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Transient monoclonal proteins in drug hypersensitivity reactions
Transient Monoclonal Proteins in Drug Hypersensitivity Reactions
JAIME DEL CARPIO, M.D. LUIS R. ESPINOZA, M.D.* STEVEN LAUTER, M.D.+ C. KIRK OSTERLAND, M.D. Montreal, Quebec, Canada
Two patients were studied in whom monoclonal (M) immunoglobulin G (IgG) proteins developed during the course of a serum sickness-like drug hypersensitivity reaction to cloxacillin (Orbenin@)and sodium cephalothin &eflin@),respectively. The clinical evidence and time sequence of events support this association. In both patients there was evidence of an active antibody response to the given antibiotic and to the benzylpenicilloyl group as well. However, protein fractions obtained by agar gel preparative electrophoresis failed to show a higher antibody concentration where the M peak was located, and absorption experiments performed with penicillin G-, cloxacillin- and cephalothin-coated red cells failed to absorb these M proteins. These transient paraproteins can be seen in association with antibiotic(s) administration in the context of a hypersensitivity reaction and do not apparently represent a specific immune response. drug reactions are common in modern medical practice, and they represent a costly aspect of health care. Penicillin is associated with a wide variety of different side effects. This drug and its derivatives often induce the formation of polyclonal immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies. The role of these antibodies in the production of hypersensitivity reactions is not clear. A protective “blocking antibody” function has been suggested for some penicillin antibodies [1,2]. In the case of cephalothin, they can be responsible for positive antiglobulin tests and possibly immune hemolytic anemia [3,4]. Ampicillin antibodies may bear important causal relation to the “ampicillin skin rash” frequently observed in patients with infectious mononucleosis [5]. The development of a monoclonal (M) protein in association with a drug reaction has been mentioned in the literature [6,7], but it is not clearly established. Recently, we have had the opportunity of seeing two patients in whom M proteins developed during the course of serum sickness-like reactions caused by a semisynthetic penicillin (cloxacillin) and a cephalosporin (sodium cephalothin], respectively. The M proteins disappeared after administration of the drug had been stopped, and clinical hypersensitivity reaction subsided.
Adverse
From the McGill University, Royal Victoria Hospital, Division of Allergy and Clinical Immunology, Montreal, Quebec, Canada. This study was supported by grants from the Canadian Arthritis and Rheumatism Society. Requests for reprints should be addressed to: Dr. Jaime Del Carpio, McGill University, Royal Victoria Hospital, Division of Allergy and Clinical Immunology, 687 Pine Avenue West, Montreal, Quebec, H3A 1Al Canada. Manuscript accepted September 27,1978. * Present address: Medical Center College of Medicine, Department of Internal Medicine, 12901 North 30th Street, University of South Florida, Tampa, Florida 33612 + Present address: Ballas Medical Office Center, 777 South New Ballas Road, Suite 305, St. Louis, Missouri 63141.
CASE REPORT Case 1. A 42 year old woman in previously good health who had no atopic background or history of penicillin hypersensitivity, received oral cloxacillin (1 g/day) for symptoms attributed to an ear infection. At the end of a week, she had fever followed by a generalized pruritic urticarial rash. Medication was stopped, but she remained ill. Five days later she became confused, and a mild weakness of the right arm was noted. She was admitted to another hospital
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Journal of Medicine
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TRANSIENT
TABLE I
MONOCLONAL
PROTEINS
AND DRUG
HYPERSENSITIVITY-DEL
CARP10
ET AL.
Summary of Immunologic Laboratory Data
Case 1 11/30/76
IgG (850-1600 mgldl) IgA (85-305 mgldl) IgM (90-285 mgldl) Antipenicillin antibodies+ (hemagglutination) Immune complexes (Clq deviation 96) IgG monoclonal protein (immunoelectrophoresis) l
1212176
1,800
394 108
12117176
1,590
382 104 1:1,024
Case2 114/77 1,040
1:512
43 %
359 85 1:18 18%
4+ (SPE N”:,
216177
(&+#3)
(SPE ;;
7128176 1,200
1:18
330 580 1:2,048
819176 1,410
215 298 1:18
(SPE #5)
(SPE ;;
(SPE #7)
Similar results obtained with cephalothin (Case 1) and cloxacillin (Case 2) sensitized red cells.
