Reprinted from Trends in Pharmacological Sciences- October 1990
`
`Vol. 11, No. 10
`
`411
`
`Clinical significance of
`pharmacological modulation of
`homocysteine metabolism
`Helga Refsum and Per Magne Ueland
`
`The metabolic fate of homocysteine is linked to vitamin 8 12, reduced folates,
`vitamin B6 and sulfur amino acids. Clinical and experimental data suggest that
`elevated plasma homocysteine is an independent risk factor for premature
`vascular disease. This is particularly significant because plasma homocysteine
`levels are altered in several diseases, including folate and vitamin 8 12
`deficiencies, and because many commonly used drugs have now been shown to
`interfere with homocysteine metabolism. In summarizing the data, Helga
`Refsum and Per Ueland highlight the clinical implications for these metabolic
`changes.
`
`Homocysteine is a sulfur amino
`acid which is not itself incorpor(cid:173)
`ated into proteins, but is import(cid:173)
`ant as an
`intermediate
`in
`the
`metabolism of methionine and
`cysteine, and because its metab(cid:173)
`olism is linked to the function of
`some vitamins.
`In the late 1960s, inborn errors
`of
`homocysteine metabolism
`(homocystinuria) were demon(cid:173)
`strated in patients with mental
`retardation, skeletal abnormali(cid:173)
`ties, lens dislocation and prema(cid:173)
`ture vascular disease 1 . Research
`into the physiological and patho(cid:173)
`logical roles of homocysteine was
`subsequently promoted. Since the
`pioneering work of Wilcken and
`coworkers in 1976, accumulated
`epidemiological and experimental
`evidence has shown that homo(cid:173)
`cysteine may provoke vascular
`lesions, and that moderate homo(cid:173)
`cysteinemia
`is an
`independent
`risk factor for premature vascular
`disease 2
`During the last few years, im(cid:173)
`proved analytical techniques have
`allowed
`the
`investigation
`of
`plasma homocysteine in healthy
`subjects and
`in disease states
`other than homocystinwia. Folate
`and cobalamin (vitamin 8 12) de(cid:173)
`ficiencies cause very high plasma
`
`Helga Refsum is Research Fellow (NAVF) and
`Per MaR"e Ueland is Professor of Clinical
`t-lead of
`I he Clinical
`Pharmacology and
`Pharmacology Unit at Haukeland University
`Hospital, N-5021 Bergen, Norway.
`
`levels of homocysteine, and plasma
`homocysteine has been est a b(cid:173)
`lished as a sensitive and respon(cid:173)
`sive
`indicator of
`intracellular
`folate and cobalamin function3
`Some drugs influence homo(cid:173)
`cysteine metabolism and plasma
`levels. This may have some im(cid:173)
`portant implications. First, plasma
`homocysteine may reflect pharma(cid:173)
`codynamic effects of some drugs,
`as most clearly demonstrated with
`methotrexate and nitrous oxide
`(see below). Secondly,
`the in(cid:173)
`creased plasma levels induced by
`some agents may have
`impli(cid:173)
`cations
`for
`their
`side-effects.
`Finally, drugs decreasing plasma
`homocysteine may reduce the risk
`of vascular disease imposed by
`homocysteinemia3 .
`
`Homocysteine metabolism
`Homocysteine holds a unique
`position in metabolic regulation.
`Its metabolism is linked to sulfur
`ammo acids, reduced folates and
`vitamins 8 12 and B6 . Its metab(cid:173)
`olism is summarized in Fig. 1.
`The only source of honwcys(cid:173)
`teine in vertebrates is the hydroly(cid:173)
`sis of 5-adenosylhomocysteine, an
`inhibitor and product of 5-aden(cid:173)
`osylmethionine-dependent trans(cid:173)
`methylabon4. The fate of intra(cid:173)
`cellular homocysteine
`is either
`salvage
`to methionine
`through
`remethylation, or conversion to
`cysteine via the trans-sulfuration
`pathway.
`In most
`tissues,
`the
`former reaction is catalysed by the
`
`ubiquitous enzyme methionine
`synthase (Fig. 1). This enzyme re(cid:173)
`quires vitamin B12 [methyl(l)cobal(cid:173)
`amin] as a cofactor and 5-methyl(cid:173)
`tetrahydrofolate as methyl donor;
`thus 5-methyltetrahydrofolate enters
`the pool of reduced folates, and
`homocysteine is remethylated to
`methioninel.
