272
Severe Adverse Reaction to Carbamazepine: Significance of Humoral and Cellular Reactions to the Drug By G. Horneff, H. G. Lenard and V. Wahn Department of Pediatrics, University of Düsseldorf, Moorenstr. 5, Düsseldorf, Germany
Abstract Hypersensitivity to carbamazepine is a wellknown phenomenon. The involvement of several organ systems including liver, kidney, bone marrow and other organs have been described. We have observed a 7-year-old boy who had been treated with carbamazepine for seizures. After 10 days of treatment he developed a severe illness with skin rash, high fever, lymphadenopathy, hepatosplenomegalyand lymphopenia. Only slightly decreased complement components and increased complement split products but no circulating immune complexes were demonstrable on admission. Anti-carbamazepine antibodies, T-cell-activation and a significant T -cell reactivity against carbamazepine were found, indicating specific hypersensitivity. Complete recovery was observed after discontinuation of the drug and steroid treatment. Keywords Carbamazepine - Hypersensitivity - Anticarbamazepine antibodies
Introduction Carbamazepine is frequently used for treatment of seizures of focal origin in adults and in children (1, 7). Side effects induced by carbamazepine are not uncommon and include rash, fever, hematological, gastrointestinal, cardiovascular, neurological symptoms and disturbances of liver and renal function (2). Although these side effects are usually mild and disappear rapidly after carbamazepine discontinuation, severe serum sickness has been reported in some cases (5). The typical clinical manifestations including fever, skin rash, petechial bleeding and lymphadenopathy were regarded as indicative of a type 111 hypersensitivity reaction (6). In this report we describe a patient suffering from severe side effects of carbamazepine associated with demonstrable anti-carbamazepine antibodies, activated T -cells and significant T -cell reactivity to carbamazepine.
Received February 18,1992; accepted May 4,1992 Neuropediatrics 23 (1992) 272-275 © Hippokrates Verlag Stuttgart
Peripheral blood mononuclear cells were separated from heparinized blood as described elsewhere (3). Phenotypic characterization was performed by two-colour immunofluorescence using fluorescein isothiocyanate (FITC) and phycoerythrin conjugated monoclonal antibodies to CD3, CD4, CD8, CD25 and HLA-DR (purchased from BectonDickinson, Mountain View, CA) in a Becton-Dickinson FACSAnalyser. For stimulation experiments, 1 x 105 mononuclear cells (PBL) were cultured per weIl in triplicates in the presence of purified phytohemagglutinin (PHA, 1Jlg/ml, Wellcome, London, UK), pokeweed mitogen (PWM, 1 Jlg/ml, Sigma, Munich, FRG), anti-CD3 antibody (5 ng/ml, Orthoclone OKT3, Cilag, FRG) and medium alone as described in detail (3). Lymphocyte proliferation induced by antigens (Tetanus-toxoid, tuberculin PPD, all Behring, Marburg, FRG; or serially diluted sera containing carbamazepine as weIl as control sera) was measured by 3H-thymidine (1 JlCi/well, Amersham, Buckinghamshire, UK) incorporation during the final 4 hours of incubation, 3 days after mitogen stimulation and 7 days after antigen stimulation. Cells from normal donors never exposed to carbamazepine served as controls. Stimulation indices were calculated by dividing the cpm of 3H-thymidine uptake in the presence of the antigen by the cpm in the absence of antigen. Single ingredients of Tegretal® (carbamazepine, cellulose, magnesiumstearat, carboxymethylcellulose and siliciumoxid) were provided by Ciba-Geigy, Wehr, FRG. All substances were incubated in RPMI 1640 containing 10 % ABserum (100 Jlg/ml)~ stirred for 1 hour at 37°C, and centrifugated at 800 g for 10 min at room temperature. Supernatants were filtered through a 0.4 Jlm filter and serial dilutions in RPMI 1640 containing 10 % AB-serum were used in the antigen assay as described above. Serial dilutions of sera af another carbamazepine treated patient and of anormal donar were used to investigate the stimulatory effect of carbamazepine and its metabolites. Anticarbamazepine antibodies in sera were quantified by an Elisa procedure: A Balb c mouse was given carbamazepin (1 mg/g) orally and bled 12 hours later. This serum was used as the antigen in the Elisa as described elsewhere (4). Serum of an untreated mouse of the same litter was used as contro!. Briefly, a 1 % dilution of sera in 10 mM carbonate buffer pH 9.6 was incubated in 96 weIl Elisa plates (NUNC, Roskilde, Denmark) for 18 hours at 4°C. The plates were washed 3 times with PBS containing 0.05 % Tween 20 (Sigma, Munich, FRG) and blocked with 10 % bovine serum albumine. Washing procedure was repeated. Serial dilutions of sera in PBS 0.05 % Tween were incubated for 2 hours, subsequently
Downloaded by: National University of Singapore. Copyrighted material.
