will be more reliable, cost-effective and less time-consuming as long as the predictive power of the test is sufficiently strong. There is a need to develop a biorelevant, predictive dissolution method that can be applied by pharmaceutical drug companies to facilitate marketing access for generic and novel drug products. In 2014, Prof. Gordon L. Amidon and his team initiated a far-ranging research program designed to integrate (1) in vivo studies in humans in order to further improve the understanding of the intraluminal processing of oral dosage forms and dissolved drug along the gastrointestinal (GI) tract, (2) advancement of in vitro methodologies that incorporates higher levels of in vivo relevance and (3) computational experiments to study the local processes underlying dissolution, transport and absorption within the intestines performed with a new unique CFD based framework. Of particular importance is revealing the physiological variables determining the variability in in vivo dissolution and GI absorption from person to person in order to address (potential) in vivo BE failures. This paper provides an introduction to this multidisciplinary project, informs the reader about current achievements and outlines future directions. ispartof: INTERNATIONAL JOURNAL OF PHARMACEUTICS vol:548 issue:1 pages:120-127 ispartof: location:Netherlands status: published

Biphenyl hydrolase-like protein (BPHL) is a novel human serine hydrolase that was originally cloned from a breast carcinoma cDNA library and shown to convert valacyclovir to acyclovir and valganciclovir to ganciclovir. However, the exclusivity of this process has not been determined and, indeed, it is possible that a number of esterases/proteases may mediate the hydrolysis of valacyclovir and similar prodrugs. The objectives of the present study were to evaluate the in situ intestinal permeability and stability of valacyclovir in wildtype (WT) and Bphl knockout (KO) mice, as well as the in vivo oral absorption and intravenous disposition of valacyclovir and acyclovir in the two mouse genotypes. We found that Bphl knockout mice had no obvious phenotype and that Bphl ablation did not alter the jejunal permeability of valacyclovir during in situ perfusions (i.e., 0.54 × 10−4 in WT vs. 0.53 × 10−4 cm/s in KO). Whereas no meaningful changes occurred between genotypes in the gene expression of proton-coupled oligopeptide transporters (i.e., PepT1, PepT2, PhT1, PhT2), enzymatic upregulation of Cyp3a11, Cyp3a16, Abhd14a and Abhd14b was observed in some tissues of Bphl knockout mice. Most importantly, we found that valacyclovir was rapidly and efficiently hydrolyzed to acyclovir in the absence of BPHL, and that hydrolysis was more extensive after the oral vs. intravenous route of administration (for both genotypes). Taken as a whole, BPHL is not obligatory for the conversion of valacyclovir to acyclovir either presystemically or systemically.

The goal of this study was to create a mass transport model (MTM) model for gastric emptying and upper gastrointestinal (GI) appearance that can capture the in vivo concentration-time profiles of the nonabsorbable drug phenol red in solution in the stomach and upper small intestine by direct luminal measurement while simultaneously recording the contractile activity (motility) via manometry. We advanced from a one-compartmental design of the stomach to a much more appropriate, multi-compartmental 'mixing tank' gastric model that reflects drug distribution along the different regions of the stomach as a consequence of randomly dosing relative to the different contractile phases of the migrating motor complex (MMC). To capture the intraluminal phenol red concentrations in the different segments of the GI tract both in fasted and fed state conditions, it was essential to include a bypass flow compartment ('magenstrasse') to facilitate the transport of the phenol red solution directly to the duodenum (fasted state) or antrum (fed state). The fasted and fed state models were validated with external reference data from an independent aspiration study using another nonabsorbable marker (paromomycin). These results will be essential for the development and optimization of computational programs for GI simulation and absorption prediction, providing a realistic gastric physiologically-based pharmacokinetic (PBPK) model based on direct measurement of gastric concentrations of the drug in the stomach. ispartof: EUROPEAN JOURNAL OF PHARMACEUTICS AND BIOPHARMACEUTICS vol:129 pages:162-174 ispartof: location:Netherlands status: published

