Review article: monitoring of immunomodulators in inflammatory bowel disease

Authors

  • F. N. Aberra,

    1. Department of Medicine, Center for Inflammatory Bowel Disease, Hospital of the University of Pennsylvania, University of Pennsylvania School of Medicine, Division of Gastroenterology, 3400 Spruce Street, 3rd floor Ravdin Building, Philadelphia, PA, USA
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  • G. R. Lichtenstein

    1. Department of Medicine, Center for Inflammatory Bowel Disease, Hospital of the University of Pennsylvania, University of Pennsylvania School of Medicine, Division of Gastroenterology, 3400 Spruce Street, 3rd floor Ravdin Building, Philadelphia, PA, USA
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Dr G. R. Lichtenstein, Department of Medicine, Center for Inflammatory Bowel Disease, Hospital of the University of Pennsylvania, University of Pennsylvania School of Medicine, Division of Gastroenterology, 3400 Spruce Street, 3rd floor Ravdin Building, Philadelphia, PA 19104-4283, USA.
E-mail: grl@uphs.upenn.edu

Summary

The armamentarium of medications for the treatment of inflammatory bowel disease is growing and becoming more complicated to use. Immunomodulators are a class of medications that have found a niche for the treatment of Crohn's disease and ulcerative colitis. Because of the mounting supporting evidence for efficacy, the most commonly-used immunomodulators are azathioprine, mercaptopurine, methotrexate and ciclosporin. These medications are being used more often due to their steroid-sparing and potentially surgery-sparing effects. Immunomodulators are also known for a significant side-effect profile and require careful monitoring. This review provides the latest information for clinicians on efficacy, side-effects, dosing and monitoring of these medications for treatment of inflammatory bowel disease.

Introduction

Treatment of inflammatory bowel diseases (IBD) commonly requires utilization of immunodulatory medications. This class of medications includes azathioprine (AZA), mercaptopurine (MP), ciclosporin, methotrexate, and on occasion mycophenolate mofetil and tacrolimus. The most commonly used immunomodulators for treatment of IBD are AZA, MP, ciclosporin and methotrexate. These medications have shown to be effective and great care must be taken in monitoring patients receiving these medications to maximize clinical benefit and reduce the risk of side-effects and toxicity. This review will provide information on clinical pharmacology, clinical indications for use, methods of dose adjusting, monitoring of metabolites for efficacy and for potential side-effects, and the adverse event profile for each medication. This review will hopefully provide guidance on utilizing immunomodulators for Crohn's disease (CD) and ulcerative colitis (UC) based on the published literature.

Methods

A Medline search was conducted from 1966 to 2004, for randomized-controlled trials of AZA, MP, ciclosporin and methotrexate for the treatment of IBD. A Medline search was also completed for studies assessing monitoring efficacy and toxicity of these medications in the setting of IBD. Additional references were obtained and reviewed from a search in BIOSIS Previews, a database, which includes published abstracts, and if pertinent literature was known to the authors or identified during searches. All searches were limited to the English language.

Mercaptopurine and azathioprine

Clinical pharmacology

Mercaptopurine and the nitroimidazole derivative of MP, AZA, are thiopurine analogues. AZA is non-enzymatically metabolized to MP. MP is metabolized to 6-thioinosine 5′-monophosphate (TIMP) by the enzyme hypoxanthine phosphoribyl transferase (HPRT). TIMP is eventually metabolized to 6-tioguanine (thioguanine) nucleotides (6-TGN) (see Figure 1). 6-TGN has been shown to bind to Rac1 instead of guanine triphosphate and with co-stimulation with CD28 leads to inhibition of Rac1 and apoptosis of T-lymphocytes.1 MP is also metabolized to 6-methylmercaptopurine (6-MMP) by the enzyme thiopurine methyltransferase (TPMT) and 6-thiouric acid by the enzyme xanthine oxidase (XO). Both 6-thiouric acid and 6-MMP are inactive metabolites of MP. The three enzymes metabolizing MP are in constant competition for substrate and the concentration of the metabolites of MP are based on the concentrations of these enzymes. About 84% of MP is quickly metabolized by XO found in high concentrations in enterocytes and hepatocytes, leaving only 16% left to be catabolized by TPMT and HPRT.2

Figure 1.

Metabolism of azathioprine (AZA) and mercaptopurine (MP). 6-TU, 6-thiouric acid; 6-MMP, 6-methyl mercaptopurine; 6-MMPR, 6-methyl mercaptopurine ribonucleotides; 6-TIMP, 6-thioinosine 5′-monophosphate; 6-TXMP, 6-thioxanthosine 5′-monophosphate; 6-TGN, 6-tioguanine nucleotides; XO, xanthine oxidase; TPMT, thiopurine methyltransferase; HPRT, hypoxanthine phosphoribosyltransferase; IMPDH, inosine monophosphate dehydrogenase; GMPS, guanosine monophosphate synthetase.

