Journal ofthe Royal Society of Medicine Volume 72 December 1979
927
a week of the first administration of transfer factor to an extremely high level of 870 units/ml (normal level up to 150 units/ml) and then rapidly fell to a level only slightly above normal. Anticandida IgM rose from 50 to 250 units (normal level up to 100 units/ml) and then fell to 40 units, and anticandida IgA was detected in the serum for the first time. Levels of all three anticandida antibodies are currently in the high normal range, lymphocyte transformation to candida antigen is present and leukocyte migration inhibitory factor is produced on exposure of leukocytes to candida antigen. Changes in nonspecific immune function have been from a PPD negative skin test to positivity after the second unit of transfer factor, and a rise in the responsiveness of peripheral blood lymphocytes to PHA stimulation. No changes in total levels of IgG, IgA or IgM have been detected during the periods of fluctuation of specific anticandida antibodies.
References Higgs J M (1973) Proceedings of the Royal Society of Medicine 66, 802 Higgs J M & Wells R S (1972) Br"itish Journal of Dermatology 86, Suppl; p 28
Kirkpatrick C H, Rich R R & Bennett J E (1971) Annals ofInternal Medicine 74, 955 Lawrence H S (1974) Transfer Factor in Cellular Immunity. Harvey Lecture Series 68. Academic Press, New York; pp 239-250 Nielsen K H, Parrat H D & White R G (1973) Journal of Immunological Methods 3, 301
Transfer factor Anne S Hamblin PhD Immunology Department, St Thomas' Hospital, London SEI 7EH
The term transfer factor is given to dialysable material present in extracts of immune peripheral blood leukocytes which apparently confers ('transfers') delayed-type hypersensitivity (or its in vitro correlates) when injected into a non-hypersensitive subject. Clinical and immunological observations following the administration of transfer factor have led to the concept that it converts non-immune to immune lymphocytes in vivo, thus acting as a mediator of adoptive sensitization (Lawrence 1974). A wide variety of clinical conditions including immunodeficiency, infectious diseases, autoimmunity and malignancy have therefore been treated with transfer factor in attempts to reconstitute cellular immunity. However, the mechanism by which transfer factor is therapeutically beneficial is complicated by clinical observations that patients may acquire cellular immune responsiveness not apparently related to donor specificity (Dupont et al. 1974, Valdimarsson et al. 1974, Spitler et al. 1972, Griscelli 1975, Kirkpatrick & Smith 1976a) suggesting that leukocyte extracts may also possess adjuvant properties. The clinical benefit derived from treatment with transfer factor may therefore depend not only on the initiation of specific immunological reactivity, but also the amplification of an existing immune response. The proposition that both the so-called specific and nonspecific properties of transfer factor may have a contribution to make to the augmentation of cellular immune responses in patients raises a number of practical issues. It is perhaps useful to remember that the term transfer factor is often applied to the whole leukocyte extract administered in vivo. However, it has been suggested that the crude material be termed 'dialysable leukocyte extract' and that the term 'transfer factor' be reserved for the unidentified factor or factors capable of transferring delayed hypersensitivity. The relative roles of the specific and nonspecific activities in the extract raise questions regarding the selection of donors for therapy. It is not clear whether a more beneficial product may be prepared from donors with specific strong delayed hypersensitivity responses or from unscreened pooled normal healthy donors. The question is
0141-0768/79/120927-03/$01.00/0
,'-.