For the last two decades, the application of physiologically based pharmacokinetic (PBPK) models has grown exponentially in the field of oral absorption and in a regulatory context. Although these models are widely used, their predictive power should be validated and optimized in order to rely on these models and to know exactly what is going on "under the hood". In this study, an automated sensitivity analysis (ASA) was performed for 11 gastrointestinal (GI) variables that are integrated into the PBPK software program Simcyp®. The model of interest was a previously validated workspace that was able to predict the intraluminal and systemic behavior of two different suspensions of posaconazole in the Simcyp® Simulator. The sensitivity of the following GI parameters was evaluated in this model: gastric and duodenal pH, gastric and duodenal bicarbonate concentrations (reflecting buffer capacity), duodenal bile salts concentration, gastric emptying, the interdigestive migrating motor complex (IMMC), small intestinal transit time (SITT), gastric and jejunal volumes, and permeability. The most sensitive parameters were gastric/duodenal pH and gastric emptying, for both suspensions. The outcome of the sensitivity analyses highlights the important GI variables that must be integrated into an in vivo predictive dissolution test to help and create a rational and scientific framework/design for product development of novel and generic drug products. ispartof: The AAPS Journal vol:20 issue:3 pages:57-57 ispartof: location:United States status: published

Supersaturating drug delivery systems (SDDS) have been put forward in the recent decades in order to circumvent the issue of low aqueous solubility. Prior to the start of clinical trials, these enabling formulations should be adequately explored in in vitro/in silico studies in order to understand their in vivo performance and to select the most appropriate and effective formulation in terms of oral bioavailability and therapeutic outcome. The purpose of this work was to evaluate the in vivo performance of four different oral formulations of posaconazole (categorized as a biopharmaceutics classification system (BCS) class 2b compound) based on the in vitro concentrations in the gastrointestinal simulator (GIS), coupled with an in silico pharmacokinetic model to predict their systemic profiles. Recently published intraluminal and systemic concentrations of posaconazole for these formulations served as a reference to validate the in vitro and in silico results. Additionally, the morphology of the formed precipitate of posaconazole was visualized and characterized by optical microscopy studies and thermal analysis. This multidisciplinary work demonstrates an in vitro-in silico-in vivo approach that provides a scientific basis for screening SDDS by a user-friendly formulation predictive dissolution (fPD) device in order to rank these formulations towards their in vivo performance. ispartof: European Journal of Pharmaceutical Sciences vol:115 pages:258-269 ispartof: location:Netherlands status: published

In vivo drug dissolution in the gastrointestinal (GI) tract is largely unmeasured. The purpose of this clinical study was to evaluate the in vivo drug dissolution and systemic absorption of the BCS class IIa drug ibuprofen under fed and fasted conditions by direct sampling of stomach and small intestinal luminal content. Expanding current knowledge of drug dissolution in vivo will help to establish physiologically relevant in vitro models predictive of drug dissolution. A multilumen GI catheter was orally inserted into the GI tract of healthy human subjects. Subjects received a single oral dose of ibuprofen (800 mg tablet) with 250 mL of water under fasting and fed conditions. The GI catheter facilitated collection of GI fluid from the stomach, duodenum, and jejunum. Ibuprofen concentration in GI fluid supernatant and plasma was determined by LC–MS/MS. A total of 23 subjects completed the study, with 11 subjects returning for an additional study visit (a total of 34 completed study visits). The subjects were primarily white (61%) and male (65%) with an average age of 30 years. The subjects had a median [min, max] weight of 79 [52, 123] kg and body mass index of 25.7 [19.4, 37.7] kg/m(2). Ibuprofen plasma levels were higher under fasted conditions and remained detectable for 28 h under both conditions. The AUC(0–24) and C(max) were lower in fed subjects vs fasted subjects, and T(max) was delayed in fed subjects vs fasted subjects. Ibuprofen was detected immediately after ingestion in the stomach under fasting and fed conditions until 7 h after dosing. Higher levels of ibuprofen were detected in the small intestine soon after dosing in fasted subjects compared to fed. In contrast to plasma drug concentration, overall gastric concentrations remained higher under fed conditions due to increased gastric pH vs fasting condition. The gastric pH increased to near neutrality after feeding before decreasing to acidic levels after 7 h. Induction of the fed state reduced systemic levels but increased gastric levels of ibuprofen, which suggest that slow gastric emptying and transit dominate the effect for plasma drug concentration. The finding of high levels of ibuprofen in stomach and small intestine 7 h post dosing was unexpected. Future work is needed to better understand the role of various GI parameters, such as motility and gastric emptying, on systemic ibuprofen levels in order to improve in vitro predictive models.

