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Early activation of human lymphocytes nuclei by isoprinosine
195
ISOPRINOSINE TRIAL IN HERPES ZOSTER. EFFECTS ON DELAYED CUTANEOUS HYPERSENSITIVITY. J, Laude, B. Lesourd, G. Rancurel, S. Doumerc, R. Moulias Service de Neurologie, HSpital de la Salp@tri&re, Paris et Laboratoire d'Immunologie, Facult@ de M@decine Piti6-Salp@tri~re, Paris - FRANCE.
26
18 cases of Herpes zoster were treated with Isoprinosine in an open study and more than 30 cases in a double blind study. In the open study, patients received 50 to 100 mg/kg/day during one or two courses of 8 days with an interval of 5 days free of therapy. In this study vesicles disappeared in all cases (except for one case) in two to six days, without new outbreak. No algy persisted after ten days (except for one case). After one month an increase of the multipuncture assay of Delayed Cutaneous Hypersensitivity (DCH) was observed in all the 15 treated patients tested. Such a variation was not observed in monthly tested normal controls. The magnitude of this increase was not in relation with the DCH before treatment but in negative relation with the duration of the disease before treatment. All the localizations of observed zosters (3 ophtalmic, 6 thoraco-abdominal (one with encephalitis), 5 dorso-abdominal, 4 geniculate ganglion) followed the same type of evolution. In patients with facial pulsy, the regression of the pulsy was slower than the improvement of cutaneous symptoms and DCH increase. In the 3 elderly patients (with ophtalmic zoster) the regression was the same as in younger ones. No residual pain was observed. In the double blind study, patients were treated either with Isoprinosine (50 mg/kg/day during 5 days) or with a placebo in a randomized study. Criteria for efficacity were quickness of vesicles dry up and of pain disappearance, modification of DCH (by multipuncture assay) and of E-rosette restoration capacity (between day I and day 30). The results of this study will be presented.
IMMUNOLOGICAL RESTORATION IN VIVO AND IN VITRO, ISOPRINOSINE THERAPY AND PROGNOSIS OF ACUTE ENCEPHALITIS. B, Lesourd, G, Rancurel, J.M. Huraux, A. Pompidou T C. Jacque, D. Denv~l, A. Buge I R. Moulias Laboratoire d'Immunologie et Service de Neurologie, CHU Piti4-Salp@tri~re, Paris - FRANCE.
27
42 acute encephalitis (19 Necrotic Acute Encephalitis (NAE) and 23 Diffuse Acute Encephalitis (DAE)) were treated with Isoprinosine in intensive care unit. Diagnosis of NAE was established on stupor and/or coma, temporal lesions on EEG and tomodensitometry (TDM), and CSF (lymphocytosis>100 cells). In 11 cases of NAE evidences of herpetic encephalitis were obtained through IF or cerebral biopsy and/or increase of herpes antibodies on CSF. All DAE presented multifocal lesions on EEG and TDM, and in 7 cases a coma (7 CDAE). All patients (NAE and CDAE) were immuno-depressed as judged on multipuncture ( p ~ O , O 0 2 for NAE, p < 0 , 0 2 for DAE), PHA stimulation ( p < O , 0 5 ) ; all of them presented a serum blocking factor for PHA stimulation and a low E-rosette restoration activity. High concentrations of protein basic encephalitogen by RAI assay were observed in CSF from NAE. The activity of Isoprinosine was tested in vitro through Lymphocyte Nuclear Refringency Test (LNRT) and in vivo by multipuncture before and after each course of Isoprinosine. Isoprinosine was administred IV and/or per os (100 mg/kg/day) for 7 to 10 days, and the patients were tested again. When incomplete restoration was observed, new courses of Isoprinosine (75 or 50 mg/kg/day per os for 5 to 7 days) were performed. Between each course there are always 5 to 7 days without Isoprinosine treatment. Correlation between the in vitro activity of Isoprinosine by LNRT and the in vivo activity by multipuncture was significant (p~O,01). Prognosis is function of initial diagnosis (4 deaths all NAE (3 herpes), 4 severe sequelae (2 NAE (I herpes), 2 CDAE), 6 benign sequelae (compatible with a normal life (5 NAE (3 herpes), I CDAE), and 28 complete recoveries (8 NAE (4 herpes), 4 CDAE and the 16 DAE without coma), but also of the precocity of treatment and of the speed of improvement of the CMI assays.