1979 the Royal Society of Medicine
928
Journal of the Royal Society of Medicine Volume 72 December 1979
under assessment in studies in which the product from normal healthy blood-bank donors is administered to patients with a wide range of diseases (Grob 1977) and in properly controlled clinical trials such as those currently under way in the United States in leprosy and coccidiomycosis. Much of the scepticism which surrounded the early work on transfer factor resulted from the consistent failure to demonstrate the transfer phenomenon in animals, particularly the guinea pig. However, recent studies in primates, guinea pigs and infectious disease models in animals have shown that it is possible to demonstrate biological properties of leukocyte extracts which may be relevant to transfer factor in humans. Such studies have revealed both general and specific stimulatory activity in leukocyte dialysates. Thus, Steele etal. (1976) have reported that human leukocyte dialysates could transfer delayed hypersensitivity responses to gnotobiotic monkeys never previously exposed to the relevant antigens. Immunological specificity was implied since recipients never converted to antigens to which the donor did not react in vivo. In contrast, Vandenbark etal. (1977) have reported that delayed hypersensitivity skin reactions could only be elicited in guinea pigs receiving human leukocyte dialysate when the animals had previously been primed with the relevant antigen, thus supporting the view that dialysates contain materials which stimulate the recipient to express sensitivity to antigens to which they have previously been exposed. The concept that transfer factor converts non-immune to immune lymphocytes raises the possibility that non-immune lymphocytes cultured with transfer factor may acquire the ability to respond to specific antigen which may be assessed by a variety of in vitro tests such as lymphocyte transformation, migration inhibition or cytotoxicity. Such specific effects have been reported and have been reviewed (Lawrence 1974, Ascher et al. 1976). However, it is clear that leukocyte dialysates also contain many substances capable of nonspecifically altering lymphocyte responses only some of which may reflect the nonspecific effects in vivo (Hamblin et al. 1976). It is, therefore, essential to bear in mind that extracts of cells may simply improve lymphocyte function in vitro by, for example, medium conditioning effects. Thus it has been reported that human leukocyte dialysate causes cyclic guanosine monophosphate (GMP) accumulation in normal leukocytes (Kirkpatrick & Smith 1976b). Cell extracts contain serotonin and ascorbate known to cause cyclic GMP accumulation. Fractionation of the crude dialysate indicated that the fraction which caused conversion of delayed skin tests also causes cyclic GMP accumulation, but did not contain ascorbate or serotonin. This example reveals the need for careful biochemical studies before conclusions may be drawn regarding the relevance of in vitro activity of the crude dialysate to its effect in vivo. To supplement these studies much effort has been made to characterize the dialysable material extracted from leukocytes biochemically. Analysis of dialysates by gel chromatography has shown that they may be separated into a number of subfractions some of which transfer specific delayed hypersensitivity responses (Zuckerman et al. 1974, Burger et al. 1976) and some of which may nonspecifically enhance delayed hypersensitivity responses in humans (Krohn et al. 1977). In addition, fractions have been described with specific and nonspecific activity in animal and in vitro models (see Ascher et al. 1976). Within leukocyte dialysates a variety of known substances have been identified which could be responsible separately or in combination for some of the reported in vitro or in vivo effects (Wilson et al. 1977, Hamblin 1979). However, the material(s) responsible for the transfer of delayed hypersensitivity has yet to be defined biochemically, and to date no subfraction of the crude dialysate has definitively been shown to be therapeutically beneficial. The lack of available information regarding the mechanisms of action and biochemical nature of transfer factor makes it difficult to describe schedules, dosage and source of material for treatment. Furthermore, a variety of activities have been reported clinically and in animal and in vitro models and the interrelationship between these has yet to be made. One view, expressed by C H Kirkpatrick in 1975, is that 'the phenomenon of transfer factor is a summation of several effects'. The results of ongoing clinical trials, together with continued biochemical and biological analysis of the dialysate, will show whether this hypothesis will be substantiated or refuted.
Journal ofthe Royal Society of Medicine Volume 72 December 1979
929
References Ascher M S Gottlieb A A & Kirkpatrick C H ed (1976) Transfer Factor: Basic Properties and Clinical Applications. Academic Press, New York Burger D R, Vandenbark A A, Daves D, Anderson W A, Vetto R M & Finke P, (1976) Journal of Immunology 117, 789 Dupont D, BaUlow M, Hansen J A, Quick C, Yunis E J & Good R A (1974) Proceedings of the National Academy of Sciences of the USA 71, 867 Grscelli C (1975) Birth Defects 11, 462 Grob P J (1977) Acta Neurologica Scandinavica 55, Suppl; p 217 Hamblin A S (1979) In: Clinical Neuroimmunology. Ed. F Clifford Rose. Blackwell Scientific Publications, London; p497 Hamblin A S, Dumonde D C & Maini R N (1976) Clinical and Experimental Immunology 23, 303 Kirkpatrick C H (1975) Birth Defects 11, 441 Kirkpatrick C H & Smith T K (1976a) Cellular Immunology 27, 232 Kirkpatrick C H & Smith T K (1976b) In: Transfer Factor: Basic Properties and Clinical Applications. Ed. M S Ascher et al. Academic Press, New York; p 161 Krohn K, Grohn P, Horsmanheimo M & VirolainenuM (1977) Medical Biology 54, 334 Lawrence H S (1974) Transfer Factor in Cellular Immunity. Harvey Lecture Series 68. Academic Press, New York; pp 239-250 Spider L E, Levin A S, Blots M S, Epstein N, Fudenberg H H, Helistrom I & Heflstrom K E (1972) Journal of Clinical Investigation 51, 92a Steele R W, Eichberg J W, Heberlung R L, Eller J J Kalter S S & Kniker W T (1976) Cellular Immunology 22, 110 Valdimarsson H, Hambleton G, Henry K & McConnell I (1974) Clinical and Experimental Immunology 16, 141 Vandenbark A A, Burger D R & Vetto R M (1977) Clinical Immunology and Immunopathology 8, 7 Wilson G B, Welch T M, Knapp D R, Horsannheimo A & Fudenberg H (1977) Clinical Immunology and Immunopathology, 8, 551 Zuckerman K S, Neidhart J A, Balcerzak S P & LoBuglio A (1974) Journal of Clinical Investigation 54, 997