`cobalamin(cid:173)
`two
`There
`are
`dependent enzymes in vertebrates:
`methionine synthase which util(cid:173)
`izes methylcob(I)alamin and
`is
`cytosolic,
`and methylmalonyl(cid:173)
`CoA mutase which is a mitochon(cid:173)
`drial enzyme containing aden(cid:173)
`osylcobalamin. A major fracbon of
`intracellular cobalamin is associ(cid:173)
`ated with these two enzymes5 .
`Homocysteine remethylation is
`also catalysed by an alternative
`enzyme,
`betaine-homocysteine
`methyltransferase, requiring be(cid:173)
`taine as methyl donor. However,
`this enzyme is generally confined
`to the liver.
`The metabolism of homocys(cid:173)
`teine along the trans-sulfuration
`pathway is catalysed by two vit(cid:173)
`amin 8 6-dependent enzymes. The
`first step
`is the cystathione ~­
`synthase reaction, where homo(cid:173)
`cysteine is condensed with serine
`to
`form
`cystathionine. Cysta(cid:173)
`thionine is then cleaved to ex-keto(cid:173)
`butyrate and cysteine, catalysed
`by cystathionine lyase 1
`
`Homocysteine and vascular
`disease
`Patients with homocystinuria
`suffer from premature vascular
`disease, localized to
`the central
`and peripheral arteries and large
`veins. This is the major cause of
`the high mortality (20-75% before
`the age of 30) in these patients.
`Thromboembolism may occur at
`any age and has even been de(cid:173)
`scribed in children 1. Both clinical
`and
`experimental
`evidence
`suggest that high homocysteine
`levels cause the vascular lesions
`(see Ref. 1).
`Even moderate homocystein(cid:173)
`emia may provoke venous throm(cid:173)
`bosis and premature vascular
`lesions in the cerebral, peripheral
`and coronary arteries (L. Bratt(cid:173)
`strom, Thesis, University of Lund,
`1989). Such a relation was suggested
`10 years ago from clinical studies
`based on measurement of acid(cid:173)
`soluble mixed disulfides in plasma
`from a small number of patients 2 .
`This has later been confirmed in
`several
`investigations, most of
`
`© 1990, Elsevier Science rublishers Lid. (UK) 0165- Gl47/'.lll/S02.DJ
`
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`
`412
`
`R
`
`Serine
`
`I. Homocysteine metabolism. AdoHcy, S-adenosylhomocysteine; Adomet,
`Fig.
`S-adenosylmethionine; 5,10-CH2 -THF, 5, 10-methylene THF: 5·CH3-THF, 5-methyl(cid:173)
`tetrahydrofolate; 5-CHO-THF, 5-formyltetrahydrofolate (folinate); CL, cystathionine
`lyase; CS, cystathionine {3-synthase; DHF, dihydrofolate; DR, dihydrofolate reductase;
`MET, methionine; MS, methionine synthase (5-methy/-THF homocysteine methy//rans(cid:173)
`ferase); SA, S-adenosylhomocysteine hydrolase; SH, serine hydroxymethyttransfer(cid:173)
`ase; THF, tetrahydrofofate.
`
`total plasma
`which determined
`homocysteine3•6· 10 . Furthermore,
`an increased incidence of hetero(cid:173)
`zygous homocystin uri a has been
`demonstrated
`in patients with
`early onset vascular disease7 .
`Other clinical and experimental
`data support an association be(cid:173)
`tween raised homocysteine levels
`and vascular lesions (reviewed in
`Ref. 3). Plasma homocysteine
`levels show age- and sex-depen(cid:173)
`dent variations resembling those
`described
`in
`arteriosclerotic
`disease. Men and postmenopausal
`women
`have higher
`plasma
`homocysteine levels during fast(cid:173)
`ing and after methionine loading
`than young women. Plasma
`homocysteine is significantly in(cid:173)
`creased in chronic renal failure,
`severe psoriasis, and
`in some
`patients with cancer. These are
`conditions associated with
`in(cid:173)
`creased risk of vascular disease
`not adequately explained by risk
`factors like smoking, lipid abnor(cid:173)
`malities, hypertension or other
`known predisposing conditions3 .
