BIOPHARMACEUTICS & DRUG DISPOSITION, VOL. 13, 31 1-320 (1992)
REVIEW ARTICLE
DRUGS AND THE LIVER. PART 111: AGEING OF THE LIVER AND THE METABOLISM OF DRUGS KEN WOODHOUSE
Professor of Geriatric Medicine, University Department of Geriatric Medicine, University of Wales College of Medicine, Cardiff Royal Infirmary, Newport Road, Cardiff CF2 ISZ, Wales, UK
ABSTRACT The clearance of many drugs by the liver is reduced in the elderly, and this accounts, at least in part, for the greater frequency of adverse drug reactions in this population. Ageing in experimental animals is accompanied by a decline in the activities of certain microsomal monooxygenase enzymes; this has been said to be the reason for decreased drug elimination. However, recent data suggests that this is probably not the case in humans. Reduced size and blood flow are important features of the ageing liver, and probably account for most of the age-related changes in drug metabolism. Major decrements in hepatic function d o occur in frail elderly individuals, although the mechanisms underlying this are at present unclear. Future studies of drug metabolism and ageing must carefully define patient groups to allow full interpretation of data.
INTRODUCTION There is now an extensive body of evidence suggesting that elderly individuals are more prone to develop pharmacologically predictable Type A (i.e. dosedependent) adverse drug reactions. l Y 2 Many factors underly this, not least increased pharmacodynamic sensitivity to drugs; but impaired hepatic clearance is another important contributory f a ~ t o r . ~ It -has ~ always been assumed that the major determinant of these age-related changes in liver function is the decreased activity of the various metabolizing enzyme^.^.^ However, over the last 5 to 10 years a considerable body of evidence has accumulated to suggest that this may not be the case and that simpler morphological and physiological variables may be more important .6,7 The hepatic clearance of a drug (CLH) may be described by the formula:
0142-2782/92/050311-10$10.00 01992 by John Wiley & Sons, Ltd.
Accepted I7 January 1991
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where C, and C, are the arterial and venous concentrations of the drug respectively; E is the steady-state hepatic extraction ratio; and Q is liver blood flow. Extraction depends upon liver size, uptake, and characteristics of drug metabolizing enzymes. The most important variables in age-related changes in drug clearance are, therefore, likely to be:
1. Liver size. 2. Liver blood flow. 3. Liver uptake. 4. Specific activities of hepatic enzymes, and their inhibition and induction. Conventionally, drug metabolism has been divided into two phases.* Phase 1 reactions comprise oxidative, reductive, hydrolytic processes, which in general render the compound more polar, but also serve to expose or add functionally reactive sites to which even more polar moieties may be conjugated (Phase 2 reactions). These include glucuronidation, sulphation, acetylation, mecaptan formation, and others. This review will: 1. examine the in vivo studies of drug clearance in relation to age which are relevant to hepatic function; 2. examine the in vitro studies of drug metabolizing enzymes; 3. discuss the morphological and physiological changes which occur in the ageing liver; 4. consider the effects of disease and environmental influence on drug metabolism.