IN VITRO ACTIVITY OF DAUNOMYCIN-ANTI-ALPHAFOETOPROTEIN CONJUGATES ON AFP PRODUCING CELLS. Michel Pag~ and ~ r t h e Belles-Isles, Department of Biochemistry, Faculty of Medicine, Universit~ Laval, Quebec, Canada. Chemotherapeutic agents currently used for antitumour therapy are selected for their toxicity towards rapidly proliferating cells. Most of them cause undesirable systemic effects such as cardiac or renal toxicity, marrow aplasia, alopecia, nausea and vomiting. During the last few years many authors have tried to eliminate these side effects by increasing the availability of the drug to the tumour site. Enzymes, radioisotopes, DNA, toxins, various macromolecules, and antibodies against fibrin or against tumour-specific
28
196
surface antigens were bound to drugs in an attempt to increase the selectivity of the chemotherapeutic agents, or to decrease their toxic effects on normal cells. The targeting of drugs to the tumour by antibodies to surface antigens may have considerable implications by increasing the therapeutic index. The use of daunomycin covalently linked to antialphafoetoprotein antibody was evaluated in vitro as a potential antitumor drug. The activity of the conjugate was compared to the one of the free drug, the free antibody or a mixture of both agents by inhibition of colony formation, inhibition of methyl-3H-thymidine incorporation and antimitotic activity. Results show that the conjugate binds to tumor cells. A positive dose-response relationship was obtained for the drug-antibody conjugate concentration. This approach suggests the potential application of antioncofoetal proteins antibody for targeting drugs in cancer treatment. (Supported by the Medical Research Council of Canada, the National Cancer Institute of Canada for a scholarship to one of us M.P. and Le Conseil de la Recherche en Sant~ du Quebec for a studentship to one of us M.B.I.)
29
EARLY ACTIVATION OF HUMAN LYMPHOCYTES NUCLEI BY ISOPRINOSINE A. Pompidou, B. Mac~, D. Esnous, C. Mayrand. C.H.U. COCHIN. 24, Fbg. St. Jacques. 75014 PARIS.
Dispersion of chromatin with decrease of nuclear refringence index reflects one of the early events of nuclei activation. Isoprinosine is a synthetic immunomodulator, obtained by combination of Inosine and Dimethyl aminopropanol-paracetamidobenzoate (~IPsalt). Isoprinosine affects the lymphocytes nuclei. Significant diminution of the nuclear refringence index is obtained by Isoprinosine alone or associated with lectins. This is not observed with Hypoxantine, Inosine or DIPsalt. The sensivity to Isoprinosine has been tested "in vitro" on 150 normal subjects, 30% of this population is insensitive. Evaluation by nuclear refringence test of the response to Con A and/or Isoprinosine enables us to measure the functiona! capacity of human blood lymphocytes and is useful in choosing an appropriate "pro host" immunotherapy. Isoprinosine is compared through nuclear refringence test to other immunomodulators such as B.C.G., C. Parvum, C. Granulosum and Muranyl Dipeptide.
3@
IN VITRO AND IN VIVO STUDY OF ISOPRINOSINE (ISO) ON I ~ U N E RESPONSES IN AN ANIMAL MODEL OF HUMAN OSTEOSARCOMA (Os). K.Y. Tsang and H.H. Fudenberg. Department of Immunology, Medical University of South Carolina, Charleston, S.C., U.S.A.
The effects of IS(] on the immune responses of normal hamsters and hamsters with Os were investigated. Os was induced in inbred hamsters (LSH/ssLAK) by techniques described by Singh and Tsang. Laparotomies were performed on pregnant (12-14 days gestation) hamsters. The uterus was exposed and the fetuses (6-10 per hamster) identified. A homogenate of 2x106 TE-85 cells (derived from a biologically proven Os) in 0.i ml of medium 199 was injected into each fetus through the intact uterine wall 2x10 ° TE-85-M-MSV cells (TE-85 cells infected with MSV-RD-II4 pseudo type virus) suspended in 0.25 ml of medium 199 were injected adjacent to the midshaft of the femur of 4 day old newborn hamsters which have previously been tolerated with TE-85 cell homogenate. In vitro, ISO increased ConA-induced proliferation of peripheral blood lymphocytes (PBL) fr--6-{normal and Os hamsters by 50-65% over controls (ConA alone). When ISO was injected i.p. on 3 successive days, increased ConA-induced PBL proliferation (50-55%) was seen 2-4 days after the last injection in both normal and Os hamsters. Similarly, the functional chemotactic index for monocytes of Os hamsters was significantly increased (mean index 49.2±6.4 to 69.4±10.1) 2-4 days after ISO. These results indicate that ISO can augment proliferation of Os PBL in vitro and can increase the functional capacity of the immune system in animals with Os. Experiments on the survival rate of Os hamsters with treatment of ISO alone and ISO in combination with transfer facter are in progress and the results will be discussed in detail.