`Moderate homocysteinemia should
`be considered as a possible cause
`of vascular disease in those cases
`(15-30%) when other risk factors
`cannot be identified. Down syn(cid:173)
`drome, on the other hand, is an
`abnormality
`characterized
`by
`low plasma homocysteine levels,
`
`probably due to increased gene
`dosage for the enzyme cystathio(cid:173)
`nine
`~-synthase residing on
`chromosome 21 (Ref. 11). In 1977
`Murdoch and co-workers suggested
`this state as an atheroma-free
`model because of the remarkable
`absence of arteriosclerotic lesions
`observed in five patients aged 44-
`66 years12.
`also more direct
`There
`is
`evidence
`that high
`levels of
`homocysteine
`mediate
`the
`thrombogenesis and accelerated
`atherogenesis observed in homo(cid:173)
`cystinuria, and that homocysteine
`may also be responsible for the
`vascular disease associated with
`moderate homocysteinemia. Homo(cid:173)
`cysteine damages human endo(cid:173)
`thelial cells in culture, possibly by
`producing hydrogen peroxide in
`an oxygen-dependent
`reaction.
`Moreover, endothelial cells from
`patients heterozygous for cyst a(cid:173)
`thionine ~-synthase gene de(cid:173)
`ficiency may have an increased
`susceptibility to injury by homo(cid:173)
`cysteine. Mechanisms linking mild
`homocysteinemia and vascular
`effects could also involve produc(cid:173)
`tion of free radicals and oxidation
`of low-density lipoprotein. Con(cid:173)
`flicting data exist on possible roles
`of platelet sequestration in
`the
`development of atherosclerotic
`lesions
`under
`conditions
`of
`
`TiPS- October 1990 [Vol. 11}
`
`elevated plasma homocys(cid:173)
`teine levels2•
`
`Folate and cobalamin
`deficiency
`Cobalamin deficiency may
`increase plasma homocys(cid:173)
`teine to levels observed in
`homocystinurics (high mino(cid:173)
`molar range); there is a nega(cid:173)
`tive
`correlation
`between
`serum cobalamin and total
`plasma homocysteine, Homo(cid:173)
`cysteine levels may also be
`elevated
`in
`cobalamin(cid:173)
`deficient patients devoid of
`like anemia,
`typical signs
`macrocytosis and
`reduced
`serum cobalamin. Levels are
`normalized following cobal(cid:173)
`amin therapy3 ,13 .
`Similarly, folate deficiency
`is a common cause of el(cid:173)
`evated plasma homocysteine
`levels, and a close negative
`correlation with serum folate
`has
`been
`demonstrated;
`again, folate therapy normal(cid:173)
`izes levels. Moderate elevation
`of plasma homocysteine is
`also observed in subjects with low
`but normal serum folate levels,
`suggesting that increased homo(cid:173)
`cysteine levels in these subjects
`mav reflect an intracellular folate
`for optimal
`con'tent insufficient
`folate-dependent remethylation of
`homocysteine3 .
`Measurement of plasma homo(cid:173)
`cysteine is therefore a promising
`laboratory
`test
`for
`evaluating
`cobalamin or
`folate deficiency
`states. It may be particularly use(cid:173)
`ful when used in conjunction with
`serum methylmalonic acid, which
`is a speci fie measure of disturb(cid:173)
`ances of cobalamin metabolism 13 .
`
`Agents decreasing homocysteine
`concentrations
`serving as co(cid:173)
`Compounds
`factors
`in homocysteine catab(cid:173)
`olism or remethylation may en(cid:173)
`hance homocysteine metabolism
`and thereby reduce plasma homo(cid:173)
`cysteine levels in inherited en(cid:173)
`zymic defects. Thus, vitamin 66
`reduces plasma homocysteine in
`homocystinurics with
`residual
`cystathionine P-synthase activity,
`and vitamin B12 acts similarly in
`some mutations of cobalamin
`metabolism. Betaine and folic acid
`have been shown to efficiently
`reduce plasma homocysteine in
`patients with cystathionine P-syn-
`
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`TiPS- October 1990 [Vol. 11]
`
`thase deficiency who are un(cid:173)
`responsive to vitamin 8 6 (Ref. 1).
`
`Folic acid
`Folic acid (5 mg daily) efficiently
`decreases plasma homocysteine
`levels. lt reduces elevated plasma
`homocysteine in renal transplant
`recipients and even in those with(cid:173)
`out overt folate deficiency. Treat(cid:173)
`ment of healthy subjects with folic
`acid for 14 days significantly re(cid:173)
`duced
`plasma
`homocysteine,
`especially in persons with high
`pretreatment leveJsH.