IN VIVO STUDIES OF DRUG METABOLISM IN HUMANS
Phase I reactions Antipyrine has been widely used as an experimental marker of hepatic drug o x i d a t i ~ n . ~ItJ ~ is well absorbed from the GI tract, has no significant presystemic metabolism, is poorly protein-bound, and is distributed in the body water. Antipyrine is metabolized to three major biotransformation products (norphenazone, 3-hydroxymethylantipyrine, and 4-hydroxyantipyrine) by the hepatic microsomal monooxygenase system. Many studies have shown that antipyrine half-life is significantlyprolonged with ageing,"-I5 and some authors suggest that these findings may be more marked in men. Some of these apparent age-related changes may be related to differences in smoking habits between young and elderly groups rather than age per se,I3but taken as a whole they suggest that the half-life of this drug may be extended by up to 45 per cent. A review of antipyrine formed the second part of this series.I6 Over the last 20 years a large number of careful pharmacokinetic studies on the elimination of drugs by hepatic Phase 1 drug metabolism have been
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Table 1 . Effect of age on the clearance of some oxidized drugs (references in brackets) No change
Decrease
Propranol01'~ Tolbutamide18
htipyrine19 Chlordiazepoxid9 Chlormethiazole2' Desmethyldiazepam" NicardipinG3 Nifedipinp Free Phenytoin19 VerapamilZS TheophyllinG6 A~etanilide2~.~*
published. Most of these have shown a decline in clearance with ageing, although this has been highly variable, ranging from 0 to 50 per cent. Examples of oxidized drugs which have been shown to be more slowly eliminated in the elderly are shown in Table 1. Recently, some in vivo studies have suggested that different cytochrome P450 isoforms may be differentially affected by ageing, and that perhaps such changes may be more pronounced in males.29Further studies are required to clarify this. Ethanol undergoes Phase 1 biotransformation by both cytosolic alcohol dehydrogenase (75-80 per cent) and by the microsomal ethanol oxidizing system MEOS (20-25 per cent). It has been shown that, although the distribution of ethanol may be modified by ageing, its hepatic clearance does not decline s~bstantially.~~
Phase 2 reactions Conjugation reactions have been studied rather less than Phase 1 processes. It is widely stated in the literature that conjugation rates are unaffected by ageing, and some work, particularly relating to conjugated benzodiazepines, is available to support this. However, other authors have reported a decline in the elimination of conjugated drugs with ageing, the most recent being a careful study of paracetamol clearance in the elderly. Clearly, further work is required to clarify this issue, but some data is summarized in Table 2. Table 2. Effect of age on the clearance of some conjugated drugs (references in brackets) No change
Decrease
Oxazeparn' ParacetamoP2
Lam~trigine~~ L~razepam~~ Morphine3s ParacetamoP6
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IN VZTRO STUDIES OF DRUG METABOLISM
Phase 1 reactions The cytochrome P450 microsomal monoxygenase enzyme system (MMO) has been most studied; an enormous body of work exists with regard to changes in this enzyme system in ageing animals,37recent reviews discussing the matter c~mprehensively.’.~ The earliest studies were in 1964 when a Japanese group demonstrated a decline in hepatic MMO activity in ageing inbred rats,37and similar findings have subsequently been reported from many other laboratories. Postulated mechanisms behind this decline have included a reduced smooth endoplasmic reticulum in aged rat liver, altered lipid profile of membranes, and, perhaps most importantly, alterations in cytochrome P450 isozyme profile^.^,^^ There is now clear evidence that most of the changes in MMO activity seen in elderly rodents are sex-related. MMO activities seem to be increased by testosterone in young males ,and a decline in the hypothalamic-pituitary-testicular axis with ageing results in a loss of male specific P450 isoforms, and a decline in drug clearance. Similar changes are obviously much less marked in The age-related changes noted in elderly animals have been widely extrapolated to primates including humans, but to date there is no good evidence of an agerelated decline in either MMO specific activities39 or in cytochrome P450 proteins. This has been investigated in macaque,40and rhesus monkeysY4’and more recently in human liver.42 Furthermore, the quality of enzyme proteins seemsto be just as good in liver from young and old rodents or humans -using a variety of probe substrates it has been demonstrated that the Michaelis constant of MMO enzymes does not increase with ageing as would be anticipated if poor quality enzyme protein were p r o d u ~ e d . ~ ~ , ~ With regard to microsomal monooxygenases, therefore, it is probably safe to say that age-related declines in activity do occur in rodents, but these seem to be sex- and species-specific. Similar reductions in ageing human or primate enzyme activities have not been shown, but the indirect evidence described above suggests that one or two specific P450 isoforms may be lost with ageing even in primates. Other Phase 1 enzyme systems have been studied much less extensively. Esterases are, however, an important group of drug metabolizing enzymes responsible for the detoxification of many compounds, and for the generation of pharmacologically active species from many prodrugs. They are found in disparate tissues, but predominantly in plasma and liver. Our studies of aspirin esterase activity in blood have clearly shown no decline in fit elderly individuals (but see below).45Furthermore, no correlation between age and enzyme activity was seen in either human or rat liver.&