ISOPRINOSINE TRIAL IN HERPES ZOSTER. EFFECTS ON DELAYED CUTANEOUS HYPERSENSITIVITY. J, Laude, B. Lesourd, G. Rancurel, S. Doumerc, R. Moulias Service de Neurologie, HSpital de la Salp@tri&re, Paris et Laboratoire d'Immunologie, Facult@ de M@decine Piti6-Salp@tri~re, Paris - FRANCE.
26
18 cases of Herpes zoster were treated with Isoprinosine in an open study and more than 30 cases in a double blind study. In the open study, patients received 50 to 100 mg/kg/day during one or two courses of 8 days with an interval of 5 days free of therapy. In this study vesicles disappeared in all cases (except for one case) in two to six days, without new outbreak. No algy persisted after ten days (except for one case). After one month an increase of the multipuncture assay of Delayed Cutaneous Hypersensitivity (DCH) was observed in all the 15 treated patients tested. Such a variation was not observed in monthly tested normal controls. The magnitude of this increase was not in relation with the DCH before treatment but in negative relation with the duration of the disease before treatment. All the localizations of observed zosters (3 ophtalmic, 6 thoraco-abdominal (one with encephalitis), 5 dorso-abdominal, 4 geniculate ganglion) followed the same type of evolution. In patients with facial pulsy, the regression of the pulsy was slower than the improvement of cutaneous symptoms and DCH increase. In the 3 elderly patients (with ophtalmic zoster) the regression was the same as in younger ones. No residual pain was observed. In the double blind study, patients were treated either with Isoprinosine (50 mg/kg/day during 5 days) or with a placebo in a randomized study. Criteria for efficacity were quickness of vesicles dry up and of pain disappearance, modification of DCH (by multipuncture assay) and of E-rosette restoration capacity (between day I and day 30). The results of this study will be presented.
IMMUNOLOGICAL RESTORATION IN VIVO AND IN VITRO, ISOPRINOSINE THERAPY AND PROGNOSIS OF ACUTE ENCEPHALITIS. B, Lesourd, G, Rancurel, J.M. Huraux, A. Pompidou T C. Jacque, D. Denv~l, A. Buge I R. Moulias Laboratoire d'Immunologie et Service de Neurologie, CHU Piti4-Salp@tri~re, Paris - FRANCE.
27
42 acute encephalitis (19 Necrotic Acute Encephalitis (NAE) and 23 Diffuse Acute Encephalitis (DAE)) were treated with Isoprinosine in intensive care unit. Diagnosis of NAE was established on stupor and/or coma, temporal lesions on EEG and tomodensitometry (TDM), and CSF (lymphocytosis>100 cells). In 11 cases of NAE evidences of herpetic encephalitis were obtained through IF or cerebral biopsy and/or increase of herpes antibodies on CSF. All DAE presented multifocal lesions on EEG and TDM, and in 7 cases a coma (7 CDAE). All patients (NAE and CDAE) were immuno-depressed as judged on multipuncture ( p ~ O , O 0 2 for NAE, p < 0 , 0 2 for DAE), PHA stimulation ( p < O , 0 5 ) ; all of them presented a serum blocking factor for PHA stimulation and a low E-rosette restoration activity. High concentrations of protein basic encephalitogen by RAI assay were observed in CSF from NAE. The activity of Isoprinosine was tested in vitro through Lymphocyte Nuclear Refringency Test (LNRT) and in vivo by multipuncture before and after each course of Isoprinosine. Isoprinosine was administred IV and/or per os (100 mg/kg/day) for 7 to 10 days, and the patients were tested again. When incomplete restoration was observed, new courses of Isoprinosine (75 or 50 mg/kg/day per os for 5 to 7 days) were performed. Between each course there are always 5 to 7 days without Isoprinosine treatment. Correlation between the in vitro activity of Isoprinosine by LNRT and the in vivo activity by multipuncture was significant (p~O,01). Prognosis is function of initial diagnosis (4 deaths all NAE (3 herpes), 4 severe sequelae (2 NAE (I herpes), 2 CDAE), 6 benign sequelae (compatible with a normal life (5 NAE (3 herpes), I CDAE), and 28 complete recoveries (8 NAE (4 herpes), 4 CDAE and the 16 DAE without coma), but also of the precocity of treatment and of the speed of improvement of the CMI assays.