`The marked effect of high doses
`of folic acid on the concentration
`of homocysteine in plasma is im(cid:173)
`portant. Since moderate homo(cid:173)
`cysteinemia may provoke vascular
`lesions, folic acid may prevent
`atherosclerotic disease in selected
`subjects. This
`intervention
`is
`particularly
`attractive
`because
`folic acid intake has essentially no
`side-effects 14 •
`Interestingly,
`the
`effect of folic acid suggests that
`the
`intracellular folate content
`is
`insufficient for an optimal
`remethylation of homocysteine.
`This may be a more common state
`than hitherto recognized, as
`it
`may not be detected by estab(cid:173)
`lished
`laboratory
`procedures,
`including determination of folate
`in serum or erythrocytes3 .
`Folic acid probably decreases
`plasma homocysteine levels by in(cid:173)
`creasing the availability of intra(cid:173)
`cellular 5-met.hyltetrahydrofolate,
`thereby enhancing homocysteine
`remethylation (Fig. 2). This re(cid:173)
`action
`forms
`tetrahydrofolate,
`which enters the pools of reduced
`folates carrying one-carbon units
`via
`the serine hydroxymethyl
`transferase reaction (Figs 1 and 2).
`In this reaction, serine is con(cid:173)
`sumed and glycine formed, as
`would be expected
`from
`the
`moderate reduction
`in plasma
`serine
`levels and
`increase
`in
`plasma glycine levels that follow
`folic acid administration 14.
`
`v-Penicillamine
`D-Penicillamine {o-~,~-dimethyl­
`cysteine) is currently used for the
`treatment of heavy metal poison(cid:173)
`ing, rheumatoid arthritis, hepato(cid:173)
`lenticular degeneration, cystinuria
`and scleroderma. It is metabolically
`stable, chelates heavy metals, pro(cid:173)
`duces disulfides and forms a thi(cid:173)
`azolidine ring with aldehydes and
`ketones. In plasma, penicillamine
`forms symmetrical penicillamine
`
`disulfides and mixed disulfides
`with cysteine, homocysteine and
`plasma proteins. The low mol(cid:173)
`ecular weight disulfides undergo
`rapid renal excretion, which ex(cid:173)
`plains the short plasma half-life
`and
`the
`therapeutic effect
`in
`cystinuria15 •
`re(cid:173)
`efficiently
`Penicillamine
`duces both
`free and protein(cid:173)
`bound plasma homocysteine in
`homocystinurics 16
`and
`total
`plasma homocysteine in patients
`with rheumatoid arthritis, a con(cid:173)
`dition with normal homocysteine
`pretreatment
`levels17 . This
`re(cid:173)
`duction in plasma levels may be
`associated with an intracellular
`homocysteine
`depletion
`suf(cid:173)
`ficiently pronounced to decrease
`homocysteine remethylation and
`thereby induce methionine de(cid:173)
`ficiency and secondary effects on
`folate metabolism. Jf such effects
`occur, penicillamine may act as an
`antifolate agent and may therefore
`interact adversely with metho(cid:173)
`trexate used in low doses in the
`management of rheumatoid ar(cid:173)
`thritis (see below)l 8 .
`A further clinical implication of
`this work is that penicillamine
`may be a useful means to reduce
`plasma homocysteine.
`
`Adenosine (nucleoside) analogs
`The cleavage of 5-adenosyl(cid:173)
`homocysteine to adenosine and
`homocysteine, catalysed by 5-
`adenosylhomocysteine hydrolase
`(Figs 1 and 2), is the only known
`source of homocysteine in ver(cid:173)
`tebrates. Several nucleoside ana(cid:173)
`logs block this reaction by serving
`either as an inactivator or in(cid:173)
`hibitor of the enzyme. In addition,
`some analogs act as substrate and
`are thus converted to the corre(cid:173)
`sponding S-adenosylhomocysteine
`analogue4 •
`Inhibition
`of
`5-
`adenosylhomocysteine hydrolase
`leads to massive accumulation of
`S-adenosylhomocysteine
`in
`iso(cid:173)
`lated cells, whole animals, and in
`patients. This is important for the
`antiviral effects of this class of
`compound4, 19 .