Phase 2 reactions Even in rodents, fewer studies of Phase 2 conjugations have been undertaken. UDP-glucuronyl transferases have been examined in Fisher 344 and Wistar
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r a t ~ , ~and ~ , ~only * trivial changes have been reported. Our own studies of paracetamol glucuronidation and sulphation in both human49 and animal’O liver have shown no age-related variables of any significance. Glutathione conjugation is an important detoxification mechanism. The concentrations of glutathione in human liver are unaffected by ageing,39but the activities of the glutathione transferases have not been studied in man. However, animal studies have demonstrated trivial age-related change^.^^^^^ MORPHOLOGICAL AND PHYSIOLOGICAL CHANGES IN THE LIVER
Liver size It has been known for half a century that liver size falls with ageing. One of the largest studies was undertaken in 1933, when 1582 subjects suffering accidental death were studied. Ages were between 20 and 80 years, and liver size fell by 24 per cent in men and 18 per cent in women.s3 Similar results have been reported in other autopsy studies. However, this kind of report is difficult to interpret in that the dead are not representative of the general population, even if death is accidental. In the elderly, for example, injury is often a manifestation of general decline and physical illness. More recently, non-invasive methods of measuring liver size have been d e ~ e l o p e d ,including ~~ radionucleotide and computerized tomographic scanning, but most usefully, ultrasound scanning. Using this last technique, several studies have been undertaken in relation to age and liver size. For example, Swift in 1978 studied 15 volunteers in their third decade and 11 elderly volunteers aged 75-86 years. Mean liver volume fell by 24 per cent from 1303 rnl in the young to 990 ml in the elderly.55Our own studies of 65 community-based volunteers aged 24-91 have shown similar findings with a fall from 1474ml at 24 to 934 ml at 91.56 More recently, these findings have been confirmed by Marchesini et Reduced liver size of between 20 and 40 per cent is comparable with the decline in the clearance of oxidized drugs with ageing, which has been reported by many authors: the most common reduction being between 30 and 40 per cent. Our studies have shown that the reduction in both acetanilidS8 and paracetam01~~ clearance with age is almost entirely due to reduced liver volume.
Liver blood flow Blood flow is an important determinant of the clearance of high extraction drugs, but because of the liver’s dual blood flow, direct measurements of hepatic blood flow tend to be invasive. Doppler methods are difficult and, in clinical practice, liver blood flow is estimated using clearance technique^.^^ The most common marker compound is indocyanine green (ICG). This is taken up into hepatocytes and secreted in bile, it has no entrapatic recirculation,
3 16
KEN WOODHOUSE
exhibits flow-dependent clearance, and has a hepatic extraction ratio of 14 per cent. A significant negative correlation between ICG clearance and age has been reported; in our own studies this comprised a 35 per cent decline between 24 to 91 years.56 We have also clearly shown that reduced ICG clearance with ageing is due to blood flow changes, and that the uptake of the compound into the liver is not age-de~endent.~~ A review of liver function tests formed the first part of this series.59 DISEASE, FRAILTY, AND ENVIRONMENTAL INFLUENCES
Physical frailty One problem which has bedevilled studies of drug metabolism in ageing is the concurrent presence of disease and physical disability. There is now good in vivo and in vitro evidence that physical frailty as defined by Woodhousem is associated with significant decrements in drug metabolism. For example, the clearance of paracetamol as the glucuronide is significantly impaired in frail patients compared with both young volunteers and fit elderly volunteers. This difference persists even when allowance for changes in liver volume are made.28 Similarly, the in vitro activity of plasma aspirin esterase is not affected by ageing per se, but is significantly impaired in the frail elderly.44These findings highlight the need to carefully define patient groups.
EnZyme inhibition It had been postulated that the elderly may be more sensitive to environmental inhibitors of drug metabolism, including other pharmacological agents. However, this appears not to be the case. Two studies have investigated the effect of cimetidine on drug oxidation; both have shown that the magnitude of inhibition is similar in old and young.61162Similarly, the inhibitory effect of alcohol on drug oxidation is comparable in young and elderly groups.27
Induction The exact relationship between ageing and the induction of drug metabolism has yet to be clarified. Several in vivo studies have suggested that the elderly respond more slowly, and to a lesser degree, to exogenous inducing agents than do the similar findings have been reported in in vitro studies in animal liver^.^^-^^ Our own studies have failed to demonstrate impaired induction of cytochrome P450 in response to benzanthracene using a human peripheral blood monocyte m ~ d e l . ’ ~Further ,~~ work is required in this area.