where eltaminationshowed
a somnolent but oriented woman with a generalized urticarial rash. Her blood pressure was 110/80 mm Hg, pulse rate 80/min and a thorough ear, nose and throat examination was within normal limits. There was no lymphadenopathy or organomegaly. Neurologic evaluation confirmed a mild weakness in the right arm and diffuse hyperflexia. Laboratory data showed a normal hemogram except for an eosinophilia of 45 per cent (3,700 of a total 8,200 white blood cells/mm3). The erythrocyte sedimentation rate was 44 (Wintrobe). Urinalysis showed a normal sediment and a trace of protein. Serum immunoglobulins were (Table I]: immunoglobulin G (IgG) 1,600, immunoglobulin A (IgA) 394, and immunoglohllin M (IgM) 106 mg/dl. On zone electrophoresis one could discern a monoclonal protein band in the gamma area. Serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, lactic dehydrogenase (LDH), alkaline phosphatase and bilirubin levels were within normal limits. Tests for hepatitis B antigen and antibodies were negative. Cerebrospinal fluid examination disclosed no abnormalities. Two days after admission, the eosinophilia had decreased to 1,600 cells/mm3, and there was no albumin in the urine. Complement determinations were not performed. The patient was investigated neurologically to rule out the presence of an intracranial abscess. Twelve days after the onset of the skin rash, the patient was fully oriented, the weakness in the right arm was resolving, and there were only a few residual urticarial lesions. kepeat laboratory studies showed the complete blood count to b? within normal limits with no eosinophilia, total hemolytic complement (CH50), the third (C3) and fourth [C4]components of complement were within normal limits. Blood cultures and viral titers, obtained during the acute and convalescent phase, were all negative. Liver functions tests showed no abnormalities, serum immunoglobulins were IgG 1,590 mg/dl, IgA 382 mg/dl and IgM 104 mg/dl. On immunoele‘ctrophoresis IgG X paraprotein was identified. Urine electrophoresis did not show any abnormalities. Cryoglobulins, cryofibrinogen, cold agglutinins, rheumatoid factor and ahtinuclear antibodies were absent. Immune complexes and hemagglutination titers are shown in Table I. Repeat lumbar puncture showed a normal cerebrospinal fluid. Brain scan and computerized axial tomography were also within normal limits. The patient continued to show improvement, and she was discharged home asymptomatic. Serial serum protein electrophoresis and immunoelectrophoresis [Figure 1, Table I)
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showed gradual disappearance of M protein. After two and a half months the IgG paraprotein was no longer detectable by immunoelectrophoresis. Hemagglutination titers against penicillin G and cloxacillin-coated red cells also showed a declining titer, and immune complex measurements decreased [Table I). Skin tests with penicilloyl-polylysine (PPL) and penicillin G one month after the urticarial rash were negative. One year later the patient was well. Case 2. A 45 year old man was first admitted to the Royal
Victoria Hospital in October 1975 for investigation of aplastic anemia. He had had acute rheumatic fever at age 17, and a heart murmur had been discovered at age 27. There was no history of drug hypersensitivities. In December 1974. he underwent a right meniscectomy at another hospital where mild pancytopenia was noted. In April 1975,a diagnosis of aplastic anemia was made. Treatment with several courses of prednisone, oxymethalone, vitamin B1zand pyridoxine was unsuccessful, and he was referred to our hospital. The diagnosis was confirmed, but no etiology was determined. Among the normal studies was a serum protein electrophoresis. He was treated with transfusions of packed red blood cells and oxymethalone 150 mg/day. He was followed as an outpatient until May 1976 when he was readmitted with fever, a hematoma on the right thigh and acute left ethmoid and maxillary sinusitis. His hemoglobin level was 5.9 g/100 ml and his white blood cell count 1,800/ mm3 with 30 per cent neutrophils and 70 per cent lymphocytes. He was treated with packed red blood cells, platelets, cloxacillin 8 g/day, gentamicin 3 mg/kg/day and ampicillin 8 g/day. Ten days later, multiple papular erythematosus lesions developed over the elbows, and later in the groin and lower extremities. Antibiotic therapy was stopped, the lesions faded, and he remained afebrile. The clinical impression confirmed by skin biopsy was leukocytoclastic angiitis. The final admission occurred in July 1976 because of fever, bleeding from buccal ulcerations and a perirectal abscess. On examination blood pressure was llO/50 mm Hg, pulse rate lOO/min and temperature 40%. Erosions were noted on the palatal and buccal mucosa. There was 2 cm jugular venous distention, bibasilar rales and a grade 3/6 holosystolic apical murmur in the anterior axillary line with an opening snap. The liver was slightly enlarged, and there was tender left inguinal adenopathy. Rectal examination revealed a large left-sided mass just inside the rectum. His white blood cell count was
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7OO/mm3,his hematocrit value 13 per cent. Liver function tests yielded normal results except for a bilirubin of 1.8 mg/lOO ml. There was a moderate growth of Escherichia coli from the rectal lesion, but repeated blood cultures were negative. The patient was given a transfusion of packed red blood cells and platelets, and treated with digoxin, Keflin@ 4 g intravenously a day, clindamycin 600 mg intravenously every 6 hours and gentamicin 5 mg/kg/day. The perirectal abscess was drained. After eight days of treatment raised, purpuric lesions developed in the patient’s left groin which spread overnight to his abdomen, chest and back with severe pruritus. Keflin therapy was discontinued, but gentamicin and clindamycin were maintained at the same dose. The lesions became more confluent, and microscopic hematuria appeared. The patient was treated with Benadryl” (diphenhydramine hydrochloride) and over the next three days the lesions faded, and his hematuria cleared. A serum protein electrophoresis on the second day of the rash revealed a total protein of 7.1 g/loo ml, albumin 3.28 g/dl, alpha, globulin 0.29 g/dl, alpha2 globulin 0.95 g/dl, beta globulin 0.68 g/d1 and gamma globulin 1.85 g/d1 with a monoclonal band in the gamma region. Quantitative immunoglobulins were IgG 1.200 mg/dl, IgA 330 mg/dl and, IgM 560 mg/dl. Immunoelectrophoresis showed an IgG h paraprotein. Australia antigen and antibody were absent. Two weeks later the rash was completely gone, and repeat serum protein electrophoresis and immunoelectrophoresis no longer displayed a paraprotein peak (Figure 1). Quantitative immunoglobulins were IgG 1,410 mg/dl. IgA 215 mg/dl and IgM 298 mg/dl (Table I). Skin tests with penicilloyl-polylysine and penicillin G yielded negative results. The patient’s clinical condition deteriorated with the appearance of a mass in the upper right lung, repeated bouts of septicemia and increasing need for transfusions of red blood cells and platelets. One month later he died. Autopsy findings revealed acute pulmonary hemorrhage secondary to fulminant Pseudomonas pneumonia and invasive Aspergillosis of the lung, but no evidence of systemic vasculitis and no evidence of plasmocytosis. METHODS Penicillin hemagglutinating antibodies were determined by the technic described by Levine et al. [8]. A similar method was used for cephalothin [4] and cloxacillin. Circulating immune complexes, were measured by the Clq deviation technic of Sobel et al. [9]. In ourlaboratory the mean inhibition of normal serum is 2.7 per cent f 5 per cent (SD) with a range of 0 to 4 per cent. The test is capable of detecting as low as 15 pg/ml of aggregated IgG. Immunoelectrophoresis was performed by micromethod of Sheidegger [lo]. The identification and serologic typing of monoclonal proteins was carried out using antiserums specific for IgG classes and light chain subclasses. Serum proteins were separated by agar gel preparative electrophoresis. Fractions eluted by freezing and thawing were assayed by Bio-Rad protein assay [II]. The location of immunoglobulin classes was ascertained by setting up agar double diffusion plates of each of the eluted fractions against specific antiserums. Fractions were either assessed directly for antibody activity or, in some cases, several
June 1979
AND DRUG HYPERSENSITIVITY-DEL
CARP10 ET AL.