Methods
Immunoreactivity to Carbamazepine lable 1 Laboratory data on admission (day 0) and follow-up examinations.
Neuropediatrics 23 (1992) Day Th rom bocytes LeukocytesjJ..l1 LymphocytesjJ..l1 CD3+* CD3+CD25+* CD3+HLA-DR+* CD4+* CD8+* CD4jCD8-Ratio PHA§ OKT3§ PWM§ medium§
1:
273
94000
2300 920 71 (72) 3 (1) 14 (4) 21 (53) 62 (26) 0.34 (2.04) 32500 (44500) (....) 7100 (28500) (~) 1700 (20500) (~) 100 (150)
7400 4440 85 (69) 4 (1) 16 (7) 55 (41) 23 (10) 2.39 (4.1)
46094 (41604) 43465 (21428) 22077 (16560) 1844 (1090)
the plates were washed and incubated with a goat-anti-human IgG alkaline phosphatase conjugate (Medac, Hamburg, FRG). Enzyme activity was measured using p-nitrophenyl-phosphate (Sigma). Case report Ten days before admission to our hospital, treatment with carbamazepine (300 mg twice daily) had been initiated in a 7-year-old boy because of complex partial seizures and pathologic EEG. Two days before admission, a skin rash developed and carbamazepine treatment was discontinued because of suspected hypersensivity. However, the skin rash persiste.d and the child became febrile. Because of his deteriorating general condition he was referred to our hospital. On admission he presented with fever up to 40.5°C, conjunctivitis, pharyngitis, generalized erythematous rash with petechial bleeding in the skin of face and trunk, generalized lymphadenopathy, hepatosplenomegaly and hematemesis. The child was drowsy and appeared severely ill. Laboratory tests showed a marked leukopenia (2100 cells/I.tl), lymphocytopenia (820 cells/I.tl), most pronounced of the T-helper-cell subset, and thrombocytopenia (93000/I·tl), anormal hemoglobin of 13 g/dl, elevated serum aspartate aminotransferase (108 V/I), alanin aminotransferase (98 V/I), y-glutamyltransferase (92 V/I), the lactic dehydrogenase (710 V/I). The erythrocyte sedimentation rate was 12/28, C-reactive protein 0.6 mg/dl, C3c was 59.5 mg/dl (slightly decreased), C4 81.9 mg/dl, the complement split product C3d 140 % (slightly increased), and CH50 140 % of pooled normal serum standard. Tests for circulating immune complexes (solid phase anti-C3 Elisa and mm IgG), antinuclear antibodies, IgM antibodies to measles, rubella, CMV or EBV were negative. However, on day 8 low amounts of circulating immune complexes were measurable. On admission, carbamazepine level in serum was 2.8 J.lg/ ml. On the second day in our hospital, the patient developed abdominal pain and decreased intestinal motility. An abdominal x-ray indicated a paralytical subileus. Treatment was started with parenteral water and salt replacement and methylprednisolone at 2 mg/kg bodyweight. On the following day the fever had disappeared and blood leukocyte counts gradually increased to reach normal levels by day 5. Subsequently, steroids were reduced. Liver enzymes decreased and were nearly normal on day 6. Splenomegaly also disappeared on the 6th day, the enlarged liver on day 10. He was then discharged without any medication.
