Originalia L. Timdr, J. Budai, G. Nyerges, R. Szigeti, I. Holt6s, I. Sonkoly

Progressive Vaccinia Immunological Aspects and Transfer Factor Therapy Summary: Progressive vaccinia is a rare arrd serious complication of smallpox vaccination. Depressed immune function can generally be found as an underlying disorder; thus adequate immuno-correction may be expected to be therapeutically effective. Humoral and cell-mediated immunity was repeatedly examined in one case throughout the course of the disease. Results indicated partial deficiency of cell-mediated immunity. No therapeutic effect was achieved by using human antivaccinia immunoglobulin and N-methylisatin /~-thiosemicarbazone. Transfer factor therapy was also attempted. Treatment with a non-specific transfer factor preparation was followed by a transitory clinical improvement. A specific transfer factor preparation given during the last month of life, however, had no therapeutic effect. The patient died on the 145th day after vaccination. Autopsy findings pointed to combined immune deficiency. Zusammenfassung: Progressive Vaccina: immunologische Aspekte und Ubertragungsfaktor-Therapie. Die progressive Vaccina ist eine seltene und schwerwiegende Komplikation der Pocken-Schutzimpfung. Als zugrunde liegende Sttirung findet man im allgemeinen einen herabgesetzten Immunmechanismus. Von angemessener Immunkorrektur kann daher therapeutische Wirksamkeit erwartet werden. In einem Fall wurde wiihrcnd des Verlaufs der Krankheit die humorale und zytergische Immunitiit mehrfach untersucht. Die Ergebnisse wiesen auf teilweise Defizienz der zytergischen Immunitiit bin. Durch Anwendung von menschlichem AntivaccinaImmunglobulin und N-Methylisatin-~-thiosemicarbazon lieB sich keine therapeutische Wirkung erzielen. Eine Behandlung mit i3bertragungsfaktor wurde ebenfalls versucht. Die Therapie mit einem unspezifischen t3bertragungsfaktor-Pr~parat hatte voriibergehende klinische Besserung zur Folge. Ein wiihrend des letzten Lebensmonats verabfolgtes spezifisches (3bertragungsfaktor-Priiparat zeitigte jedoch keine therapeutische Wirkung. Der Patient verstarb am 145. Tag nach der Impfung. Die Autopsiebefunde wiesen auf kombinierte Immundefizienz hin.

Introduction Progressive vaccinia (vaccinia necrosum) is an extremely serious complication of smallpox vaccination. It was invariably fatal before the introduction of immunotherapy (1). Vaccinia necrosum may develop after both primary vaccination and revaccination with an overall frequency of 0.7--0.9 per million vaccinations (2). During the seventies 7--8 persons died every year of smallpox vaccination complications in the USA. The mortality rate of postvaccinat encephalitis and vaccinia necrosum was about equal (3). The clinical picture of vaccinia necrosum is characterized by progressive necrosis of the vaccination site and, occasionally, by metastatic lesions. The vaccination lesion showed no evidence of healing even after the 15th day following vaccination (4). I n patients with this complication immunological deficiency can usually be

demonstrated. Keidan (5) was the first to report the lack of humoral immunity in this disease, while O'ConelI (6) and Hansson (7) observed isolated cellular immunodeficiency in patients with vaccinia necrosum. Hypogammaglobulinaemia or a malignant haematological disease was found as an underlying condition in the majority of cases (1, 2, 7, 8, 9). The first case of vaccinia necrosum with complete recovery was described by Barbero (10). The patient was given vaccinia immunoglobulin. The effectiveness of the vaccinia immunoglobulin therapy was confirmed by Kempe (1). Successful treatment with N-methylisatin-/3thiosemicarbasone (Marboran ®) was reported by Hansson (7). The administration of immunocompetent human lymphocytes was found effective in a number of cases (1, 6). Transfer factor therapy was suggested by Conolly (11) as early as 1962. However, the first attempt to use transfer factor in vaccinia was mentioned by Grob in 1975 (12).

