Michael R. Moore

International review of drugs in acute porphyria—1980

In the years following the very successful review of the effects of drugs in acute intermittent porphyria by Wetterberg (1976), many new items of information have accrued on this subject. The aim of this current review (initiated at the International Meeting on Porphyrins and the Porphyrias in Buenos Aires, 1979) is to up-date this information with especial reference to the potential clinical effects of drugs. It will contain no information on the effects of drugs in the non-acute porphyrias, since in these diseases only circumscribed groups of drugs may be linked with exacerbation of the disease, such as exacerbation of cutaneous hepatic porphyria (porphyria cutanea tarda) by ethanol and chlorinated hydrocarbons or a similar exacerbation of erythropoietic protoporphyria by ethanol (MathewsRoth, 1980). The importance of this review may be understood in the description of the acute porphyrias as a group of pharmacogenetic diseases, that is a group of genetically defined diseases which show an idiosyncratic reaction to many common drugs. The porphyrias may thus be induced into attack in the presence of certain drugs especially drugs known to be inducers of cytochrome P-450 mediated oxidation although other mechanisms are undoubtedly present (Maxwell & Meyer, 1976; Meyer, 1978). Because of the multiplicity of drug structures, it is usually very difficult to predict whether or not a drug may induce excess porphyrin synthesis although it has reasonably been suggested that all lipophilic drugs must be suspect in this respect (De Matteis, 1971; Smith & De Matteis, 1980; Sweeney, 1980). Certain chemical groups have been linked with induction of porphyria, such as ally1 groups (Goldberg & Rimington, 1954), the basic nucleus of the barbiturates (De Matteis, 1967) and certain steroids (Paxton et al., 1976). Typically, drugs such as the barbiturates and diphenylhydantoin act by early depletion of free haem and thus induce delta-aminolaevulinic acid synthase. Although depletion of free haem is probably the principal mechanism in action, it has been noted that the porphyrinogenic barbiturates may decrease the activity of steroid 5-a-reductase (Kappas et al., 1977), thus mimicking the ratio difference of 5-p to S-a steroids found in acute porphyrias (Moore er al., 1973). In recent years, a number of very useful reviews of porphyrinogenic effects of specific categories of drugs have been produced, especially drugs which might potentially be used during an acute porphyric attack, such as anti-hypertensive drugs (Anderson, 1978), anaesthetic drugs (Parikh & Moore, 1978) and anticonvulsant drugs (McCall, 1980; Larson et al.. 1978). For further information on these drugs and the methods used to arrive at the classification, the reader should consult the appropriate papers. There are three means by which drugs may be implicated in induction of attacks of porphyria. Firstly, there is analysis of the effects of the drugs in various types of cell culture on either production of porphyrins, or induction of ALA synthase activity. This is a very sensitive means of assessment and will often produce false positive results. Secondly, there is a similar assessment in the effects of drugs either on their own or in combination with proven porphyrinogens in whole animals. This suffers from the physiological differences in drug metabolism routes between rats and man and thus in differences in drug sensitivity between these two species. Lastly, there is the anecdotal evidence of porphyrinogenicity in man. either through induction of acute attacks by drugs or by measurement of ALA synthase activity or porphyrins in human tissues and excreta. This information may be culled from the literature and from the personal experience of workers in the field. This current review has concentrated upon this last approach. Information has been requested of a large number of investigators in the world who have experience of the porphyrias and their treatment. From the volumes of evidence that they have submitted and from lists of drugs considered by them to be safe and unsafe for use in the porphyrias, the following tables have been generated. In many cases, the information depends not only on the third approach, but also on the first and second forms of approach. For this reason, it is likely that all drugs listed as “safe” for use in the porphyrias are safe, but that drugs listed as “unsafe” in the porphyrias are not definitively so and that in many cases, porphyric patients may have been treated with one or more of these drugs with no ill effect. Equally, one can say that none of the drugs listed as unsafe for use in the porphyrias should be used in these diseases without extreme care, lest one potentiate an acute attack of porphyria.

Abnormal haem biosynthesis in chronic alcoholics.