Two months later, the patient was seen in our outpatient clinic. Clinical examination was unremarkable. A skin prick test with emulsions of all drug ingredients showed no immediate type I reactivity in the presence of a positive control reaction to histamin. Table 1 shows the time course of thrombocytes, leukocytes, lymphocytes and lymphocyte subpopulations. Circulating lymphocytes demonstrated a markedly elevated proportion of activated T -cells. The CD4/CD8 ratio was decreased (0.34) and returned to normal during steroid therapy. In contrast, lymphocyte activation markers were still demonstrable at this time but had returned to normal 8 weeks after hospitalization. On admission, lymphocytes showed a decreased proliferation upon stimulation with PHA, PWM and OKT3. The T -cell reactivity towards carbamazepine was tested by stimulation of the lymphocytes with carbamazepine in vitro. The stimulatory effect of another patient's serum who had taken carbamazepine without any side effect could possibly reflect T -cell reactivity towards carbamazepine and its metabolites. As outlined in Figure 1, a stimulation index of 4 was noted either by incubaStim.lndex 100 [§ill] Tetanu8 toxoid ~ Tubercullne PPD _
Carbamazeplne
_
Serum cont. C.
10
Kon
BH
Fig. 1 T-cell reactivity to carbamazepine: Lymphocytes were stimulated with tetanus toxoid, tuberculine PPD, carbamazepine or a patient's serum containing carbamazepine and its metabolites.
Downloaded by: National University of Singapore. Copyrighted material.
* % of PBL, §counts per minute, control values are shown in brackets.
G. Horneffet al
274 Neuropediatrics 23 (1992) 1000
900
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800
B.H. 6/27/91
00405
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•
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600
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O+------r------r----~-----.-------.
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O-+------r------.----~-----.--------.
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dilution of serum -1
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700
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600
o
Control
600
o
Control
500
500
400
400
300
300
200
200
100
100 0 80
40
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320
640
Fig. 2 IgG-reactivity towards carbamazepine and its metabolites: Serial dilutions of patients sera were incubated with Elisa plates coated with murine serum containing carbamazepine (black circles) as weil as control
L1
20
40
80
160
320
640
dilution of serum -1
dilution of serum -1
murine serum (open circles). Higher binding activity towards serum containing the drug compared to control serum demonstrates drug-specific IgG-reactivity.
00
tion of patients' lymphocytes with carbamazepine or with the serum containing also the metabolites.
600
I DOther Patlents
-
ND
-
Index-Patient
I
500 400 300 200
1
2
3
4
5
6
7
8
9 10 11 12
I
11
11I IV 0
8 56
day
Fig. 3 IgG-reactivity towards carbamazepine and its metabolites: Differences in IgG-reactivity towards murine serum containing carbamazepine compared to control serum were also seen in patients who were treated with carbamazepine without side effects (open bars). However, drugspecific IgG reactivity is most pronounced in the index patient reported here (black bars). Hatched bars: sera of healthy controls never exposed to carbamazepine.
Antibodies of the IgG class reactive with carbamazepine or metabolites were measured by Elisa (Fig. 2). Serial dilutions of the patients sera (Figs.2 A-C) showed a markedly higher binding to mouse serum containing carbamazepine than to control mouse serum. No carbamazepine specific binding of IgG was noticed with serum of a healthy contral (Fig. 2 D). Experiments were performed to investigate the humoral anti-carbamazepine reactivity in sera of patients wha were treated with carbamazepine without side effects. In Figure 3, differences in IgG-reactivity towards mouse serum containing carbamazepine and mouse serum alone were shown in 12 patients and 4 normal donors. However, a low probably non-specific IgG reactivity toward the serum containing carbamazepine was also demonstrable in some individuals.
Downloaded by: National University of Singapore. Copyrighted material.