Therapy of candidiasis Yvonne Clayton PhD Institute of Dermatology, Lisle Street, London WC2
Until relatively recently, the polyene antibiotics, particularly nystatin, amphotericin B and natamycin have been the drugs of choice for the treatment of candida infections. These polyenes are macrolide antibiotics produced mainly by Streptomyces species. They interact with the membranes of sensitive organisms causing leakage of internal constituents and ultimately cell death. Nystatin is very effective for topical treatment and there are virtually no side effects related to its use. Amphotericin B, unlike either nystatin or natamycin, may be used not only topically but can also be given by the intravenous route. Neither primary resistance nor the development of laboratory demonstrable resistant strains of yeasts during therapy has presented problems during the often prolonged use of these polyene antibiotics. In recent years, the realization of the -antifungal properties of imidazole derivatives has produced three compounds, clotrimazole, miconazole and econazole, which are not only effective against candida and other yeasts, but also against other fungus species pathogenic to man.
Clotrimazole is a tritylimidazole derivative which acts on the cell membrane of fungi and inhibits protein synthesis. The drug is very active in vitro against C. albicans and clinical trials have established that it is as effective as the polyenes when applied to the skin or mucosal surfaces (Clayton & Connor 1973). Miconazole, a phenethalcohol imidazole derivative, has a similar spectrum of activity to clotrimazole and is as effective as clotrimazole when applied topically to treat fungal infections of the skin and mucous membranes (Clayton & Knight 1976). It may also be administered orally or intravenously. There are, as yet, comparatively few reports in the literature on the treatment of systemic candida infections by oral or intravenous miconazole and its value has still to be established. The presence of primary resistance or the development of resistance to imidazoles during treatment has not been a therapeutic problem. Recently, however, Holt & Azmi (1978) 0141-0768/79/120929-02/$01.00/0
X-1 1979 The Royal Society of Medicine
927
a week of the first administration of transfer factor to an extremely high level of 870 units/ml (normal level up to 150 units/ml) and then rapidly fell to a level only slightly above normal. Anticandida IgM rose from 50 to 250 units (normal level up to 100 units/ml) and then fell to 40 units, and anticandida IgA was detected in the serum for the first time. Levels of all three anticandida antibodies are currently in the high normal range, lymphocyte transformation to candida antigen is present and leukocyte migration inhibitory factor is produced on exposure of leukocytes to candida antigen. Changes in nonspecific immune function have been from a PPD negative skin test to positivity after the second unit of transfer factor, and a rise in the responsiveness of peripheral blood lymphocytes to PHA stimulation. No changes in total levels of IgG, IgA or IgM have been detected during the periods of fluctuation of specific anticandida antibodies.
References Higgs J M (1973) Proceedings of the Royal Society of Medicine 66, 802 Higgs J M & Wells R S (1972) Br"itish Journal of Dermatology 86, Suppl; p 28
Kirkpatrick C H, Rich R R & Bennett J E (1971) Annals ofInternal Medicine 74, 955 Lawrence H S (1974) Transfer Factor in Cellular Immunity. Harvey Lecture Series 68. Academic Press, New York; pp 239-250 Nielsen K H, Parrat H D & White R G (1973) Journal of Immunological Methods 3, 301
Transfer factor Anne S Hamblin PhD Immunology Department, St Thomas' Hospital, London SEI 7EH
The term transfer factor is given to dialysable material present in extracts of immune peripheral blood leukocytes which apparently confers ('transfers') delayed-type hypersensitivity (or its in vitro correlates) when injected into a non-hypersensitive subject. Clinical and immunological observations following the administration of transfer factor have led to the concept that it converts non-immune to immune lymphocytes in vivo, thus acting as a mediator of adoptive sensitization (Lawrence 1974). A wide variety of clinical conditions including immunodeficiency, infectious diseases, autoimmunity and malignancy have therefore been treated with transfer factor in attempts to reconstitute cellular immunity. However, the mechanism by which transfer factor is therapeutically beneficial is complicated by clinical observations that patients may acquire cellular immune responsiveness not apparently related to donor specificity (Dupont et al. 1974, Valdimarsson et al. 1974, Spitler et al. 1972, Griscelli 1975, Kirkpatrick & Smith 1976a) suggesting that leukocyte extracts may also possess adjuvant properties. The clinical benefit derived from treatment with transfer factor may therefore depend not only on the initiation of specific immunological reactivity, but also the amplification of an existing immune response. The proposition that both the so-called specific and nonspecific properties of transfer factor may have a contribution to make to the augmentation of cellular immune responses in patients raises a number of practical issues. It is perhaps useful to remember that the term transfer factor is often applied to the whole leukocyte extract administered in vivo. However, it has been suggested that the crude material be termed 'dialysable leukocyte extract' and that the term 'transfer factor' be reserved for the unidentified factor or factors capable of transferring delayed hypersensitivity. The relative roles of the specific and nonspecific activities in the extract raise questions regarding the selection of donors for therapy. It is not clear whether a more beneficial product may be prepared from donors with specific strong delayed hypersensitivity responses or from unscreened pooled normal healthy donors. The question is
0141-0768/79/120927-03/$01.00/0
,'-.
