Q J Med 2000; 93: 223-229
© 2000 Association of Physicians
Clinical versus genetic diagnosis of familial Mediterranean fever
From the Service de Médecine Interne, l'Hôtel-Dieu, Paris, 1 Service de Biochimie Génétique, Hôpital Cochin, Assistance Publique-Hôpitaux de Paris, Paris, 2 Service de Biochimie Génétique et de Génétique Moléculaire Hôpital Henri Mondor, Créteil, and 3 Service d'Hépato-Gastro-entérologie, Hôpital de Villeneuve Saint-Georges, France
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The diagnosis of familial Mediterranean fever (FMF) has until recently been based on clinical signs alone. Discovery of the MEFV gene has enabled a molecular approach to diagnosis, which is already well established for diagnosing typical clinical forms of FMF. We evaluated the utility of this molecular approach in a large series of patients with various clinical presentations and ethnic origins. We looked for mutations in the MEFV gene in 303 unselected consecutive patients with a variable (from high to low) clinical suspicion of FMF. Two mutations were found in 133 patients (44%). In 22 patients (7%), the clinical diagnosis of FMF was unlikely according to the Tel Hashomer clinical criteria. Our results suggest that the spectrum of FMF-associated signs is broader than previously believed. Wider indications for genotyping should lead to more frequent diagnosis of FMF.
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Introduction |
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Familial Mediterranean fever (FMF MIM 249100) is an autosomic recessive disease of the inflammatory pathway. FMF is the most frequent of the hereditary fevers, which include the hyperimmunoglobulinaemia D with periodic fever syndrome (HIDS) (MIM 260920)1,2 and autosomal dominant recurrent fevers such as Tumour-Necrosis-Factor-Receptor-1-associated Periodic Syndrome (TRAPS) (MIM 142680 and 134610).3–6 FMF mainly affects populations of the Mediterranean basin, such as Arabs, Armenians, Sephardic Jews and Turks. Carrier frequency has been estimated to reach 1 : 7 to 1 : 5 in Armenians and Sephardic Jews, respectively.7–8 FMF has also been described in other European populations from Europe.9–11 The disease typically presents as recurrent episodes of fever accompanied by topical signs of inflammation, mainly involving the peritoneal, pleural and articular cavities.12 Importantly, the severity of the disease is linked to the occurrence of AA amyloidosis.12 Given the absence of pathognomical clinical or easily available biochemical abnormalities, the diagnosis of FMF was until recently based on clinical suspicion and the use of criteria. The diagnosis of FMF may be extremely difficult to establish in the presence of atypical signs, and with late onset and absence of family history or at-risk background. However, it is crucial to establish the diagnosis of FMF, since it leads to the beginning of a daily and lifelong administration of colchicine, which is an efficient preventive treatment of both the attacks and amyloidosis.13
The gene responsible for FMF (designated MEFV) situated on the short arm of chromosome 16 (16p13.3) has been identified by positional cloning,14,15 and encodes a protein named marenostrin or pyrin. The spectrum of MEFV mutations responsible for FMF has been regularly widening, and about fifteen mutations have now been discovered.9,11, 16–18 These molecular methods now constitute a powerful tool to establish the diagnosis of FMF in patients with clinical criteria. However, the diagnostic value of the molecular tests in patients with atypical or mild signs, especially in those who do not satisfy clinical criteria, is unknown.
We prospectively investigated a large series of unselected patients, some of whom did not satisfy the clinical criteria of FMF, in order to evaluate the involvement of the MEFV gene in this population.
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Inclusion criteria
After the discovery of the MEFV gene, routine molecular diagnosis of FMF in our laboratory began in November 1997, since when, the MEFV genotypes of 303 patients have been determined. Samples were sent by physicians from various medical specialities including internal medicine, gastroenterology, pediatrics, rheumatology, and nephrology. The clinical presentations for which molecular diagnosis was requested were diverse, ranging from ‘textbook’ clinical observations of FMF to clinical situations with a very low suspicion of FMF, including patients with organ involvement limited to one system. Main clinical data were registered on a standard form: age, sex, origin of both parents, consanguinity, familial history of FMF, age of onset of the inflammatory attacks, duration of attacks, organ involvement, frequency of the attacks, splenomegaly, amyloidosis and efficacy of colchicine. In order to detect mild or atypical clinical presentations of the disease, we did not require the physician to use a specific set of clinical criteria. The use of prerequisite criteria would have be an obstacle to detecting such clinical presentations.
When the diagnosis of HIDS or TRAPS was considered in patients without any identified mutations in the MEFV gene, we measured serum IgD levels and searched for TNFR1 mutations.