Case Report The patient was a boy, 14 months old on admission. His birthweight was 3500 g. Apart from mild episodes of recurrent bronchitis, his history was not contributory. Previous vaccinations with BCG, triple vaccine Sabin and live measles (Leningrad-16 strain) vaccines caused no abnormal reactions. Smallpox vaccination with a Lister-strain calf-lymph-vaccine was carried out at i3 months of age. The initial course of the vaccination reaction was normal. The pustule, however, continued to enlarge and no hyperaemic area developed. No complete scabbing could be seen even on the 15th day after vaccination. On the 16th day a distant metastasis appeared followed by several others during the successive days. The child had no temperature. On the 29th day after vaccination he was given 2000 I U human antivaccinia immunoglobutin (HAVIG, "Human", Budapest). He was admitted to our department on the 33rd day after vaccination, on 28 July 1975. On admission the patient was in good general condition. His weight was 8500 g. At the vaccination site over the left deltoid Received: 6 June 1977 Dr. L. Timdr, Dr. 1. Budai, Dr. G. Nyerges, Central Municipal Hospital for Infectious Diseases, H-1097 Gyfili u 5/7, Budapest, Hungary; Dr.R. Szigeti, Second Department of Pediatrics, Semmelweis University Medical School, Budapest, H-1094, Tiizolt6 u 7/9, Budapest, Hungary; Dr. I. Hollds, National Institute of Hygiene, H-1966, Gy~tli u 4/6, Budapest, Hungary; Dr. I. Sonkoly, Fourth Department of Medicine, University Medical School, H-4012, Nagyerdei krt. 98, Debrecen, Hungary.

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L. Timdr et at.: Immunology and Therapy of Progressive Vaccinia area there was a lesion of 3-4 cm in diameter with raised edges and covered by a slightly oozing yellowish scab. Eighteen similar lesions, measuring 1-2 cm in diameter appeared on the face, the left arm, the lower extremities and on the mucous m e m b r a n e of the mouth. Several other lesions of vesicular or pustular character without hyperaemic area were also noted. There was a BCG scar on the left shoulder above the smallpox vaccination site. A lymphnode was palpable in the left armpit. Laboratory examinations showed increased erythrocyte sedimentation rate (40 mm/hr), anaemia, lymphocytopenia and moderate eosinophilia. No thymus was visible on the chest X-ray. The child was given two h u m a n antivaccinia immunoglobulin injections, b o t h 2000 ILl. Thiosemicarbazone (Marboran ®, Borroughs Wellcome Co, London) treatment was introduced with a daiIy dose of 50 mg/kg body weight. Therapy was supplemented by antibiotics and vitamins. Further data of the therapy are given in Figure 2. On the 40th day after vaccination a new lesion appeared on the face while earlier lesions continued to enlarge. On the 47th day after vaccination treatment with transfer factor was introduced. The transfer factor preparation (Transferfaktor Nr. 178, Kantonsapotheke, Zurich) was obtained through the I m m u n o firm, Vienna. The patient's condition improved remarkably during the subsequent days. The scabs dried and fell off. Seven lesions healed, ten others became nearly epithelized although small slightly indurated focuses still remained on their edges. However, three of the lesions (on the face, on the left upper arm and on the right foot) showed further spreading in spite of the transfer factor treatment repeated one week later (Figure 1). There was no remarkable change during the subsequent six weeks in spite of the introduction of a new antiviral drug, methisoprinol (Isoprinosine, Newport Pharmaceuticals International, Costa Rica) in a total dose of 6.0 g. Three lesions showed small progression while the others remained practically, unchanged. A considerable worsening ensued around the 100th day after vaccination. Active focuses showed further enlargement and new metastatic lesions appeared on the so far intact parts of the skin. The rapidly extending facial lesions covered twothirds of the skin of the face. The eyelids and the lips were also involved. The lesion on the upper lip and the one inside

Figure 1: Progressive vaccinia on the 52rd day after vaccination. the mouth fused. Feeding the patient was difficult and his general condition rapidly deteriorated. He became feverish. The temperature curve assumed a septic character. Several transfusions of fresh blood were given because of the ensuing anaemia and atrophy, without any sign of "graft versus host" reaction. ParenteraI feeding was introduced.

Abs, lymphocyte 2 5O0 count

• : Secunderlesions

2 000 15O0

I Wooinouoil I

,0oo'

Admission..5~ 0

,,

J

~ ' ' 0 16 3 3033 ~0 Therap_£ TF: Howg : 2000 U daily

Morboran

~'7 5~

I_.I L . ~

:

I

I

o o iu,

I

I

I

I

,

-

1~5 Postmccinattbn dw

HHHHHH L_.I L..-I

~OOmg doily Isoprinosine: kOg doily Idox-&/dine eye drops

A tibioU s :

I

100 106 113 723 137

'

I

I Erythro

-

I Vibra- F Gentomycin I,

Figure 2: The changes in the absolute number of lymphocytes and the therapy during the course of the disease.