`immediate conse(cid:173)
`The other
`quence of S-adenosylhomocys(cid:173)
`teine hydrolase
`inhibition, re(cid:173)
`duction of homocysteine
`for(cid:173)
`mation (Fig. 2), has been studied
`only
`recently.
`Homocysteine
`depletion and inhibition of homo(cid:173)
`cysteine export have been demon(cid:173)
`strated in isolated cells exposed to
`nucleoside analogs 20,21 . A reduc-
`
`413
`
`tion in plasma homocysteine was
`found
`in patients with acute
`leukemia treated with 2-deoxy(cid:173)
`coformycin, which indirectly in(cid:173)
`activates S-adenosylhomocysteine
`hydrolase22.23.
`Inhibition
`of
`homocysteine formation plays an
`important role in the cytostatic
`action of some nucleoside analogs
`against some 24,25 but not alJ2~>-2s
`cell types, and probably mediates
`the differentiation of HL-60 cells
`induced by adenosine dialde(cid:173)
`hyde29. The consequences of cellular
`homocysteine deficiency are two(cid:173)
`fold. First, nucleoside analogs may
`induce severe methionine de(cid:173)
`ficiency, since homocysteine sal(cid:173)
`vage is a significant source of
`methionine in humans30·31 . Sec(cid:173)
`ondly, lack of homocysteine may
`trap reduced folate as 5-methyl(cid:173)
`tetrahydrofolate because homo(cid:173)
`cysteine is the methyl acceptor in
`the methionine synthase reaction
`catalysing
`the
`conversion
`of
`5-methyltetrahydrofolate to tetra(cid:173)
`hydrofolate. In this way, lack of
`tetrahydrofolate may ensue, and
`thereby inhibit folate-dependent
`purine and thymidylate synthesis.
`Both mechanisms have been
`demonstrated in cultured cells24.32.
`Induced deficiencies of meth(cid:173)
`ionine and
`folates might be
`avoided clinically by appropriate
`supplementation which would be
`expected
`to
`reduce associated
`side-effects. Moreover, adverse
`interactions of adenosine ana(cid:173)
`logues with drugs that also inter(cid:173)
`fere with folate or methionine
`metabolism such as nitrous oxide
`or methotrexate should be con(cid:173)
`sidered. This has been suggested
`for the antiviral agent vidarabine
`(9-~-o-ara binofuranosyladeni ne)
`and high-dose methotrexate with
`folinic acid rescue3°. Reduction in
`plasma homocysteine levels might
`however have beneficial vascular
`effects.
`
`Inhibitors of homocysteine
`remethylation and degradation
`Several important drugs interfere
`with homocysteine metabolism.
`
`Nitrous oxide
`The anesthetic agent nitrous
`oxide was used for a century be(cid:173)
`fore it was discovered that long(cid:173)
`term exposure caused megalo(cid:173)
`blastic and aplastic bone marrow
`changes, anemia and myelopathy.
`
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`
`414
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`TiPS- October 1990 [Vol. 11]
`
`Methorrex+ate
`DHF
`.
`
`,. /
`
`TI-IF
`
`Adenosine analogues
`
`Folic acid
`
`' /
`AdoH~·lOC~:: _::;
`. )""
`l ~ THF
`Ado ~
`Hey
`Met
`B12
`~H3 cob(l)alamin
`Nitrous oxide
`
`~Folic acid
`/
`
`5- CH -THF
`3
`~HF
`B12
`
`Hey
`
`~et
`
`DHF -f!ID:~~~•IIIE"· --1110 ... THF
`
`Folic acid
`
`"
`
`6
`" '
`Cystathionine
`B 6~
`Hcy-SH
`~ (extracellular)
`Cysteine
`
`R-S-S-R i
`
`Ado He~
`)(
`
`Ado
`
`H cy
`
`6-Azauridine triacetate
`
`~Hey
`/8
`\
`)tD
`
`Cystathionine
`
`Cysteine
`
`Hcy-S-S-R
`
`D-Peniciltamine
`(P-SH)
`
`~ Hey-S-S-P
`
`5- CJ-13 -THF
`• •. )
`',,,~HF·
`Hey Cll 3wb(ll)alamin
`inactive B 12
`
`Me<
`
`P-S-S-R -1 ~
`R-S-5-R i
`
`Hcy-SH
`(extracellular)
`
`Renal
`excretion
`
`Fig. 2. Proposed or established mechan(cid:173)
`isms for 111e effect of various agents on
`the metabolism and plasma level of
`homocysteine. The figure in the center
`indicates normal metabolism and tile six
`panels in the boxes show the effects of
`various drugs on different pathways.