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CONCLUSIONS In summary, the following conclusions can be drawn in relation to hepatic drug metabolism and ageing: 1. The in vivo clearance of a large number of drugs declines with age, and this seems to be true for drugs metabolized by both Phase 1 and Phase 2 processes. 2. The activity of drug metabolizing enzymes in the liver does not appear to decline with age. This would appear to be true for most cytochrome P450 species; for esterase; and for glucuronyl and sulphotransferases. 3. There is indirect evidence that a few highly specific cytochrome P450 forms may be impaired with ageing in humans, although further work is needed. This may reflect the position in animals where the male-specific cytochrome P450s are lost with ageing. 4. Liver size and liver blood flow fall significantly with ageing; these alterations are probably the main determinants of reduced hepatic drug clearance in the elderly. 5 . Physical frailty is a major determinant of impaired drug clearance in the elderly, and this appears to be an independent variable in addition to age. 6. The response of hepatic drug metabolism in the elderly to inhibitors is similar to that in the young. 7. Enzyme induction appears to be decreased in vivo in the elderly in some studies, but this has not as yet been confirmed in humans using in vitro methodology.
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40. M. A. Sutter, W. G. Wood, L. S . Williamson ef al. Comparison of the hepatic mixed function oxidase system of young adults, and old non-human primates (Macca nemesfrina).Biochem. Pharmacol., 34, 2983-2987 (1985). 41. A. G. Maloney, D. L. Schmucker, D. S . Vessey et al. The effects of ageing on the hepatic micros o d mixed-function oxidax system of male and female monkeys. Hepfology,6,282-287 (1986). 42. D. L. Schmucker, K. W. Woodhouse, R. K. Wang, H. Wynne, 0. F. W. James ef al. Effect of age and gender on in vitro properties of human liver microsomal monooxygenases. Clin. Pharm. Ther. 48, 365-374 (1990). 43. H. Wynne, E. Mutch, 0. F. W. James, M. Rawlins and K. W. Woodhouse, Effect of age on monooxygenase enzyme kinetics in rat liver microsomes. Age Ageing, 16: 153-158 (1987). 44. H. Wynne, E. Mutch, 0. F. W. James, I. P. Wright, M. Rawlins and K. W. Woodhouse, The effect of age upon the affinity of microsomal monooxygenase enzymes for substrate in human liver. Age Ageing, 17, 401-405 (1988). 45. F. M. Williams, H. Wynne, K. W. Woodhouse and M. D. Rawlins, Plasma aspirin esterase: The influence of old age and frailty. Age Ageing, 18, 39-42 (1989). 46. C. Yelland, E. Nicholson, H. Wynne and K. W. Woodhouse, The effect of age upon aspirin esterase activity in human liver. Age Ageing, 20, 16-18 (1991). 47. M. ALi, P. J. Nicholls and A. Yoosuf, The influence of old age and of renal failure on hepatic glucuronidation in the rat. Proceedings of the BPS, April, 1979, pp. 498-499. 48. S. J. Borghoff and L. S. Bimhaum, Age-related changes in glucuronidation and deglucuronidation in liver, small intestine, lung and kidney of male Fischer rats. Drug Metab. Dkpos., 13,62-67 (1985). 49. B. Herd and K. W. Woodhouse, The effect of age and gender on glucuronidation and sulphation
in rat liver. Arch Geronfol Geriafr (in press). 50. B. Herd, H. Wynne, P. Wright, 0. F. W. James and K. W. Woodhouse, The effect of age on glucuronidation and sulphation of paracetamol by human liver fractions. Br. J. Clin. Pharmacol., 32, 768-770 (1991). 51. M. E. Spearman and K.C. Leibman, Effects of aging on hepatic and pulmonary glutathioneS-transferase activities in male and female Fischer 344 rats. Biochem. Pharmacol., 33, 1309-1313 (1984). 52. D. J. Sweeny and M. Weiner, Metabolism of acetaminophen in hepatocytes isolated from mice and rats of various ages. Drug Metab., 13, 377-379 (1985). 53. E. Boyd, Normal variability of weight of the adult human liver and spleen. Arch. Pafhol., 16, 350-372 (1933). 54. K. W. Woodhouse and H. Wynne, Age-related changes in hepatic function: Implications for drug therapy. Drugs and Aging, 1991 (in press).