Mutation analysis
Our diagnostic strategy was first based on direct sequencing of exon 10 of MEFV after amplification by polymerase chain reaction (PCR) which allows the detection of most of known mutations. Mutations were subsequently searched for in exons 2, 3 and 5 with denaturing gradient gel electrophoresis (DGGE), restriction enzyme analysis and direct sequencing.18
Mutations were searched for in the seven TNFR1 exons encoding the complete extracellular part of the TNFR1 by direct sequencing.
Clinical criteria tested versus genotype analysis results
We then compared the results of the molecular analysis to the probability of the clinical diagnosis of FMF according to a set of criteria. We chose to use the Tel Hashomer criteria as they were easily obtained from our standard form.19 Major criteria are: (i) recurrent febrile episodes accompanied by peritonitis, synovitis or pleuritis; (ii) amyloidosis of the AA type without predisposing disease; and (iii) favourable response to continuous colchicine treatment. Minor criteria are: (i) recurrent febrile episodes; (ii) erysipelas-like erythema; and (iii) FMF in a first-degree relative. Definite diagnosis requires two major, or one major and two minor criteria; ‘probable’ diagnosis requires one major and one minor.
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Results |
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The main clinical characteristics of the 303 patients were as follows: 164 patients were male, fever was observed in 254 patients (83.3%), abdominal signs in 225 (74.0%), thoracic signs in 73 (24%), joint signs in 153 (50.3%), erysipela-like erythema in 25 (8.2%), splenomegaly in 26 (8.5%), amyloidosis in 12 (4%). The age of onset of the disease was below 30 years in 84% of the patients. Of the patients investigated, 30% were Sephardic Jews, 10% Armenians, 16% Arabs, 14% Turks, 9% French without ethnic background typically associated with FMF, and 21% belonged to other ethnic groups, or only one of their parents belonged to an at-risk ethnic group.
Two mutations were found in 133 patients (44%). Among them, 85 were homozygous and 48 compound heterozygous. Forty-six patients (15%) were found to be heterozygous. In 124 patients (41%), no mutations was found. The results of the different mutations identified in these different ethnic groups were consistent with those of the literature (data not shown).
Table 1
shows the results of the genotypes according to the clinical probability of FMF using clinical criteria. Among the 127 patients with a definite clinical diagnosis of FMF, two mutations were found in 85 patients (67%), and no mutation was identified in 24 patients (19%). Conversely, among the 137 patients in whom the clinical diagnosis of FMF was unlikely, no mutation was identified in 92 patients (67%) and two mutations were found in 22 (16%).
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Table 2
shows the clinical data for the 22 patients with an ‘unlikely’ diagnosis of FMF and who carried two MEFV mutations. The age of onset varied from 1 to 48 years. Some patients were newly diagnosed, while others had a long disease history. In 16 of the 22 patients (72%), clinical signs were limited to recurrent febrile episodes with peritonitis. A family history of FMF was found in four of them (18%). The 22 patients belonged to diverse populations usually affected by FMF (i.e. Sephardic Jews, Armenians, Arabs, Lebanese and Turks). Ten patients were homozygous for the M694V mutation, and seven other genotypes were present in the 12 remainder patients. Five patients had the M694V/E148Q genotype. In three of them, parental analysis confirmed that the two mutations were not located on the same chromosome. One patient, who was homozygous for the M694V mutation, had amyloidosis without the typical history of recurrent inflammatory attacks.
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Table 3
shows the clinical data of the 24 patients with a definite diagnosis of FMF and without any identified mutation in the MEFV gene. The age of onset varied from 2 to 43 years. Recurrent febrile episodes accompanied by abdominal signs were present in 20 patients (83%). A family history was found in five (21%). In nine of the 24 patients, both parents belonged to a population not usually affected by FMF. For three patients only one parent belonged to the population usually affected by FMF (Jews, Arabs, and Turks). Two patients had reactive AA amyloidosis and no identified inflammatory disease.
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In six patients with no identified mutation in the MEFV gene, serum IgD levels were measured and were found to be normal. In seven patients, the search for mutations in the TNRF1 gene was negative.
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Discussion |
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The diagnosis of FMF has until recently been based on clinical signs alone, and can be difficult in case of mild or atypical forms of the disease. The discovery of the MEFV gene has led to a molecular approach to diagnosis, aiming at improving the global diagnosis of the disease. The value of the molecular approach has been established in patients with a high suspicion of clinical diagnosis and who satisfied proposed criteria.14–18 However, the utility of the molecular approach to FMF diagnosis in patients of varying ethnic origins with atypical clinical presentations has not yet been addressed.
