Recent developments in immunomodulatory therapy

THE JOURNAL OF

ALLERGY AND

CLINICA L IMM UNOLOGY VOLUME 75

NUMBER 4

Continuing Medical Education This continuing medical education self-assessment program is sponsored by The American Academy of Allergy and Immunology and supported by a grant-in-aid from Fisons Corporation.

Recent developments in immunomodulatory therapy Emil J. Bardana, Jr., M.D. Portland, Ore.

The remarkable gains in basic immunologic knowledge have outpaced improvements in available agents or techniques that can be used to clinically modify the immune response. Many biologic and synthetic immunomodifiers, adjuvants, and drugs with both specific and/or nonspecific effects on immunity have been investigated. Many of the therapies currently available are still in a developmental stage. However, it appears very likely that the extraordinary gains of the next decade will come in the sphere of "immunomodulation." The development of monoclonal antibodies has opened several avenues where immunochemistry can be applied to modify biologic responses. One concept enjoying a revival in recent years is that of the "magic b u l l e t " - - a n idea similar to that initially proposed by Paul Ehrlich nearly a century ago. The modem bullet will use the high selectivity of monoclonal antibodies to direct cytotoxic agents to abnormal tissue. There is a need to develop agents that can selectively enhance or suppress one specific class or subclass of immunocyte. It appears probable that when the mechanism of action of the many immunomodFrom the Division of Immunology,Allergy and Rheumatology, Oregon Health Sciences University, Portland, Ore. Reprintrequests: E. J. Bardana,Jr., M.D., 3181 S. W. SamJackson Park Rd, L-329, Portland, OR 97201.

Abbreviations used

ISG: HSG: AIDS: CMV: MHC: HLA: SCID: GVHD: DLE: TF: RNA: CMC: ISO: BCG: ITP: TTP:

Immune serum globulin Hyperimmune serum globulin Acquired immunodeficiency syndrome Cytomegalovirus Major histocompatibility complex Human leukocyte antigens Severe combined immunodeficiency Graft-versus-host disease Dialyzable leukocyte extracts Transfer factor Ribonucleic acid Chronic mucocutaneous candidiasis Isoprinosine, lnosiplex, Methisoprinol Bacillus Calmette-Gu~rin

Immune thrombocytopenic purpura Thrombotic thrombocytopenic purpura

ulatory agents is more completely understood, specific sequential combinations will be apparent to achieve a desired clinical result. This article will highlight recent developments in immunomodulatory therapies and will discuss them under three broad headings: (1) agents that facilitate a normal immune response, i.e., immunorestoration, (2) agents that stimulate the immune response, i.e., immunostimulation, and (3) non-

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cytotoxic agents that suppress immune response, i.e., immunosuppression.

IMMUNORESTORATION The concept of restoring a dysfunctional or failing immune system has many parallels to other organ systems in the body. The closest parallels in immunology relate to immunoglobulin replacement, interferon therapy, administration of thymic hormones, and bone marrow and fetal liver transplantation. In addition to the latter, there are a variety of other T cell-produced soluble mediators, lymphokines, that have great relevancy to immunologic reconstitution but that are beyond the scope of this review. The same could be said of the recently identified interleukins that transmit growth and differentiation signals among immunocytes.

Development of gamma globulin Treatment with immunoglobulin is on the basis of the principle of passively immunizing patients with primary and secondary humoral immunodeficiency disorders with exogenous antibody. This concept was developed by Von Behring and Kitasato ~ and Ehrlich 2 who used immune horse serum to treat patients with diphtheria and tetanus. The serum sickness reactions that resulted on repeated exposures stimulated the search to secure human serum for the same purpose. The first major advance came in 1933 when ammonium sulfate precipitation of serum was used to isolate gamma globulins. 3 This was supplanted by cold ethanol fractionation. 4 By examining multiple fractions derived by this technique, IgG activity was found to reside in fraction II, and this material became known as ISG. ISG is produced either from a large pool of healthy donor plasma (polyvalent ISG) or from a limited number of convalescent or actively immunized subjects yielding products known as HSG.

Intramuscular ISG The availability of standardized preparations of ISG provided the obvious vehicle to treat patients with primary B cell deficiencies, i.e., congenital and acquired agammaglobulinemia, the Wiskott-Aldrich syndrome, and the hyper-IgM syndrome. In the initial phases of replacement, ISG had to be administered by intramuscular route because of the frequency of severe reactions if it was infused intravenously: 6 Although extremely helpful in the treatment of these disorders, intramuscular ISG was not an ideal therapy. The recommended dose of 0.7 ml/kg of 16.5 gm/dl preparation on a monthly basis was designed to maintain serum IgG levels above 200 mg/dl. These recommendations are arbitrary and are currently being

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reevaluated with a goal of providing guidelines for optimal doses in individual patients. Intramuscular injection of ISG is painful caused by the amount of very viscous material that is delivered to a limited muscle mass. Injections can result in nerve and muscle damage as well as formation of sterile abscesses. These physical drawbacks are often accentuated during episodes of sepsis since IgG metabolism is accelerated and since more frequent ISG doses are required. In addition, serum antibody levels achieved by this route are unpredictable because (1) absorption from muscle is slow and may take 4 to 7 days to complete, (2) proteolytic enzymes may degrade variable amounts of ISG at the injection site, and (3) once ISG is adsorbed, there is considerable variation of its circulating half-life. 7 Intramuscular injection of ISG can be associated with a variety of systemic vasomotor symptoms believed to be secondary to complement activation, eg., nausea, vomiting, anxiety, cyanosis, flushing, facial swelling, abdominal cramping, and loss of consciousness. Anaphylactic reactions may result from (1) trace lgA present in ISG when it is administered to hypogammaglobulinemic patients with selective lgA deficiency who make IgG and/or IgE antibody to IgA, and (2) generation of antiatlotypic antibodies in hypogammaglobulinemic patients with formation of immune complexes.

Plasma infusion The second phase of IgG replacement started in the mid-1960s with emphasis directed to infusion of fresh plasma rather than injection of ISG. 8 This had great clinical appeal in that infusion of large amounts of all immunoglobulin classes could be achieved fairly painlessly. Selected volunteer donors could be immunized to supply specific antibodies needed by the patient. Serum IgG levels could be raised to near normal levels. However, plasma infusion on a regular basis also has its drawbacks. Even with sophisticated radioassays, undetectable amounts of hepatitis virus can be transmitted, and there exists no screening test for the carrier state in non-A and non-B hepatitis. There is about 1% risk of infection with this virus with every unit of fresh or frozen plasma infused. Transmission of AIDS is also possible and discussed in greater detail below. These risks can be minimized by the use of the "buddy" system, i.e., several carefully selected individuals are screened for hepatitis and selected as donors. Occasional anaphylactoid-like reactions of unknown cause are noted with plasma infusion and can be abbrogated by use of small doses of antihistamine or aspirin. For plasma therapy, a minor cross match of the donor's plasma with the patient's erythrocytes should be negative. For patients with severe

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defects in cellular immunity, the plasma should be irradiated to eliminate viable donor lymphocytes capable of inducing GVHD.