150

Infection 6 (1978) Nr. 4

L. Timdr et al.: Immunology and Therapy of Progressive Vaccinia On the 106th day after vaccination treatment with a specific transfer factor preparation was initiated. Transfer factor was produced from the pooled leucocytes of freshly revaccinated donors according to the method of Grob (13). The extract obtained through the rapid freezing and thawing of the cells was dialysed and lyophilised, One dose of the preparation corresponded to 5 X 10 ~ cells. F o u r injections of transfer factor were given at one week intervals. F r o m the 120th day after vaccination 2000 I U h u m a n antivaccinia immunoglobulin was administered daiIy. N o n e of these treatments helped the patient. He became unconscious and died of circulatory insufficiency on the 145th day after vaccination.

The results of the immunological investigations are presented in Tables 1 and 2. Table 1: Tests of cell-mediated immunity. (Transfer factor

was administered on the 47---49th, 54-56th, 106th, ll3th, 123th and 1Mst days after vaccination.) Day Percentage after spontaneous vaccina- rosettetion forming cells (no./100 Iymphocytes)

Laboratory Findings F r o m the fresh pustules vaccinia virus was isolated on the chorioallantoic m e m b r a n e of chick-embryos on f o u r occasions. The first isolations yielded pocks similar to those produced by the Lister strain. F r o m the materials taken during the third and fourth month, however, haemorrhagic pocks developed. The absolute lymphocyte count was low throughout the period observed. Occasional changes correlated welt with the clinical course of the disease (Figure 2). On admission the absolute count of lymphocytes was 1240/#1. At the time of the appearance of new lesions this value was 640/#1, and in the course of the clinical improvement the value increased to 2100//A. During the period when the disease was slowly progressing the absolute count of lymphocytes was invariably low about 1200/~1. A transitory rise followed by a steep fall was noted at the time of rapid progression, o n the day of the death an extremely low count (320//A) was found.

37

38

44

25

5I

20

61

20

72 85

31 30

111 121

23 20

PHAstimulation index ~

Leukocyte Delayed migration type skin index reactions

6.4 (control 7.2, 8.5) 5.9 (control 6.9, 7.9)

1.54

negative b negative b

2.7 (control 14.6)

0.54 negative c

0.42 °

negative c

count per minute stimulated: count per minute nonstimulated (controls healthy children of same age) b with PPD ° with P H A

Table 2: Tests on humoral immunity. (Results are influenced by the human antivaccinia immunoglobuIin given on the 29th, 34th and 39th days.) Results Day after vaccination

Tests 34

42

71

85

119

Percentage surface

IgG

receptor-bearing

IgM 14

2

IgM / 8

cells (B-cells)

IgA

IgA

2

IgG

(no./100 lymphocyte) Serum immunoglobulin

IgG

1134

1476

1476

880

concentrations

IgM

151

128

68

58

(mg/100 mI)

IgA

161

119

137

120

Titers of the specific antibodies: Vaccinia neutralising antibody Isohaemagglutinin

1:16 1 : 256 0

anti-A anti-B

Diphtheria antitoxin (IU)

0.03

Staphylococcus alpha-antitoxin (IU)

0.24

I :25

1:64 0 0.03 0.19

Measles haemagglutinin inhibition

1:20

1:20

0

Rubella haemagglutinin inhibition Polio type 1 type 2 type 3

1:32 1:12 1 : 128 1:64

1:16

0

Pertussis agglutinin

Echerichia coli 026

1 : 128

1:64 1:16

1:4

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L. Timdr et al.: Immunology and Therapy of Progressive Vaccinia T cells were estimated using the spontaneous rosette forming technique (14). The stimulation index was determined by the measurement of 3H-thymidine incorporation (15). Leucocyte migration inhibition by phytohaemagglutinin (PHA) and purified protein derivate (PPD) was tested. B cells were quantitated by staining for surface immunglobuIin with the immunfluorescent method (16). Skin tests were carried out using PPD and purified PHA. The serum immunglobulin concentrations and the specific antibody titers against different antigens were also measured. Necropsy revealed considerable hypoplasia of the thymus and the lymphatic system. In the unusually small thymus no Hassal bodies were found and no true lymphocytes could be recognized. Lymph node was devoid of true folliculi and germinal centres. In the spleen and intestine mainly reticular elements were seen instead of the lymph follicles, however, some lymphocyte-like but larger cells were also found. There were no other pathological alterations.