55. C. G. Swift, M. Homeida, M. Halliwell and C. J. Roberts, Antipyrine disposition and liver size in the elderly. Eur. J. Clin. Pharmacol., 14, 149-152 (1978). 56. H. Wynne, L. Cope, E. Mutch, M.D. Rawlins, K. W. Woodhouse and 0. F. W. James, The effect of age upon liver size and liver blood flow in man. Hepatology, 9, 297-301 (1989). 57. G. Marchesini, G. P. Bianchi, A. Fabbri ef al. Synthesis of urea after a protein-rich meal in normal man in relation to ageing. Age Ageing, 19, 4-10 (1990). 58. H. Wynne, J. Goudouvenous, P. Adams, M. D. Rawlins, 0. F. W. James and K. W. Woodhouse. Hepatic extraction of drugs: The effect of age using indocyanine green as a model substrate. Br. J. Clin. Pharmacol., 30, 634-637 (1990). 59. R. W e , J. H. Tredger and R. Williams, Drugs and the liver I. Testing liver function. Biopharm. Drug. Disposif., 12, 251-259 (1991). 60. K. W. Woodhouse, H. Wynne, S. Baillie, 0. F. W. James and M. D. Rawlins, Who are the frail elderly? Q. J. Med., 68, 505-506 (1988). 61. J. Feely, L. Pereira, E. Guy and N. Hockings, Factors affecting the response to inhibition of drug metabolism to cimetidine-dose response and sensitivity of elderly and induced subjects. Br. J. Clin. Pharmacol., 17, 77-81 (1984). 62. N. Divoll, D. Greenblatt, D. Abernethy and R. Shader, Cimetidine impairs clearance of antipyrine and desmethyldiazepam in elderly. J. Am. Geriafr. SOC.,30, 684-689 (1982). 63. S. A. M. Salem, P. Rajjayabu, A. M. M. Sheperd and I. H. Stevenson, Reduced induction of drug metabolism in the elderly. Age Ageing, 7, 68-73 (1978). 64. M. W. Pearson and C. J. Roberts, Drug induction of hepatic enzymes in the elderly. Age Ageing, 13, 313 (1984).
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Y. Twum-Barima, T. Finnigan, A. I. Habash. R. D. Cape and S. G. Carruthers, Impaired enzyme induction by rifampicin in the elderly. Br. J. Clin. Pharmacol., 17, 595-597 (1984). R. Wang. Effects of aging and phenobarbital on the rat liver microsomal drug metabolising system. Mech. Ageing Dev.,15, 189-202 (1981). D. McMartin, J. O’Connor, M. Fasco and I. Kaminsky, Influence of aging and induction on rat liver and kidney microsomal mixed function oxidase systems. Toxicol. Appl. Pharmacol., 54, 411-419 (1980). L. Birnbaum and M. Baird, Induction of hepatic mixed function oxidases in senescent rodents. Exp. Gerontol., 13, 299-303 (1978). L. Birnbaum and M. Baird, Induction of hepatic mixed function oxidases in senescent rodents. 11. Effect of polychlorinated biphenyls. Exp. Gerontol., 13, 469-477 (1978). G. George, H. A. Wynne and K. W. Woodhouse, The association of age with the induction of drug metabolising enzymes in human monocytes. Age Ageing, 19, 364-367 (1990). G. George, H.Wynne and K. W. Woodhouse, Age and the rate of induction of aryl hydrocarbon hydroxylase in isolated peripheral blood monocytes. Age Ageing, 1991 (in press).
66. D. Schmucker and 67. 68. 69. 70. 71.
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REVIEW ARTICLE
DRUGS AND THE LIVER. PART 111: AGEING OF THE LIVER AND THE METABOLISM OF DRUGS KEN WOODHOUSE
Professor of Geriatric Medicine, University Department of Geriatric Medicine, University of Wales College of Medicine, Cardiff Royal Infirmary, Newport Road, Cardiff CF2 ISZ, Wales, UK
ABSTRACT The clearance of many drugs by the liver is reduced in the elderly, and this accounts, at least in part, for the greater frequency of adverse drug reactions in this population. Ageing in experimental animals is accompanied by a decline in the activities of certain microsomal monooxygenase enzymes; this has been said to be the reason for decreased drug elimination. However, recent data suggests that this is probably not the case in humans. Reduced size and blood flow are important features of the ageing liver, and probably account for most of the age-related changes in drug metabolism. Major decrements in hepatic function d o occur in frail elderly individuals, although the mechanisms underlying this are at present unclear. Future studies of drug metabolism and ageing must carefully define patient groups to allow full interpretation of data.