Intravenous ISG To preserve the biologic function of IgG, the World Health Organization (WHO) recommended that intravenous ISG be modified as little as possible, contain at least 90% intact IgG with no fragments, and have a normal distribution of IgG subclasses. 9 During the last three decades scientists have been exploring a variety of chemical alterations of ISG that would permit safe and effective intravenous use. These have included enzyme treatment,~~ 13-propiolactone treatment, ,2 reduction and alkylation, t3. ,4 sulphonation, ~ addition of polyethylene glycol, 16 brief treatment at pH 4.0 with traces of pepsin, s' t7 and stabilization with albumin. ~ Only the latter three modifications yield a product consistent with WHO guidelines ~7' ~9; nevertheless, several preparations have been extensively studied in this country and have proved to be effective in the management of primary humoral immunodeficiency disorders. The most extensively studied preparation in this country is that manufactured by Cutter Biological Di'vision of Miles Laboratories, Inc., Emeryville, Calif., i.e., Gamimune. This is prepared by mild disulfide reduction and alkylation of ISG to prevent globulin aggregation. Ten percent maltose and 0.1 mol/L of glycine are added to stabilize the preparation. It contains at least 90% immunoglobulin with traces of IgA and is marketed as a 5% solution. It was released for general use in 1981, but the first therapeutic observations were reported nearly a decade ago 2~ and subsequently augmented by extensive studies at a variety of centersfl t-23 Buckley 22 compared equal amounts of intramuscular and intravenous ISG delivered at doses of 100 mg/kg in primary humoral deficiency disease and failed to demonstrate significant differences in serum IgG. Ochs et al. 23 made similar observations by use of three separate protocols with doses ranging between 100 and 150 mg/kg. Pirofsky et al. 24reported significantly higher serum IgG levels with use o f intravenous ISG at doses of between 150 and 200 mg/ kg compared to intramuscular dosing at 100 mg/kg on a monthly basis. These investigators also observed a rapid loss of serum IgG in the first 7 days after infusion with more than 60% decrease in serum IgG. After 7 days the rate of breakdown followed an exponential curve with preinfusion levels being reached at between 3 and 4 wk. 25 This was consistent with prior reports of a 22-day half-life with intravenously administered ISG at doses of 100 to 150 mg/kg. 26 The reason for the rapid drop within the first week

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after infusion is unclear, but possible explanations include rapid equilibration in extravascular fluids, rapid clearance of denatured or aggregated lgG, and formation and clearance of immune complexes composed of infused IgG and circulating in vivo antigens. Marked individual variations in the rate of IgG catabolism were noted by all investigators. Pirofsky et al. noted about a third of patients receiving infusions of 150 to 200 mg/kg had a slower rate of IgG breakdown with a tendency to plateau at increased preinfusion levels. 24' 25 Most patients would not have this staircase effect and would revert to baseline preinfusion levels within 3 to 4 wk. Additional studies with the use of higher doses of 500 mg/kg amplified these initial observations in about the same percentage of patients. 27 As a result, serum IgG levels above 350 mg/dl could be maintained for 8 to 12 wk in a limited number of patients. Ochs et al. 28 also noted this stepwise increase in both post- and preinfusion levels of serum IgG by use of doses of 400 mg/kg. These investigators noted this staircase phenomenon in most patients receiving these higher doses of ISG. However, they did not observe it in any patient receiving doses of 100 mg/kg. On the average serum lgG levels rose by about 250 mg/dl for each 100 mg/kg of ISG infused. 28Marked individual variation was demonstrated by Buckley 22after carrying out half-life studies. In four patients the IgG half-life varied between 18 V2 and 76 days that translated into maintenance infusion doses ranging between 50 and 300 mg/kg per month. 22 In comparing 100 mg/kg per month doses of Gamastan (Cutter Biological Division) intramuscularly with intravenous infusion of the same dose of Gamimune, B u c k l e r : observed a slight increase in the incidence of minor upper respiratory infection with intravenous administration. Ochs et al. 23 made similar observations by use o f identical doses of ISG intramuscularly and intravenously. Comparing Gamimune infusion at 150 to 200 mg/kg per month with Gamastan 100 mg/kg per month intramuscularly, Pirofsky et al. 24 observed a significant reduction of acute respiratory infection with intravenous ISG. Differences in the infection rate among these investigators was attributed to variation in the infusion dose of ISG. Nevertheless, there remains considerable doubt regarding the value of higher doses of intravenous ISG to achieve better protection against infection. Sorensen and Polmar 29 failed to observe a reduced infection rate by the use of infusion doses of 250 mg/kg per month. Similar observations were made by Berger et al. 3~ who continued to record recurrent infections in pregnancy despite normal serum lgG maintained by frequent subcutaneous infusion. Even at doses of

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400 mg/kg per month, Ochs et al. 28 did not demonstrate significant improvement in development of acute infection when these doses were compared to doses of 100 mg/kg per month. There was agreement that intravenous infusion was equally protective against infection as the intramuscular route and that it clearly was the preferred modality among patients.22-24 Most investigators who use intravenous ISG have reported a heightened sense of well being in their patients. The use of high-dose intravenous ISG in life-threatening infections in patients with primary immunodeficiency syndromes have yielded mixed results. Mease et al. 3~ successfully treated a patient with ECHO virus meningoencephalitis with six daily infusions of 160 mg/kg for 6 days followed by 160 mg/ kg every 10 days. Serum IgG levels were maintained at normal levels, the virus disappeared, and chronic symptoms of infection cleared entirely. On the other hand, Buckley22 used a similar approach in two patients with the same type of infection without success. Cunningham-Rundles et al. ~7recently reported their experience with an extensively studied European intravenous preparation, i.e., Sandoglobulin. The material is produced by Sandoz Laboratories, Basel, Switzerland, with the gamma globulin being provided by the Swiss Red Cross in Bern, Switzerland. Cohn fraction II is briefly brought to pH 4.0 by the addition of HC1 in the presence of traces of pepsin and then neutralized and lyophilized. The material is higher than 90% 7S IgG on reconstitution, contains less than 4% dimers, contains less than 0.3% aggregates, and lgG subclasses are present in normal proportions. ~9 IgA is present in low concentrations averaging 0.72 mg/ml. These investigators treated 21 patients with primary immunodeficiency by use of two infusions of 150 mg/kg, two infusions at 200 mg/kg all at 2-week intervals, followed by infusions of 300 mg/kg at 3week intervals. Three patients were withdrawn from the study, two because of lack of benefit and one because of preexisting anti-IgA antibody. All but one patient achieved preinfusion serum IgG levels that were within one standard deviation of the mean normal value after 3 mo of treatment. Patients with associated T cell defects derived as much improvement as those with normal T cell function. Certain infections including upper respiratory tract infections, bronchitis, and pneumonia were significantly reduced in the second 6 mo of therapy, whereas the incidence of sinusitis, arthritis, and conjunctivitis appeared to decrease immediately. Aside from the obvious differences in dose, the authors believed their lower infection rate as compared to prior studies of similar size was related to the manner in which the ISG was prepared.

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Adverse reactions The incidence of adverse reactions to infused ISG varies considerably. The incidence is much higher in patients with primary antibody deficiency syndromes as opposed to nondeficient patients with severe infections, z5 Adverse reactions can be divided into two broad categories, i.e., mild to moderate vasomotor reactions and severe anaphylactic reactions. In patients with antibody deficiency syndromes, Gamimune has been associated with an approximate 15% incidence of mild vasomotor reactions at doses of 150 mg/kg. 25 Ochs et al. 28 compared side effects during infusions of either low or high dose Gamimune. The reactions ranged from mild to moderately severe and were similar with doses of 100 mg/kg or 400 mg/kg. An almost equal number of reactions were observed during ISG infusion as were noted 2 to 6 hours after infusion. Adverse symptoms included chills, flushing, headache, abdominal cramps, nausea, emesis, muscle pain, chest tightness, anxiety, and dizziness. The frequency and characteristics of side effects between the low-dose and high-dose groups were 18% and 15%, respectively. Similar reactions were observed with Sandoglobulin, but the frequency was much lower, i.e., 2.5%.~7 They were usually immediate in nature but occasionally delayed 2 to 6 hr after infusion. Regardless of whether Gamimune or Sandoglobulin were used, most of the reactions were mild and could be eliminated by slowing or temporarily stopping the infusion. Occasionally reactions precluded the continuation of 1SG infusion. ~4, ~7,28Most severe reactions could be attributed to lgG aggregates, dimeric IgG, aggregated IgA, and mixed IgG-IgA complexes in the ISG preparation, all of which are capable of activating both the classic and alternative complement pathways. 32-34 Day et a134 demonstrated that although changes of complement and complement components occurred with ISG infusion, they were not usually associated with clinically detectable adverse reactions. Circulating immune complexes were usually detectable in the immediate postinfusion period without symptoms and usually disappeared before the next infusion. Patients with severe adverse reactions to infused ISG may have serum autoantibodies that interact with antigens present in the ISG preparation, e.g., IgA, 13-1ipoprotein, etc. 34 In addition to the reported adverse effects, there are other drawbacks to the use of intravenous 1SG, not the least of which is its considerable expense. The recent media coverage of AIDS as well as hepatitis virus transmission by blood products has made this a major concern of many patients requiring regular infusion of ISG. Although most cases of AIDS continue to be reported in previously identified high-risk