Genetic variation of metabolism.  Mutations of the gene encoding TPMT leading to varying functional activity of the enzyme have been identified in the population. There are at least 12 mutant alleles responsible for TPMT deficiency and several silent and intronic mutations have been described for the TPMT gene located on chromosome 6.3, 4 The most common of these variants is the normal TPMT*1 or wild-type allele. TPMT*3A is the most common mutant allele and seen predominantly in Caucasians. TPMT*3C is the most prevalent variant in African-American and Asian populations.5–10 Homozygous mutations of the TPMT gene produce enzyme with minimal activity and accounts for 0.3% of the general population.11, 12 Heterozygous mutations yield enzyme with moderate activity and accounts for 11% of the general population.11 A recent study in patients with CD has shown the frequency of homozygous and heterozygous TPMT mutations parallels the frequency in the general population.12 TPMT level is measured from lysed erythrocytes and the TPMT level corresponds to that in liver, kidney and lymphocytes.13 TPMT activity appears to be inversely correlated with drug response. TPMT genotyping is also available but limited to the allelic variants that are known. More detail is given utilizing TPMT activity for tailoring treatment with AZA or MP in the following sections.

Indication and dosing

There have been several clinical trials supporting the use of AZA and/or MP for the treatment of the following indications: active CD, maintenance of remission of CD, fistulizing CD, prevention of post-operative recurrence of CD, maintenance of remission of UC, and as a steroid-sparing agent. In a meta-analysis of randomized-controlled trials from 1966 to 1994 of AZA and MP in active and quiescent CD the odds ratio (OR) for a clinical response to therapy of active CD was 3.09 (95% CI: 2.45–3.91), 1.45 (95% CI: 1.12–1.87) without the one clinical trial with MP.14 For quiescent CD the OR for maintaining remission if on AZA was 2.27 (95% CI: 1.76–2.93). Both longer duration and higher dose of AZA/MP was associated with improved response and a steroid-sparing effect was seen in subjects with active and quiescent disease. Several studies have also shown a steroid dose reduction with the addition of MP and AZA to therapy.15–22 MP and AZA have also been found to be useful in the treatment of fistulizing CD with several studies showing a higher response of complete closure or decreased drainage of fistulas.17, 21, 23 There is also emerging data supporting MP and AZA use for CD post-operative prevention of recurrence of in those with previous history of surgeries and perforating disease.24–28 In UC, AZA and MP appear to be most useful in maintaining remission and as a steroid-sparing agent.29

The mercaptopurine and AZA have a significantly delayed onset of action with several studies showing clinical benefit after 2–3 months of treatment.17, 21 The best approach if rapid clinical response is needed is to start another form of therapy such as corticosteroids or infliximab until the period of clinical onset is achieved. It is not yet clear what the ideal length of therapy for maintenance of remission should be. A study by Bouhnik et al. showed a low relapse rate of <20% in patients in remission who used AZA for 4 years or more, 2 years after completing therapy. The relapse rate was similar to those that continued AZA therapy.30 In a recent study published in abstract form by Lemann et al., 19% relapsed at 18 months and 60% relapsed at 54 month after AZA was stopped for maintenance of remission of CD.31

For CD and UC the most effective doses appear to be 2.5 mg/kg of AZA and 1.5 mg/kg of MP, although there has not yet been a head-to-head comparison at various dose levels, or a comparative trial evaluating efficacy of MP vs. AZA in patients with IBD. There are two methods for starting therapy, dose escalating to the weight-based dose vs. starting immediately at the weight-calculated dose. Dose escalating came into practice due to the fear of dose-related toxicities such as leucopoenia, thrombocytopenia and hepatitis. Whether leucopoenia or other dose-related side-effects are less severe with a gradual escalation algorithm compared with starting at the weight-calculated dose has yet to be determined. Neither method prevents toxicity. In a study by Connell et al., the range of time from starting AZA to onset of leucopoenia was 2–11 years with a median of 9 months.32 Whereas in a study by Present et al. most of the cases of leucopoenia occurred within 1 month of starting MP.33 The dose escalating method chosen is arbitrary, but a common practice is to start at 50 mg everyday and increase the dose by 25 mg every 1–2 weeks and monitor for leucopoenia and other potential adverse events.