00405 1000
Neuropediatrics 23 (1992)
Immunoreactivity to Carbamazepine
Discussion
Clinically, our patient showed symptoms of severe carbamazepine allergy. This was associated with demonstrable both humoral and cellular reactivity in vitro. Measurement of complement components, complement split products and circulating immune complexes indicated participation of the complement system in this carbamazepine induced serum sickness-like disease. As shown in Figure 3, an IgG response to carbamazepine or its metabolites is also demonstrable in patients tolerating the drug therapy, however in a lower degree than in our patient. Thus, antibodies to carbamazepine per se are not indicative of immunological drug intolerance and the importance and frequency of antibody production specific to carbamazepine remains to be determined. Skin tests showed no immediate type hypersensitivity. Besides lymphocytopenia, marked changes in the CD4/CD8 ratio and elevated T-cell activation markers T-cell reactivityto carbamazepine was measurable in the patient reported here. In conclusion, drug-specific antibodies and type IV hypersensitivity mediated by antigenspecific T -lymphocytes should also be investigated in patients demonstrating severe adverse reactions to carbamazepine.
Acknowledgment We thank Christiane Zubrod for skillful technical assistance.
References Andersen, E. B., A. Philbert,]. G. Klee: Carbamazepine monotherapy in epileptic outpatients. Acta Neurol. Scand. (Suppl) 94 (1983) 29-34 2 Askmark, H., B. Wilholm: Epidemiology of adverse reactions to carbamazepine as seen in a spontaneous reporting system. Acta N eurol. Scand.81 (1990) 131-140 3 Bunnester, G. R.,]. R. Kalden, H. H. Peter, 1. Schedel, P. Beck, A. Wittenborg: Immunological and functional characteristics of peripheral blood and synovial fluid lymphocytes from patients with rheumatoid arthritis. Scand. J. Immunol. 7 (1978) 405-417 4 Horneff, G., T Winkler,]. R. Kalden, F. Emmrich, G. R. Bunnester: Human antimouse antibody reponse induced by anti-CD4 monoclonal antibody therapy in patients with rheumatoid arthritis. Clin. Immunol. Immunopathol. 59 (1991) 89-103 5 Hosoda, N., W. Sunaoshi, H. Shirai, Y. Bando, H. Miura, M. Igarashi: Anticarbamazepine antibody induced by carbamazepine in a patient.with severe serum sickness. Arch. Dis. Child 66 (1991) 722-723 6 Mullick, F. G., H. A. McAllister, B. M. Wagner,].]. Fenoglio: Drugrelated vasculitis. Clinicopathological correlations in 30 patients. Hum. Pathol. 10 (1979) 313-325 7 Ochinsky, 5., O. Chosinow, C. Prost, P. ]oly,]. C. Roujeau, ]. Revuz: Carbamazepine update. Lancet 981 (1989) 8669 8 Pacifici, R., L. Paris, S. Di Carlo, S. Pichini, P. Zuccaro: Immunological aspects of carbamazepine treatment in epileptic patients. Epilepsia 1 (1991) 122-127 9 Zakrzewska, ]. M., L. Ivanyi: In vitro lymphocyte proliferation by carbamazepine, carbamazepine-1 0, 11-epoxyde, and oxcarbamazepine in the diagnosis of drug-induced hypersensitivity. J. Allergy Clin. Immunol. 82 (1988) 110-115 1
Prof Dr. V. Wahn University of Düsseldorf Moorenstr.5 D-4000 Düsseldorf, Germany
Downloaded by: National University of Singapore. Copyrighted material.
The anticonvulsant carbamazepine is wellknown to cause side effects, both dose-dependent but more often dose-independent (2). Most frequently, rashes are observed, followed by hematological abnormalities (1). However, immunological reactions to carbamazepine have rarely been analyzed. Several immunological secondary phenomena like decreased serum IgA and IgM levels, decreased T-helperl suppressor lymphocyte ratio, diminished PHA induced lymphocyte proliferation and increased killer cell activity have been reported (8). Because of their low molecular size, carbamazepine or its metabolites alone are hardly immunogenic, but they may act as haptens to become immunogenic in association with a carrier protein. Specific immunity is usually demonstrable 7 to 10 days after the first exposure to the drug and has recently been reported in a child with severe serum sickness (5). In vitro lymphocyte reactivity to carbamazepine has been shown in a study including 9 patients with carbamazepine induced skin rashes (9).