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groups, there has been a definite increase in the number of transfusion-associated cases. 35' 36 The occurrence of AIDS after transfusion in 35 adults with no other identified risk factors supports the probability that blood components from single-unit donations also transmit the disease. Blood donors with mild or completely inapparent illness are believed to be the source of most transfusion-associated A I D S . 36 Although the intravenous infusion of ISG has been associated with immunologic abnormalities including increase of suppressor cell function 34 and reduction in circulating B cells,29 reversal of the helper: suppressor cell ratio and changes in the natural killer cell number and function have not been encountered. Similar immune abnormalities have been observed in individuals receiving multiple blood transfusions. 37 In contrast to plasma, blood, and coagulation factors, ISG preparations have not been associated with hepatitis transmission. There has been one recent article of acute, short-incubation, non-A, non-B hepatitis in 12 patients with primary hypogammaglobulinemia. 38 The ISG used was prepared by the British Blood Products Laboratory by use of conventional alcohol fractionation. Most patients were asymptomatic, but 10 mo after onset of hepatitis, 10 of the 12 still had abnormal liver function. Other currently available ISG preparations do not appear to have a problem with transmitted hepatitis, probably because the viruses are inactivated by the preparation procedures used.

Recommendations The presence of a primary humoral deficiency does not necessarily mandate intravenous ISG. There are patients who tolerate monthly intramuscular injections reasonably well and remain relatively free of severe recurrent infections. Such patients should be maintained on 100 mg/kg per month intramuscularly. Intravenously administered ISG should be held in reserve to treat acute, severe infections. In addition, there are patients whose preinfusion levels of serum IgG are so low and/or metabolism of infused ISG so rapid as to preclude significant therapeutic benefit from ISG. ~7 Judicious use of antibiotics is the most reasonable alternative treatment. An occasional patient may either develop or have preexisting autoantibodies that interact with antigens within the infused ISG preventing its further use. zS' 34 When a decision has been made to use intravenous ISG and the antibody deficiency has been diagnosed early, before the establishment of chronic sinopulmonary infection, patients will probably do quite well with infusions of 100 mg/kg per month. Since there are wide individual variations in the catabolism of ISG, measurement of peak and trough levels of serum IgG is encouraged to

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customize the dosage. In those patients with established disease and chronic infection, an initial dosage of 150 to 200 mg/kg per month appears more appropriate. The reason for this lies in the fact that IgG is metabolized more rapidly in the presence of active infection. Every effort to eradicate chronic respiratory disease with antibiotics should be made. It appears likely that once chronic infection has been controlled, either the infusion dose or the frequency of infusion can be decreased. There is no evidence that intravenous infusion of larger doses substantially increases therapeutic benefits with regard to either incidence or duration of infection. In fact, there is some pharmacokinetic data that suggest that increasing IgG serum concentrations is likely to shorten the half-life of infused immunogtobulin) 9

Intravenous ISG in secondary immunodeficiency Secondary immunodeficiency is far more commonly encountered in the practice of medicine than any or a combination of all the primary immunodeficiencies. 4~ The availability of intravenous ISG will facilitate treatment of secondary antibody deficiencies associated with excessive loss of immunoglobulin and increased frequency of pyogenic infection. 4j' 42 This includes protein-losing enteropathy, nephrotic syndrome irrespective of causation, exfoliative dermatitis, intestinal lymphangiectasia, and excessive bums. Lymphoproliferative neoplasia is especially associated with disordered immunity, and the lowest immunoglobulin levels are ~und in those B cell disorders that have the most protracted course, e.g., giant follicular lymphoma and chronic lymphocytic leukemia. Thymoma and multiple myeloma are also associated with humoral deficiency. Perhaps the best example of a secondary immunodeficiency in which intravenous ISG may be effective is in extensive thermal injury. Burn victims have a depressed immune capacity and often die of overwhelming sepsis despite heroic use of topical and systemic antibiotics. 43 Circulating levels of IgG remain subnormal for days to weeks after the initial injury. 44 Shirani et a l . 45 demonstrated that infusion of ISG at doses of 500 mg/kg twice weekly maintains normal circulating IgG concentrations in burn patients without obvious adverse reactions. It remains to be observed whether such infusions significantly reduce morbidity, but data assimilated from animal models suggest a definite prophylactic role for infused ISG. a6 CMV infection is a common complication of bone marrow transplantation in immunocompromised patients and is frequently associated with posttransplant interstitial pneumonia. 47 Treatment of established

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CMV interstitial pneumonia is uniformly unrewarding, but passive immunization with CMV immune plasma or hyperimmune globulin is effective in its prevention/8-5~ Because of the limited availability of high-titered CMV plasma and hyperimmune globulin, several lots of Gamimune were studied and found to have comparable antibody titers, if it was used in sufficient amounts. 5~ Preliminary results of a controlled trial in bone marrow transplant patients demonstrated that weekly doses of 20 ml/kg modified the severity of CMV infection and prevented interstitial pneumonia. 52 Despite the large volumes needed to confer protection, very few individuals experienced adverse effects.

HSG for prophylaxis These preparations are derived from recently convalescent or actively immunized patients who are in a state of hyperimmunity. HSG is available for pertussis, vaccinia, rabies, CMV, varicella zoster, and hepatitis B. In general, these preparations are expensive to produce and available in limited quantities. Pooled 1SG may also be used to attenuate certain infections if it is administered during the incubation period. The degree of protection varies from prevention of clinical symptoms to minor modification, depending on a large extent on the infectious agent, the potency of the preparation, and timing of administration relative to exposure to the organism. Since a future self-assessment topic will address the entire issue of immunization, it will not be discussed further here.

Interferon Interferon is a natural and broad spectrum antiviral glycoprotein that is species specific and produced by virtually all nucleated animal cells. Although it is usually referred to as a single molecule, three different types of interferon have been identified, i.e., or-, 13-, and -y-interferon. There are 14 subtypes of a-interferon produced mainly by mononuclear leukocytes, 13-interferon is made by fibroblasts in connective tissue, and ~/-interferon is produced by T cells. Production of interferon takes place immediately after instillation of live or attenuated viruses into cultured cells or intact animals. Production can also occur after exposure to synthetic polynucleotides, e.g., poly inosinic-polycytidylic acid, 53 by antigens, 54 and by endotoxin2 5 Patients with defects in interferon production may manifest frequent or severe viral infections. 56 Recently, T cells from patients with AIDS were found deficient in interferon production. 57 The manner by which interferon exerts antiviral action is very unique to the generally specific immune system. On invasion by a virus, the infected cell re-