Monitoring

Use of metabolites: 6-TGN and 6-MMP. The metabolites 6-TGN and 6-MMP have been used to determine likelihood of therapeutic response. See Table 1 for metabolite levels and the associated response and/or toxicity. Non-responders to MP and AZA can be categorized into three groups based on levels of the metabolites 6-TGN and 6-MMP. (i) Low levels of both metabolites are likely a consequence of under dosing or non-compliance. (ii) Low 6-TGN and high 6-MMP may signify increased likelihood for hepatotoxicity and also suggest metabolic resistance to MP/AZA, shunting away from 6-TGN production.34 Early recognition of this pattern may lead to early replacement of AZA with another medication. (iii) A normal 6-TGN level likely represents refractoriness to AZA or MP.

Table 1.  Metabolite levels and possible clinical correlations*
6-TGN level6-MMP levelClinical response
  1. * Low 6-TGN is defined as <235 pmol/8 × 108 and high >450 pmol/8 × 108. High 6-MMP is defined as >5700 pmol/8 × 108.

  2. MMP, 6-methylmercaptopurine; MP, mercaptopurine.

LowLowSubtherapeutic dosing or non-compliance
LowHighMP resistant
NormalHighMP resistant or high risk for abnormal liver chemistries
NormalLowResponders or refractory
HighLowTrue refractory, responder or risk of leucopoenia
HighHighOver-dosed

The initial studies of metabolite levels and associated clinical response were studied in paediatric IBD patients predominately with CD with various disease severities, type of disease and disease distribution. The application of measuring metabolite levels in UC studies has not been as well evaluated. Discriminant levels of 235–250 pmol/8 × 108 have been considered predictive of response. Yet, several studies have shown subjects that were in remission receiving MP or AZA with 6-TGN levels lower than 235–250.35–39Table 2 provides the sensitivity, specificity and positive predictive value of using discriminant levels (>230, 235 and 250 pmol/8 × 108) to predict clinical remission based on data from published studies. The positive predictive value for remission of 6-TGN level >230–250 are poor to fair at best ranging from 24 to 83%. Overall, therapeutic drug monitoring with measuring 6-TGN levels in patients treated with AZA or MP may be most useful and can be considered in the following selected settings: patients suspected of non-compliance and possibly patients who are failing to respond to standard doses of drug. The latter has yet to be proven in a prospective controlled fashion. There is now an ongoing trial comparing the benefit of dosing based upon 6-TG levels vs. weight-based dosing.

Table 2.  Utility of 6-TGN level for determining remission of inflammatory bowel disease
Study and 6-TGN levelTotal number of subject (n)IBD populationSensitivity (%)Specificity (%)Positive predictive value (%)Negative predictive value (%)
  1. * pmol/8 × 108

  2. † Studies may have multiple measurements of 6-TGN.

Belaiche et al.36
 >250*28Adult CD26888336
 >23032787535
Cuffari et al.38
 >25082Adult CD, UC69868670
Dubinsky et al.39
 >23593Paediatric CD, UC65737060
Goldenberg et al.116
 >23574Adult CD, UC47822482
Gupta et al.117
 >235101Paediatric CD, UC54706467
Mardini and Arnold118
 >23044Adult CD, UC1006165100
Su et al.11929Adult CD, UC55734183
>230Subgroups    
 Luminal CD86806084
 Fistulizing CD33661783
 UC40713679
Cuffari earlier study12010Paediatric CD80838083

TPMT.  The TPMT enzyme activity and genotype are tests that are available on a commercial basis for testing. An increasing number of TPMT mutations leading to reduced enzyme are being identified. Normal levels of enzyme activity appear to be associated with more response than high levels of TPMT enzyme activity but both groups have a widely variable clinical response.40, 41 Low enzyme levels can lead to development of severe myelosuppression in the setting of MP/AZA therapy.42, 43

The relationship of leucopoenia and various TPMT genotypes in the setting of AZA or MP treatment has been the focus of several research studies. In a study by Colombel et al. of 75 patients whom developed leucopoenia while treated with AZA or MP, 10% of patients were homozygous for TPMT mutant alleles and 17% were heterozygous for TPMT mutant alleles.44 The median time to leucopoenia was significantly shorter in those with homozygous TMPT mutations, 1 month (range: 0–1.5) compared with 4 months (range: 1–18) in those with one mutant allele and 3 months (range: 0.5–87) in those with normal TPMT.44 It is not yet clear whether patients heterozygous for a TPMT mutation are more likely to develop leucopoenia compared to those with normal TPMT. In a study by Seddik et al. published in abstract form 75 subjects with CD that were starting AZA/MP treatment were assessed for TPMT mutations. Six (8%) of the subjects were heterozygous for a TPMT mutation and none was homozygous of a TPMT mutation. Seven (9%) of the subjects developed leucopoenia and only one subject was heterozygous for a TPMT mutation.45 In a case–control study by Curvers et al. published as an abstract subjects with TPMT variant alleles were compared to subjects with normal alleles for the risk of developing leucopoenia and the OR was 1.5 (95% CI: 0.7–3.2) for developing leucopoenia.46 In this study and others, the majority of myelosuppression cases occur with normal levels of TPMT enzyme activity.