275
Severe Adverse Reaction to Carbamazepine: Significance of Humoral and Cellular Reactions to the Drug By G. Horneff, H. G. Lenard and V. Wahn Department of Pediatrics, University of Düsseldorf, Moorenstr. 5, Düsseldorf, Germany
Abstract Hypersensitivity to carbamazepine is a wellknown phenomenon. The involvement of several organ systems including liver, kidney, bone marrow and other organs have been described. We have observed a 7-year-old boy who had been treated with carbamazepine for seizures. After 10 days of treatment he developed a severe illness with skin rash, high fever, lymphadenopathy, hepatosplenomegalyand lymphopenia. Only slightly decreased complement components and increased complement split products but no circulating immune complexes were demonstrable on admission. Anti-carbamazepine antibodies, T-cell-activation and a significant T -cell reactivity against carbamazepine were found, indicating specific hypersensitivity. Complete recovery was observed after discontinuation of the drug and steroid treatment. Keywords Carbamazepine - Hypersensitivity - Anticarbamazepine antibodies
Introduction Carbamazepine is frequently used for treatment of seizures of focal origin in adults and in children (1, 7). Side effects induced by carbamazepine are not uncommon and include rash, fever, hematological, gastrointestinal, cardiovascular, neurological symptoms and disturbances of liver and renal function (2). Although these side effects are usually mild and disappear rapidly after carbamazepine discontinuation, severe serum sickness has been reported in some cases (5). The typical clinical manifestations including fever, skin rash, petechial bleeding and lymphadenopathy were regarded as indicative of a type 111 hypersensitivity reaction (6). In this report we describe a patient suffering from severe side effects of carbamazepine associated with demonstrable anti-carbamazepine antibodies, activated T -cells and significant T -cell reactivity to carbamazepine.
Received February 18,1992; accepted May 4,1992 Neuropediatrics 23 (1992) 272-275 © Hippokrates Verlag Stuttgart
Peripheral blood mononuclear cells were separated from heparinized blood as described elsewhere (3). Phenotypic characterization was performed by two-colour immunofluorescence using fluorescein isothiocyanate (FITC) and phycoerythrin conjugated monoclonal antibodies to CD3, CD4, CD8, CD25 and HLA-DR (purchased from BectonDickinson, Mountain View, CA) in a Becton-Dickinson FACSAnalyser. For stimulation experiments, 1 x 105 mononuclear cells (PBL) were cultured per weIl in triplicates in the presence of purified phytohemagglutinin (PHA, 1Jlg/ml, Wellcome, London, UK), pokeweed mitogen (PWM, 1 Jlg/ml, Sigma, Munich, FRG), anti-CD3 antibody (5 ng/ml, Orthoclone OKT3, Cilag, FRG) and medium alone as described in detail (3). Lymphocyte proliferation induced by antigens (Tetanus-toxoid, tuberculin PPD, all Behring, Marburg, FRG; or serially diluted sera containing carbamazepine as weIl as control sera) was measured by 3H-thymidine (1 JlCi/well, Amersham, Buckinghamshire, UK) incorporation during the final 4 hours of incubation, 3 days after mitogen stimulation and 7 days after antigen stimulation. Cells from normal donors never exposed to carbamazepine served as controls. Stimulation indices were calculated by dividing the cpm of 3H-thymidine uptake in the presence of the antigen by the cpm in the absence of antigen. Single ingredients of Tegretal® (carbamazepine, cellulose, magnesiumstearat, carboxymethylcellulose and siliciumoxid) were provided by Ciba-Geigy, Wehr, FRG. All substances were incubated in RPMI 1640 containing 10 % ABserum (100 Jlg/ml)~ stirred for 1 hour at 37°C, and centrifugated at 800 g for 10 min at room temperature. Supernatants were filtered through a 0.4 Jlm filter and serial dilutions in RPMI 1640 containing 10 % AB-serum were used in the antigen assay as described above. Serial dilutions of sera af another carbamazepine treated patient and of anormal donar were used to investigate the stimulatory effect of carbamazepine and its metabolites. Anticarbamazepine antibodies in sera were quantified by an Elisa procedure: A Balb c mouse was given carbamazepin (1 mg/g) orally and bled 12 hours later. This serum was used as the antigen in the Elisa as described elsewhere (4). Serum of an untreated mouse of the same litter was used as contro!. Briefly, a 1 % dilution of sera in 10 mM carbonate buffer pH 9.6 was incubated in 96 weIl Elisa plates (NUNC, Roskilde, Denmark) for 18 hours at 4°C. The plates were washed 3 times with PBS containing 0.05 % Tween 20 (Sigma, Munich, FRG) and blocked with 10 % bovine serum albumine. Washing procedure was repeated. Serial dilutions of sera in PBS 0.05 % Tween were incubated for 2 hours, subsequently
Downloaded by: National University of Singapore. Copyrighted material.
