Brian C. Tiegs

<h3>Background</h3> Interaction tables are restricted to unspecified isoenzymes of the families CYP1, CYP2, and CYP3. Data on gender impact has been required by the FDA only after 1993. <h3>Purpose</h3> The aim is to assess the effect of gender and age on pharmacokinetics and to explain inconsistent data reported so far. <h3>Materials and methods</h3> A systematic online literature research was performed on the usual platforms. 168 references could be evaluated. <h3>Results</h3> Ontogeny, peri- and postnatal phase: CYP450 inducibility begins in the earliest embryonic stage and reaches high rates before birth. Drugs are not distributed freely to all parts of the body in newborns. Childhood: preadolescent fasting boys absorb 35.2%, girls 45% of an oral iron loading dose. This explains the higher prevalence of iron anaemia in boys aged 11-15 (12.1%) compared to equally aged girls (6.1%). Adulthood: mean gastric fasting pH is 2.15 for men and 2.8 for women corresponding to a fivefold H⁺ activity in men. Women secrete gastrin and bicarbonate at the moment of substrate afflux, men more steadily. Gastric emptying, small intestine motility and colon transit times are downregulated by oestrogen and progesterone. In men, isoenzymes CYP1A2, CYP2C9, CYP2E1 are more active (CYP1A2 up to 40-fold), in women CYP3A4,5,7, CYP2A6, CYP2B6, and CYP2D6 (CYP2D6 only in the fertile phase). CYP3A4,5,7 activity depends on the menstrual cycle and peaks before ovulation and in pregnancy. Hepatic and intestinal P-gp (permeability glycoprotein) is expressed more in men than in women. Confused reports arise if the authors do not account for any mutually opposed effects of P-gp and CYP3A4,5,7. Pharmacokinetics change markedly in pregnancy due to slow motility, haemodynamics, cardiac output, etc. Incomplete protein digestion due to PPI treatment in pregnancy is a documented risk factor for predisposition to immune responses and asthma of the child (5.6% vs 3.7% in the untreated population). Copper absorption is higher in women aged 20-59 (71%) than in men of the same age (64%). This difference between the genders does not exist in the 60-83 age range. <h3>Conclusions</h3> Inconsistent data arise from crossed effects of co-localised P-gp and CYP3A4,5,7. Thus, only studies involving drugs that are not transported by P-gp are appropriate in CYP3A4,5,7 studies and vice versa. Interaction tables are limited tools. They do not distinguish between special patient groups or age ranges and thus need improvement.

Inhaled β-agonists are effective at reversing bronchoconstriction in asthma, but the mechanism by which they exert this effect is unclear and controversial. PKA is the historically accepted effector, although this assumption is made on the basis of associative and not direct evidence. Recent studies have asserted that exchange protein activated by cAMP (Epac), not PKA, mediates the relaxation of airway smooth muscle (ASM) observed with β-agonist treatment. This study aims to clarify the role of PKA in the prorelaxant effects of β-agonists on ASM. Inhibition of PKA activity via expression of the PKI and RevAB peptides results in increased β-agonist-mediated cAMP release, abolishes the inhibitory effect of isoproterenol on histamine-induced intracellular calcium flux, and significantly attenuates histamine-stimulated MLC-20 phosphorylation. Analyses of ASM cell and tissue contraction demonstrate that PKA inhibition eliminates most, if not all, β-agonist-mediated relaxation of contracted smooth muscle. Conversely, Epac knockdown had no effect on the regulation of contraction or procontractile signaling by isoproterenol. These findings suggest that PKA, not Epac, is the predominant and physiologically relevant effector through which β-agonists exert their relaxant effects.

Airway remodeling is a hallmark feature of asthma and chronic obstructive pulmonary disease. Clinical studies and animal models have demonstrated increased airway smooth muscle (ASM) mass, and ASM thickness is correlated with severity of the disease. Current medications control inflammation and reverse airway obstruction effectively but have limited effect on remodeling. Recently we identified the expression of bitter taste receptors (TAS2R) on ASM cells, and activation with known TAS2R agonists resulted in ASM relaxation and bronchodilation. These studies suggest that TAS2R can be used as new therapeutic targets in the treatment of obstructive lung diseases. To further establish their effectiveness, in this study we aimed to determine the effects of TAS2R agonists on ASM growth and promitogenic signaling. Pretreatment of healthy and asthmatic human ASM cells with TAS2R agonists resulted in a dose-dependent inhibition of ASM proliferation. The antimitogenic effect of TAS2R ligands was not dependent on activation of protein kinase A, protein kinase C, or high/intermediate-conductance calcium-activated K(+) channels. Immunoblot analyses revealed that TAS2R agonists inhibit growth factor-activated protein kinase B phosphorylation without affecting the availability of phosphatidylinositol 3,4,5-trisphosphate, suggesting TAS2R agonists block signaling downstream of phosphatidylinositol 3-kinase. Furthermore, the antimitogenic effect of TAS2R agonists involved inhibition of induced transcription factors (activator protein-1, signal transducer and activator of transcription-3, E2 factor, nuclear factor of activated T cells) and inhibition of expression of multiple cell cycle regulatory genes, suggesting a direct inhibition of cell cycle progression. Collectively, these findings establish the antimitogenic effect of TAS2R agonists and identify a novel class of receptors and signaling pathways that can be targeted to reduce or prevent airway remodeling as well as bronchoconstriction in obstructive airway disease.