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sponds by synthesizing and releasing molecules of interferon. The secreted interferon binds to receptor sites on the surface of uninfected neighboring cells and stimulates these cells to produce a group of protective proteins. These proteins inhibit viral reproduction in a manner not fully understood. However, the replication of the virus is prevented. Hence, unlike antibody, interferon molecules interact with other cells of the body, and their effect is not limited to the specific virus that initially stimulated them. Interferon also modulates the activity of various immune cells, e.g., by activating macrophages and by increasing the destructive capacity of cytotoxic T cells and of a class of immune cells known as natural killer cells. Interferon can also inhibit the division of cells, including tumor cells. In the past interferon for clinical trials had been produced in human leukocytes by the Cantell method as an intentional byproduct of blood banking. 58 This method was quite complex and had a relatively low yield with 1 unit of blood producing 1 million units or the equivalent of 4 Ixg of interferon. More recently, Goeddel et al. 59produced human leukocytic interferon (et2) in bacteria with the use of DNA recombinant techniques. This method has permitted major production of interferon with initiation of extensive clinical trials. Smith et al. 6~ have extensive experience with the use of interferon in chronic hepatitis B infection. With the use of recombinant e~-interferon, they noted greater suppression of viremia with doses in excess of 50 million units than with doses under 10 million units administered daily by intramuscular route. Interferon has also been effective in the treatment of herpes zoster in patients with cancer6~"62 and in CMV viremia after marrow transplantation. 63 It has been successfully used in juvenile papilloma, herpetic keratitis, and in the prevention of experimental rhinovirus infection in humans. It should be considered in the setting of any catastrophic viral illness. Interferon has produced variable results in the treatment of cancer with the exception of its consistent and dramatic effectiveness in hairy cell leukemia. 64 The clinical application of interferon is presently limited by its many side effects. 65 The influenza-like picture that often develops suggests that much of the constitutional symptoms we experience with viral infection may be as a result of interferon release. All three classes of interferon are pyrogenic and enhance the potential of human monocytes to produce the endogenous pyrogen, interleukin-1. Other side effects include anorexia, malaise, fatigue, nausea, vomiting, diarrhea, leukopenia, thrombocytopenia, proteinuria, mild transaminitis, headache, numbness, confusion, seizures, and arrhythmias.

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Thymic hormones The central role of the thymus gland in the development and maintenance of the immune system has been recognized for many years. 66 Several peptides have been isolated from thymic tissues and were found to circulate in the peripheral blood, establishing the role of the thymus as an endocrine gland. Thymosin has been demonstrated to stimulate the release of pituitary neuropeptides establishing a direct link between the thymus and the neuroendocrine system. Thymic hormones induce the maturation of T cell precursors and promote the differentiated and proliferative functions of mature T cells. 67 A number of hormones have been identified including thymosin fraction 5, thymosin oq, thymopoietin, thymulin, and thymic humoral factor, among others. Of these, thymosin oq, thymopoietin, and thymulin have been purified, sequenced, and synthesized, either chemically or by genetic engineering techniques. All these humoral factors except for thymic humoral factor are produced in thymic epithelial cells. The presence of these thymic hormones in the circulation and correlation of their declining levels with aging and various immunologic diseases lends support to their role as humoral regulators. 68 Encouraged by research carried out in animal models, a limited number of trials have been undertaken to treat primary and secondary immunodeficiency diseases as well as malignancy with thymic f a c t o r s . 67"69 Although this initial work is encouraging, the role of thymic hormones in treating human disease will require further well-controlled studies.

Marrow, thymus, and fetal tissue transplantation Initial attempts at bone marrow transplantation were generally unsuccessful unless an identical twin was available as a donor. It was only after the importance of MHC became known in transplantation that HLA tissue-matched sibling grafts succeeded in a variety of disease settings, eg., all varieties of SCID, neutrophil defects, and Wiskott-Aldrich syndrome among others. 70-73Unlike other forms of transplant procedures in which the immunologically intact host rejects the graft, in bone marrow transplantation, the grafted cells can reject the host producing GVHD. 74 Strict D-locus MHC identity between donor and recipient is necessary to avoid severe GVHD. Fetal liver cells have been found to be nearly as effective as bone marrow in restoring immunologic function. 75In the most successful cases liver cells were obtained from fetal donors below the age of 14 wk gestation. The use of combined fetal liver and thymus transplants are said to provide a better result and have

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also been used successfully. Their major drawback is the time required for fetal thymus and/or liver cells to establish themselves, i.e., 6 to 12 mo. There are many problems in keeping young recipients alive and free of fatal infection during this period of time. 75The use of fetal thymus transplants alone has not been very effective. The transient improvement noted in some patients may be secondary to production of thymic hormones. Advances have been made in transplanting across major histocompatibility barriers by the use of separated bone marrow-enriched, stem-cell fractions. 76

IMMUNOSTIMULATION A variety of immunomodulating agents are being evaluated as potentiators of the immune response. Most of these materials act at several points in the immune response without a specific target cell identified. These stimulatory agents can be divided into those that are antigen specific, eg., transfer factor, and those that are entirely nonspecific in their mode of action. The latter can be further divided into agents that exert their major effect on a disordered immune system, eg., levamisole and ISO and agents that require a functional immune system to operate such as BCG or Corynebacterium. There are many other synthetic compounds with immunotherapeutic potential that can only be mentioned briefly.

DLE TF It is almost 30 yr since Lawrence 77 prepared TF by use of a dialysis method and transferred delayed hypersensitivity from an immune to a nonimmune subject. In fact the crude DLE prepared by Lawrence has been demonstrated to contain approximately 160 separate moieties. DLE is the current designation for such preparations, and TF is now reserved for those components with antigen-specific activity. 78 TF activity in crude DLE preparations is believed to reside in several nucleopeptides containing both RNA and protein. 79 Crude DLE preparations also contain a nonspecific adjuvant activity along with one or more inhibitory activities.8~The mechanism of action of the TF portion of DLE is not yet known. TF may act on an uncommitted stem cell to induce specificity for an antigen or group of antigens, or it may assist in the recruitment of specific antigen-sensitive cells. The nonspecific, adjuvant portions of DLE appear to act by enhancing preexisting reactivity of recipient lymphocytes. The DLE is nonantigenic and can be lyophylized and stored indefinitely without loss of potency. It does not transmit any infectious disease, has no HLA antigens, and causes no serious side effects. It does not appear to affect humoral immunity.

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A variety of in vitro tests can be used to assess the immunologic status of recipients as they may have been effected by DLE. Such studies include facilitated chemotaxis, increased intracellular nucleotides, augmentation of antigen and mitogen responses, and increased formation of active T cell rosettes. The leukocyte migration inhibition assay in agarose, which measures production of granulocytic migration inhibitory factor, determines both the specific and nonspecific effects of DLE in vitro and is the favored method for assessing DLE for most clinical applications. 8o The primary impetus for the clinical use of TF during the last several decades has been its capacity to induce antigen-specific immunity and its low toxicity. It has therefore been administered to a large number of patients with a myriad of conditions. Most earlier studies were uncontrolled and involved very small numbers of patients with inadequate immunologic documentation.8~. 82 Although TF has been used in a variety of primary immunodeficiency diseases, the most consistent clinical successes have been reported in patients with CMC and the Wiskott-Aldrich syndrome. CMC represents an excellent choice for treatment with TF, since many of these patients have a selective defect in T cell immunity. It would appear that TF can prolong remission in CMC provided it is used in conjunction with conventional chemotherapy. The best results are observed in patients with positive conversion of delayed hypersensitivity to C a n d i d a . 83 In Wiskott-Aldrich syndrome excellent results were observed in over half the patients treated with TF. Although mortality is not significantly affected, the associated eczema improves, and incidence of infections decreases. Patients who responded very favorably included a subgroup characterized by a defect in monocyte receptors for IgG. 84 A variety of infectious and parasitic diseases caused by agents modulated by T cells have been treated with TF.81, 82 Dramatic improvements have been reported in disseminated vaccinia, measles pneumonia, congenital herpes simplex, herpes zoster, and CMV as well as others. Several refractory mycobacterial disorders have been treated successfully. Clinical improvement has been reported in disseminated candidiasis, disseminated coccidioidomycosis, and disseminated histoplasmosis initially refractory to conventional antifungal therapy. A controlled study has demonstrated TF to be very effective in the treatment of chronic cutaneous leishmaniasis. 85 It is not possible to reach any definite conclusions about the use of TF in the management of malignant disease. Most studies have been uncontrolled and involved only small numbers of patients with an inadequate follow-up period. A significant problem has

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been the selection of appropriate leukocyte donors. Nevertheless, TF has been of some reported benefit in patients with metastatic melanoma and osteosarcoma. In other tumor types the use of TF has proved generally disappointing.