INTRODUCTION There is now an extensive body of evidence suggesting that elderly individuals are more prone to develop pharmacologically predictable Type A (i.e. dosedependent) adverse drug reactions. l Y 2 Many factors underly this, not least increased pharmacodynamic sensitivity to drugs; but impaired hepatic clearance is another important contributory f a ~ t o r . ~ It -has ~ always been assumed that the major determinant of these age-related changes in liver function is the decreased activity of the various metabolizing enzyme^.^.^ However, over the last 5 to 10 years a considerable body of evidence has accumulated to suggest that this may not be the case and that simpler morphological and physiological variables may be more important .6,7 The hepatic clearance of a drug (CLH) may be described by the formula:
0142-2782/92/050311-10$10.00 01992 by John Wiley & Sons, Ltd.
Accepted I7 January 1991
312
KEN WOODHOUSE
where C, and C, are the arterial and venous concentrations of the drug respectively; E is the steady-state hepatic extraction ratio; and Q is liver blood flow. Extraction depends upon liver size, uptake, and characteristics of drug metabolizing enzymes. The most important variables in age-related changes in drug clearance are, therefore, likely to be:
1. Liver size. 2. Liver blood flow. 3. Liver uptake. 4. Specific activities of hepatic enzymes, and their inhibition and induction. Conventionally, drug metabolism has been divided into two phases.* Phase 1 reactions comprise oxidative, reductive, hydrolytic processes, which in general render the compound more polar, but also serve to expose or add functionally reactive sites to which even more polar moieties may be conjugated (Phase 2 reactions). These include glucuronidation, sulphation, acetylation, mecaptan formation, and others. This review will: 1. examine the in vivo studies of drug clearance in relation to age which are relevant to hepatic function; 2. examine the in vitro studies of drug metabolizing enzymes; 3. discuss the morphological and physiological changes which occur in the ageing liver; 4. consider the effects of disease and environmental influence on drug metabolism.
IN VIVO STUDIES OF DRUG METABOLISM IN HUMANS
Phase I reactions Antipyrine has been widely used as an experimental marker of hepatic drug o x i d a t i ~ n . ~ItJ ~ is well absorbed from the GI tract, has no significant presystemic metabolism, is poorly protein-bound, and is distributed in the body water. Antipyrine is metabolized to three major biotransformation products (norphenazone, 3-hydroxymethylantipyrine, and 4-hydroxyantipyrine) by the hepatic microsomal monooxygenase system. Many studies have shown that antipyrine half-life is significantlyprolonged with ageing,"-I5 and some authors suggest that these findings may be more marked in men. Some of these apparent age-related changes may be related to differences in smoking habits between young and elderly groups rather than age per se,I3but taken as a whole they suggest that the half-life of this drug may be extended by up to 45 per cent. A review of antipyrine formed the second part of this series.I6 Over the last 20 years a large number of careful pharmacokinetic studies on the elimination of drugs by hepatic Phase 1 drug metabolism have been
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Table 1 . Effect of age on the clearance of some oxidized drugs (references in brackets) No change
Decrease
Propranol01'~ Tolbutamide18
htipyrine19 Chlordiazepoxid9 Chlormethiazole2' Desmethyldiazepam" NicardipinG3 Nifedipinp Free Phenytoin19 VerapamilZS TheophyllinG6 A~etanilide2~.~*
published. Most of these have shown a decline in clearance with ageing, although this has been highly variable, ranging from 0 to 50 per cent. Examples of oxidized drugs which have been shown to be more slowly eliminated in the elderly are shown in Table 1. Recently, some in vivo studies have suggested that different cytochrome P450 isoforms may be differentially affected by ageing, and that perhaps such changes may be more pronounced in males.29Further studies are required to clarify this. Ethanol undergoes Phase 1 biotransformation by both cytosolic alcohol dehydrogenase (75-80 per cent) and by the microsomal ethanol oxidizing system MEOS (20-25 per cent). It has been shown that, although the distribution of ethanol may be modified by ageing, its hepatic clearance does not decline s~bstantially.~~
Phase 2 reactions Conjugation reactions have been studied rather less than Phase 1 processes. It is widely stated in the literature that conjugation rates are unaffected by ageing, and some work, particularly relating to conjugated benzodiazepines, is available to support this. However, other authors have reported a decline in the elimination of conjugated drugs with ageing, the most recent being a careful study of paracetamol clearance in the elderly. Clearly, further work is required to clarify this issue, but some data is summarized in Table 2. Table 2. Effect of age on the clearance of some conjugated drugs (references in brackets) No change
Decrease
Oxazeparn' ParacetamoP2
Lam~trigine~~ L~razepam~~ Morphine3s ParacetamoP6
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IN VZTRO STUDIES OF DRUG METABOLISM
