D. Elewaut, Department of Rheumatology, University Hospital Ghent, De Pintelaan 185, 9000 Gent Belgium
E-mail: Dirk.Elewaut{at}Ugent.be
Since its original synthesis more than half a century ago [1], methotrexate (MTX) has become an established drug in several medical specialties. Developed as a specific antagonist of folic acid, this drug was shown to inhibit proliferation of malignant cells primarily by inhibiting de novo synthesis of purines and pyrimidines [2]. Hence its original use primarily as a chemotherapeutic agent, but later it was also evaluated for the treatment of rheumatoid arthritis (RA) because of its presumed antiproliferative activities [3]. For more than 15 yr now it has been an established DMARD with pronounced anti-inflammatory activity that goes beyond inhibition of folic acid metabolism [2, 4]. In fact, several studies have indicated that its anti-inflammatory effects may be due to other mechanisms, such as adenosine release, which can induce immunosuppression [5]. Although MTX is definitely one of the most effective drugs for the treatment of RA and other rheumatic diseases, such as psoriatic arthritis, in recent years it become apparent that, besides its toxicity profile, MTX has several other shortcomings.
First, although several studies have indicated that MTX therapy induces a considerable clinical remission rate, it does not halt the progression of erosions in RA. In fact, Lipsky et al. demonstrated that more than 30% of patients with active RA treated with 12.5 mg MTX per week demonstrated major radiological progression after 54 weeks of therapy [6]. In addition, about 35% of RA patients do not respond sufficiently to MTX alone and require adjustment of their treatment [6]. This could, however, be partly due to starting therapy too late. Indeed, it has become clear that the early use of DMARDs in the treatment of RA is efficacious and that delaying its initiation reduces the impact of this therapy [7]. Moreover, female gender, prior DMARD use, disease functional class and disease activity may also affect the response rate [7]. In addition, genetic factors are likely to play a critical role in determining the response rate in RA patients. Therefore, the pharmacogenetic study of genetic differences (polymorphisms) in the enzymes involved in the metabolic pathways of MTX that could affect drug efficacy is currently an area of intense investigation [8, 9]. Polymorphisms in drug transporters (such as P glycoprotein) and drug targets (such as receptors) could also influence treatment results.
Finally, the pharmacokinetic properties of MTX are rather unsatisfactory and may result in inadequate clinical response rates. After uptake, more than 80% of MTX is rapidly eliminated unchanged via glomerular filtration and tubular secretion [10]. A smaller part is excreted through the biliary tract. The half-life of MTX is only 7 h, although this may extend indefinitely in renal failure. As a result of this fast elimination, the drug concentration in the target tissues is low. Most MTX is delivered to cells as the parent compound; a small fraction is hydroxylated in the liver and circulates as 7-OH-MTX [11], a biologically active metabolite which is eliminated more slowly than MTX [10]. A portion of the intracellular MTX and 7-OH MTX is metabolized to polyglutamates in the same manner as naturally occurring folates [12]. MTX polyglutamates are long-lived metabolites that retain some of the antifolate activities of MTX, although the potency for inhibition of various folate-dependent enzymes is altered [12, 13]. These metabolites are considered to contribute significantly to the clinical efficacy of MTX as an anti-rheumatic drug.
The disadvantageous pharmacokinetic profile of MTX certainly contributes to its unsatisfactory treatment efficacy in some RA patients. To circumvent these problems, MTX could be covalently coupled to a suitable drug carrier to improve its pharmacokinetic properties. From a theoretical point of view, optimal drug carriers allow high accumulation in target tissues but low uptake in normal tissues. They should also have low toxicity and have the chemical property of being able to be coupled to the drug of choice. The plasma protein albumin has been shown to be an appropriate carrier molecule [14, 15]. Therefore, a group of German investigators developed an MTXhuman serum albumin (MTX-HSA) compound drug in which MTX was covalently coupled to human serum albumin initially as a chemotherapeutic agent [16, 17]. A series of preclinical studies demonstrated that MTX-HSA accumulates actively within tumours in mice by a process of endocytosis. This is followed by the lysosomal breakdown of albumin, after which MTX is liberated within the tumour cells. Because of the high accumulation rate in tumour tissue as opposed to normal tissue, this compound drug preferentially targets the malignant tissues. Interestingly, MTX-HSA was found to be superior to MTX in the suppression of tumour growth and, remarkably, the two agents were shown to act synergistically [18, 19].