One of the main obstacles for cancer therapies is to deliver medicines effectively to target sites. Since stroma cells are developed around tumors, chemotherapeutic agents have to go through stroma cells in order to reach tumors. As a method to improve drug delivery to the tumor site, a prodrug approach for gemcitabine was adopted. Amino acid and dipeptide monoester prodrugs of gemcitabine were synthesized and their chemical stability in buffers, resistance to thymidine phosphorylase and cytidine deaminase, antiproliferative activity, and uptake/permeability in HFF cells as a surrogate to stroma cells were determined and compared to their parent drug, gemcitabine. The activation of all gemcitabine prodrugs was faster in pancreatic cell homogenates than their hydrolysis in buffer, suggesting enzymatic action. All prodrugs exhibited great stability in HFF cell homogenate, enhanced resistance to glycosidic bond metabolism by thymidine phosphorylase, and deamination by cytidine deaminase compared to their parent drug. All gemcitabine prodrugs exhibited higher uptake in HFF cells and better permeability across HFF monolayers than gemcitabine, suggesting a better delivery to tumor sites. Cell antiproliferative assays in Panc-1 and Capan-2 pancreatic ductal cell lines indicated that the gemcitabine prodrugs were more potent than their parent drug gemcitabine. The transport and enzymatic profiles of gemcitabine prodrugs suggest their potential for delayed enzymatic bioconversion and enhanced resistance to metabolic enzymes, as well as for enhanced drug delivery to tumor sites, and cytotoxic activity in cancer cells. These attributes would facilitate the prolonged systemic circulation and improved therapeutic efficacy of gemcitabine prodrugs.

In this study, we determined the pH and buffer capacity of human gastrointestinal (GI) fluids (aspirated from the stomach, duodenum, proximal jejunum, and mid/distal jejunum) as a function of time, from 37 healthy subjects after oral administration of an 800 mg immediate-release tablet of ibuprofen (reference listed drug; RLD) under typical prescribed bioequivalence (BE) study protocol conditions in both fasted and fed states (simulated by ingestion of a liquid meal). Simultaneously, motility was continuously monitored using water-perfused manometry. The time to appearance of phase III contractions (i.e., housekeeper wave) was monitored following administration of the ibuprofen tablet. Our results clearly demonstrated the dynamic change in pH as a function of time and, most significantly, the extremely low buffer capacity along the GI tract. The buffer capacity on average was 2.26 μmol/mL/ΔpH in fasted state (range: 0.26 and 6.32 μmol/mL/ΔpH) and 2.66 μmol/mL/ΔpH in fed state (range: 0.78 and 5.98 μmol/mL/ΔpH) throughout the entire upper GI tract (stomach, duodenum, and proximal and mid/distal jejunum). The implication of this very low buffer capacity of the human GI tract is profound for the oral delivery of both acidic and basic active pharmaceutical ingredients (APIs). An in vivo predictive dissolution method would require not only a bicarbonate buffer but also, more significantly, a low buffer capacity of dissolution media to reflect in vivo dissolution conditions.