Levamisole Levamisole is a synthetic derivative of tetramisole and one of the first chemically derived immunostimulants. 86 It has been used extensively as a veterinary antihelminthic drug. It acts directly on lymphocytes, macrophages, and granulocytes to modify their mobility, secretion, and proliferation. The principal mechanism of action appears to be facilitation of monocyte chemotaxis. 87 It can be immunostimulatory or immunosuppressive in its effect depending on the dose and timing of administration. When levamisole is administered clinically, it acts on the cellular limb of the immune response and does not affect antibody production. Its action is generally that of an immunopotentiator with positive effects usually requiring the parallel administration of a primary stimulus such as an antigen. The magnitude of its effect appears to depend on the degree of abnormality in the responding immune system. Levamisole can restore impaired cell-mediated immune responses to normal, but it will not hyperstimulate a normal immune system. 86 In clinical trials levamisole has been very inconsistent in its effect on malignancy and a variety of autoimmune disorders. This unpredictable efficacy in addition to its side effects of producing metallic taste, nausea, flu-like malaise, dermatitis, and severe granulocytopenia have made levamisole a less desireable agent.

Isoprinosine Isoprinosine (Newport Pharmaceuticals, Inc., Newport Beach, Calif.) is an extensively used and recently approved synthetic immunomodulatory agent with antiviral properties. ISO inhibits the replication of both DNA and RNA viruses in tissue culture and has documented clinical efficacy in a wide spectrum of viral disorders including subacute sclerosing panencephalitis, cutaneous herpes, aphthous stomatitis, CMV, and warts. 88' 89 In a double-blind study, this drug accelerated restoration of immune functions after depression by radiotherapy in patients with solid tumors. 9~ Preliminary data demonstrated similar improvement of T cell function in patients with AIDS. 9~ Its mechanism of action is uncertain, but ISO increases cell-mediated function, including T cell proliferative responses to both mitogen and antigen, active Trosette formation, and macrophage activation. This may be mediated by a direct augmentation of interleukin-2 production.92 Toxicologic, teratogenicity, and

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caracinogenicity studies have demonstrated ISO is safe, well tolerated, and remarkably free of side effects except for occasional hyperuricemia. It appears to have a remarkable potential in the future armamentarium of the clinical immunologist.

BCG BCG is a viable, attenuated strain of Mycobacterium boris obtained by progressive reduction of virulence in a culture medium. It is a nonspecific immunostimulant believed to act by stimulating the reticuloendothelial cell system. This may be secondary to T cell activation and lymphokine production. BCG also stimulates natural killer cells that can nonspecifically kill malignant cells. Some investigators have found that BCG cross-reacts immunologically with hepatoma, melanoma, and leukemic cells, an observation that may account for some of its apparently specific inhibitory effects on these tumors. The effectiveness of BCG in the treatment of malignancy has been demonstrated most dramatically by use of the scarification technique in the treatment of malignant melanoma. 93 In most other clinical cancer trials, the effectiveness of BCG has been equivocal.

Corynebacterium parvum A gram-positive bacterium, like BCG, Corynebacterium parvum induces macrophage activation. However, it paradoxically depresses T cell function. This adjuvant has been most successful in inducing tumor regression after intralesional injection. Serious side effects hamper its widespread use. Many individuals develop very high fever, headache, and vomiting. Some also develop hypertension associated with peripheral vasoconstriction.

Bestatin Bestatin is a nontoxic, orally active immunostimulating compound extracted from Streptomyces olivoreticuli. 94 This agent stimulates both humoral and cell-mediated immune responses with cellular targets including the macrophage, granulocyte precursors, natural killer cells, and possibly T cells. It also induces release of interleukin-2. Human toxicity has been negligible.

Tuftsin Tuftsin represents amino acid residues 289 to 292 of the constant region of IgG heavy chain. Tuftsin acts to stimulate motility, phagocytosis, antigen processing, and tumoricidal activity of macrophages. As such it demonstrates tremendous promise as an antineoplastic agent. In addition, it also increases neutrophil chemotaxis. It is a biologic peptide that is nontoxic and without significant side effects.

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IMMUNOSUPPRESSION Since the application of corticosteroids in the late 1940s, a variety of agents have been used in a large number of treatment protocols to abbrogate excessive immune responses. After the discovery of azathioprine in 1961 and the introduction of heterologous antilymphocyte serum in 1967, very little innovation has occurred in the area of immunosuppression. During the previous several decades cytotoxic drugs have been used to induce immune tolerance by destroying replicating cells. The cytotoxic drugs are divided into cell-cycle drugs that destroy rapidly multiplying cells (antimetabolites and folic acid antagonists) and the noncycle drugs that are injurious to all cells and usually result in a depletion of small lymphocytes (alkylating agents). 95' 96 The true advances in this field have been the recent introduction of several noncytotoxic techniques and agents. The most dramatic discovery has been the introduction of cyclosporine. This is the first immunosuppressive agent that allows selective alteration of the immune system and is unlike all other available immunosuppressants. Apheresis is being increasingly used to accomplish extracorporeal manipulation of the body's immune system. Finally, there is preliminary evidence regarding the use of ISG to abrogate autoimmune disorders.

Cyclosporine Cyclosporine is a cyclic endecapeptide first extracted in 1970 by Sandoz Laboratories in Basel, Switzerland, from fungi found in soil samples from Norway and Wisconsin. It was originally isolated as part of a search for biologically produced antifungal agents. After its rejection as an antibiotic, Borel et al. 97 agreed to test it for its potential as an immunosupressant. During the previous decade the compound has been extensively studied. 98 Its mechanism of action in both in vitro and in vivo studies relates to its inhibition of lymphokine production by impairing interleukin-2 production by both quiescent and activated helper T cells. This blockade of T cell help required for B cell activation, cytolytic T cell generation, and further expansion of helper/inducer T cell subpopulations underscores the potent mechanism of action of cyclosporine. There is also a Simultaneous expansion of suppressor T cell populations. It does not interrupt primary antibody responses unless introduced during the early steps of B cell activation. Although cyclosporine is very potent, it has not been found to have specific toxic effects on lymphocytes or other myelocytes. However, it does have significant nephrotoxicity that is occasionally associated with glomerular thrombosis and arteriolar hyaline deposition. Significant interstitial fibrosis has been noted in selected patients. Some of these nephrotoxic

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changes may be permanent, but reversal is observed in most patients when therapy is converted from cyclosporine to azathioprine. Other side effects include reversible hepatotoxicity, hirsuitism, gingival hyperplasia, tremor, seizures, and gastrointestinal intolerance. All of the latter appear to be dose related and reversible with cessation of the drug. As of early 1984, more than 5550 patients had been treated with cyclosporine with development of 33 lymphoma and seven additional lymphoproliferative lesions. 99 This represents an overall incidence of 0.7% that is not significantly different from the risk noted with conventional immunosuppression. These malignancies occurred in the setting of allogeneic organ transplantation in which there is a higher incidence of malignancy possibly related to the foreign graft. Cyclosporine has been demonstrated to have remarkable benefit in organ transplantation. It has been used extensively in kidney, liver, bone marrow, pancreas, heart, lung, and heart-lung transplantation. Beyond its obvious benefit in organ transplantation, the high degree of T cell specificity suggest it may have application in treatment of T cell malignancies. There are very limited data in its application toward treatment of autoimmune disorders, but experience derived from the rapid taper protocols with use of lower doses of cyclosporine (which blocks interleukin-2 production) in concert with corticosteroids (which block interleukin-1 production) may provide the impetus for its wider application in autoimmune disease.

