Postnatal Hepatobiliary and Gastrointestinal Systems Development and Impact on ADME

Abstract

Toxicity can result from variable target organ sensitivity and exposure based on postnatal development. Changes in the gastrointestinal tract (GIT) in neonates are driven by initial enteral feedings. These are important for nutrient uptake as well as drug disposition and include motility, expansion of enzyme and transporter function, permeability, intestinal microbiome, and species-specific maturation. Some aspects of GIT function do not mature until driven by increased dietary complexity. As with the GIT, postnatal hepatic maturation in the rat includes a variety of anatomic and functional changes that include refinements in the activities or expression of drug transporters and drug-metabolizing enzymes. These changes may impact rodent pharmacokinetics, nonclinical toxicity profiles, and estimation of safe pediatric doses. Pilot or dose range finding studies can help characterize and mitigate toxicity related to drug disposition, especially in juvenile rodents. Interpretation of developmental toxicity requires knowledge of developing systems in humans and nonclinical models.

The following is a brief synopsis of lectures presented by Drs April Kluever and Armando R. Irizarry Rovira during the STP 40th Annual Symposium session, “ADME and Clinical Pathology Considerations in Juvenile Animals.” Dr Kluever provided an overview of “lessons learned” from the medical and nutritional literature related to gastrointestinal tract (GIT) maturation in humans and across various nonclinical species. Much of the relevant content was derived from a recent Health and Environmental Sciences Institute (HESI) working group manuscript and references therein. ¹ Dr Kluever’s discussion was followed by Dr Irizarry providing a practical overview of the developing hepatobiliary system in the rat, with emphasis on relevance to the disposition of small molecule pharmaceuticals. Both lectures provided a review of considerations relevant to the practicing toxicologic pathologist and identified data gaps of potential future relevance. Highlights are summarized from each lecture below.

The Developing Gastrointestinal System Responds Dynamically to Feeding

In neonatal humans, including both term and premature infants, initiation of enteral feeds influences motor function, epithelial differentiation, growth, decreased permeability of the intestinal epithelium, and changes in nutrient receptors and enzymes. ^1,2 In contrast, nonnutritive feedings such as water, or total parenteral nutrition, can delay these maturation processes. ^1,2 Developmental changes in the GIT during the newborn period are important for nutrient uptake as well as the disposition of orally administered medications. Importantly, some aspects of gastrointestinal function do not mature until driven by increased dietary complexity and nutritional demands (eg, weaning). ^1,3,4 The expression and activity of most digestive enzymes are driven by the nutrient composition of enteral feeds and differ across species. ^1,3,4 For example, enzymatic digestion is minimal in the rat prior to weaning, and humans have a greater capacity to digest carbohydrates at birth as compared to rats. ^1,3,4 Gastrointestinal (GI) motility, including gastric emptying and intestinal transit, is also influenced by nutritive feeds in neonates, as is gastric pH. ¹ Although there are a number of publications that emphasize a relatively high gastric pH in the postnatal period, this probably reflects the transient elevation of gastric pH associated with the postprandial state. Although the gastric pH at birth is neutral, like amniotic fluid, it drops quickly to 2.0 and achieves a typical fasting pH of 2 to 3.5 by 1 week of age in term or preterm human neonates. ¹

Neonatal Gut Permeability Is Transient

In most species, there is a brief perinatal period of increased intestinal absorption of intact macromolecules, which allows for direct uptake of immunoglobulins (Igs), growth factors, and hormones as well as nutrients from the milk. ¹ There may be a transient period of enhanced update of macromolecular xenobiotics as well. Active transport of Igs occurs via the neonatal fragment crystallizable receptor, while this receptor persists through adulthood in humans, the GI uptake of Igs is limited and species specific. ⁵ For example, Igs are absorbed intact for weeks in rats and rabbits, but Ig absorption declines over hours to days in most other species such as the guinea pig, hamster, dog, and pig. ^1,3,5 The decline of macromolecular absorption has been insufficiently studied in humans, but the current consensus is that the direct absorption of macromolecules in terms of neonates is more similar to the piglet than to the rat, and that absorption of specific macromolecules such as Igs is greater in preterm infants than term infants. ^1,3,5

The Gut Is Not Sterile in Neonates

It is now generally accepted that initial colonization of the GIT occurs in utero. This is, however, greatly expanded postnatally, including vertical transmission during the birth process. In humans, the diversity of neonatal GIT flora shows some dependence on the mode of delivery (vaginal vs C-section), but overall there is a clear correlation of the flora to the mother’s gut, vaginal, and aureolar microbiota. ^1,6 Neonates tend to have more aerobes in their gut, which is linked to higher levels of dissolved O₂ in the neonatal gut. In addition, gut flora patterns differ for preterm versus term neonates, and for formula versus breastfed neonates. ^1,6 Ultimately, there is additional co-maturation of commensal gut flora with the developing intestinal immune system. ^1,6