igure 1. Sequence of serum protein kectrophoresis (zone). Case 1 (cloxacillin hypersensitivity): electrophoresis 2-3-4-5. Case 2 (cephalothin hypersensitivity): electrophoresis 6-7. Electrophoresis 1: normal serum for comparison. (For dates see Table I.)
fractions were pooled and concentrated by pressure dialysis prior to further testing. Serum from the patient (Case 2) was filtered through a Sephadex@ G 200 column. The top tubes from each of the three G 200 peaks were pooled and concentrated prior to testing. Serum from both patients containing M peaks was used for absorption experiments. Red cells sensitized with penicillin G. cephalothin and cloxacillin were used to absorb patient’s serum at 37’C for 2 hours and overnight at 4% After each absorption, the serums were retested by zone electrophoresis and serologically. The absorptions were repeated sequentially eight times. RESULTS
protein electrophoreses (acetate cellulose], in both patients and a normal control subject for comparison, are shown in Figure 1. Laboratory results on both patients are summarized in Table I. In Case 1 (cloxacillin hypersensitivity) IgG, hemagglutinating titers against penicillin G and cloxacillin-coated red cells and immune complexes levels, deSerum
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-
HA
CARP10 ET AL.
+
I 200
I
150
I 250
I 300
’
ELUTION VOLUME (ml) Figure 2. Case 2: Sephadex G 200 column fractionation. HA = hemagglutination; HA-ME = hemagglutinations with mercaptoethanol-treated samples. Positive HA: 7s peak.
clined in parallel with IgG monoclonal protein, which was no longer present 10 weeks after the onset of clinical hypersensitivity reactions. In Case 2 (Keflin hypersensitivity) the variations observed in total immunoglobulin values seen over a two week period are not explained, but the hemagglutinating titers of whole serum with penicillin G and Keflincoated red cells did show strikingdecrease in the convalescent phase of hypersensitivity reaction, coincidental with the disappearance of IgG monoclonal protein. Furthermore, hemagglutination performed with protein fractions and Sephadex G 200 peaks (Figure 2), proved that the hemagglutinating antibody belonged to the IgG class. In both cases, hemagglutination performed on protein fractions obtained by agar gel preparative electrophoresis showed a broad distribution of positivity with similar titers where M peak was localized. Repeated absorption experiments failed to show disappearance of the h4 peak when red cells sensitized with either penicillin, cephalothin or cloxacillin were used. COMMENTS
The cases of transient paraproteins reported in the literature are frequently associated with conditions in which an active antibody response might be expected. Acute and chronic infective illnesses are listed as a
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The American Journal of Medicine
major diagnosis in approximately 50 per cent of the cases. Among other conditions associated with this finding are collagen vascular diseases, cirrhosis of the liver, genetically determined immune deficiency states, successfully treated nonreticular neoplasms, leukemias, immunoproliferative disorders, prosthetic heart valve insertion associated with multiple blood transfusions and a miscellaneous group that includes hyperglobulinemic purpura and folic acid deficiency associated with sprue and cold sensitivity [12]. In a series of 154 cases of monoclonal gammopathies, nine patients had infective diseases such as chronic bronchitis, pneumonia, cellulitis, etc.-conditions undoubtedly associated with antibiotic use [13]. In a report on 59 patients with M components, a patient had the diagnosis of penicillin hypersensitivity, but no further details were given and it was not mentioned whether the paraprotein was transient or not [7]. Osserman’s review on plasma cell dyscrasias [6] included the case of an elderly man in whom an M-protein developed associated with pancytopenia and marrow plasmacytosis after gantrisin (3-4 dimethyl -5 sulfamido-isoxazole) therapy. Discontinuation of the drug resulted in prompt recovery from pancytopenia. About three months later, while the patient had a hepatitis B infection, the M protein was significantly decreased in quantity; approximately six months after, it was no longer detectable. Immunoelectrophoresis was not performed.