Of our 303 patients, we were able to detect at least one mutation in 59% of the patients, a percentage value that is lower than that of previous studies.11,18 As our strategy includes the screening of almost all mutations known to date, it is unlikely that several new mutations underlie the disease phenotype of our patients. This low percentage is more likely explained by our clinical recruitment of the patients for whom a molecular diagnosis was done, which includes many patients without true FMF. In the absence of a simple confirmatory clinical test, FMF diagnosis relied until recently on clinical features. Early authors proposed sets of criteria in order to standardize the diagnosis of FMF.12 More recently, other authors established their own criteria,9 but until now the sole criteria to be validated were those of Livneh et al.20 Other sets of criteria have been proposed, but none of these are validated in our unselected population. Among the different sets of criteria currently proposed to establish the diagnosis of FMF,9,12,20 we chose the Tel Hashomer criteria, which are the easiest to use. Our results show that if we had used these criteria to select the patients for molecular diagnosis, we would have missed 22 patients with two mutations. These criteria are thus not sensitive enough to allow the diagnosis of mild forms of the disease. However, it is of note that we considered, from a pragmatic point of view, that the third major criteria (i.e. efficiency of colchicine) was absent in the 22 patients with two mutations, as none of them was reported to have a complete response to this treatment (treatment not used or partially efficient). If more detailed data regarding the treatment by colchicine had been available, more patients with a positive third major criteria would probably have been registered. Colchicine therapy was not undertaken in most of our patients, because the diagnosis was not made, in the absence of complete clinical presentation and family history, even in presence of a typical ethnic background. Thus, in our series of 303 patients, wide indications for genotyping based on broader clinical criteria led to more frequent diagnosis of FMF.
The theoretical view that some homozygous patients may remain asymptomatic even in advanced age is now supported by the presence of two mutations in several patients with late onset, and of asymptomatic parents in some families.18 The outcome of amyloidosis in such asymptomatic patients remains a subject of discussion. One of our patients, a 12-year-old Turkish patient carrying the homozygous M694V genotype, has had amyloidosis with only atypical articular signs in the past. He can be considered as having a quasi type 2 phenotype. There is increasing evidence that rare type 2 FMF phenotype, although controversial, probably exists.21 How many of our 21 patients with a mild form of the disease, and of their asymptomatic parents, are likely to develop amyloidosis is unknown. These data raise the question of genetic screening of asymptomatic parents of affected subjects and of treatment with colchicine of asymptomatic patients with two mutations, especially those who are M694V homozygotes, as this status seems to be particularly associated with amyloidosis.18,21,22
The specificity of the tested criteria is also limited, as 24 patients with positive criteria had no identified mutations. The following explanations could account for this observation. First, some of these patients are not of Mediterranean extraction, and some so-far-unknown mutations may exist in unexplored regions, though our mutation screening was near-exhaustive. Secondly, another as-yet-unidentified FMF locus may exist, as suggested in the Turkish population.23 Thirdly, there is the question of the true diagnosis in these patients. Many diseases can present as fever of recurrent origin. A series of 45 patients with recurrent fever of undetermined origin, including two patients with FMF, underlined the pitfalls of this diagnosis.24 Four patients in this series had a final diagnosis of Still's disease, which can present with recurrent fever arthritis, skin lesions and a strong acute-phase response. However there is no confirmative test for this disease, whose diagnosis is based on a set of criteria including the exclusion of other diagnoses, requiring a lot of invasive investigation. Three patients of the same series had a final diagnosis of inflammatory bowel disease, which can also present as recurrent attacks of abdominal pain with fever, and histological evidence of the disease can take a long time to obtain. Both diseases can mimic FMF and were discussed in some of our patients in whom no mutation of the MEFV gene was found.
Other forms of recurrent hereditary fevers are probably rare in our population. Indeed, most of our cases of recurrent fever appeared to be sporadic. In patients lacking familial history but a clinical suspicion of HIDS or TRAPS, there was no biochemical or genetic evidence for these syndromes.
The molecular approach to FMF diagnosis now enables confirmation of FMF in most typical cases of the disease. However it also shows that the MEFV gene is implicated in atypical clinical presentations, including patients who do not satisfy clinical criteria. Thus, the clinical spectrum regarded as indicative of molecular testing for FMF should be broadened.
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Notes |
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Address correspondence to Dr G. Grateau, Service de Médecine Interne, l'Hôtel-Dieu, 1 Place du Parvis Notre-Dame, 75181 Paris cedex 04, France. e-mail: gilles.grateau{at}htd.ap\|[hyphen]\|hop\|[hyphen]\|paris.fr
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