The prevalence (one of 300) of low TPMT enzyme is high enough and the potential complications of myelosuppression severe enough for us to recommend obtaining a TPMT enzyme activity level prior to starting therapy. A few small studies have suggested that TPMT-deficient patients can safely use low doses of MP/AZA, but until more studies verify safety other immunomodulators should be considered.42, 47

In patients with intermediate TPMT enzyme activity, the risk of myelosuppression is not clearly increased when compared to those with normal TPMT enzyme activity. Until more studies are performed formal evidence-based recommendations to vary treatment (e.g. maximal dose used or rapidity of achieving maximal dose) based upon whether an individual has the wild type (full TPMT enzyme activity) or intermediate TPMT enzyme activity cannot be formally established. It is not yet clear whether TPMT genotype or phenotype can also be used to predict other adverse effects of AZA such as nausea, vomiting, hepatitis and pancreatitis because of conflicting data.48

Mean corpuscular volume.  Several studies have suggested that mean corpuscular volume (MCV) correlates with 6-TGN levels and perhaps may be used as surrogate marker for monitoring therapeutic dosing.36,49,50 A study by Thomas et al., showed that a change of MCV (mean change in MCV was 7.5 ± 6.3 fL) from baseline during treatment with AZA or MP was directly correlated with 6-TGN levels (r = 0.33, P < 0.001) and inversely correlated with leucocyte counts (r = 0.26, P = 0.001).50 A study by Belaiche et al. also showed a correlation of MCV with 6-TGN concentrations (r = 0.38, P = 0.048).36 Further studies are needed to determine the MCV level equivalent to a 6-TGN level.

Adverse events

Several side-effects have been associated with MP/AZA use and include allergic reactions, pancreatitis, myelosuppression, nausea, infections, hepatoxicitiy and malignancy. Bone marrow suppression is related to levels of 6-TGN and may occur at any time during the duration of treatment.32 Therefore, monitoring of complete blood counts at regular intervals throughout the duration of therapy is suggested. It has been described that leucopoenia can occur in the setting of 6-TGN levels <230 pmol/8 × 108, but has also been associated with non-response to MP/AZA.39 Hepatoxicity is a rare complication and the pathophysiological mechanism of hepatic injury is unknown. Possibilities include drug-induced hepatitis, cholestasis, nodular regenerative hyperplasia and peliosis.33, 51 6-MMP levels >5700 pmol/8 × 108 is associated with hepatotoxicity, but patients may have high levels with normal liver chemistries and at low 6-MMP levels hepatotoxicity may occur.39 Routine measurement of 6-MMP for hepatotoxicity is not currently recommended, although the routine measurement of serum liver chemistries is recommended. A baseline measurement of serum liver chemistries should be obtained, then periodic measurements at least once every year thereafter. In patients developing nausea may be switched from AZA to MP or vice versa.52 Patients developing biochemical hepatitis or pancreatitis should have AZA or MP treatment discontinued. There have been several cases of lymphoma reported in AZA/MP users and may be partially related to infection with Epstein–Barr virus, but it is still unclear if there is a definitive increased risk.53–55

Several categories of drugs have been shown to possibly interact with MP and AZA metabolism and include medications that have 5-aminosalicylates (5-ASA) as the active moiety [sulfasalazine, mesalazine (mesalamine), olsalazine and balsalazide], allopurinol, aspirin and furosemide. In a study by Lowry et al. RBC 6-TGN levels were slightly higher but not statistically significant in subjects taking mesalazine, sulfasalazine or olsalazine concurrently compared with subjects not on these medications, 182 vs. 153 pmol/8 × 108 respectively (P = 0.10). The pathway of the drug interaction between MP/AZA and 5-ASA appears to be by inhibition of the TPMT enzyme which may lead to a higher risk for leucopoenia.56–59 Genetic variants of an intestinal epithelial enzyme arylamine-N-acetyl transferase, involved in 5-ASA metabolism, also appear to increase the risk for adverse events of MP therapy.60 Subjects on these medications should be more carefully monitored for myelosuppression.