Apheresis Apheresis is a process by which blood is withdrawn from a subject and its constituents such as plasma, buffy coat, or platelets are separated and removed and the desired components are reinfused. Plasmapheresis is the specific removal of large volumes of plasma. Replacement is undertaken with combinations of saline solutions, albumin, human donor plasma, or ISG. The buffy coat fraction of blood containing both lymphocytes and other leukocytes can be selectively removed, and this process is variably termed lymphapheresis or leukapheresis. With the current technology it is also possible to collect single donor platelets, granulocytes, and young erythrocytes. During the previous decade there has been an explosion in the number of proposed applications of apheresis in the management of a variety of immunologic disorders. 10o.m, Despite its wide application, its absolute indications are few and include hyperviscosity syndrome, Goodpasture's syndrome, cryoglobulinemia, TTP, and lifethreatening complications of immunologic disorders refractory to conventional treatment. On the basis of early work demonstrating the ef-

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ficacy of thoracic duct drainage as a means of inducing immunosuppression via lymphocyte depletion, lymphapheresis has been used in a variety of rheumatic disorders with very limited success as the sole therapy. Lymphoplasmapheresis combined with cytotoxic immunosuppressive therapy is more likely to result in significant clinical benefit. Unfortunately, most studies in this area have been with small numbers of patients and are largely uncontrolled. In general plasmapheresis has been believed to have some potential benefit in the following categories of disease: (1) diseases associated with abnormal (or excess) plasma proteins, toxins, or metabolic products, eg., hyperviscosity syndrome, multiple myeloma, cryoglobulinemia, paraquat poisoning, etc., (2) diseases associated with autoantibodies, eg., Goodpasture's syndrome, Guillain-Barr6 syndrome, myasthenia gravis, ITP, etc., (3) diseases associated with alloantibodies, e.g., factor VIII and IX inhibitors, hemolytic disease of the newbom, renal transplant rejection, etc., (4) diseases associated with immune complexes, eg., rheumatoid arthritis, systemic lupus erythematosus, and (5) selected miscellaneous disorders that do not fit any of the above categories such as TTP. In considering the effectiveness of plasmapheresis in treating antibody or immune complex-mediated disease, it is important to remember that many of the autoantibodies associated with these diseases belong to the IgG class of immunoglobulin that is distributed in both the intra- and extravascular spaces. The less than expected decrement in serum autoantibody titers is related to the rapid equilibration between the extravascular and intravascular compartments. Even with successful reduction of autoantibody levels, many observers report "antibody rebound" as soon as the procedure is discontinued. This is believed to be related to diminished feedback inhibition. Because of the above considerations, therapeutic apheresis may become a selective adjunctive therapy to effect a shortterm improvement but definitely not indicated as a chronic therapy in most disease states. The best results have been reported in association with cytotoxic immunosuppressive therapy, m2. m3 There are some very preliminary studies suggesting synergism between plasmapheresis and intravenous infusion of ISG. ~~ Plasmapheresis has been associated with a variety of complications. The dysequilibrium syndrome, similar to a vasovagal episode, is the most frequent complication. This originates from rapid volume shifts during the procedure. Other complications include edema, hypotension, hypokalemia, citrate reactions, anemia, thrombocytopenia, and increased risk of infection. Vascular access remains a significant limiting variable in many patients. The technology of apheresis is very

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adaptable and represents an exciting approach to the selective removal of autoantibodies, activated immunocytes, immune complexes, and other inflammatory mediators. Its true place in the management of immunologic disease will require further careful study. Intravenous ISG as an imrnunosuppressant Chronic ITP is an autoimmune disorder resulting from an IgG antibody reactive with glycoprotein antigens on the platelet membrane.~~ Jo6 While children with Wiskott-Aldrich syndrome were being treated with prophylactic intravenous ISG, a Swiss group noted a striking increase in platelet counts. Later, Imbach et a1.1~ reported their observations in 13 individuals with acute and chronic ITP treated with 400 mg/kg per day of Sandoglobulin for 5 consecutive days. Several patients underwent complete remission of disease, whereas others required maintenance ISG infusions to remain stable. A few patients experienced an initial good response that could not be sustained. These observations were confirmed by several other groups in both children and adult patients, lo8. lo9 The mechanism by which ISG induced increases in platelet counts was not completely understood. Several hypotheses have been suggested including (1) inhibition of reticuloendothelial macrophage Fc-receptor function by infused ISG with resultant survival of antibody-coated platelets,~l~ (2) coating of red cell blood group antigens by specific antibody (anti-A and antiB) leading to premature erythrocyte destruction, subsequent reticuloendothelial overload with increased platelet survival,l~! and (3) feedback inhibition of antibody synthesis by ISG. 39' 107.112,ij3 High-dose ISG has also been useful in the treatment of myasthenia gravis in which autoantibodies are directed against various muscle antigens, particularly the postsynoptic membrane acetylcholine receptor. 114, ~J~ The mechanism of action was believed to be similar to that operative in ITP. A recent multicenter article indicates that high-dose ISG, i.e., Venilon (Tijin, Ltd. Japan), administered at 400 mg/kg for 5 days, was acting with aspirin to prevent development of coronary artery lesions in Kawasaki disease. 116Since the cause of this condition is not known, the authors could only speculate on how ISG was suppressing the coronary artery lesion. It was believed that if immune complexes were playing a role in the pathogenesis of Kawasaki disease, there might be competitive inhibition of Fc receptors within blood vessels by IgG in the infused ISG. Other possibilities included negative feedback and infusion of some undefined antitoxin or neutralizing antibody against an as yet unknown pathogen. Last, there is some evidence

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in one patient with autoimmune neutropenia that antibody synthesis may be inhibited with intravenous ISG. t~7 Perhaps the most intriguing observation to date on the immunosuppressant capacities of ISG is the recent report by Sultan et al. j'~ that high-dose intravenous ISG, i.e., Sandoglobulin, resulted in rapid and prolonged suppression of autoantibody to antihemophilic factor (VIIIc). These authors proposed the mechanism of action to be an autoantibody interaction with factor VIII through an idiotype: anti-idiotype interaction, i.e., the ISG contained anti-idiotypic antibodies against idiotypes expressed by anti-factor VIII autoantibodies. If this is true, ISG may have a much broader application in the treatment of autoimmune disease. REFERENCES 1. von Behring E, Kitasato O: Ueber das zustandekommender diphtherie-immunitat und der tetanus-immunitat bei thieren. Dtsch Med Wochenschr 16:1113, 1890 2. Ehrlich P: Die Wertbemessungdes diphtherieheilserums und deren theoretische grundlagen. Klin Jahrbuch 6:299, 1897 3. McKhann CF, Chu FT: Antibodies in placental extracts. J Infect Dis 52:266, 1933 4. Ordman CW, Jenning CG, Janeway CA: Chemical, clinical, and immunologicalstudies on the products of human plasma fractionation. XII. The use of concentrated normal human serum gamma globulin (human immune serum globulin) in the prevention and attenuation of measles. J Clin Invest 23:541, 1944 5. Janeway CA: The development of clinical uses of immunoglobulins: a review. In Merler E, editor: Immunoglobulins. Washington, D.C., 1970, National Academyof Sciences, pp 3-14 6. Barandun S, Kistler P, Jeunet F, Isliker H: Intravenous administration of human gammaglobulin. Vox Sang 7:157, 1962 7. Smith GN, Griffiths B, Mollison D, Mollison PL: Uptake of IgG after intramuscular and subcutaneous injection. Lancet 1:1208, 1972 8. Stiehm ER, VaermanJP, Fudenberg HH: Plasma infusions in immunologic deficiency states: metabolic and therapeutic studies. Blood 28:918, 1966 9. Immunodeficiency:report of a WHO Scientific Group. Geneva, Switzerland, 1978, World Health Organization, WHO Technical Report Series no. 630, pp 3:80 10. Schultze HE, Schwick G: Uber neue moglich-keitan intravenoser gammaglobulin-applikation.Dtsch Med Wochenschr 87:1643, 1962 11. SqourisJT: The preparationof plasmin-treated immuneserum globulin for intravenous use. Vox Sang 13:71, 1967 12. Stephan W: Hepatitis-free and stable human serum for intravenous therapy. Vox Sang 20:442, 1971 13. Nolte MT, Pirofsky B, Gerritz GA, Golding B: Intravenous immunoglobulin therapy for antibody deficiency. Clin Exp Immunol 36:237, 1979 14. AmmannAJ, AshmanRF, BuckleyRH, KrantmannHJ, Ochs H, Stiehm ER, Tiller TL, Wara DW, WedgewoodR: Use of intravenous gammaglobulin in antibody immunodeficiency: results of a multi-center controlled trial. Clin Immunol Immunopathol 22:60, 1982