Abstract. The activities of six of the enzymes of haem biosynthesis have been examined in eleven chronic alcoholics admitted to hospital for alcohol withdrawal. The mitochondrial enzymes delta‐aminolae‐vulinic acid (ALA) synthase, coproporphyrinogen oxidase and ferrochelatase were monitored in peripheral leucocytes and the cytosolic enzymes ALA dehydratase, uroporphyrinogen‐1 ‐synthase and uro‐porphyrinogen decarboxylase in peripheral erythro‐cytes. Compared with control subjects the activity of the initial and rate controlling enzyme of the pathway, ALA synthase, was increased (P < 0.01) and the activities of ALA dehydratase and uroporphyrinogen decarboxylase depressed (P < 0.01, P < 0.02 respectively) on the day after admission but all returned to normal by the tenth to twentieth days after alcohol withdrawal. This stimulation of ALA synthase and inhibition of uroporphyrinogen decarboxylase explains the mechanism by which chronic alcohol ingestion may precipitate cutaneous hepatic porphyria.

Sex differences in haem biosynthesis and porphyrin content in the harderian gland of the golden hamster

Methods are described for the measurement of seven haem biosynthetic enzymes in Harderian gland tissue from male and female golden hamsters. Sex differences were found in five of the seven enzymes. In each case, female tissue exhibited higher activity than male tissue. These differences in enzyme activity are sufficient to account for the major sex difference in porphyrin content in the Harderian gland of this species.

Detection of seven point mutations in the porphobilinogen deaminase gene in patients with acute intermittent porphyria, by direct sequencing of in vitro amplified cDNA.

SummaryDirect cDNA sequencing has been performed on asymmetrically amplified transcripts from the human porphobilinogen deaminase gene. Lymphocytes from 30 patients with acute intermittent porphyria were the source of mRNA; of the seven separate point mutations detected, three were silent, whereas four resulted in amino acid changes. Three of these changes involved highly conserved amino acids, and the remaining one a conserved charge. One of these mutations was predicted to cause structural alterations in the protein product. The application of this method to affected families allows the direct identification of these heterogeneous mutations, thus permitting the unequivocal detection of carriers.

Tin protoporphyrin prolongs the biochemical remission produced by heme arginate in acute hepatic porphyria

BACKGROUND In acute porphyria, repletion of intrahepatic heme, with exogenously administered heme, suppresses the overproduction of delta-aminolaevulinic acid (ALA) and porphobilinogen (PBG). The effect of reducing heme breakdown has been assessed by administering tin protoporphyrin, a competitive inhibitor of heme oxygenase. METHODS The effect of tin protoporphyrin, 1 mumol/kg, and heme arginate, 3 mg/kg, individually and combined was compared with placebo in patients with an acute porphyric crisis. The treatments were given by intravenous infusion on three successive mornings. Thirty-four attacks were studied in 8 patients (9 placebo, 10 heme arginate alone, 4 tin protoporphyrin alone, and 11 combination treatments). RESULTS Placebo and tin protoporphyrin alone had little effect on ALA and PBG excretion. Following heme arginate alone or combined with tin protoporphyrin, there was a marked and similar suppression of both ALA and PBG excretion (P < 0.005 for each, compared with pretreatment values). However, on the 5th day after discontinuing treatment, the excretion of ALA and PBG were both lower following combination therapy than following heme arginate alone (P < 0.005 and P < 0.01, respectively). CONCLUSIONS These findings suggest that inhibition of heme oxygenase by tin protoporphyrin prolongs the biochemical remission induced by heme arginate in the porphyric crisis.

Detection of a high mutation frequency in exon 12 of the porphobilinogen deaminase gene in patients with acute intermittent porphyria

Direct cDNA sequencing was performed on asymmetrically amplified transcripts from the porphobilinogen deaminase (PBG-D) gene of thirteen unrelated individuals with acute intermittent porphyria. Four different mutations and a polymorphic site were detected in exon 12 of the gene, four being the result of single base substitutions and one being caused by dinucleotide deletion. All of these mutations are located in domain 3 of the PBG-D molecule, with the single base substitutions affecting the hydrophobic interfaces between domains 1 and 3. The dinucleotide deletion results in a frame-shift producing a premature stop codon.

HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC ANALYSES OF PORPHYRINS IN HAMSTER HARDERIAN GLANDS

Porphyrin content and 5-aminolaevulinate synthase activity of the Harderian gland were measured in intact and gonadectomized male and female hamsters; porphyrin profiles were analysed by high-pressure liquid chromatography. The total porphyrin content of the two female groups was similar, but enzyme activity in females ovariectomised for 20 weeks significantly decreased. Intact males have low porphyrin content and enzyme activity, while in castrates (6 weeks) both increased to female levels. Protoporphyrin IX formed 93% of total porphyrins in intact females, compared with 70% of total porphyrins in intact males. The remainder in both sexes was chiefly penta- and hexacarboxylic porphyrins and coproporphyrin and (in females) Harderoporphyrin. Gonadectomy in either sex resulted in protoporphyrin levels intermediate between male and female values.

Successful abatement of lead exposure from water supplies in the West of Scotland

A major problem has existed in the West of Scotland for at least the past century associated with lead uptake by water from leaden water distribution systems. Initial studies in Glasgow from 1969 to 1976 and in Ayr in 1980/1981 showed that not only were water supplies soft, acid in consequence highly plumbosolvent, and that water lead levels were, on average, unacceptably high but that blood lead concentrations were also in excess of acceptable limits. A decision was therefore made by Strathclyde Water Department to carry out remedial water treatment to adjust the water pH. The success of this venture was proven by the parallel rapid falls in blood lead and water lead concentrations in the city of Glasgow. This encouraged the Water Department to institute a similar scheme in the town of Ayr. Work on this commenced in 1981, and in a study of the participants in a previous blood lead survey, a highly significant fall in blood lead concentrations was observed. The equation linking these two parameters was found to follow a curvilinear relationship where blood lead varied as the cube root of the water lead with a highly significant coefficient of correlation. This relationship has been shown to hold across a wide range of water lead concentrations down to 1 microgram/liter. This detailed information allows accurate calculation of acceptable limits of lead exposure from specific sources based upon acceptable blood lead concentrations.

Elevation of blood lactate and pyruvate levels in acute intermittent porphyria — A reflection of haem deficiency?

Blood lactate concentrations after glucose loading were significantly higher in 6 patients with acute intermittent porphyria in clinical remission than in 6 control subjects and the percentage rise in glucose pyruvate and lactate concentrations were greater in the porphyric subjects than in the control group. It is postulated that the raised lactate levels in the porphyric patient group may reflect haem deficiency affecting the cytochromes of the terminal respiratory chain.

Regulation of haem biosynthesis in normoblastic erythropoiesis: role of 5-aminolaevulinic acid synthase and ferrochelatase

The development of haem biosynthetic enzyme activity during normoblastic human erythropoiesis was examined in seven patients. The first and last enzymes of the haem biosynthetic pathway, ALA synthase and ferrochelatase, were assayed by radiochemical/high performance liquid chromatographic (HPLC) methods. An assay for ferrochelatase activity in human bone marrow was developed. Enzyme substrates were protoporphyrin IX and 59Fe2+ ions. 59Fe-labelled haem was isolated by organic solvent extraction/sorbent extraction followed by reversed-phase HPLC. Optimal activity occurred at pH 7.3 in the presence of ascorbic acid, in darkness and under anaerobic conditions. Haem production was proportional to cell number and was linear with time to 30 min. The assay was sensitive to the picomolar range of haem production. ALA synthase and ferrochelatase activity was assayed in four highly purified age-matched erythroid cell populations. ALA synthase activity was maximal in the most immature erythroid cells and diminished as the cells matured with an overall five fold loss of activity from proerythroblast to late erythroblast development. Ferrochelatase activity was, however, more stable with less than a two fold change in activity observed during the same period of erythroid differentiation. Maximal activity occurred in erythroid fractions enriched with intermediate erythroblasts. These results support sequential rather than simultaneous appearance of these enzymes during normoblastic erythropoiesis. Quantitative analysis of relative enzyme activity however indicates that at all times during erythroid differentiation ferrochelatase activity is present in excess to that theoretically required relative to ALA synthase activity since ALA and haem are not produced in stoichiometric amounts. The lability of ALA synthase versus the stability and gross relative excess of ferrochelatase activity indicates a far greater role for ALA synthase in the regulation of erythroid haem biosynthesis than for ferrochelatase.