Ciclosporin

Clinical pharmacology

Ciclosporin is a lipophilic cyclic polypeptide that inhibits cytosolic enzyme calcineurin that leads to selective inhibition of interleukin (IL)-2 and interferon-γ production by T-helper cells, IL-3, IL-4, IL-5, tumour necrosis factor (TNF)-α and TNF-β.61 Bioavailability ranges from 10 to 89% and can be reduced by a fat-rich meal.62, 63 Ciclosporin is primarily metabolized by CYP3A4/5 in liver and the intestine and >90% is excreted in the bile.

Indication and dosing

Ciclosporin is reserved for patients with severe UC that are refractory to steroid therapy. This is generally defined as persistently active colitis despite 7–10 days of high dose (equivalent to 40–60 mg of prednisone daily) intravenous steroids. The primary use is to induce remission and serve as a ‘bridge’ for MP/AZA therapy in UC patients with severe colitis.

This clinical use in UC was defined in a landmark article by Lichtiger et al. where hospitalized subjects with severe UC who failed intravenous steroids after 7 days were randomized to 4 mg/kg of intravenous ciclosporin or placebo. About 82% of subjects receiving ciclosporin responded compared with none that received placebo after 14 days of treatment (P < 0.001).64 In a later study, intravenous ciclosporin 4 mg/kg/day was compared with intravenous steroids 40 mg/day for the treatment of hospitalized subjects with UC for 8 days. About 64% of subjects receiving ciclosporin compared with 53% receiving intravenous steroids responded (P = N.S.).65 A more recent study has shown no significant difference in response of 2 mg/kg/day when compared with 4 mg/kg/day with fewer side-effects.66 There has not been a randomized placebo-controlled trial of ciclosporin for the maintenance of remission of UC.

There have been several controlled trials of intravenous ciclosporin for the treatment of CD with only one of three studies showing possible efficacy.67–69 Because of insufficient evidence of efficacy, ciclosporin is not recommended for the treatment of CD.

Monitoring

The initial studies of ciclosporin for treatment of IBD revealed a dose–response relationship between 200 and 400 ng/mL [monoclonal radioimmunoassay (RIA)] of whole blood ciclosporin. Non-responders were more likely if target blood concentrations were <200 ng/mL, whereas no additional benefit in efficacy has been observed with blood levels >400 ng/mL.64, 70

Baseline medical history, physical examination and laboratory studies are needed prior to initiation of therapy. Patients with a history of seizures, poor compliance to medical therapy, poorly controlled hypertension, and cancer are relative contraindications to starting ciclosporin.71 Pregnancy is also a relative contraindication to starting ciclosporin. In addition, a patient's medication list should be carefully reviewed for possible drug interactions. Active infection should be excluded prior to therapy and hypertension treated. Ciclosporin has a propensity to induce seizures in the setting of low serum cholesterol (≤120–140 mg/dL) or hypomagnesaemia.72 Baseline laboratory tests should include a serum creatinine, potassium, magnesium, cholesterol, complete blood count, liver chemistries, and a pregnancy test in women of child-bearing age. During the first hour of infusion it is advised to watch for signs of an allergic response such as wheezing, urticaria, hypotension every 15 min. Blood work except for serum cholesterol should be taken at minimum every second day. If blood work is abnormal including serum cholesterol, then blood work should be checked daily. Intravenous lipids have been used to boost borderline cholesterol levels. Corticosteroids and 5-ASA can be continued during therapy and MP/AZA discontinued.

A ciclosporin trough level should be obtained every 2 days with a goal blood level of 200–400 ng/mL assessed by high-performance liquid chromatography (HPLC), RIA or immunoassay. Ciclosporin level determined by HPLC is preferred because of immunoassays cross-reacting with ciclosporin metabolites.73–75 If the trough levels are above 500 ng/mL for 2 consecutive days, then ciclosporin dose should be reduced by at least 25%. Medications to be cognizant of that may increase serum levels due to a reduction of ciclosporin metabolism from inhibition of CYP3A4/5 include ketoconazole, erythromycin, grapefruit juice, calcium-channel blockers, bromocriptine, metoclopramide and danazol.76, 77 On the contrary, low levels of ciclosporin may be a result of medications that increase excretion and include medications such as rifampicin, phenobarbital, phenytoin, carbamazepine and St John's wort.78, 79 Other potential reasons for reducing the ciclosporin dose are if serum creatinine increases by 30% over baseline, serum liver enzymes double or diastolic blood pressure exceeds 90 mmHg (12 kPa) or systolic blood pressure exceeds 150 mmHg (20 kPa) despite antihypertensive treatment.