Gastrointestinal (GI) fluid volume and its dynamic change are integral to study drug disintegration, dissolution, transit, and absorption. However, key questions regarding the local volume and its absorption, secretion, and transit remain unanswered. The dynamic fluid compartment absorption and transit (DFCAT) model is proposed to estimate in vivo GI volume and GI fluid transport based on magnetic resonance imaging (MRI) quantified fluid volume. The model was validated using GI local concentration of phenol red in human GI tract, which was directly measured by human GI intubation study after oral dosing of non-absorbable phenol red. The measured local GI concentration of phenol red ranged from 0.05 to 168 μg/mL (stomach), to 563 μg/mL (duodenum), to 202 μg/mL (proximal jejunum), and to 478 μg/mL (distal jejunum). The DFCAT model characterized observed MRI fluid volume and its dynamic changes from 275 to 46.5 mL in stomach (from 0 to 30 min) with mucus layer volume of 40 mL. The volumes of the 30 small intestine compartments were characterized by a max of 14.98 mL to a min of 0.26 mL (0–120 min) and a mucus layer volume of 5 mL per compartment. Regional fluid volumes over 0 to 120 min ranged from 5.6 to 20.38 mL in the proximal small intestine, 36.4 to 44.08 mL in distal small intestine, and from 42 to 64.46 mL in total small intestine. The DFCAT model can be applied to predict drug dissolution and absorption in the human GI tract with future improvements.

First introduced in the second half of the 19th century, enteric coatings are commonly used to protect acid-labile drugs, reduce the risk of gastric side effects due to irritating drugs, or for local drug delivery to the lower gastrointestinal (GI) tract. The currently available enteric-coatings are based on pH-sensitive weakly acidic polymers. Despite the long history of their use, the causes behind their performance often being unpredictable have not been properly investigated with most of the attention being focused only on the gastric emptying. However, little attention has been given to the postgastric emptying disintegration and dissolution of these dosage forms. This lack of attention has contributed to the difficulty in predicting the in vivo behavior of these dosage forms and to cases of bioavailability problems with some enteric-coated products. Therefore, increased attention needs to be given to this issue.

As an orally administered, locally acting gastrointestinal drug, mesalamine products are designed to achieve high local drug concentration in the gastrointestinal (GI) tract for the treatment of ulcerative colitis. The aim of this study was to directly measure and compare drug dissolution of three mesalamine formulations in human GI tract and to correlate their GI concentration with drug concentration in plasma. Healthy human subjects were orally administered Pentasa, Apriso, or Lialda. GI fluids were aspirated from stomach, duodenum, proximal jejunum, mid jejunum, and distal jejunum regions. Mesalamine (5-ASA) and its primary metabolite acetyl-5-mesalamine (Ac-5-ASA) were measured using LC-MS/MS. GI tract pH was measured from each GI fluid sample, which averaged 1.82, 4.97, 5.67, 6.17, and 6.62 in the stomach, duodenum, proximal jejunum, middle jejunum, and distal jejunum, respectively. For Pentasa, high levels of 5-ASA in solution were observed in the stomach, duodenum, proximal jejunum, mid jejunum, and distal jejunum from 1 to 7 h. Apriso had minimal 5-ASA levels in stomach, low to medium levels of 5-ASA in duodenum and proximal jejunum from 4 to 7 h, and high levels of 5-ASA in distal jejunum from 3 to 7 h. In contrast, Lialda had minimal 5-ASA levels from stomach and early small intestine. A composite appearance rate (CAR) was calculated from the deconvolution of individual plasma concentration to reflect drug release, dissolution, transit, and absorption in the GI tract. Individuals dosed with Pentasa had high levels of CAR from 1 to 10 h; individuals dosed with Apriso had low levels of CAR from 1 to 4 h and high levels of CAR from 5 to 10 h; Lialda showed minimal levels of CAR from 0 to 5 h, then increased to medium levels from 5 to 12 h, and then decreased to further lower levels after 12 h. In the colon region, Pentasa and Apriso showed similar levels of accumulated 5-ASA excreted in the feces, while Lialda showed slightly higher 5-ASA accumulation in feces. However, all three formulations showed similar levels of metabolite Ac-5-ASA in the feces. These results provide direct measurement of drug dissolution in the GI tract, which can serve as a basis for investigation of bioequivalence for locally acting drug products.