Discussion In our patient the clinical picture of vaccinia necrosum was characterized by the continuous progression of the intentional vaccination and appearance of new lesions. The slowly progression process lead to the exhaustion of the organism and finally to death within 21 weeks. The results of the investigations suggested that the humoral defence mechanism could not be seriously impaired at the time of the smallpox vaccination. However, the results may have been influenced by the repeated human antivaccinia immunoglobulin injections given previously. Unfortunately, we could not carry out immunological investigations at the beginning of the disease. However, the nearly normal immunoglobulin levels measured six weeks after the human antivaccinia immunoglobulin injection, the normal level of anti-A isohaemagglutinin as well as the presence of antivaccinia antibodies twelve weeks after the human antivaccinia immunoglobulin injection all indicated that the patient's humoral immunity was not severely impaired. The decrease of antibacterial and antiviral titers might have been caused either by the rapid elimination of antibodies given in human antivaccinia immuoglobulin or by the progressive exhaustion of the immune system as a consequence of the long lasting virus infection. On the other hand, the low titer of diphtheria antitoxin and that of the Staphylococcus alpha antitoxin as well as the lack of measles and rubella antibodies on the l l 9 t h day pointed to a certain extent of humoral insufficiency. Tests for cell-mediated immunity revealed an impairment of the cellular defence functions. Although the P H A stimulation test yielded approximately normal values in two out of three occasions, the low percentage of rosette forming lymphocytes and the negative skin tests indicated a definite depression of the cellular immunity. The negativity of P H A skin tests can be considered as one of the most reliable indicators of the impairment of cellular immune mechanism (17). N o inhibition of leucocyte migration by PPD was seen on the 44th day after vaccination. Later on, however, the leucocyte migration inhibition test became positive. It seems very likely that

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Infection 6 (1978) Nr. 4

the change was brought about by the administration of transfer factor on the 47th day. The absolute lymphocyte count was invariably low during the course of the disease, from which it woutd appear that there was a permanent deficiency in the effector cells of cellular immunity. The fact that occasional changes in the lymphocyte count correlated with the changes in the clinical course also speaks for the pathogenic role of the impairment of cellular immunity. In the case history of the patient no indication of a depressed immune function could be found. Results of investigations carried out during the course of the disease showed a light impairment of immune function. Although the failure seemed to be primarily of a cellular type, one could not preclude the possibility of a combined immune deficiency, either. Finally autopsyrevealed the picture of serious combined immune deficiency. Treatment with antiviral drugs (thiosemicarbazone) and immunotherapy (vaccinia immunoglobulin) have been recommended for the treatment of vaccinia necrosum (4). During the time the antiviral drug thiosemicarbazone was given new lesions appeared and methisoprinol treatment also proved to be a failure. The progression was n o t stopped by vaccinia immunoglobulin. A high dose given during the last weeks of the disease was also unsuccesful. Since in our case the deficiency of cell-mediated immunity was thought to be an underlying disorder, therapeutic results were expected from drugs having effect on cellular immunity. Since the first therapeutic attempts by Lawrence (18), transfer factor was tried in several disorders with impaired cell-mediated immunity (20, 21). The medical centers preparing the transfer factor used in ot~ patient reported good therapeutic effect in certain disorders (12, 19). The transfer factor preparation given in daily doses from the 47th to the 49th day had a remarkable effect, which, however, could not be repeatedly produced in spite of the reiterated treatment one week later. The beneficial effect of transfer factor manifested itself in a definite slowing down of the process and could be observed for about two months. After this period a rapid progression of the disease ensued. Transfer factor treatment was therefore repeated using a specific preparation. But this time, no therapeutic effect whatsoever could be observed. The failure of specific transfer factor treatment might be ascribed to the increasing shortage of antigen sensitive T cells due to continuous virus invasion (22). In this case therapeutic effect might have been expected only from thymus transplantation (20, 22) or from thymosin (23). Thus, the correct therapy would have been thymosin treatment followed by the administration of transfer factor and, in case of ineffectiveness, thymus transplantation. Unfortunately, the results of the immunological examinations, especially of those carried out at the early stage of the disease did not indicate the need for such therapeutical measures,

L. Timdr et aI.: Immunology and Therapy of Progressive Vaccinia Acknowledgements We thank the Institute for ~erobacteriological Production and Research H U M A N , the National Institute of Hygiene, the National Institute of Haematology and Bloodtransfusion, Budapest, and the Firm Immuno, Vienna, for helping us in the laboratory investigations and therapy.

Literature 1. Kempe, C. H.: Studies on smallpox and complications of smallpox vaccination. Pediatrics 26 (1960) 176-189.

2. Lane, J. M., Ruben, F. L., Neff, J. M., Millar, J. D.: Complications of smallpox vaccination, 1968. N. EngL J. Med. 281 (1969) 1201-1208.