If patients do not respond to therapy after 10 days or their symptoms become worse during that time, then they should be referred to surgery. Otherwise, patients who respond to at least 7 days of intravenous ciclosporin should be switched to oral ciclosporin. Details about switching to oral ciclosporin are suggested in a recently published user's guide to ciclosporin use in severe UC.71 If ciclosporin is continued, then patients should also be started on trimethoprim/sulfamethoxazole 160/80 mg for prevention of pneumocystitis pneumonia. Thus far, there are no published randomized-controlled trial conducted evaluating an oral ciclosporin, for the treatment of active UC or maintenance of remission.

Adverse events

Many of the potential adverse events were mentioned in the section on monitoring. Potential side-effects that have been reported in clinical trials for treatment of IBD of ciclosporin include hypertension, renal insufficiency, seizure, headache, nausea/vomiting, paresthesias, hypertrichosis, gingival hyperplasia and opportunistic infections. Adverse events that occurred in the intravenous ciclosporin trials for the treatment of UC are listed in Table 3. The mechanism of ciclosporin nephrotoxicity may be due to renal arteriole vasoconstriction. If hypertension were to develop calcium-channel blockers are preferred for the treatment of hypertension treatment because of their arteriole vasodilatation effects and protection from ciclosporin-induced nephrotoxicity. Pneumocystitis pneumonia has been reported during treatment with ciclosporin for other non-IBD indications and because of this potential risk trimethoprim/sulfamethoxazole prophylaxis is recommended in those patients receiving ciclosporin >7 days.

Table 3.  Adverse events in trials of intravenous ciclosporin for ulcerative colitis*
 Lichtiger and Present121Lichtiger et al.64Hermida-Rodriguez et al.122†*D'Haens et al.65Van Assche et al.66
  1. † No control group.

  2. ‡ Ten of 11 subjects with mean blood levels provided, based on radioimmunoassay technique.

  3. § Enzyme multiplied immuno test.

  4. ¶ Monoclonal antibody-based fluorescence polarization immunoassay technique.

  5. NR, not reported.

  6. * Also included Crohn's disease patients.

Duration of study (days)10≤147–1088
Ciclosporin dose (mg/kg)44442 vs. 4
Ciclosporin mean blood levels (ng/mL)NR482‡250§376¶237/332¶
Response ciclosporin/control (%)7382/07064/5386/84
Paresthesias ciclosporin/control (%)40¶36/000/76/8
Hypertension ciclosporin/control (%)1336/1107/09/24
Nausea/vomiting ciclosporin/control (%)09/1147/0NR
Nephrotoxicity ciclosporin/control (%)70/090/017/18
Hepatoxicity ciclosporin/control (%)00/00NRNR

Methotrexate

Clinical pharmacology

Methotrexate is a folic acid antagonist that leads to the inhibition of purine synthesis, DNA and RNA formation, and eventually inhibition of the S-phase of the cell cycle. Intracellular, methotrexate is converted to the active metabolite methotrexate polyglutamate by the enzyme folate polyglutamase synthase. Dihydrofolate reductase (DHFR), an enzyme involved in converting folate to tetrahydrofolate, is inhibited by binding to both methotrexate and methotrexate polyglutmate.80–82 Increased DHFR transcription is associated with decreased responsiveness to methotrexate and recently SNPT829C located in the untranslated region of the DHFR gene has been shown to be associated with increased levels of DHFR mRNA.83

Methotrexate polyglutamate is also capable of inhibiting other enzymes in the pathway of folate metabolism leading to purine nucleotide synthesis. The actual cell targets of methotrexate involved in the suppression of inflammation in chronic inflammatory conditions are not known. Potential target cells include cells within the lamina propria of the intestine, intestinal intraepithelial lymphocytes, leucocytes, monocytes-macrophages and intestinal epithelial cells. In vitro and in vivo studies have shown increased levels of extracellular adenosine associated with methotrexate therapy, however, in a small clinical study of 10 IBD patients plasma adenosine and rectal tissue adenosine levels were not found to be elevated after receiving methotrexate 15 or 25 mg subcutaneously once every week.84 Neutrophils, lymphocytes and macrophages all have receptors for adenosine that appears to suppress cytokine release from these cell line.85 Adenosine also appears to inhibit the production of reactive oxygen metabolites, adhesion to and injury of endothelial cells, synthesis and release of leukotriene B4, and production of TNF-α in neutrophils.82 In macrophages/monocytes TNF-α, IL-6 and IL-8 production are inhibited and there is increased promotion the IL-1 receptor antagonist and secretion of IL-10.82