1979 the Royal Society of Medicine
928
Journal of the Royal Society of Medicine Volume 72 December 1979
under assessment in studies in which the product from normal healthy blood-bank donors is administered to patients with a wide range of diseases (Grob 1977) and in properly controlled clinical trials such as those currently under way in the United States in leprosy and coccidiomycosis. Much of the scepticism which surrounded the early work on transfer factor resulted from the consistent failure to demonstrate the transfer phenomenon in animals, particularly the guinea pig. However, recent studies in primates, guinea pigs and infectious disease models in animals have shown that it is possible to demonstrate biological properties of leukocyte extracts which may be relevant to transfer factor in humans. Such studies have revealed both general and specific stimulatory activity in leukocyte dialysates. Thus, Steele etal. (1976) have reported that human leukocyte dialysates could transfer delayed hypersensitivity responses to gnotobiotic monkeys never previously exposed to the relevant antigens. Immunological specificity was implied since recipients never converted to antigens to which the donor did not react in vivo. In contrast, Vandenbark etal. (1977) have reported that delayed hypersensitivity skin reactions could only be elicited in guinea pigs receiving human leukocyte dialysate when the animals had previously been primed with the relevant antigen, thus supporting the view that dialysates contain materials which stimulate the recipient to express sensitivity to antigens to which they have previously been exposed. The concept that transfer factor converts non-immune to immune lymphocytes raises the possibility that non-immune lymphocytes cultured with transfer factor may acquire the ability to respond to specific antigen which may be assessed by a variety of in vitro tests such as lymphocyte transformation, migration inhibition or cytotoxicity. Such specific effects have been reported and have been reviewed (Lawrence 1974, Ascher et al. 1976). However, it is clear that leukocyte dialysates also contain many substances capable of nonspecifically altering lymphocyte responses only some of which may reflect the nonspecific effects in vivo (Hamblin et al. 1976). It is, therefore, essential to bear in mind that extracts of cells may simply improve lymphocyte function in vitro by, for example, medium conditioning effects. Thus it has been reported that human leukocyte dialysate causes cyclic guanosine monophosphate (GMP) accumulation in normal leukocytes (Kirkpatrick & Smith 1976b). Cell extracts contain serotonin and ascorbate known to cause cyclic GMP accumulation. Fractionation of the crude dialysate indicated that the fraction which caused conversion of delayed skin tests also causes cyclic GMP accumulation, but did not contain ascorbate or serotonin. This example reveals the need for careful biochemical studies before conclusions may be drawn regarding the relevance of in vitro activity of the crude dialysate to its effect in vivo. To supplement these studies much effort has been made to characterize the dialysable material extracted from leukocytes biochemically. Analysis of dialysates by gel chromatography has shown that they may be separated into a number of subfractions some of which transfer specific delayed hypersensitivity responses (Zuckerman et al. 1974, Burger et al. 1976) and some of which may nonspecifically enhance delayed hypersensitivity responses in humans (Krohn et al. 1977). In addition, fractions have been described with specific and nonspecific activity in animal and in vitro models (see Ascher et al. 1976). Within leukocyte dialysates a variety of known substances have been identified which could be responsible separately or in combination for some of the reported in vitro or in vivo effects (Wilson et al. 1977, Hamblin 1979). However, the material(s) responsible for the transfer of delayed hypersensitivity has yet to be defined biochemically, and to date no subfraction of the crude dialysate has definitively been shown to be therapeutically beneficial. The lack of available information regarding the mechanisms of action and biochemical nature of transfer factor makes it difficult to describe schedules, dosage and source of material for treatment. Furthermore, a variety of activities have been reported clinically and in animal and in vitro models and the interrelationship between these has yet to be made. One view, expressed by C H Kirkpatrick in 1975, is that 'the phenomenon of transfer factor is a summation of several effects'. The results of ongoing clinical trials, together with continued biochemical and biological analysis of the dialysate, will show whether this hypothesis will be substantiated or refuted.