While MTX-HSA is currently being investigated in clinical trials for its use in cancer treatment, its effect as a potential anti-rheumatic drug had not been evaluated until recently. Wunder et al. evaluated the pharmacokinetics of MTX-HSA in comparison with MTX in murine collagen-induced arthritis (CIA), a mouse model of arthritis, by comparing the distribution of intravenously administered radiolabelled albumin with that of MTX in healthy and arthritic mice [20]. Marked differences were observed in tracer uptake and circulation times. Neither albumin nor MTX accumulated in non-inflamed paws. However, in arthritic paws there was significant uptake of radiolabelled albumin but not of MTX [20]. By contrast, the uptake in liver and kidney was found to be three times lower for albumin than for MTX. However, the authors did not directly compare the pharmacokinetics of 3H-labelled MTX with those of 3H-labelled MTX-HSA, which would be required for a direct comparison between the two compounds. In addition, synovial fibroblasts from RA patients were found to actively take up fluorescently labelled albumin in vitro as well as in vivo when co-implanted with human articular cartilage in SCID mice [20]. These findings indicate that albumin might be a useful drug-carrier, allowing specific targeting of MTX to sites of inflammation.
Next, the authors initiated a preclinical prophylactic efficacy study in CIA, applying two doses of MTX vs MTX-HSA based on the maximum tolerated dose of the two compounds [20]. The therapeutic effect of both compounds was determined by clinical assessment of paw inflammation. Interestingly, at least a five-fold higher dose of MTX was required to achieve the same level of efficacy, expressed as incidence of arthritis. The severity of the disease in mice that developed arthritis, however, was similar regardless of the treatment regimen [20].
In this issue of Rheumatology, Fiehn et al. extend the knowledge of MTX-HSA as an anti-arthritic drug by comparing the preclinical prophylactic efficacies of MTX, MTX-HSA and the combination of both in the CIA model [21]. Curiously, when lower doses of both drugs were administered as a combination therapy, they were found to act synergistically and markedly reduced the incidence of arthritis in a preventive protocol. Combination therapy, however, did not influence the degree of clinical paw swelling in this particular mouse model [21]. However, the authors did not perform a histological assessment to evaluate the effects of MTX and/or MTX-HSA on joint inflammation. Therefore, it remains to be determined whether the effects of the two drugs on synovial inflammation and particularly bone erosions and cartilage damage are comparable. Given the observed synergistic effects of both drugs, one would expect that their respective modes of action may show subtle differences. Of interest are the recent results from the same investigators assessing the effects of MTX and MTX-HSA on cartilage invasion and degradation by RA synovial fibroblasts when co-implanted with human cartilage in SCID mice. In this particular model, both MTX and MTX-HSA effectively inhibited cartilage invasion and degradation to a similar degree [22].
In order to define more precisely the putative target cells of MTX-HSA, Fiehn et al. also compared the uptake of fluorescently labelled albumin by peripheral blood mononuclear cells (PBMC) of RA patients vs healthy controls using flow cytometry [21]. Albumin was found to be taken up by monocytes, granulocytes and B cells in both RA patients and controls. Whereas in normal circumstances only a minority of T cells take up albumin, this fraction was significantly increased in RA patients, probably reflecting T-cell activation. Unfortunately, no direct comparison between the uptake of MTX vs MTX-HSA by PBMC has been performed to date. Therefore it remains to be determined whether the cell types in the circulation targeted by MTX-HSA differ from those targeted by MTX.
Despite its improved pharmacokinetics and superior preclinical prophylactic effect on the incidence of arthritis in CIA, several questions remain to be answered about MTX-HSA. What is the precise reason for improved clinical efficacy? Is this solely due to the altered pharmacokinetic properties or are other mechanisms, such as changes in drug metabolites, also involved? What is the nature of the observed synergistic effect of MTX and MTX-HSA? Is there a therapeutic effect on established disease? Will there still be a therapeutic advantage in treating with MTX-HSA rather than with MTX once clinical remission has been achieved, given that MTX-HSA preferentially accumulates in inflamed but not in non-inflamed joints? These are only a few of the questions that will have to be addressed in future studies.
In summary, MTX-HSA is a promising example of the importance of drug carriers in developing a new generation of targeted therapeutic drugs. Targeting inflamed joints by this MTX-HSA compound drug might be an attractive strategy in the treatment of chronic inflammatory rheumatic diseases such as RA.
DE is supported by grants from the Fund for Scientific ResearchFlanders, the Research Council of the University of Ghent and the Marató Foundation.
The author has declared no conflicts of interest.
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