Oral methotrexate is well absorbed, but this appears to be in dose-dependent fashion.86 Lower doses of methotrexate are better absorbed than higher doses (>25 mg/week).86, 87 This dose effect may be related to metabolism by intestinal bacteria. Intramuscular (I.M.) and subcutaneous (S.Q.) formulations of methotrexate have near complete bioavailability and have been shown to have high concentrations in intestinal mucosa.88 It has been suggested that methotrexate is primarily delivered to the intestinal mucosa by the blood stream rather than direct intestinal absorption into sites of inflammation because of higher concentrations of methotrexate in rectal mucosa given in the I.M. or S.Q. form compared with oral methotrexate. Methotrexate is primarily excreted by the kidneys.

Indication and dosing

Methotrexate was initially used in rheumatoid and psoriatic arthritis, but has been found to be useful for many chronic inflammatory conditions. Methotrexate has been shown to be beneficial in the treatment of steroid refractory CD.89

There have been several uncontrolled studies and randomized-control studies suggesting a benefit for the induction of remission, steroid-sparing effects, and maintenance of remission for CD.90–96 There have been a small number of uncontrolled studies and one randomized-control study evaluated the efficacy of methotrexate for UC.89, 91, 95, 97, 98 In the initial study, five of seven patients were induced into remission showing possible benefit was by Kozarek et al.89 In a recent study by Paoluzi et al., 10 patients whom could not tolerate or were resistant to AZA were given methotrexate 12.5 mg/week I.M. for active UC. Six subjects were induced into remission and four subjects had significant improvement based on clinical, endoscopic and histological examinations after 6 months of therapy.98 In the one randomized-controlled study by Oren et al., methotrexate 12.5 mg/week per oral (PO) was compared with placebo after 9 months for inducing active UC into remission. About 47% of methotrexate subjects after a median of 4.1 ± 1.9 months went into remission compared with 49% of placebo subjects after 3.4 ± 1.7 months.91 Although this study did not show a benefit perhaps I.M. or SQ and higher doses may be effective. The role of methotrexate in UC still remains unclear.

For the treatment of active CD, methotrexate doses of 15–25 mg S.Q./I.M. for 12–16 weeks may be considered. Based on clinical trial results oral methotrexate has been shown to be modestly superior to placebo at best at doses from 12.5 to 22.5 mg.92 Oral bioavailability of methotrexate has been reported approximately 73% (95% CI: 0.62–0.86) compared with S.Q. administration with minimal change of bioavailability when given with folate.99 If patients are switched to an oral formulation of methotrexate, then the equivalent S.Q. dose needs to be determined. Once a patient is in remission, maintenance treatment is recommended with either continuation of methotrexate at 15 mg S.Q./I.M. or starting AZA, MP or 5-ASA.

Monitoring

There are no specific metabolites available to monitor therapy for efficacy or side-effects. Because of the potential myelosuppression and hepatotoxicity total blood cell counts and liver enzymes should be monitored regularly every 1–3 months. In a study by Feagan et al., seven of 94 patients treated with methotrexate in a 16-week trial for active CD had transaminases greater than twice the upper limits of normal and persisted after 1 week despite cessation of methotrexate.90 In this study, no patients were withdrawn because of leucopoenia. In a 40-week maintenance study of methotrexate in CD in remission there were no patients withdrawn due to leucopoenia or abnormal liver enzymes.96 A single point in time liver enzyme elevation has been shown to be a poor predictor of severity of liver, whereas persistent enzyme elevation on serial measurement appears to correlate with liver histology.100 The Psoriasis Task Force suggest a liver biopsy pre-treatment, if a patient will be on methotrexate long-term and after a patient has received a cumulative methotrexate dose of 1.5 g.101 Indications for liver biopsy from the American College of Rheumatology are (i) pre-treatment biopsy for a history of excessive alcohol consumption, persistently abnormal aspartate aminotransferase (AST) or chronic hepatitis B or C infection, (ii) during treatment biopsy for persistently abnormal liver function tests defined as elevations of AST in five of nine determinations within a given 12-month interval or a decrease in serum albumin below the normal range. In a recent case series of 20 IBD patients taking methotrexate (mean accumulated dose of 2633 mg) only one had liver fibrosis by liver biopsy.102 This suggests that IBD patients may be at lower risk for liver fibrosis than psoriasis or rheumatoid patients. If liver enzymes are abnormal, we recommend rechecking liver enzymes after a short interval, i.e. 1-month interval. If the liver enzymes remain elevated, then a liver biopsy should be considered as well as a work-up for other possible aetiologies. Whether to obtain a liver biopsy after a methotrexate cumulative dose of 1.5 g is unclear taking into consideration the possible lower likelihood for hepatic fibrosis in IBD patients when compared with psoriasis and rheumatoid arthritis. Further studies are needed in the IBD population to determine the need for liver biopsy.