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15. Yamanaka T, Abo W, Chiba S, Nakao T, Masuho Y, Tomibe K, Noguchi T: Clinical effect and metabolism of 5-sulfonated immunoglobulin in 7 patients with congenital humoral immunodeficiency. Vox Sang 37:14, 1979 16. Eibl M: Intravenous immunoglobulins: clinical and experimental studies. In Alving BM, Finlayson JS, editors: Immunoglobulins: characteristic and uses of intravenous preparations. Washington, D.C., 1980, U.S. Government Printing Office, USDHHS Publication no. (FDA)-80-9005, p 23 17. Cunningham-Rundles C, Siegal FP, Smithwick EM, LionBoule A, Cunningham-Rundles S, O'Malley J, Barandun S, Good RA: Efficacy of intravenous immunoglobulin in primary humoral immunodeficiency disease. Ann Intern Med 101:435, 1984 18. Hanson LA, Bjorkander J, Wadsworth C, Bake B: Intravenous immunoglobulin in antibody deficiency syndromes. Lancet 1:396, 1982 19. Romer J, Morgenthaler JJ, Scherz R, Skvaril F: Characterization of various immunoglobulin preparations for intravenous application. I. Protein composition and antibody content. Vox Sang 42:62, 1981 20. Nolte MT, Bardana EJ, Pirofsky B: Intravenous serum immune globulin in agammaglobulinemia. J ALLERGYCLIN IMMtJNOL 55:114, 1975 (abst) 21. Pirofsky B, Anderson CJ, Bardana EJ: Therapeutic and detrimental effects of intravenous immunoglobulin therapy. In Alving BM, Finlayson, JS, editors: Immunoglobulins: characteristics and uses of intravenous preparations. USDHHS Publication, 1980, no. (FDA)-80-9005, pp 15-20 22. Buckley RH: Long-term use of intravenous immune globulin in patients with primary immunodeficiency diseases: inadequacy of current dosage, practices, and approaches to the problem. J Clin lmmunol 2(suppl): 15, 1982 23. Ochs HD, Fischer SH, Wedgewood RJ: Modified immune globulin: its use in the prophylactic treatment of patients with immune deficiency. J Clin lmmunol 2(suppl):225, 1982 24. Pirofsky B, Campbell SM, Montanaro A: Individual patient variations in the kinetics of intravenous immune globulin administration. J Clin Immunol 2(suppl):75, 1982 25. Pirofsky B: Intravenous immune globulin therapy in hypogammaglobulinemia: a review. Am J Med 76(3A):53, 1984 26. Wells JV, Stites DP, Wybren J, Fudenberg HH: New preparation of therapeutic intravenous gammaglobulin Clin Res 20:521, 1972 (abst) 27. Montanaro A, Pirofsky B: Prolonged interval high-dose intravenous immunoglobulin in patients with primary immunodeficiency states. Am J Med 76(3A):67, 1984 28. Ochs HD, Fischer SH, Wedgewood RJ, Wara DW, Cowan MJ, Ammann AJ, Saxon A, Budinger MD, Allred RN, Rousell RH: Comparison of high-dose and low-dose intravenous immunoglobulin therapy in patients with primary immunodeficiency diseases. Am J Med 76(3A):78, 1984 29. Sorensen RU, Polmar SH: Efficacy and safety of high-dose intravenous immune globulin therapy for antibody deficiency syndromes. Am J Med 76(3A):83, 1984 30. Berger M, Cupps TR, Fauci AS: High-dose immunoglobulin replacement therapy by slow subcutaneous infusion during pregnancy. JAMA 247:2824, 1982 31. Mease PJ, Ochs HD, Wedgewood RJ: Successful treatment of ECHO viral meningoencephalitis and myositis-fasciitis with intravenous immune globulin therapy in a patient with X-linked agammaglobulinemia. N Engl J Med 304:1278, 1981 32. Romer J, Spath PJ, Skvaril F, Nydegger UE: Characterization of various immunoglobulin in preparations for intravenous

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application. 11. Complement activation and binding to staphylococcus protein A. Vox Sang 42:74, 1981 Wright JK: Reduced immunoglobulin G activates complement system with decreased cooperativity. Biochem Biophys Res Commun 83:1284, 1978 Day NK, Good RA, Wahn V: Adverse reactions in selected patients following intravenous infusions of gamma globulin. Am J Med 76(3A):25, 1984 Jett JR, Kuritsky JN, Katzman JA, Homburger HA: Acquired immunodeficiency syndrome associated with blood-product transfusions. Ann Intern Med 99:621, 1983 Chamberland ME, Castro KG, Haverkos HW, Miller B1, Thomas PA, Reiss R, Walker J, Spira TJ, Jaffe HW, Curran JW: Acquired immunodeficiency syndrome in the United States: an analysis of cases outside high-incidence groups. Ann Intern Med 101:617, 1984 Gascow P, Zoumbos NC, Young NS: Immunologic abnormalities in patients receiving multiple blood transfusions. Ann Intern Med 100:173, 1984 Lever AM, Webster ADB, Brown D, Thomas HC: Non-A, non-B hepatitis occurring in agammaglobuliuemic patients after intravenous immunoglobulin. Lancet 2:1062, 1984 Waldmann TA, Strober B: Metabolism of immunoglobulins. Prog Allergy 13:1, 1969 Bardana EJ: A conceptual approach to immunodeficiency. Med Clin North Am 65:959, 1981 Rousell RH, Collins MS, Dobkin MB, Louie RE, Roby RE, Sweet BH: Antibody levels in reduced/alkalated intravenous immune globulin. Am J Med 76(3A):40, 1984 Dwyer JM: Thirty years of supplying the missing link: history of gamma globulin therapy for immunodeficient diseases. Am J Med 76(3A):46, 1984 Pruitt BA Jr, McManus AT: Opportunistic infections in severely burned patients. Am J Med 76(3A):146, 1984 Munster AM, Hoagland HC, Pruitt BA Jr: The effect of thermal injury on serum immunoglobulins. Ann Surg 172:965, 1970 Shirani KZ, Vaughan GM, McManus AT. Amy BW, McManus WF, Pruitt BA Jr, Mason AD Jr: Replacement therapy with modified immunoglobulin G in burn patients: preliminary kinetic studies. Am J Med 76(3A):175, 1984 Holder IA, Naglich JG: Experimental studies of the pathogenesis of infections due to Pseudomonas aeruginosa: treatment with intravenous immune globulin. Am J Med 76(3A): 161, 1984 Marker SC, Howard RJ, Simmons RL, Kalis JM, Connelly DP, Najarian JS, Balfour HH: Cytomegalovirus infection: a quantitative prospective study of three hundred twenty consecutive renal transplants. Surgery 89:660, 1981 Condie RM, O'Reilly RJ: Prophylaxis of cytomegalovirns infection in bone marrow transplant recipients by hyperimmune cytomegalovirus gamma globulin. Dev Biol Stand 52:501, 1982 Winston DJ, Pollard RB, Ho WG, Rasmussen LE, Nan-Ying S, Lin CH, Gossett TG, Merigan TC, Gale RP, Gallagher JG: Cytomegalovirns immune plasma in bone marrow transplant recipients. Ann Intern Med 97:11, 1982 Nicholls AJ, Brown CB, Edward N, Cuthbertson B, Yap PL, McClelland DBL: Hyperimmune immunoglobulin for cytomegalovirus infections. Lancet 1:532, 1983 Winston DJ, Ho WG, Rasmussen LE, Lin CH, Chu CL, Merigan TC, Gale RP: Use of intravenous immune globulin in patients receiving bone marrow transplants. J Clin Immunol 2:425, 1982 Winston DJ, Ho WG, Lin CH, Budinger MD, Champlin RE,