Triple negative breast cancer (TNBC) typically exhibits rapid progression, high mortality and faster relapse rates relative to other breast cancer subtypes. In this report we examine the combination of taxanes (paclitaxel or docetaxel) with a breast cancer stem cell (CSC)-targeting agent sulforaphane for use against TNBC. We demonstrate that paclitaxel or docetaxel treatment induces IL-6 secretion and results in expansion of CSCs in TNBC cell lines. Conversely, sulforaphane is capable of preferentially eliminating CSCs, by inhibiting NF-κB p65 subunit translocation, downregulating p52 and consequent downstream transcriptional activity. Sulforaphane also reverses taxane-induced aldehyde dehydrogenase-positive (ALDH+) cell enrichment, and dramatically reduces the size and number of primary and secondary mammospheres formed. In vivo in an advanced treatment orthotopic mouse xenograft model together with extreme limiting dilution analysis (ELDA), the combination of docetaxel and sulforaphane exhibits a greater reduction in primary tumor volume and significantly reduces secondary tumor formation relative to either treatment alone. These results suggest that treatment of TNBCs with cytotoxic chemotherapy would be greatly benefited by the addition of sulforaphane to prevent expansion of and eliminate breast CSCs.

The goal of this study was to improve the intestinal mucosal cell membrane permeability of the poorly absorbed guanidino analogue of a neuraminidase inhibitor, oseltamivir carboxylate (GOC) using a carrier-mediated strategy. Valyl amino acid prodrug of GOC with isopropyl-methylene-dioxy linker (GOC-ISP-Val) was evaluated as the potential substrate for intestinal oligopeptide transporter, hPEPT1 in Xenopus laevis oocytes heterologously expressing hPEPT1, and an intestinal mouse perfusion system. The diastereomers of GOC-ISP-Val were assessed for chemical and metabolic stability. Permeability of GOC-ISP-Val was determined in Caco-2 cells and mice. Diastereomer 2 was about 2 times more stable than diastereomer 1 in simulated intestinal fluid and rapidly hydrolyzed to the parent drug in cell homogenates. The prodrug had a 9 times-enhanced apparent permeability (P-app) in Caco-2 cells compared with the parent drug. Both diastereomer exhibited high effective permeability (P-eff) in mice, 6.32 +/- 3.12 and 5.20 +/- 2.81 x 10(-5) cm/s for diastereomer 1 and 2, respectively. GOC-ISP-Val was found to be a substrate of hPEPT1. Overall, this study indicates that the prodrug, GOC-ISP-Val, seems to be a promising oral anti-influenza agent that has sufficient stability at physiologically relevant pHs before absorption, significantly improved permeability via hPEPT1 and potentially rapid activation in the intestinal cells. (C) 2016 American Pharmacists Association (R). Published by Elsevier Inc. All rights reserved.

Dipeptide monoester prodrugs of floxuridine and gemcitabine were synthesized. Their chemical stability in buffers, enzymatic stability in cell homogenates, permeability in mouse intestinal membrane along with drug concentration in mouse plasma, and anti-proliferative activity in cancer cells were determined and compared to their parent drugs. Floxuridine prodrug was more enzymatically stable than floxuridine and the degradation from prodrug to parent drug works as the rate-limiting step. On the other hand, gemcitabine prodrug was less enzymatically stable than gemcitabine. Those dipeptide monoester prodrugs exhibited 2.4- to 48.7-fold higher uptake than their parent drugs in Caco-2, Panc-1, and AsPC-1 cells. Floxuridine and gemcitabine prodrugs showed superior permeability in mouse jejunum to their parent drugs and exhibited the higher drug concentration in plasma after in situ mouse perfusion. Cell proliferation assays in ductal pancreatic cancer cells, AsPC-1 and Panc-1, indicated that dipeptide prodrugs of floxuridine and gemcitabine were more potent than their parent drugs. The enhanced potency of nucleoside analogs was attributed to their improved membrane permeability. The prodrug forms of 5¢-L-phenylalanyl-l-tyrosyl-floxuridine and 5¢-L-phenylalanyl-L-tyrosyl-gemcitabine appeared in mouse plasma after the permeation of intestinal membrane and the first-pass effect, suggesting their potential for the development of oral dosage form for anti-cancer agents. This work was supported by grants NIGMD-2R01GM037188.