Journal ofthe Royal Society of Medicine Volume 72 December 1979
929
References Ascher M S Gottlieb A A & Kirkpatrick C H ed (1976) Transfer Factor: Basic Properties and Clinical Applications. Academic Press, New York Burger D R, Vandenbark A A, Daves D, Anderson W A, Vetto R M & Finke P, (1976) Journal of Immunology 117, 789 Dupont D, BaUlow M, Hansen J A, Quick C, Yunis E J & Good R A (1974) Proceedings of the National Academy of Sciences of the USA 71, 867 Grscelli C (1975) Birth Defects 11, 462 Grob P J (1977) Acta Neurologica Scandinavica 55, Suppl; p 217 Hamblin A S (1979) In: Clinical Neuroimmunology. Ed. F Clifford Rose. Blackwell Scientific Publications, London; p497 Hamblin A S, Dumonde D C & Maini R N (1976) Clinical and Experimental Immunology 23, 303 Kirkpatrick C H (1975) Birth Defects 11, 441 Kirkpatrick C H & Smith T K (1976a) Cellular Immunology 27, 232 Kirkpatrick C H & Smith T K (1976b) In: Transfer Factor: Basic Properties and Clinical Applications. Ed. M S Ascher et al. Academic Press, New York; p 161 Krohn K, Grohn P, Horsmanheimo M & VirolainenuM (1977) Medical Biology 54, 334 Lawrence H S (1974) Transfer Factor in Cellular Immunity. Harvey Lecture Series 68. Academic Press, New York; pp 239-250 Spider L E, Levin A S, Blots M S, Epstein N, Fudenberg H H, Helistrom I & Heflstrom K E (1972) Journal of Clinical Investigation 51, 92a Steele R W, Eichberg J W, Heberlung R L, Eller J J Kalter S S & Kniker W T (1976) Cellular Immunology 22, 110 Valdimarsson H, Hambleton G, Henry K & McConnell I (1974) Clinical and Experimental Immunology 16, 141 Vandenbark A A, Burger D R & Vetto R M (1977) Clinical Immunology and Immunopathology 8, 7 Wilson G B, Welch T M, Knapp D R, Horsannheimo A & Fudenberg H (1977) Clinical Immunology and Immunopathology, 8, 551 Zuckerman K S, Neidhart J A, Balcerzak S P & LoBuglio A (1974) Journal of Clinical Investigation 54, 997
Therapy of candidiasis Yvonne Clayton PhD Institute of Dermatology, Lisle Street, London WC2
Until relatively recently, the polyene antibiotics, particularly nystatin, amphotericin B and natamycin have been the drugs of choice for the treatment of candida infections. These polyenes are macrolide antibiotics produced mainly by Streptomyces species. They interact with the membranes of sensitive organisms causing leakage of internal constituents and ultimately cell death. Nystatin is very effective for topical treatment and there are virtually no side effects related to its use. Amphotericin B, unlike either nystatin or natamycin, may be used not only topically but can also be given by the intravenous route. Neither primary resistance nor the development of laboratory demonstrable resistant strains of yeasts during therapy has presented problems during the often prolonged use of these polyene antibiotics. In recent years, the realization of the -antifungal properties of imidazole derivatives has produced three compounds, clotrimazole, miconazole and econazole, which are not only effective against candida and other yeasts, but also against other fungus species pathogenic to man.
Clotrimazole is a tritylimidazole derivative which acts on the cell membrane of fungi and inhibits protein synthesis. The drug is very active in vitro against C. albicans and clinical trials have established that it is as effective as the polyenes when applied to the skin or mucosal surfaces (Clayton & Connor 1973). Miconazole, a phenethalcohol imidazole derivative, has a similar spectrum of activity to clotrimazole and is as effective as clotrimazole when applied topically to treat fungal infections of the skin and mucous membranes (Clayton & Knight 1976). It may also be administered orally or intravenously. There are, as yet, comparatively few reports in the literature on the treatment of systemic candida infections by oral or intravenous miconazole and its value has still to be established. The presence of primary resistance or the development of resistance to imidazoles during treatment has not been a therapeutic problem. Recently, however, Holt & Azmi (1978) 0141-0768/79/120929-02/$01.00/0
X-1 1979 The Royal Society of Medicine