IN VITRO ACTIVITY OF DAUNOMYCIN-ANTI-ALPHAFOETOPROTEIN CONJUGATES ON AFP PRODUCING CELLS. Michel Pag~ and ~ r t h e Belles-Isles, Department of Biochemistry, Faculty of Medicine, Universit~ Laval, Quebec, Canada. Chemotherapeutic agents currently used for antitumour therapy are selected for their toxicity towards rapidly proliferating cells. Most of them cause undesirable systemic effects such as cardiac or renal toxicity, marrow aplasia, alopecia, nausea and vomiting. During the last few years many authors have tried to eliminate these side effects by increasing the availability of the drug to the tumour site. Enzymes, radioisotopes, DNA, toxins, various macromolecules, and antibodies against fibrin or against tumour-specific
28
196
surface antigens were bound to drugs in an attempt to increase the selectivity of the chemotherapeutic agents, or to decrease their toxic effects on normal cells. The targeting of drugs to the tumour by antibodies to surface antigens may have considerable implications by increasing the therapeutic index. The use of daunomycin covalently linked to antialphafoetoprotein antibody was evaluated in vitro as a potential antitumor drug. The activity of the conjugate was compared to the one of the free drug, the free antibody or a mixture of both agents by inhibition of colony formation, inhibition of methyl-3H-thymidine incorporation and antimitotic activity. Results show that the conjugate binds to tumor cells. A positive dose-response relationship was obtained for the drug-antibody conjugate concentration. This approach suggests the potential application of antioncofoetal proteins antibody for targeting drugs in cancer treatment. (Supported by the Medical Research Council of Canada, the National Cancer Institute of Canada for a scholarship to one of us M.P. and Le Conseil de la Recherche en Sant~ du Quebec for a studentship to one of us M.B.I.)
29
EARLY ACTIVATION OF HUMAN LYMPHOCYTES NUCLEI BY ISOPRINOSINE A. Pompidou, B. Mac~, D. Esnous, C. Mayrand. C.H.U. COCHIN. 24, Fbg. St. Jacques. 75014 PARIS.
Dispersion of chromatin with decrease of nuclear refringence index reflects one of the early events of nuclei activation. Isoprinosine is a synthetic immunomodulator, obtained by combination of Inosine and Dimethyl aminopropanol-paracetamidobenzoate (~IPsalt). Isoprinosine affects the lymphocytes nuclei. Significant diminution of the nuclear refringence index is obtained by Isoprinosine alone or associated with lectins. This is not observed with Hypoxantine, Inosine or DIPsalt. The sensivity to Isoprinosine has been tested "in vitro" on 150 normal subjects, 30% of this population is insensitive. Evaluation by nuclear refringence test of the response to Con A and/or Isoprinosine enables us to measure the functiona! capacity of human blood lymphocytes and is useful in choosing an appropriate "pro host" immunotherapy. Isoprinosine is compared through nuclear refringence test to other immunomodulators such as B.C.G., C. Parvum, C. Granulosum and Muranyl Dipeptide.
3@
IN VITRO AND IN VIVO STUDY OF ISOPRINOSINE (ISO) ON I ~ U N E RESPONSES IN AN ANIMAL MODEL OF HUMAN OSTEOSARCOMA (Os). K.Y. Tsang and H.H. Fudenberg. Department of Immunology, Medical University of South Carolina, Charleston, S.C., U.S.A.
The effects of IS(] on the immune responses of normal hamsters and hamsters with Os were investigated. Os was induced in inbred hamsters (LSH/ssLAK) by techniques described by Singh and Tsang. Laparotomies were performed on pregnant (12-14 days gestation) hamsters. The uterus was exposed and the fetuses (6-10 per hamster) identified. A homogenate of 2x106 TE-85 cells (derived from a biologically proven Os) in 0.i ml of medium 199 was injected into each fetus through the intact uterine wall 2x10 ° TE-85-M-MSV cells (TE-85 cells infected with MSV-RD-II4 pseudo type virus) suspended in 0.25 ml of medium 199 were injected adjacent to the midshaft of the femur of 4 day old newborn hamsters which have previously been tolerated with TE-85 cell homogenate. In vitro, ISO increased ConA-induced proliferation of peripheral blood lymphocytes (PBL) fr--6-{normal and Os hamsters by 50-65% over controls (ConA alone). When ISO was injected i.p. on 3 successive days, increased ConA-induced PBL proliferation (50-55%) was seen 2-4 days after the last injection in both normal and Os hamsters. Similarly, the functional chemotactic index for monocytes of Os hamsters was significantly increased (mean index 49.2±6.4 to 69.4±10.1) 2-4 days after ISO. These results indicate that ISO can augment proliferation of Os PBL in vitro and can increase the functional capacity of the immune system in animals with Os. Experiments on the survival rate of Os hamsters with treatment of ISO alone and ISO in combination with transfer facter are in progress and the results will be discussed in detail.