The improvement of therapeutic efficacy for cancer agents has been a big challenge which includes the increase of tumor selectivity and the reduction of adverse effects at non-tumor sites. In order to achieve those goals, prodrug approaches have been extensively investigated. In this report, the potential activation enzymes for 5¢-amino acid/dipeptide monoester floxuridine prodrugs in pancreatic cancer cells were selected and the feasibility of enzyme specific activation of prodrugs was evaluated. All prodrugs exhibited the range of 3.0–105.7 min of half life in Capan-2 cell homogenate with the presence and the absence of selective enzyme inhibitors. 5¢-O-L-Phenylalanyl-L-tyrosyl-floxuridine exhibited longer half life only with the presence of pepstatin A. Human cathepsin B and D selectively hydrolized 5¢-O-L-phenylalanyl-L-tyrosylfloxuridine and 5¢-O-L-phenylalanyl-L-glycylfloxuridine compared to the other tested prodrugs. The wide range of growth inhibitory effect by floxuridine prodrugs in Capan-2 cells was observed due to the different affinities of prodrug promoieties to enyzmes. In conclusion, it is feasible to design prodrugs which are activated by specific enzymes. Cathepsin D might be a good candidate as a target enzyme for prodrug activation and 5¢-O-L-phenylalanyl-L-tyrosylfloxuridine may be the best candidate among the tested floxuridine prodrugs.

Dipeptide monoester prodrugs of floxuridine were synthesized, and their chemical stability in buffers, resistance to glycosidic bond metabolism, affinity for PEPT1, enzymatic activation and permeability in cancer cells were determined and compared to those of mono amino acid monoester floxuridine prodrugs. Prodrugs containing glycyl moieties were the least stable in pH 7.4 buffer ( t 1/2 < 100 min). The activation of all floxuridine prodrugs was 2- to 30-fold faster in cell homogenates than their hydrolysis in buffer, suggesting enzymatic action. The enzymatic activation of dipeptide monoester prodrugs containing aromatic promoieties in cell homogenates was 5- to 20-fold slower than that of other dipeptide and most mono amino acid monoester prodrugs ( t 1/2 approximately 40 to 100 min). All prodrugs exhibited enhanced resistance to glycosidic bond metabolism by thymidine phosphorylase compared to parent floxuridine. In general, the 5'-O-dipeptide monoester floxuridine prodrugs exhibited higher affinity for PEPT1 than the corresponding 5'-O-mono amino acid ester prodrugs. The permeability of dipeptide monoester prodrugs across Caco-2 and Capan-2 monolayers was 2- to 4-fold higher than the corresponding mono amino acid ester prodrug. Cell proliferation assays in AsPC-1 and Capan-2 pancreatic ductal cell lines indicated that the dipeptide monoester prodrugs were equally as potent as mono amino acid prodrugs. The transport and enzymatic profiles of 5'- l-phenylalanyl- l-tyrosyl-floxuridine, 5'- l-phenylalanyl- l-glycyl-floxuridine, and 5'- l-isoleucyl- l-glycyl-floxuridine suggest their potential for increased oral uptake, delayed enzymatic bioconversion and enhanced resistance to metabolism to 5-fluorouracil, as well as enhanced uptake and cytotoxic activity in cancer cells, attributes that would facilitate prolonged systemic circulation for enhanced therapeutic action.

A series of amino acid monoester prodrugs of floxuridine was synthesized and evaluated for the improvement of oral bioavailability and the feasibility of target drug delivery via oligopeptide transporters. All floxuridine 5'-amino acid monoester prodrugs exhibited PEPT1 affinity, with inhibition coefficients of Gly-Sar uptake (IC50) ranging from 0.7 - 2.3 mM in Caco-2 and 2.0 - 4.8 mM in AsPC-1 cells, while that of floxuridine was 7.3 mM and 6.3 mM, respectively. Caco-2 membrane permeabilities of floxuridine prodrugs (1.01 - 5.31 x 10(-6 )cm/sec) and floxuridine (0.48 x 10(-6 )cm/sec) were much higher than that of 5-FU (0.038 x 10(-6) cm/sec). MDCK cells stably transfected with the human oligopeptide transporter PEPT1 (MDCK/hPEPT1) exhibited enhanced cell growth inhibition in the presence of the prodrugs. This prodrug strategy offers great potential, not only for increased drug absorption but also for improved tumor selectivity and drug efficacy.