Methods
Immunoreactivity to Carbamazepine lable 1 Laboratory data on admission (day 0) and follow-up examinations.
Neuropediatrics 23 (1992) Day Th rom bocytes LeukocytesjJ..l1 LymphocytesjJ..l1 CD3+* CD3+CD25+* CD3+HLA-DR+* CD4+* CD8+* CD4jCD8-Ratio PHA§ OKT3§ PWM§ medium§
1:
273
94000
2300 920 71 (72) 3 (1) 14 (4) 21 (53) 62 (26) 0.34 (2.04) 32500 (44500) (....) 7100 (28500) (~) 1700 (20500) (~) 100 (150)
7400 4440 85 (69) 4 (1) 16 (7) 55 (41) 23 (10) 2.39 (4.1)
46094 (41604) 43465 (21428) 22077 (16560) 1844 (1090)
the plates were washed and incubated with a goat-anti-human IgG alkaline phosphatase conjugate (Medac, Hamburg, FRG). Enzyme activity was measured using p-nitrophenyl-phosphate (Sigma). Case report Ten days before admission to our hospital, treatment with carbamazepine (300 mg twice daily) had been initiated in a 7-year-old boy because of complex partial seizures and pathologic EEG. Two days before admission, a skin rash developed and carbamazepine treatment was discontinued because of suspected hypersensivity. However, the skin rash persiste.d and the child became febrile. Because of his deteriorating general condition he was referred to our hospital. On admission he presented with fever up to 40.5°C, conjunctivitis, pharyngitis, generalized erythematous rash with petechial bleeding in the skin of face and trunk, generalized lymphadenopathy, hepatosplenomegaly and hematemesis. The child was drowsy and appeared severely ill. Laboratory tests showed a marked leukopenia (2100 cells/I.tl), lymphocytopenia (820 cells/I.tl), most pronounced of the T-helper-cell subset, and thrombocytopenia (93000/I·tl), anormal hemoglobin of 13 g/dl, elevated serum aspartate aminotransferase (108 V/I), alanin aminotransferase (98 V/I), y-glutamyltransferase (92 V/I), the lactic dehydrogenase (710 V/I). The erythrocyte sedimentation rate was 12/28, C-reactive protein 0.6 mg/dl, C3c was 59.5 mg/dl (slightly decreased), C4 81.9 mg/dl, the complement split product C3d 140 % (slightly increased), and CH50 140 % of pooled normal serum standard. Tests for circulating immune complexes (solid phase anti-C3 Elisa and mm IgG), antinuclear antibodies, IgM antibodies to measles, rubella, CMV or EBV were negative. However, on day 8 low amounts of circulating immune complexes were measurable. On admission, carbamazepine level in serum was 2.8 J.lg/ ml. On the second day in our hospital, the patient developed abdominal pain and decreased intestinal motility. An abdominal x-ray indicated a paralytical subileus. Treatment was started with parenteral water and salt replacement and methylprednisolone at 2 mg/kg bodyweight. On the following day the fever had disappeared and blood leukocyte counts gradually increased to reach normal levels by day 5. Subsequently, steroids were reduced. Liver enzymes decreased and were nearly normal on day 6. Splenomegaly also disappeared on the 6th day, the enlarged liver on day 10. He was then discharged without any medication.