Adverse events

Methotrexate has been used for diseases such as psoriasis and rheumatoid arthritis long before its use in IBD, so its side-effect profile has been well-documented. Major adverse events reported include hepatotoxicity, myelosuppression, pneumonitis in 3–12%, infertility and foetal malformation, and enteritis/colitis. Minor adverse events reported include nausea, vomiting, diarrhoea, stomatitis, alopecia, rash and neurological changes.87 Hepatoxicity is a well-known side-effect of treatment. Although studied in a non-IBD population, the changes in liver histology include macrovescicular steatosis, nuclear variability, chronic inflammatory infiltrates in the portal tracts, hepatocyte necrosis, fibrosis and cirrhosis. Hepatic fibrosis and cirrhosis are the most concerning long-term potential toxicities and tend to occur with accumulated methotrexate use of >1.5 g. Myelosuppression is known potential side-effect of methotrexate, but it is uncommon if the duration of therapy is <1 year.103 Infections reported in methotrexate users include pneumocystitis carinii pneumonia, herpes zoster, cytomegalovirus, Epstein–Barr virus and Listeria monocytogenes.104–107 There have been several case reports of lymphoma in rheumatoid arthritis patients and other connective tissue disorders treated with methotrexate with resolution of lymphoma once methotrexate is stopped.108–110 Many of the lymphomas reported have been associated with Epstein–Barr virus. Although, there has yet been a controlled study verifying an increased risk for lymphoma in subjects using methotrexate.

In the four randomized trials of methotrexate for the treatment of CD for durations ranging from a minimum of 4– 12 months and doses ranging from a minimum of 12.5 mg PO/week to 25 mg i.m./week, the most common side-effects were nausea and elevation of liver enzymes.90, 92, 93, 96 In the rheumatoid arthritis population folate and folinic acid supplementation to methotrexate therapy has been found effective in reducing side-effects in a systematic review, meta-analysis and a randomized-controlled trial.111–113 Folate and folinic acid have not been found to be different in their effectiveness for reducing side-effects from methotrexate therapy, but folinic acid is more expensive than folate. To avoid potential toxicities folate supplementation is recommended, either 1 mg per mg of methotrexate divided into daily doses or 0.25–0.50 mg per mg of methotrexate given 4–24 h after administration of weekly methotrexate.114, 115

Conclusion

Drug monitoring for effectiveness and side-effects of immunomodulators is paramount. Table 4 provides a guide to pre-treatment and during treatment drug monitoring. With recent biomedical technology advances our understanding of the metabolic consequence of this class of drugs in vivo has grown. Yet we are still at the infancy stage of understanding and fine tuning these advances. As a consequence, there is still a dearth of information on evidence for monitoring for effectiveness and side-effects of these medications within the IBD population. The algorithms and suggestions provided are partly subjective but encompass our best-educated advice due to lack of literature in this area. Studies are still needed to: (i) clarify the role of 6-TGN for measuring efficacy of AZA and MP, (ii) cost-effectiveness of TPMT testing prior to starting AZA and MP therapy, (iii) determine the best starting dose in individuals with intermediate TMPT activity, (iv) determine the optimal frequency of surveillance blood testing in out-patients (such as complete blood count and liver function test) in individuals treated with immunomodulators and (v) determine the role of liver biopsy in the IBD population after receiving a cumulative dose of >1.5 g of methotrexate. As the indications for use of immunomodulators broadens for the treatment of IBD, monitoring for efficacy and side-effects will be even more critical.

Table 4.  Laboratory monitoring of immunomodulators*
ImmunomodulatorPre-treatment assessmentTreatment assessment
  1. * TPMT, thiopurine methyltransferase; 6-MMP, 6-methylmercaptopurine; CBC, complete blood count; LFT, liver function test.

Azathioprine/ MercaptopurineTPMTCBC
CBCLFT
LFTMetabolites (6-TGN, 6-MMP)
CiclosporinCBCCBC
LFTLFT
Serum electrolytes (potassium, magnesium)Serum electrolytes (potassium, magnesium)
Serum creatinineSerum creatinine
Serum cholesterolSerum cholesterol
 Ciclosporin level
MethotrexateCBCCBC
LFTLFT

Acknowledgement

GR Lichtenstein has received honorarium from Promethius Inc and Salix Pharmaceuticals.

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