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Gale RP: Intravenous immunoglobulin for modification of cytomegalovirus infections associated with bone marrow transplantation. Am J Med 76(3A):128, 1984 53. Field AK, Tytell AA, Lampson GP, Hilleman MR: Inducers of interferon and host resistance. II. Mulfistranded synthetic polynucleotide complexes. Proc Natl Acad Sci USA 58:1004, 1967 54. Green JA, Cooperband SR, Kibrick S: Immune specific induction of interferon production in cultures of human blood lymphocytes. Science 164:1415, 1969 55. Ho M: Interferon-like viral inhibitor in rabbits after intravenous administration of endotoxin. Science 146:1472, 1964 56. Isaacs D, Clarke JR, Tyrrell DAJ, Webster ADB, Valman HB: Deficient production of leukocyte interferon (interferonalpha) in vitro and in vivo in children with recurrent respiratory tract infections. Lancet 2:950, 1981 57. Murray HW, Rubin BY, Masur H: Impaired production of lymphokines and immune (gamma) interferon in the acquired immunodeficiency syndrome. N Engl J Med 310:883, 1984 58. Cantell K, Hirvonen S: Preparation of human leukocyte interferon for clinical use. Tex Rep Biol Med 35:138, 1977 59. Goeddel DV, Yelverton E, Ullrich A, Heyneker HL, Miozzari G, Holmes W, Seeburg PH, Dull T, May L, Stebbing N, Crea R, Maeda S, McCandliss R, Sloma A, Tabor JM, Gross M, Familletti PC, Pestka S: Human leukocyte interferon produced by E. coli is biologically active. Nature 287:411, 1980 60. Smith CI, Weissberg J, Bernhardt PB, Gregory PB, Robinson WS, Merigan TC/ Acute Dane particle suppression with recombinant leukocyte A interferon in chronic hepatitis B infection. J Infect Dis 148:907, 1983 61. Merigan TC, Rand KH, Pollard RB, Abdallah PS, Jordan GW, Fried RP: Human leukocyte interferon for the treatment of herpes zoster in patients with cancer. New Engl J Med 298:981, 1978 62. Hirsch MS, Schooley RT: Treatment of herpes virus infections. N Engl J Med 309:963, 1983 63. Meyers JD, McGuffin RW, Neiman PE, Singer JW, Thomas ED: Toxicity and efficacy of human leucocyte interferon for treatment of cytomegalovirus pneumonia after marrow transplantation. J Infect Dis 141:555, 1980 64. Quesada JR, Reuben J, Manning JT, Hersh EM, Gutterman JV: Alpha interferon for induction of remission in hairy cell leukemia. N Engl J Med 310:15, 1984 65. Postic B, Arroyo JC: Prospects for the medical use of interferon in 1984. Henry Ford Hosp Med J 32:116, 1984 66. Good RA, Dalmaseo AP, Martinez C, Archer OK, Pierce JC, Papermaster BW: The role of thymus in the development of immunological capacity in rabbits and mice. J Exp Med 116:773, 1962 67. Wara DW: Thymic hormones in primary immunodeficiency. Clin Immunol Allergy 3:169, 1983 68. Incefy GS: Effect of thymic hormones on human lymphocytes. Clin Immunol Allergy 3:95, 1983 69. Aiuti F, Businco L: Effects of thymic hormones on immunodeficiency. Clin Immunol Allergy 3:187, 1983 70. Bach FH, Albertini RJ, Joo P, Anderson JL, Bortin MM: Bone marrow transplantation in a patient with WiskottAldrich syndrome. Lancet 2:1364, 1968 71. Bortin MM, Rimm AA: Severe combined immunodeficiency disease. Characterization of the disease and results of transplantation. JAMA 238:591, 1977 72. Kenny AB, Hitzig W: Bone marrow transplantation for severe combined immunodeficiency. Eur J Pediatr 131:155, 1979 73. Rappeport JM, Parkman R, Newburger P, Camitta BM, Chusid MJ: Correction of infantile agranulocytosis (Kostmann's

Syndrome) by allogeneic bone marrow transplantation. Am J Med 68:605, 1980

74. Ramsey NK, Kersey JH, Robison LL, McGlave PB, Woods WG, Krivit W, Kim TH, Goldman A1, Nesbit ME Jr: A randomized study of the prevention of acute graft-versus-host disease. New Engl J Med 306:392, 1982 75. Touraine JL: T-lymphocyte maturation after bone marrow, fetal, liver, or thymus transplantation in immunodeficiencies. Transplant Proc 11:494, 1979 76. Clift RA, Hansen JA, Thomas ED, Bucker CD, Sanders JE, Mickelson EM, Storb R, Johnson FL, Singer JW, Goodell BW: Marrow transplantation from donors other than HLA identical siblings. Transplantation 28:235, 1979 77. Lawrence HS: The transfer in humans of delayed skin sensitivity to streptococcal M substance and to tuberculin with disrupted leukocytes. J Clin Invest 34:219, 1955 78. Wilson GB, Fudenberg HH: Is controversy about "transfer factor therapy" nearing an end? Immunol Today 4:157, 1983 79. Wilson BG, Paddock GV, Fudenberg HH: Effects of dialyzable leukocyte extracts with transfer factor activity on leukocyte migration in vitro. V. Antigen-specific lymphocyte responsiveness can be initiated by two structurally distinct polyribonucleotides. Thymus 2:257, 1980 80. Wilson GB, Smith CL, Fudenberg HH: Effects of dialyzable leukocyte extracts with transfer factor activity on leukocyte migration in vitro. III. Characterization of the antigenindependent migration inhibition factor in DLEs as a neutrophil immobilizing factor. J ALLERGYCLIN IMMUNOL64:56, 1979 81. Basten A, Croft S: Transfer factor: clinical usage and experimental studies. In Jirsch DW, editor: Immunological Engineering. London, 1978, Medical and Technical Publications, p 83 82. Arala-Chaves MP, Horsmanheimo M, Goust JM, Fudenberg HH: Biological and clinical aspects of transfer factor. In Jirsch DW, editor: Immunological Engineering. London, 1978, Medical and Technical Publications, p 35 83. Petersen EA, Kirkpatrick CH: Nature and activities of transfer factor. Ann NY Acad Sci 332:216, 1979 84. Spitler LE: Transfer factor therapy in the Wiskott-Aldrich syndrome. Results of long-term follow-up in 32 patients. Am J Med 67:59, 1979 85. Sharma M, Anaraki F, Ala F: Preliminary results of transfer factor therapy of persistent cutaneous leishmania infection. Clin Immunol Immunopathol 12:183, 1979 86. Symoens J, Rosenthal M: Levamisole in the modulation of the immune response: the current experimental and clinical state. J Reticuloendothel Soc 21:175, 1977 87. Pike MC, Daniels CA, Synderman R: Influenza-induced depression of monocyte chemotaxis: reversal by levamisole. Cell Immunol 32:234, 1977 88. Bradshaw LJ, Summer HL: In vitro studies on cell-mediated immunity in patients treated with Inosiplex for herpes virus infection. Ann NY Acad Sci 284:190, 1977 89. Wybran J, Famaey JP, Gortz R, Dab I, Malfroot A, Appleboom T: Inosiplex (lsoprinosine): a review of its immunological and clinical effects of disease. Adv Pharmacol Therapeut II 6:123, 1982 90. Fridman H, Calle R, Morin A: Double-blind study of lsoprinosine influence on immune parameters in solid tumorbearing patients treated by radiotherapy, lnt J Immunopharmacol 2:194, 1980 91. Grieco MH, Reddy MM, Manvar D, Ahuja KK, Moriarty ML: In vivo immunomodulation by lsoprinosine in patients

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