`Sizes of type and arrows indicate the
`concentration of a particular metabolite
`and the flux through the pathway, re-
`spectively Folic acid probably increases
`the cellular content of 5-methyltetrahydrofolate (5-CH 3-THF), which increases the homocysteine (Hey) remethylation catalysed by
`methionine synthase (MS). a-Penicillamine (P-SH) forms a mixed disulfide with homocysteine (Hey-S-S-P); this disulfide has a high renal
`clearance. Adenosine analogs are inhibitors of S-adenosylhomocysteine hydrolase (SA). Nitrous oxide oxidizes methylcob(l)alamin Ia
`methytcob{ll)alamin and thereby Irreversibly inhibits methtonine synthase (MS). Methotrexate inhibits dihydrotolate reductase (DR),
`thereby inhibiting regeneration of tetrahydrofolate (THF) from dihydrofolate (DHF). Tetrahydrofolate supplies 5-methyl/elra!Jydrofolate via
`5, 10-methytenetetrahydrofolate (5,10-CH2-THF). Methotrexate may therefore induce depletion of 5-methylletrahydrofolate, and inhibit
`homocysteine remethylation catalysed by methionine synthase. Azauridine is a vitamin 86 antagonist, and may inhibit some vitamin 8 6 -
`dependent enzymes, including cystathionine {3-synthase (CS).
`Hey, homocysteine; Met, methionine; Ado, adenosine; P·SH, a-penicillamine reduced form; Hcy-S-S-R, homocysteine mixed disulfide;
`Hey-S-S-P, a mixed disulfide between homocysteine and a-penicillamine; P-S-5-R, a-penicillamine mixed disulfide.
`For other abbreviations, see Fig. 1.
`
`Hcy-5-S-R
`
`These side-effects are similar to
`the symptoms of vitamin 8 12
`deficiency33 .
`Nitrous oxide oxidizes the cob(cid:173)
`alamine
`species methylcob(I)(cid:173)
`alamine, inactivating specifically
`the enzyme methionine synth<lse,
`
`without affecting the adenosyl(cid:173)
`cobalamine cofactor of methyl(cid:173)
`malonyl-CoA mutase33 .
`Inacti(cid:173)
`vation of methionine synthase
`causes a cascade effect on folate
`metabolism including trapping of
`reduced folates such as 5-methyl-
`
`tetrahydrofolate, Joss of folate in
`the urine, and reduction in tissue
`is a sub(cid:173)
`levels. There
`folate
`sequent decrease of folate-depen(cid:173)
`dent purine and
`thymidylate
`synthesis, as demonstrated by the
`deoxyuridine suppression
`(dU)
`
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`TiPS- October 1990 [Vol. 11 J
`
`in
`test, which reveals changes
`human bone marrow within 5-6
`hours. In humans, 50% of meth(cid:173)
`ionine synthase is inactivated in
`about two hours, and there is a
`reduction in plasma methionine
`after 8-24 hours of exposure.
`Megaloblastic
`bone marrow
`changes can be detected after 12-
`24 hours of exposure33.
`Data on the effect of nitrous
`oxide on homocysteine metab(cid:173)
`olism are sparse, but this drug has
`been reported
`to
`increase
`the
`urinary excretion of homocysteine
`in sheep, and increase plasma
`homocysteine levels in fruit bats3 .
`We have recently demonstrated
`that nitrous oxide
`induces a
`marked increase in plasma homo(cid:173)
`cysteine within 90 minutes, with a
`concurrent
`increase
`in urinary
`homocysteine excretion. In patients
`receiving nitrous oxide for 3-6
`hours, plasma homocysteine re(cid:173)
`mained above normal for at least 7
`days (Ermens, A. A. M. et al., un(cid:173)
`published data). The homocysteine
`response evolved before other
`early signs of nitrous oxide(cid:173)
`induced cobalamine inactivation 33.