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The clinical course of the two patients presented here strongly suggests a relationship between the hypersensitivity reaction to drugs and the development of the monoclonal protein. The first patient had a serum sickness type drug reaction that started on the ninth day after a seven day course of oral cloxacillin therapy, without any objective evidence of infection. The acute but transient development of fever, urticaria, monoparesis, eosinophilia, albuminuria, associated with the presence of circulating immune complexes, hemagglutinating antibodies against pencillin and cloxacillin, and an IgG monoclonal paraprotein that gradually decreased in concentration until 10 weeks later when it was no longer present, suggest that this paraprotein was part of the hypersensitivity reaction. The second patient had a more complicated clinical problem. He had an aplastic marrow and a localized rectal abscess, and he received blood transfusions and multiple drugs before the appearance of the monoclonal protein. However, there is evidence linking the M protein with the drug hypersensitivity reaction: two months preceding the detection of the M peak and after having received semisynthetic penicillins (ampicillin and cloxacillin), a skin rash developed with histologic evidence of vasculitis. The second episode related to sodium cephalothin was more severe and associated with hematuria. IgG hemagglutinating antibodies appeared against penicillin and Keflin-coated red cells and decreased strikingly with the disappearance of the IgG monoclonal globulin. The occasional demonstration of free light chains in the serum, peripheral blood plasmacytosis and hypergammaglobulinemia provides evidence of increased antibody production in serum sickness reactions [14]. The development of a specific monoclonal antibody in response to antigenic stimulation is unusual, but it does occur in approximately 10 per cent of rabbits experimentally hyperimmunized with streptococcal group specific bacterial vaccines [l5] and with type III and VI
CARP10 ET AL.
pneumococcal polysaccharides [16]. The biologic process which gives rise to exceptionally high levels of electrophoretically homogeneous antibody is not clear. The mechanism leading to the stimulation of a restricted population of antibody producing cell clones may depend upon several factors, such as the chemical nature of the antigen or hapten, its physical state, the route of immunization and the genetic background of the immunized animals [l7]. In clinical situations in which an M protein has been identified as probably related to some antigenic stimulation, e.g., hepatitis B, pneumococcal infection, attempts to prove antibody specificity in the M protein have not usually been successful [7,12]. We were able to show evidence of an immune response against the benzylpenicilloyl group in the serums of both patients presented and against cloxacillin and cephalothin in Case 1 and Case 2, respectively. We could not demonstrate that the antibody activity was confined to the M band in either case, and the paraprotein was not removed from the serum by repeated absorptions with antigen-sensitized red cells. It is conceivable, however, that the M protein antibody activity is directed against a different antigen, such as occurs on a drug metabolite. The M band did disappear with cessation of the drug and clinical resolution of the hypersensitivity reaction. The recognition of the association between transient monoclonal paraprotein and hypersensitivity drug reactions widens the spectrum of manifestations of drug hypersensitivity and the causes of benign monoclonal gammopathies. Awareness of such an association may save some costly diagnostic investigations for some patients, provided an adequate follow-up is assured. ACKNOWLEDGMENT
We are grateful for the technical assistance of Susan Gaylord, Elizabeth Cormier and to Silvia Delli Colli and Linda Passalalpi for the preparation of the manuscript.
REFERENCES 1. Levine BB, Zolov DM: Prediction of penicillin allergy by immunological tests. J Allergy 43: 231. 1969. 2. Atkinson NF: Quantitative studies of the IgE and IgG immune response to penicillin administration in man [abstract). Abstracts of The American Congress of Allergy and Immunology meeting, New York, 1977. 3. Spath P, Garratty G, Petz L: Studies on the immune response to penicillin and cephalothin in humans. J Immunol 107: 860.1971. 4.
Gralnick HR, McGinniss M, Elton W, et al.: Hemolytic anemia associated with cephalothin. JAMA 217: 1193,1971. 5. McKenzie H. Parrat D, White RG: IgM and IgG antibody levels to ampicillin in patients with infectious mononucleosis. Chn Exp Immunol26: 214.1976. 6. Osserman EF: Plasma cell dyscrasias. Am J Med 44: 256, 1968. 7. Abramson N, Shattil SJ: M components. JAMA 223: 156, 1973.
June 1676
8. 9.
Levine BB, Fellner MJ, Levitska V: Benzylpenicilloyl specific serum antibodies to penicillin in man. J Immunol96: 707, 3966. Sobel AT, Borish VAA, Miiller-Eberhard HJ: Clq deviation test for the detection of immune complexes, aggregates of IgG. and bacterial product, in human serum. J Exp Med 142: 139.1975.
lo.
Scheidegger JJ: Une micromethode d’immunoelectrophorese. Int Arch Allergy Appl Immunol 7: 163.1955. 11. Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of Protein-Dye binding. Anal Biochem 72: 248, 1976. 12. Young VH: Transient paraproteins. Proc R Sot Med 62: 30, 1969. 13. Benbassat J, Fluman N. Zlotnick A: Monoclonal immunoglobulin disorders: a report of 154 cases. Am J Med Sci 271: 324.1976.
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14. 15.
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Freedman SO: Anaphylaxis and serum sickness, Chap. 3. Clinical Immunology (Freedman SO. Gold P, eds) New York, Harper & Row, Inc. 1976. Miller EJ. Osterland CK, Davie JM, et al.: Electrophoretic analysis of polypeptide chains isolated from antibodies in the serum of immunized rabbits. J Immunol 98: 716, 1967
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16.
17.
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Pincus JH, Jaton JC, Block KJ, et al.: Antibodies to type VIII pneumonococcal polysaccharides: evidence for restricted structural heterogeneity in hyperimmunized rabbits. J Immunol104: 1143,197O. Osterland CK, Miller EJ, Karakawa W, et al.: Characteristics of streptococcal group-specific antibody isolated from hyperimmune rabbits. J Exp Med 123: 599.1966.