ABSTRACT Arrestins have been shown to regulate numerous G protein‐coupled receptors (GPCRs) in studies employing receptor/arrestin overexpression in artificial cell systems. Which arrestin isoforms regulate which GPCRs in primary cell types is poorly understood. We sought to determine the effect of β‐arrestin‐1 or β‐arrestin‐2 inhibition or gene ablation on signaling and function of multiple GPCRs endogenously expressed in airway smooth muscle (ASM). In vitro [second messenger (calcium, cAMP generation)], ex vivo (ASM tension generation in suspended airway), and in vivo (invasive airway resistance) analyses were performed on human ASM cells and murine airways/whole animal subject to β‐arrestin‐1 or ‐2 knockdown or knockout (KO). In both human and murine model systems, knockdown or KO of β‐arrestin‐2 relative to control missense small interfering RNA or wild‐type mice selectively increased (40‐60%) β2‐adrenoceptor signaling and function. β‐arrestin‐1 knockdown or KO had no effect on signaling and function of β2‐adrenoceptor or numerous procontractile GPCRs, but selectively inhibited M3 muscarinic acetylcholine receptor signaling (∼50%) and function (∼25% ex vivo , &gt;50% in vivo ) without affecting EC 50 values. Arrestin subtypes differentially regulate ASM GPCRs and β‐arrestin‐1 inhibition represents a novel approach to managing bronchospasm in obstructive lung diseases.—Pera, T., Hegde, A., Deshpande, D. A., Morgan, S. J., Tiegs, B. C., Theriot, B. S., Choi, Y. H., Walker, J. K. L., Penn, R. B. Specificity of arrestin subtypes in regulating airway smooth muscle G protein‐coupled receptor signaling and function. FASEB J. 29, 4227‐4235 (2015). www.fasebj.org

To clarify the potential utility of targeting GRK2/3‐mediated desensitization as a means of manipulating airway smooth muscle (ASM) contractile state, we assessed the specificity of GRK2/3 regulation of procontractile and relaxant G‐protein‐coupled receptors in ASM. Functional domains of GRK2/3 were stably expressed, or siRNA‐mediated GRK2/3 knockdown was performed, in human ASM cultures, and agonist‐induced signaling was assessed. Regulation of contraction of murine tracheal rings expressing GRK2 C terminus was also assessed. GRK2/3 knockdown or expression of the GRK2 C terminus caused a significant (~30–90%) increase in maximal β‐agonist and histamine [phosphoinositide (PI) hydrolysis] signaling, without affecting the calculated EC 50 . GRK2 C‐terminal expression did not affect signaling by methacholine, thrombin, or LTD4. Expression of the GRK2 N terminus or kinase‐dead holo‐GRK2 diminished (~30–70%) both PI hydrolysis and Ca 2+ mobilization by every G q ‐coupled receptor examined. Under conditions of GRK2 C‐terminal expression, β‐agonist inhibition of methacholine‐stimulated PI hydrolysis was greater. Finally, transgenic expression of the GRK2 C terminus in murine ASM enabled ~30–50% greater β‐agonist‐mediated relaxation of methacholine‐induced contraction. Collectively these data demonstrate the relative selectivity of GRKs for the β 2 AR in ASM and the ability to exploit GRK2/3 functional domains to render ASM hyporesponsive to contractile agents while increasing responsiveness to bronchodilating β‐agonist.—Deshpande, D. A., Yan, H., Kong, K.‐C., Tiegs, B. C., Morgan, S. J., Pera, T., Panettieri, R. A., Eckhart, A. D., Penn, R. B. Exploiting functional domains of GRK2/3 to alter the competitive balance of pro‐ and anticontractile signaling in airway smooth muscle. FASEB J. 28, 956–965 (2014). www.fasebj.org