Two months later, the patient was seen in our outpatient clinic. Clinical examination was unremarkable. A skin prick test with emulsions of all drug ingredients showed no immediate type I reactivity in the presence of a positive control reaction to histamin. Table 1 shows the time course of thrombocytes, leukocytes, lymphocytes and lymphocyte subpopulations. Circulating lymphocytes demonstrated a markedly elevated proportion of activated T -cells. The CD4/CD8 ratio was decreased (0.34) and returned to normal during steroid therapy. In contrast, lymphocyte activation markers were still demonstrable at this time but had returned to normal 8 weeks after hospitalization. On admission, lymphocytes showed a decreased proliferation upon stimulation with PHA, PWM and OKT3. The T -cell reactivity towards carbamazepine was tested by stimulation of the lymphocytes with carbamazepine in vitro. The stimulatory effect of another patient's serum who had taken carbamazepine without any side effect could possibly reflect T -cell reactivity towards carbamazepine and its metabolites. As outlined in Figure 1, a stimulation index of 4 was noted either by incubaStim.lndex 100 [§ill] Tetanu8 toxoid ~ Tubercullne PPD _
Carbamazeplne
_
Serum cont. C.
10
Kon
BH
Fig. 1 T-cell reactivity to carbamazepine: Lymphocytes were stimulated with tetanus toxoid, tuberculine PPD, carbamazepine or a patient's serum containing carbamazepine and its metabolites.
Downloaded by: National University of Singapore. Copyrighted material.
* % of PBL, §counts per minute, control values are shown in brackets.
G. Horneffet al
274 Neuropediatrics 23 (1992) 1000
900
900
800
B.H. 6/27/91
00405
B.H. 6/20/91
•
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O+------r------r----~-----.-------.
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Control
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500
400
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100 0 80
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Fig. 2 IgG-reactivity towards carbamazepine and its metabolites: Serial dilutions of patients sera were incubated with Elisa plates coated with murine serum containing carbamazepine (black circles) as weil as control
L1
20
40
80
160
320
640
dilution of serum -1
dilution of serum -1
murine serum (open circles). Higher binding activity towards serum containing the drug compared to control serum demonstrates drug-specific IgG-reactivity.
00
tion of patients' lymphocytes with carbamazepine or with the serum containing also the metabolites.
600
I DOther Patlents
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-
Index-Patient
I
500 400 300 200
1
2
3
4
5
6
7
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9 10 11 12
I
11
11I IV 0
8 56
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Fig. 3 IgG-reactivity towards carbamazepine and its metabolites: Differences in IgG-reactivity towards murine serum containing carbamazepine compared to control serum were also seen in patients who were treated with carbamazepine without side effects (open bars). However, drugspecific IgG reactivity is most pronounced in the index patient reported here (black bars). Hatched bars: sera of healthy controls never exposed to carbamazepine.
Antibodies of the IgG class reactive with carbamazepine or metabolites were measured by Elisa (Fig. 2). Serial dilutions of the patients sera (Figs.2 A-C) showed a markedly higher binding to mouse serum containing carbamazepine than to control mouse serum. No carbamazepine specific binding of IgG was noticed with serum of a healthy contral (Fig. 2 D). Experiments were performed to investigate the humoral anti-carbamazepine reactivity in sera of patients wha were treated with carbamazepine without side effects. In Figure 3, differences in IgG-reactivity towards mouse serum containing carbamazepine and mouse serum alone were shown in 12 patients and 4 normal donors. However, a low probably non-specific IgG reactivity toward the serum containing carbamazepine was also demonstrable in some individuals.
Downloaded by: National University of Singapore. Copyrighted material.