JAIME DEL CARPIO, M.D. LUIS R. ESPINOZA, M.D.* STEVEN LAUTER, M.D.+ C. KIRK OSTERLAND, M.D. Montreal, Quebec, Canada
Two patients were studied in whom monoclonal (M) immunoglobulin G (IgG) proteins developed during the course of a serum sickness-like drug hypersensitivity reaction to cloxacillin (Orbenin@)and sodium cephalothin &eflin@),respectively. The clinical evidence and time sequence of events support this association. In both patients there was evidence of an active antibody response to the given antibiotic and to the benzylpenicilloyl group as well. However, protein fractions obtained by agar gel preparative electrophoresis failed to show a higher antibody concentration where the M peak was located, and absorption experiments performed with penicillin G-, cloxacillin- and cephalothin-coated red cells failed to absorb these M proteins. These transient paraproteins can be seen in association with antibiotic(s) administration in the context of a hypersensitivity reaction and do not apparently represent a specific immune response. drug reactions are common in modern medical practice, and they represent a costly aspect of health care. Penicillin is associated with a wide variety of different side effects. This drug and its derivatives often induce the formation of polyclonal immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies. The role of these antibodies in the production of hypersensitivity reactions is not clear. A protective “blocking antibody” function has been suggested for some penicillin antibodies [1,2]. In the case of cephalothin, they can be responsible for positive antiglobulin tests and possibly immune hemolytic anemia [3,4]. Ampicillin antibodies may bear important causal relation to the “ampicillin skin rash” frequently observed in patients with infectious mononucleosis [5]. The development of a monoclonal (M) protein in association with a drug reaction has been mentioned in the literature [6,7], but it is not clearly established. Recently, we have had the opportunity of seeing two patients in whom M proteins developed during the course of serum sickness-like reactions caused by a semisynthetic penicillin (cloxacillin) and a cephalosporin (sodium cephalothin], respectively. The M proteins disappeared after administration of the drug had been stopped, and clinical hypersensitivity reaction subsided.
Adverse
From the McGill University, Royal Victoria Hospital, Division of Allergy and Clinical Immunology, Montreal, Quebec, Canada. This study was supported by grants from the Canadian Arthritis and Rheumatism Society. Requests for reprints should be addressed to: Dr. Jaime Del Carpio, McGill University, Royal Victoria Hospital, Division of Allergy and Clinical Immunology, 687 Pine Avenue West, Montreal, Quebec, H3A 1Al Canada. Manuscript accepted September 27,1978. * Present address: Medical Center College of Medicine, Department of Internal Medicine, 12901 North 30th Street, University of South Florida, Tampa, Florida 33612 + Present address: Ballas Medical Office Center, 777 South New Ballas Road, Suite 305, St. Louis, Missouri 63141.
CASE REPORT Case 1. A 42 year old woman in previously good health who had no atopic background or history of penicillin hypersensitivity, received oral cloxacillin (1 g/day) for symptoms attributed to an ear infection. At the end of a week, she had fever followed by a generalized pruritic urticarial rash. Medication was stopped, but she remained ill. Five days later she became confused, and a mild weakness of the right arm was noted. She was admitted to another hospital
June 1676
The American
Journal of Medicine
Volume 66
1051
TRANSIENT
TABLE I
MONOCLONAL
PROTEINS
AND DRUG
HYPERSENSITIVITY-DEL
CARP10
ET AL.
Summary of Immunologic Laboratory Data
Case 1 11/30/76
IgG (850-1600 mgldl) IgA (85-305 mgldl) IgM (90-285 mgldl) Antipenicillin antibodies+ (hemagglutination) Immune complexes (Clq deviation 96) IgG monoclonal protein (immunoelectrophoresis) l
1212176
1,800
394 108
12117176
1,590
382 104 1:1,024
Case2 114/77 1,040
1:512
43 %
359 85 1:18 18%
4+ (SPE N”:,
216177
(&+#3)
(SPE ;;
7128176 1,200
1:18
330 580 1:2,048
819176 1,410
215 298 1:18
(SPE #5)
(SPE ;;
(SPE #7)
Similar results obtained with cephalothin (Case 1) and cloxacillin (Case 2) sensitized red cells.