Phase 1 reactions The cytochrome P450 microsomal monoxygenase enzyme system (MMO) has been most studied; an enormous body of work exists with regard to changes in this enzyme system in ageing animals,37recent reviews discussing the matter c~mprehensively.’.~ The earliest studies were in 1964 when a Japanese group demonstrated a decline in hepatic MMO activity in ageing inbred rats,37and similar findings have subsequently been reported from many other laboratories. Postulated mechanisms behind this decline have included a reduced smooth endoplasmic reticulum in aged rat liver, altered lipid profile of membranes, and, perhaps most importantly, alterations in cytochrome P450 isozyme profile^.^,^^ There is now clear evidence that most of the changes in MMO activity seen in elderly rodents are sex-related. MMO activities seem to be increased by testosterone in young males ,and a decline in the hypothalamic-pituitary-testicular axis with ageing results in a loss of male specific P450 isoforms, and a decline in drug clearance. Similar changes are obviously much less marked in The age-related changes noted in elderly animals have been widely extrapolated to primates including humans, but to date there is no good evidence of an agerelated decline in either MMO specific activities39 or in cytochrome P450 proteins. This has been investigated in macaque,40and rhesus monkeysY4’and more recently in human liver.42 Furthermore, the quality of enzyme proteins seemsto be just as good in liver from young and old rodents or humans -using a variety of probe substrates it has been demonstrated that the Michaelis constant of MMO enzymes does not increase with ageing as would be anticipated if poor quality enzyme protein were p r o d u ~ e d . ~ ~ , ~ With regard to microsomal monooxygenases, therefore, it is probably safe to say that age-related declines in activity do occur in rodents, but these seem to be sex- and species-specific. Similar reductions in ageing human or primate enzyme activities have not been shown, but the indirect evidence described above suggests that one or two specific P450 isoforms may be lost with ageing even in primates. Other Phase 1 enzyme systems have been studied much less extensively. Esterases are, however, an important group of drug metabolizing enzymes responsible for the detoxification of many compounds, and for the generation of pharmacologically active species from many prodrugs. They are found in disparate tissues, but predominantly in plasma and liver. Our studies of aspirin esterase activity in blood have clearly shown no decline in fit elderly individuals (but see below).45Furthermore, no correlation between age and enzyme activity was seen in either human or rat liver.&
Phase 2 reactions Even in rodents, fewer studies of Phase 2 conjugations have been undertaken. UDP-glucuronyl transferases have been examined in Fisher 344 and Wistar
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r a t ~ , ~and ~ , ~only * trivial changes have been reported. Our own studies of paracetamol glucuronidation and sulphation in both human49 and animal’O liver have shown no age-related variables of any significance. Glutathione conjugation is an important detoxification mechanism. The concentrations of glutathione in human liver are unaffected by ageing,39but the activities of the glutathione transferases have not been studied in man. However, animal studies have demonstrated trivial age-related change^.^^^^^ MORPHOLOGICAL AND PHYSIOLOGICAL CHANGES IN THE LIVER
Liver size It has been known for half a century that liver size falls with ageing. One of the largest studies was undertaken in 1933, when 1582 subjects suffering accidental death were studied. Ages were between 20 and 80 years, and liver size fell by 24 per cent in men and 18 per cent in women.s3 Similar results have been reported in other autopsy studies. However, this kind of report is difficult to interpret in that the dead are not representative of the general population, even if death is accidental. In the elderly, for example, injury is often a manifestation of general decline and physical illness. More recently, non-invasive methods of measuring liver size have been d e ~ e l o p e d ,including ~~ radionucleotide and computerized tomographic scanning, but most usefully, ultrasound scanning. Using this last technique, several studies have been undertaken in relation to age and liver size. For example, Swift in 1978 studied 15 volunteers in their third decade and 11 elderly volunteers aged 75-86 years. Mean liver volume fell by 24 per cent from 1303 rnl in the young to 990 ml in the elderly.55Our own studies of 65 community-based volunteers aged 24-91 have shown similar findings with a fall from 1474ml at 24 to 934 ml at 91.56 More recently, these findings have been confirmed by Marchesini et Reduced liver size of between 20 and 40 per cent is comparable with the decline in the clearance of oxidized drugs with ageing, which has been reported by many authors: the most common reduction being between 30 and 40 per cent. Our studies have shown that the reduction in both acetanilidS8 and paracetam01~~ clearance with age is almost entirely due to reduced liver volume.