`Thus, significant cobalamine oxi(cid:173)
`dation and methionine synthase
`inact!·.ration occurs even after
`short-term exposure, which pre(cid:173)
`viously has not been regarded as
`harmful. The resultant increase in
`plasma homocysteine observed in
`some patients may in itself be
`detrimental. Furthermore,
`the
`homocysteine response may re(cid:173)
`flect loss of functional cobalamine
`and folate, and may enhance sen(cid:173)
`sitivity towards antifolate drugs
`such as methotrexate34 (Ermens,
`A. A. M., PhD Thesis, University
`of Rotterdam, 1990). Since an in(cid:173)
`crease in plasma homocysteine
`levels is both an early and sensi(cid:173)
`tive measure of cobalamine oxi(cid:173)
`dation,
`plasma
`homocysteine
`monitoring could be useful in the
`detection of such effects in the
`clinic.
`
`Methotrexate
`is an antifolate
`Methotrexate
`drug which has been used exten(cid:173)
`sively in intermediate and high
`doses in the treatment of leukemia
`and some solid tumors. Low-dose
`methotrexate
`is used
`in
`the
`management of some non-malig(cid:173)
`nant diseases such as rheumatoid
`arthritis and psoriasis.
`Methotrexate acts by inhibiting
`dihydrofolate reductase, thereby
`
`blocking the regeneration of tetra(cid:173)
`hydrofolate
`from dihydrofolate
`(see Ref. 35). This leads to deple(cid:173)
`tion of reduced folates, including
`5-methylteh·ahydrofolate36,37 . Thus,
`methotrexate may also inhibit the
`folate-dependent remethylation of
`homocysteine catalysed by methio(cid:173)
`nine synthase. This would explain
`the increased homocysteine export
`from cultured cells exposed
`to
`methotrexate, and the methotrex(cid:173)
`ate-induced homocysteinemia and
`urinary homocysteine excretion in
`patients38.
`Low-dose methotrexate (25 mg
`daily) given to psoriatics induced
`increased plasma homocysteine
`levels, which maximized after
`about two days, and normalized
`within one week39 . This shows
`that plasma homocysteine is a
`sensitive measure of the antifolate
`effect.
`(1-13.6 g)
`Intermediate doses
`given
`to patients with
`solid
`tumors, induced a rapid increase
`in plasma homocysteine within
`hours, which was reversed on
`administration of folinic acid 24
`hours after start of infusion. This
`response was observed following
`several methotrexate doses in a
`single patient40 . High doses of
`methotrexate (8-33.6 m2) given to
`children gave a similar response,
`i.e. a rapid increase a few hours
`after start of administration and
`a
`decline
`following
`'rescue'
`therapy 41 • This is analogous to the
`results obtained with cultured
`ce!ls38
`in
`levels
`Basal homocysteine
`patients with acute lymphoblastic
`leukemia were often above normal
`before
`treatment, and declined
`markedly
`following
`treatment
`with cytotoxic agents including
`methotrexate (Refsum, H. et al.,
`unpublished). This may be due to
`eradication of proliferating cells
`exporting large amounts of homo(cid:173)
`cysteine.
`The high-dose methotrexate
`regimen also induced a transient
`but marked reduction in plasma
`methionine41 which may contrib(cid:173)
`ute to the killing of tumor cells or
`toxicity of methotrexate. Plasma
`homocysteine
`response
`and
`methionine depletion may corre(cid:173)
`late with the therapeutic as well as
`the side-effects of methotrexate;
`including liver toxicity42 and an
`increased incidence of thrombo(cid:173)
`embolism43; plasma homocysteine
`measurements could provide a
`
`415
`
`useful adjunct to serum metho(cid:173)
`trexate determination
`in
`the
`management
`of methotrexate
`therapy.
`
`Vitamin B6 antagonists
`Azauridine is an antimetabolite
`interfering with de novo synthesis
`of uridine-5'-monophosphate. 1t
`was initially used for the treat(cid:173)
`ment of refractory psoriasis, but
`was withdrawn by the FDA in
`1976 because its use was associ(cid:173)
`ated with an increased incidence
`of vascular episodes44 . This may
`be due to effects on homocysteine
`metabolism. Azauridine causes
`homocysteinemia, abnormal homo(cid:173)
`cysteine excretion and a signifi(cid:173)
`cant increase in serum methionine
`levels in patients. Studies in rab(cid:173)
`bits suggest that it functions as a
`pyridoxa!S' -phosphate antagonist
`and causes homocysteinemia by
`inhibiting vitamin B6-dependent
`cystathionine
`synthesis44 . This
`suggests
`that
`supplementing
`vitamin B6 would prevent the in(cid:173)
`hibition of homocysteine catab(cid:173)
`olism; determination of plasma
`homocysteine may
`identify
`patients at risk of vascular epi(cid:173)
`sodes.