where eltaminationshowed
a somnolent but oriented woman with a generalized urticarial rash. Her blood pressure was 110/80 mm Hg, pulse rate 80/min and a thorough ear, nose and throat examination was within normal limits. There was no lymphadenopathy or organomegaly. Neurologic evaluation confirmed a mild weakness in the right arm and diffuse hyperflexia. Laboratory data showed a normal hemogram except for an eosinophilia of 45 per cent (3,700 of a total 8,200 white blood cells/mm3). The erythrocyte sedimentation rate was 44 (Wintrobe). Urinalysis showed a normal sediment and a trace of protein. Serum immunoglobulins were (Table I]: immunoglobulin G (IgG) 1,600, immunoglobulin A (IgA) 394, and immunoglohllin M (IgM) 106 mg/dl. On zone electrophoresis one could discern a monoclonal protein band in the gamma area. Serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, lactic dehydrogenase (LDH), alkaline phosphatase and bilirubin levels were within normal limits. Tests for hepatitis B antigen and antibodies were negative. Cerebrospinal fluid examination disclosed no abnormalities. Two days after admission, the eosinophilia had decreased to 1,600 cells/mm3, and there was no albumin in the urine. Complement determinations were not performed. The patient was investigated neurologically to rule out the presence of an intracranial abscess. Twelve days after the onset of the skin rash, the patient was fully oriented, the weakness in the right arm was resolving, and there were only a few residual urticarial lesions. kepeat laboratory studies showed the complete blood count to b? within normal limits with no eosinophilia, total hemolytic complement (CH50), the third (C3) and fourth [C4]components of complement were within normal limits. Blood cultures and viral titers, obtained during the acute and convalescent phase, were all negative. Liver functions tests showed no abnormalities, serum immunoglobulins were IgG 1,590 mg/dl, IgA 382 mg/dl and IgM 104 mg/dl. On immunoele‘ctrophoresis IgG X paraprotein was identified. Urine electrophoresis did not show any abnormalities. Cryoglobulins, cryofibrinogen, cold agglutinins, rheumatoid factor and ahtinuclear antibodies were absent. Immune complexes and hemagglutination titers are shown in Table I. Repeat lumbar puncture showed a normal cerebrospinal fluid. Brain scan and computerized axial tomography were also within normal limits. The patient continued to show improvement, and she was discharged home asymptomatic. Serial serum protein electrophoresis and immunoelectrophoresis [Figure 1, Table I)
1052
June 1979 The American Journal of Medicine
showed gradual disappearance of M protein. After two and a half months the IgG paraprotein was no longer detectable by immunoelectrophoresis. Hemagglutination titers against penicillin G and cloxacillin-coated red cells also showed a declining titer, and immune complex measurements decreased [Table I). Skin tests with penicilloyl-polylysine (PPL) and penicillin G one month after the urticarial rash were negative. One year later the patient was well. Case 2. A 45 year old man was first admitted to the Royal
Victoria Hospital in October 1975 for investigation of aplastic anemia. He had had acute rheumatic fever at age 17, and a heart murmur had been discovered at age 27. There was no history of drug hypersensitivities. In December 1974. he underwent a right meniscectomy at another hospital where mild pancytopenia was noted. In April 1975,a diagnosis of aplastic anemia was made. Treatment with several courses of prednisone, oxymethalone, vitamin B1zand pyridoxine was unsuccessful, and he was referred to our hospital. The diagnosis was confirmed, but no etiology was determined. Among the normal studies was a serum protein electrophoresis. He was treated with transfusions of packed red blood cells and oxymethalone 150 mg/day. He was followed as an outpatient until May 1976 when he was readmitted with fever, a hematoma on the right thigh and acute left ethmoid and maxillary sinusitis. His hemoglobin level was 5.9 g/100 ml and his white blood cell count 1,800/ mm3 with 30 per cent neutrophils and 70 per cent lymphocytes. He was treated with packed red blood cells, platelets, cloxacillin 8 g/day, gentamicin 3 mg/kg/day and ampicillin 8 g/day. Ten days later, multiple papular erythematosus lesions developed over the elbows, and later in the groin and lower extremities. Antibiotic therapy was stopped, the lesions faded, and he remained afebrile. The clinical impression confirmed by skin biopsy was leukocytoclastic angiitis. The final admission occurred in July 1976 because of fever, bleeding from buccal ulcerations and a perirectal abscess. On examination blood pressure was llO/50 mm Hg, pulse rate lOO/min and temperature 40%. Erosions were noted on the palatal and buccal mucosa. There was 2 cm jugular venous distention, bibasilar rales and a grade 3/6 holosystolic apical murmur in the anterior axillary line with an opening snap. The liver was slightly enlarged, and there was tender left inguinal adenopathy. Rectal examination revealed a large left-sided mass just inside the rectum. His white blood cell count was
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7OO/mm3,his hematocrit value 13 per cent. Liver function tests yielded normal results except for a bilirubin of 1.8 mg/lOO ml. There was a moderate growth of Escherichia coli from the rectal lesion, but repeated blood cultures were negative. The patient was given a transfusion of packed red blood cells and platelets, and treated with digoxin, Keflin@ 4 g intravenously a day, clindamycin 600 mg intravenously every 6 hours and gentamicin 5 mg/kg/day. The perirectal abscess was drained. After eight days of treatment raised, purpuric lesions developed in the patient’s left groin which spread overnight to his abdomen, chest and back with severe pruritus. Keflin therapy was discontinued, but gentamicin and clindamycin were maintained at the same dose. The lesions became more confluent, and microscopic hematuria appeared. The patient was treated with Benadryl” (diphenhydramine hydrochloride) and over the next three days the lesions faded, and his hematuria cleared. A serum protein electrophoresis on the second day of the rash revealed a total protein of 7.1 g/loo ml, albumin 3.28 g/dl, alpha, globulin 0.29 g/dl, alpha2 globulin 0.95 g/dl, beta globulin 0.68 g/d1 and gamma globulin 1.85 g/d1 with a monoclonal band in the gamma region. Quantitative immunoglobulins were IgG 1.200 mg/dl, IgA 330 mg/dl and, IgM 560 mg/dl. Immunoelectrophoresis showed an IgG h paraprotein. Australia antigen and antibody were absent. Two weeks later the rash was completely gone, and repeat serum protein electrophoresis and immunoelectrophoresis no longer displayed a paraprotein peak (Figure 1). Quantitative immunoglobulins were IgG 1,410 mg/dl. IgA 215 mg/dl and IgM 298 mg/dl (Table I). Skin tests with penicilloyl-polylysine and penicillin G yielded negative results. The patient’s clinical condition deteriorated with the appearance of a mass in the upper right lung, repeated bouts of septicemia and increasing need for transfusions of red blood cells and platelets. One month later he died. Autopsy findings revealed acute pulmonary hemorrhage secondary to fulminant Pseudomonas pneumonia and invasive Aspergillosis of the lung, but no evidence of systemic vasculitis and no evidence of plasmocytosis. METHODS Penicillin hemagglutinating antibodies were determined by the technic described by Levine et al. [8]. A similar method was used for cephalothin [4] and cloxacillin. Circulating immune complexes, were measured by the Clq deviation technic of Sobel et al. [9]. In ourlaboratory the mean inhibition of normal serum is 2.7 per cent f 5 per cent (SD) with a range of 0 to 4 per cent. The test is capable of detecting as low as 15 pg/ml of aggregated IgG. Immunoelectrophoresis was performed by micromethod of Sheidegger [lo]. The identification and serologic typing of monoclonal proteins was carried out using antiserums specific for IgG classes and light chain subclasses. Serum proteins were separated by agar gel preparative electrophoresis. Fractions eluted by freezing and thawing were assayed by Bio-Rad protein assay [II]. The location of immunoglobulin classes was ascertained by setting up agar double diffusion plates of each of the eluted fractions against specific antiserums. Fractions were either assessed directly for antibody activity or, in some cases, several
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igure 1. Sequence of serum protein kectrophoresis (zone). Case 1 (cloxacillin hypersensitivity): electrophoresis 2-3-4-5. Case 2 (cephalothin hypersensitivity): electrophoresis 6-7. Electrophoresis 1: normal serum for comparison. (For dates see Table I.)