00405 1000
Neuropediatrics 23 (1992)
Immunoreactivity to Carbamazepine
Discussion
Clinically, our patient showed symptoms of severe carbamazepine allergy. This was associated with demonstrable both humoral and cellular reactivity in vitro. Measurement of complement components, complement split products and circulating immune complexes indicated participation of the complement system in this carbamazepine induced serum sickness-like disease. As shown in Figure 3, an IgG response to carbamazepine or its metabolites is also demonstrable in patients tolerating the drug therapy, however in a lower degree than in our patient. Thus, antibodies to carbamazepine per se are not indicative of immunological drug intolerance and the importance and frequency of antibody production specific to carbamazepine remains to be determined. Skin tests showed no immediate type hypersensitivity. Besides lymphocytopenia, marked changes in the CD4/CD8 ratio and elevated T-cell activation markers T-cell reactivityto carbamazepine was measurable in the patient reported here. In conclusion, drug-specific antibodies and type IV hypersensitivity mediated by antigenspecific T -lymphocytes should also be investigated in patients demonstrating severe adverse reactions to carbamazepine.
Acknowledgment We thank Christiane Zubrod for skillful technical assistance.
References Andersen, E. B., A. Philbert,]. G. Klee: Carbamazepine monotherapy in epileptic outpatients. Acta Neurol. Scand. (Suppl) 94 (1983) 29-34 2 Askmark, H., B. Wilholm: Epidemiology of adverse reactions to carbamazepine as seen in a spontaneous reporting system. Acta N eurol. Scand.81 (1990) 131-140 3 Bunnester, G. R.,]. R. Kalden, H. H. Peter, 1. Schedel, P. Beck, A. Wittenborg: Immunological and functional characteristics of peripheral blood and synovial fluid lymphocytes from patients with rheumatoid arthritis. Scand. J. Immunol. 7 (1978) 405-417 4 Horneff, G., T Winkler,]. R. Kalden, F. Emmrich, G. R. Bunnester: Human antimouse antibody reponse induced by anti-CD4 monoclonal antibody therapy in patients with rheumatoid arthritis. Clin. Immunol. Immunopathol. 59 (1991) 89-103 5 Hosoda, N., W. Sunaoshi, H. Shirai, Y. Bando, H. Miura, M. Igarashi: Anticarbamazepine antibody induced by carbamazepine in a patient.with severe serum sickness. Arch. Dis. Child 66 (1991) 722-723 6 Mullick, F. G., H. A. McAllister, B. M. Wagner,].]. Fenoglio: Drugrelated vasculitis. Clinicopathological correlations in 30 patients. Hum. Pathol. 10 (1979) 313-325 7 Ochinsky, 5., O. Chosinow, C. Prost, P. ]oly,]. C. Roujeau, ]. Revuz: Carbamazepine update. Lancet 981 (1989) 8669 8 Pacifici, R., L. Paris, S. Di Carlo, S. Pichini, P. Zuccaro: Immunological aspects of carbamazepine treatment in epileptic patients. Epilepsia 1 (1991) 122-127 9 Zakrzewska, ]. M., L. Ivanyi: In vitro lymphocyte proliferation by carbamazepine, carbamazepine-1 0, 11-epoxyde, and oxcarbamazepine in the diagnosis of drug-induced hypersensitivity. J. Allergy Clin. Immunol. 82 (1988) 110-115 1
Prof Dr. V. Wahn University of Düsseldorf Moorenstr.5 D-4000 Düsseldorf, Germany
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The anticonvulsant carbamazepine is wellknown to cause side effects, both dose-dependent but more often dose-independent (2). Most frequently, rashes are observed, followed by hematological abnormalities (1). However, immunological reactions to carbamazepine have rarely been analyzed. Several immunological secondary phenomena like decreased serum IgA and IgM levels, decreased T-helperl suppressor lymphocyte ratio, diminished PHA induced lymphocyte proliferation and increased killer cell activity have been reported (8). Because of their low molecular size, carbamazepine or its metabolites alone are hardly immunogenic, but they may act as haptens to become immunogenic in association with a carrier protein. Specific immunity is usually demonstrable 7 to 10 days after the first exposure to the drug and has recently been reported in a child with severe serum sickness (5). In vitro lymphocyte reactivity to carbamazepine has been shown in a study including 9 patients with carbamazepine induced skin rashes (9).
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