Data Gaps Remain

The postnatal structural and functional development of the GIT across species has been reviewed, with extensive references therein. ^1,3,4,7 In this area, nutrition science has established a better characterization of receptors and transporters than pharmaceutical sciences. ^1,4 There remains a lack of consistent data across species for pharmaceutical-relevant intestinal transporter and enzyme ontogeny; study methodologies vary in the assessment of transcript expression, protein levels, or target activity. ¹ Overall, data for the rat are the most readily available, although it is not a good model for the neonatal human GIT. ³ It is clear that the neonatal rat absorbs more nutrients and macromolecules, even in the absence of active transport and/or enzymatic digestion, than other nonrodent species. ^{1,3 –5} There is also extensive data for the pig, which represents a better physiological model for gut maturation than the rat but is not a standard species for regulatory projects. ^1,3 Ultimately, there is uncertainty with any model, which can be contextualized based on existing knowledge to improve interpretation, such as continued refinement of pharmaco- or toxicokinetic models for juvenile animals and humans.

Dr Kluever’s talk was followed by a lecture by Dr Armando R. Irizarry Rovira, providing a detailed look at the postnatal development of the hepatobiliary system in the rat, with particular emphasis on relevance to the disposition of small molecule pharmaceuticals. It is well established that children are not simply small adults with regard to drug disposition, although use in pediatrics is primarily guided by data from adults. When juvenile animal studies (JAS) are warranted, the rat has historically been the most commonly used species, and therefore, postnatal development of the rat is important to understand. ⁸

Hepatic Effects on Drug Disposition

With regard to drug disposition, the liver is the most important site of metabolism and xenobiotic transformation. ^{7,9 –11} Due to the dual vascular supply, xenobiotics can be delivered to the liver directly from the intestine via the portal vein, or via the hepatic artery following parenteral administration; within the liver, there is then diffusion or transporter-mediated uptake. ^{9 –11} Hepatic biotransformation occurs when enzymes mediate chemical reactions to facilitate excretion. Traditionally, these are divided into phase 1 reactions mediated by enzymes such as cytochrome P450 enzymes, and phase 2 conjugation reactions. Larger xenobiotics and conjugates are excreted in bile, or efflux transporters excrete metabolic products into the blood. In both instances, enterohepatic recirculation can also contribute. ^{9 –11}

The Rodent Liver Matures Postnatally

As has been extensively reviewed, the rat hepatobiliary system undergoes substantial postnatal maturation with regard to both structure and function. ^8,9,12 For example, the maturation of the ductal plates to bile ducts occurs over the first 4 weeks postnatal in the rat. ^7,12 During that same time frame, there is decreased hepatic extramedullary hematopoiesis, maturation of hepatic cords, and a decrease in the number of hepatocytes per gram of liver tissue as hepatocytes and lobules grow in size. ^7,12,13 The vascular system matures in parallel, including fenestrations and formation of portal triads, and there are concomitant increases in extracellular matrix and serum albumin. Finally, there is a complex system of interrelationships that drive the maturation of hepatic biotransformation components. ^9,10

Data Gaps Remain in the Knowledge Base Captured in the Literature

Similar to the GIT, data on enzyme and transporter maturation reflect different experimental approaches. Some investigators focus on enzyme activity, others on messenger RNA expression or protein levels in tissue, and there are also sex, species, and strain differences. In a recent review, Van Groen et al detail the existing published information and have identified different general patterns of enzyme and transporter maturation: (1) some increase over time to adult levels, (2) some begin high and decrease over time to adult levels, (3) some have complex or cyclical changes, and (4) some do not show age-related changes. ⁹ Importantly, even enzymes or transporters within the same class or family can have different patterns of maturation.

Developmental Differences in Drug Disposition Are Not All the Same

A number of high impact examples have been summarized, including the “gray syndrome” of cyanosis and cardiovascular collapse with chloramphenicol administration to babies, ⁹ decreased susceptibility to acetaminophen toxicity at some ages of both children and rodents, ^9,14 and increased toxicity of darunavir in preweaning rats potentially linked to increased blood and brain exposures. ¹⁵ These examples illustrate different mechanisms of developmental toxicity and further highlight the complexity of predicting and translating toxicity signals in juvenile rodents. For this reason, when juvenile toxicity assessments of small molecule therapeutics are warranted in rodents, conduct of pilot or dose range finding studies can help characterize the developmental impact of drug transporter and drug metabolic enzyme maturation on pharmacokinetics and toxicity. Ultimately, this can also further support appropriate toxicity monitoring and dose selection for pediatric use.

In summary, the structural and functional maturation of the gastrointestinal and hepatobiliary systems can critically contribute to drug disposition and toxicity. There are species, strain, and sex differences, in addition to environmental factors that can drive or influence the maturation process, all of which must be considered for appropriate planning and interpretation of JAS, as well as support for pediatric use of pharmaceuticals. Data gaps remain, and there are opportunities for collaborative study to further characterize and document postnatal ontologic processes.