fractions were pooled and concentrated by pressure dialysis prior to further testing. Serum from the patient (Case 2) was filtered through a Sephadex@ G 200 column. The top tubes from each of the three G 200 peaks were pooled and concentrated prior to testing. Serum from both patients containing M peaks was used for absorption experiments. Red cells sensitized with penicillin G. cephalothin and cloxacillin were used to absorb patient’s serum at 37’C for 2 hours and overnight at 4% After each absorption, the serums were retested by zone electrophoresis and serologically. The absorptions were repeated sequentially eight times. RESULTS
protein electrophoreses (acetate cellulose], in both patients and a normal control subject for comparison, are shown in Figure 1. Laboratory results on both patients are summarized in Table I. In Case 1 (cloxacillin hypersensitivity) IgG, hemagglutinating titers against penicillin G and cloxacillin-coated red cells and immune complexes levels, deSerum
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-
HA
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+
I 200
I
150
I 250
I 300
’
ELUTION VOLUME (ml) Figure 2. Case 2: Sephadex G 200 column fractionation. HA = hemagglutination; HA-ME = hemagglutinations with mercaptoethanol-treated samples. Positive HA: 7s peak.
clined in parallel with IgG monoclonal protein, which was no longer present 10 weeks after the onset of clinical hypersensitivity reactions. In Case 2 (Keflin hypersensitivity) the variations observed in total immunoglobulin values seen over a two week period are not explained, but the hemagglutinating titers of whole serum with penicillin G and Keflincoated red cells did show strikingdecrease in the convalescent phase of hypersensitivity reaction, coincidental with the disappearance of IgG monoclonal protein. Furthermore, hemagglutination performed with protein fractions and Sephadex G 200 peaks (Figure 2), proved that the hemagglutinating antibody belonged to the IgG class. In both cases, hemagglutination performed on protein fractions obtained by agar gel preparative electrophoresis showed a broad distribution of positivity with similar titers where M peak was localized. Repeated absorption experiments failed to show disappearance of the h4 peak when red cells sensitized with either penicillin, cephalothin or cloxacillin were used. COMMENTS
The cases of transient paraproteins reported in the literature are frequently associated with conditions in which an active antibody response might be expected. Acute and chronic infective illnesses are listed as a
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major diagnosis in approximately 50 per cent of the cases. Among other conditions associated with this finding are collagen vascular diseases, cirrhosis of the liver, genetically determined immune deficiency states, successfully treated nonreticular neoplasms, leukemias, immunoproliferative disorders, prosthetic heart valve insertion associated with multiple blood transfusions and a miscellaneous group that includes hyperglobulinemic purpura and folic acid deficiency associated with sprue and cold sensitivity [12]. In a series of 154 cases of monoclonal gammopathies, nine patients had infective diseases such as chronic bronchitis, pneumonia, cellulitis, etc.-conditions undoubtedly associated with antibiotic use [13]. In a report on 59 patients with M components, a patient had the diagnosis of penicillin hypersensitivity, but no further details were given and it was not mentioned whether the paraprotein was transient or not [7]. Osserman’s review on plasma cell dyscrasias [6] included the case of an elderly man in whom an M-protein developed associated with pancytopenia and marrow plasmacytosis after gantrisin (3-4 dimethyl -5 sulfamido-isoxazole) therapy. Discontinuation of the drug resulted in prompt recovery from pancytopenia. About three months later, while the patient had a hepatitis B infection, the M protein was significantly decreased in quantity; approximately six months after, it was no longer detectable. Immunoelectrophoresis was not performed.