`Several other drugs also inter(cid:173)
`fere with the function of vitamin
`isoniazid, cycloserine, hy(cid:173)
`8 6 :
`dralazine, penicillamine, phenel(cid:173)
`zine and procarbazine44 . Pertur(cid:173)
`bation of homocysteine metab(cid:173)
`olism in patients has been demon(cid:173)
`strated with isoniazid45 • In one
`out of six patients given 300 mg
`isoniazid daily for one month,
`urinary homocysteine excretion
`was fivefold higher than normal.
`Inhibition
`of
`cystathionine
`metabolism in these patients is
`supported by increased excretion
`of
`this compound after meth(cid:173)
`ionine loading45.
`
`Other agents
`Premenopausal women have
`lower plasma homocysteine than
`men
`and
`postmenopausal
`women46 , and plasma levels are
`low during pregnancy. However
`there is no conclusive evidence
`that homocysteine metabolism
`and plasma homocysteine levels
`are under the influence of estro(cid:173)
`gens. Preliminary data in women
`given contraceptive steroids or the
`antiestrogen tamoxifen suggest a
`polymorphic response. In some
`women, altered estrogen status
`may cause a decrease, and in
`
`NEPTUNE GENERICS 1012- 00005
`APOTEX 1012- 0005
`
`

`
`416
`
`in plasma
`increase
`others, an
`homocysteine levels. Because of
`the widespread use of contracep(cid:173)
`tives, and the suggested use of
`tamoxifen as prophylactic inter(cid:173)
`vention in healthy women at high
`risk of developing breast cancer,
`the effect on plasma homocysteine
`from altered estrogen status is a
`question of major concern
`to
`public heaJth3
`Various
`antiepileptic drugs,
`particularly phenytoin but also
`phenobarbital, primidone, carba(cid:173)
`mazepine, and valproic acid, may
`induce folate deficiency47 • The
`activity of methylenetetrahydro(cid:173)
`folate
`reductase,
`the
`enzyme
`producing
`5-methyltetrahydro(cid:173)
`folate, is altered in mouse liver
`following exposure to these drugs.
`Preliminary data suggest
`that
`phenytoin and possibly carbama(cid:173)
`zepine may
`increase plasma
`homocysteine, but its relation to
`overt folate deficiency has not
`been established3.
`Several other drugs are known
`to
`interfere with folate metab(cid:173)
`olism or function. These include
`some phenothiazines and tricyclic
`antidepressants, oral contracep(cid:173)
`tives, possibly some tuberculo(cid:173)
`static drugs, and antifolate drugs
`such as trimethoprim47 . Thiols or
`disulfide forming drugs, such as
`cysteamine and N-acetylcysteine
`are actual candidates as modifiers
`of plasma homocysteine levels,
`due to a possibly unique dis(cid:173)
`position of the mixed disulfides
`with homocysteine, as demon(cid:173)
`strated with penicillamine
`in
`humans 16.
`
`0
`
`0
`
`0
`
`Various pharmacological agents
`have been shown
`to enhance
`homocysteine remethylation and
`urinary
`excretion, or
`inhibit
`homocysteine
`production,
`re(cid:173)
`methylation or catabolism thereby
`affecting plasma homocysteine
`levels. Modulation of homocys(cid:173)
`teine metabolism and plasma
`concentrations may be an import(cid:173)
`ant component of drug action.
`
`Agents that reduce plasma homo(cid:173)
`cysteine (e.g. folic acid, penicill(cid:173)
`amine) may prevent vascular
`disease, while agents increasing
`plasma homocysteine (e.g. nitrous
`oxide, methotrexate, azauridine)
`may provoke vascular episodes.
`Also for some drugs (e.g. nitrous
`oxide and methotrexate), elevation
`is an
`of plasma homocysteine
`early and sensitive measure of
`drug action and plasma homocys(cid:173)
`teine has been shown to be use(cid:173)
`ful in the diagnosis and follow-up
`of some diseases, in particular
`homocystinuria, folate deficiency
`and cobalamine deficiency.
`
`(1989) Haemostasis 19
`
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