3. Karzon, D. T.: Smallpox vaccination: The end of an era. Acta Med. Scand. Suppl. 576 (1975) 29-38.

4. GoIdstein, ]. A., Neff, ]. M., Lane, J. M., Koplan, J. P.: Smallpox vaccination reactions, prophylaxis and therapy of complications. Pediatrics 55 (1975) 342-347.

5. Keidan, S. E., McCarthy, K., Haworth, J. C.: Fatal generalized vaccinia with failure of antibody production and absence of serum gamma globulin. Arch. Dis. Child. 28 (1953) 110-116.

6. O'ConeIl, C. 1., Karzon, D. T., Barron, A. L , Plaut, M. E., All, V. M.: Progressive vaccinia with normal antibodies. A

10. Barbero, G. J., Gray, A., Scott, T. F., Kempe, C. H.: Vaccinia gangrenosa treated with hyperimmun vaccinal gamma globulin. Pediatrics 16 (1955) 609. 11. Conolly, ]. H., Dick, G. W. A., Field, C. M. B.: A fatal case of progressive vaccinia. Br. Med. J. 1 (1962) 1315-1317. 12. Grob, P. J., Reymond, J. F., Hiicki, M. A., Frey-Wettstein, M.: Transferfaktor. Miinch. Med. Wschr. 117 (1975) 517-524. 13. Grob, P. J., BIiiker, F., SchuIz, K. H.: Immunfunktion und Transferfaktor. Dtsch. Med. Wschr. 98 (I973) 446-451. 14. Wybran, J., Carr, M. C., Fudenberg, H. H.: The human rosette4orming cell as a marker of a population of thymusderived cells. J. Clin. Invest. 51 (1972) 2537-2543. 15. Rdvdsz, T , Szigeti, R., Schuler, D.: The role of serum factors in the lymphocyte transformation test of children with acute leukaemia. Acta Paediatr. Scand. 63 (1974) 715-720. 16. Johnson, G. D., Halborow, E. J.: Immtmfluorescence. In: Weir, D. M. (ed.): Handbook of experimental immunology, Vol. 1: Immunochemistry, p. 18-20 Blackwell Sci. Publ., Oxford 1973. 17. Lawlor, G. J., Stiehm, E. R., Kaplan, M. S., Dharmendra, M. D., Sengar, P. S., Terasaki, P. I.: Phytohemagglutinin skin test in the diagnosis of cellular immunodeficiency. J. Allergy Clin. Immunol. 52 (1973) 31-37.

case possibly due to deficient cellular immunity. Ann. Intern. Med. 60 (1964) 282-289.

18. Lawrence, H. S.: Transfer factor. Adv. Immnnol. 2 (1969) 195-266.

7. Hansson, 0., Johansson, S. G. 0., Vahlquist, B.: Vaccinia

19. Balogh, E., Szegedi, Gy., Karrnazsin, L., Szab6, G., Szabolcsi, M.: Immunologischer Aspekt der Inklination fiir

gangrenosa with normal humoral antibodies: a case possibly due to defficient cellular immunity treated with N-methylisatin-fi-thiosemicarbazone (compound 33T57 Marboran). Acta Paediatr. Scand. 55 (1966) 264-2:72.

8. Lane, Y. M., Millar, J. D.: Risk of smallpox vaccination complications in the United States. Am. J. EpidemiC. 93 (1971) 238-240.

9. Tympner, K. D., StickI, H., PrechteI, K., Flamm, U.: Vaccinia generalisata progressiva bei AntikSrpermangel-Syndrom (prim~ire kombinierte immunologische Defizienz). Msehr. Kinderheilk. 124 (1976) 684-688.

mykotische Infektionen. Dermatol. Mschr. 162 (1976) 100-103. 20. Bach, F. H., Good, R. A. (ed.): Clinical immunbiology Vol. II. Academic Press, New York, 1974. 21. Ascher, M. S., Gotttieb, A. A., Kirkpatrick, C. H. (ed.): Transfer factor: Basic properties and clinical applications. Academic Press, New York 1976. 22. Lawrence, H. S.: Personal communication, by letter (1977). 23. Wara, D. W., Amman, A. J.: Thymic cells and humoral factors as therapeutic agents. Pediatrics 57 (1976) 634-646.

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Progressive vaccinia: immunological aspects and transfer factor therapy.

Originalia L. Timdr, J. Budai, G. Nyerges, R. Szigeti, I. Holt6s, I. Sonkoly Progressive Vaccinia Immunological Aspects and Transfer Factor Therapy S...
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