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The clinical course of the two patients presented here strongly suggests a relationship between the hypersensitivity reaction to drugs and the development of the monoclonal protein. The first patient had a serum sickness type drug reaction that started on the ninth day after a seven day course of oral cloxacillin therapy, without any objective evidence of infection. The acute but transient development of fever, urticaria, monoparesis, eosinophilia, albuminuria, associated with the presence of circulating immune complexes, hemagglutinating antibodies against pencillin and cloxacillin, and an IgG monoclonal paraprotein that gradually decreased in concentration until 10 weeks later when it was no longer present, suggest that this paraprotein was part of the hypersensitivity reaction. The second patient had a more complicated clinical problem. He had an aplastic marrow and a localized rectal abscess, and he received blood transfusions and multiple drugs before the appearance of the monoclonal protein. However, there is evidence linking the M protein with the drug hypersensitivity reaction: two months preceding the detection of the M peak and after having received semisynthetic penicillins (ampicillin and cloxacillin), a skin rash developed with histologic evidence of vasculitis. The second episode related to sodium cephalothin was more severe and associated with hematuria. IgG hemagglutinating antibodies appeared against penicillin and Keflin-coated red cells and decreased strikingly with the disappearance of the IgG monoclonal globulin. The occasional demonstration of free light chains in the serum, peripheral blood plasmacytosis and hypergammaglobulinemia provides evidence of increased antibody production in serum sickness reactions [14]. The development of a specific monoclonal antibody in response to antigenic stimulation is unusual, but it does occur in approximately 10 per cent of rabbits experimentally hyperimmunized with streptococcal group specific bacterial vaccines [l5] and with type III and VI
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pneumococcal polysaccharides [16]. The biologic process which gives rise to exceptionally high levels of electrophoretically homogeneous antibody is not clear. The mechanism leading to the stimulation of a restricted population of antibody producing cell clones may depend upon several factors, such as the chemical nature of the antigen or hapten, its physical state, the route of immunization and the genetic background of the immunized animals [l7]. In clinical situations in which an M protein has been identified as probably related to some antigenic stimulation, e.g., hepatitis B, pneumococcal infection, attempts to prove antibody specificity in the M protein have not usually been successful [7,12]. We were able to show evidence of an immune response against the benzylpenicilloyl group in the serums of both patients presented and against cloxacillin and cephalothin in Case 1 and Case 2, respectively. We could not demonstrate that the antibody activity was confined to the M band in either case, and the paraprotein was not removed from the serum by repeated absorptions with antigen-sensitized red cells. It is conceivable, however, that the M protein antibody activity is directed against a different antigen, such as occurs on a drug metabolite. The M band did disappear with cessation of the drug and clinical resolution of the hypersensitivity reaction. The recognition of the association between transient monoclonal paraprotein and hypersensitivity drug reactions widens the spectrum of manifestations of drug hypersensitivity and the causes of benign monoclonal gammopathies. Awareness of such an association may save some costly diagnostic investigations for some patients, provided an adequate follow-up is assured. ACKNOWLEDGMENT
We are grateful for the technical assistance of Susan Gaylord, Elizabeth Cormier and to Silvia Delli Colli and Linda Passalalpi for the preparation of the manuscript.
REFERENCES 1. Levine BB, Zolov DM: Prediction of penicillin allergy by immunological tests. J Allergy 43: 231. 1969. 2. Atkinson NF: Quantitative studies of the IgE and IgG immune response to penicillin administration in man [abstract). Abstracts of The American Congress of Allergy and Immunology meeting, New York, 1977. 3. Spath P, Garratty G, Petz L: Studies on the immune response to penicillin and cephalothin in humans. J Immunol 107: 860.1971. 4.
Gralnick HR, McGinniss M, Elton W, et al.: Hemolytic anemia associated with cephalothin. JAMA 217: 1193,1971. 5. McKenzie H. Parrat D, White RG: IgM and IgG antibody levels to ampicillin in patients with infectious mononucleosis. Chn Exp Immunol26: 214.1976. 6. Osserman EF: Plasma cell dyscrasias. Am J Med 44: 256, 1968. 7. Abramson N, Shattil SJ: M components. JAMA 223: 156, 1973.
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8. 9.
Levine BB, Fellner MJ, Levitska V: Benzylpenicilloyl specific serum antibodies to penicillin in man. J Immunol96: 707, 3966. Sobel AT, Borish VAA, Miiller-Eberhard HJ: Clq deviation test for the detection of immune complexes, aggregates of IgG. and bacterial product, in human serum. J Exp Med 142: 139.1975.
lo.
Scheidegger JJ: Une micromethode d’immunoelectrophorese. Int Arch Allergy Appl Immunol 7: 163.1955. 11. Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of Protein-Dye binding. Anal Biochem 72: 248, 1976. 12. Young VH: Transient paraproteins. Proc R Sot Med 62: 30, 1969. 13. Benbassat J, Fluman N. Zlotnick A: Monoclonal immunoglobulin disorders: a report of 154 cases. Am J Med Sci 271: 324.1976.
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14. 15.
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Freedman SO: Anaphylaxis and serum sickness, Chap. 3. Clinical Immunology (Freedman SO. Gold P, eds) New York, Harper & Row, Inc. 1976. Miller EJ. Osterland CK, Davie JM, et al.: Electrophoretic analysis of polypeptide chains isolated from antibodies in the serum of immunized rabbits. J Immunol 98: 716, 1967
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17.
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Pincus JH, Jaton JC, Block KJ, et al.: Antibodies to type VIII pneumonococcal polysaccharides: evidence for restricted structural heterogeneity in hyperimmunized rabbits. J Immunol104: 1143,197O. Osterland CK, Miller EJ, Karakawa W, et al.: Characteristics of streptococcal group-specific antibody isolated from hyperimmune rabbits. J Exp Med 123: 599.1966.