Liver blood flow Blood flow is an important determinant of the clearance of high extraction drugs, but because of the liver’s dual blood flow, direct measurements of hepatic blood flow tend to be invasive. Doppler methods are difficult and, in clinical practice, liver blood flow is estimated using clearance technique^.^^ The most common marker compound is indocyanine green (ICG). This is taken up into hepatocytes and secreted in bile, it has no entrapatic recirculation,
3 16
KEN WOODHOUSE
exhibits flow-dependent clearance, and has a hepatic extraction ratio of 14 per cent. A significant negative correlation between ICG clearance and age has been reported; in our own studies this comprised a 35 per cent decline between 24 to 91 years.56 We have also clearly shown that reduced ICG clearance with ageing is due to blood flow changes, and that the uptake of the compound into the liver is not age-de~endent.~~ A review of liver function tests formed the first part of this series.59 DISEASE, FRAILTY, AND ENVIRONMENTAL INFLUENCES
Physical frailty One problem which has bedevilled studies of drug metabolism in ageing is the concurrent presence of disease and physical disability. There is now good in vivo and in vitro evidence that physical frailty as defined by Woodhousem is associated with significant decrements in drug metabolism. For example, the clearance of paracetamol as the glucuronide is significantly impaired in frail patients compared with both young volunteers and fit elderly volunteers. This difference persists even when allowance for changes in liver volume are made.28 Similarly, the in vitro activity of plasma aspirin esterase is not affected by ageing per se, but is significantly impaired in the frail elderly.44These findings highlight the need to carefully define patient groups.
EnZyme inhibition It had been postulated that the elderly may be more sensitive to environmental inhibitors of drug metabolism, including other pharmacological agents. However, this appears not to be the case. Two studies have investigated the effect of cimetidine on drug oxidation; both have shown that the magnitude of inhibition is similar in old and young.61162Similarly, the inhibitory effect of alcohol on drug oxidation is comparable in young and elderly groups.27
Induction The exact relationship between ageing and the induction of drug metabolism has yet to be clarified. Several in vivo studies have suggested that the elderly respond more slowly, and to a lesser degree, to exogenous inducing agents than do the similar findings have been reported in in vitro studies in animal liver^.^^-^^ Our own studies have failed to demonstrate impaired induction of cytochrome P450 in response to benzanthracene using a human peripheral blood monocyte m ~ d e l . ’ ~Further ,~~ work is required in this area.
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CONCLUSIONS In summary, the following conclusions can be drawn in relation to hepatic drug metabolism and ageing: 1. The in vivo clearance of a large number of drugs declines with age, and this seems to be true for drugs metabolized by both Phase 1 and Phase 2 processes. 2. The activity of drug metabolizing enzymes in the liver does not appear to decline with age. This would appear to be true for most cytochrome P450 species; for esterase; and for glucuronyl and sulphotransferases. 3. There is indirect evidence that a few highly specific cytochrome P450 forms may be impaired with ageing in humans, although further work is needed. This may reflect the position in animals where the male-specific cytochrome P450s are lost with ageing. 4. Liver size and liver blood flow fall significantly with ageing; these alterations are probably the main determinants of reduced hepatic drug clearance in the elderly. 5 . Physical frailty is a major determinant of impaired drug clearance in the elderly, and this appears to be an independent variable in addition to age. 6. The response of hepatic drug metabolism in the elderly to inhibitors is similar to that in the young. 7. Enzyme induction appears to be decreased in vivo in the elderly in some studies, but this has not as yet been confirmed in humans using in vitro methodology.
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