A comparison of the acute effects of radiation therapy, including or excluding the thymus, on the lymphocyte subpopulations of cancer patients.

Radiation therapy to either mediastinum or pelvis causes a rapid decrease in circulating lymphocytes of both B and T types and in addition an impairment in the function of the remaining lyphocytes, as measured by their ability to proliferate in response to mitogens. The acute depression is short-lived. Substantial recovery is apparent within 3 wk after cessation of therapy; however, most patients show a modest, chronic depression in both numbers and functional capacities of circulating lymphocytes. T cells are somewhat more sensitive than B cells, but both are affected. Irradiation of the thymus per se seems to have little influence on the acute changes which occur, as patients receiving pelvic and mediastinal (including thymic) radiotherapy show a similiar degree of lymphopenia and depression of lymphocyte responsiveness.

The protective actions of some anticholinergic drugs in sarin poisoning.

1. The central and peripheral anticholinergic activities of a series of drugs comprising atropine, hyoscine, caramiphen and one of its analogues, and three glycollic acid esters, have been measured.2. The ability of the same drugs used alone, and in conjunction with N-methyl pyridinium-2-aldoxime methanesulphonate (P2S), to protect mice, rats and guinea-pigs from the lethal effects of sarin has been assessed.3. No correlation was found to exist between central or peripheral anticholinergic activity and ability to protect from sarin.4. On the indirectly stimulated isolated rat phrenic nerve-diaphragm preparation all drugs with the exception of hyoscine caused potentiation of responses to low frequency stimulation but partial block of responses to high frequency stimulation. The drugs did not reverse the effects of sarin on the phrenic nerve-diaphragm preparation.6. It is concluded that a pharmacological action other than an anticholinergic one is involved, in part, in the protective actions against sarin of some of the drugs studied. Whether their effects on skeletal muscle are of any relevance in this respect is unresolved.

Relationship of serum CA125 and lipid-associated sialic acid tumor-associated antigen levels to the disease status of patients with gynecologic malignancies.

Serum samples obtained from 133 patients with gynecologic malignancies were assayed for two tumor-associated antigen markers: CA125, an ovarian marker, and lipid-associated sialic acid, a nonspecific marker. In the patient population, there were 77 papillary serous and 19 unspecified ovarian adenocarcinomas, and 24 miscellaneous ovarian carcinomas. Thirteen patients had nonovarian malignancies. Sixty-nine percent (74 of 108) of the patients with known disease had abnormal CA125 levels, whereas only 32% (20 of 63) had abnormal lipid-associated sialic acid levels. Changes in CA125 serum levels reflected the disease status of the patients for whom there were serial serum samples. Normal levels of CA125 corresponded to no evidence of disease in 100% (six of six) surgically evaluated patients and 75% (30 of 40) of clinically evaluated patients. Changes in CA125 levels from normal to abnormal corresponded to disease progression in 80% (12 of 15) of the patients. Decreases in CA125 levels from abnormal to normal corresponded to complete clinical response in 55% (11 of 20), and partial clinical response in 45% (nine of 20). No such correlations were available for lipid-associated sialic acid antigen levels. For tumors that express CA125 antigen, serum levels appear to be a good marker for the extent of malignant gynecologic disease. Levels of CA125 that rose from normal to abnormal were usually associated with recurrent disease.

Effect of maternal parasitic disease on the neonate.

Parasitic disease is the most common infectious disease complication of pregnancy worldwide, resulting in maternal debilitation and fetal prematurity and low birth weight. The increasing incidence of these diseases in our population led to the present study of 125 patients, 34 of whom were found to be infected with at least one intestinal parasite. In contrast to studies in developing countries, no significant differences in either maternal anemia, or fetal birth weight, or prematurity were found between the infected and non-infected groups. However, there was a three-fold increase in the incidence of significant neonatal hyperbilirubinemia in the parasitized group. Parasitic disease complicating pregnancy in our population does not appear to exert the same adverse effect on mother and fetus as that described in other countries. In view of the limited pathology associated with parasitic disease, treatment, other than with iron and vitamin supplementation, is not routinely indicated in pregnancy in populations similar to ours. However, due to the increased incidence of neonatal jaundice and morbidity we would recommend close observation of the neonates in the immediate postpartum period.

Effect of oral contraceptives on leukocyte phagocytic activity and plasma levels of prostaglandin E2 and thromboxane B2 in normal menstruating women.

Mononuclear and polymorphonuclear cells were isolated from the peripheral blood of normal menstruating women. Four of the subjects were not using oral contraceptives and five were taking various formulations. The women were tested once a week for 12 consecutive weeks. Plasma levels of 6-keto-prostaglandin F2 alpha (6-KF), prostaglandin E2 (PGE2), thromboxane B2 (TxB2), estrogen, and progesterone were measured by specific radioimmunoassays. The phagocytic activity of the mononuclear and polymorphonuclear cells isolated from the peripheral blood was measured with a bacterial phagocytosis and killing assay. The phagocytic activity of both types of cells was depressed perimenstrually in both groups of women. However, examination of individuals showed that those subjects not taking oral contraceptives had a worsening of phagocytic activities with approaching menses while the oral contraceptive subjects generally had an improving of these activities at this time. We were unable to correlate the phagocytic activities with either hormone or prostaglandin levels in the plasma of these subjects. However, the subjects on oral contraceptives had significantly lower levels of PGE2 and TxB2 than those women who were not using oral contraceptives.

The protection of primates against soman poisoning by pretreatment with pyridostigmine.

The effectiveness of pyridostigmine pretreatment against soman poisoning has been determined in rhesus monkeys and marmosets receiving atropine therapy. Pretreatment with the maximum sign-free dose (200 microgram kg-1, i.v.) raised the subcutaneous LD50 of soman by a factor of 28 in rhesus monkeys and 15 in marmosets. The protection afforded by a quarter of the sign-free dose of pyridostigmine was not significantly less. These levels of protection are higher than any reported in non-primate species.

Effect of immunomodulating factors present in ascitic fluids and sera from cancer patients on the responses of cultured mononuclear cells from normal subjects.

Ascitic fluids and sera from patients with malignant tumors were tested for their ability to modulate the mitogen-induced blastogenic responses of normal subjects' peripheral blood mononuclear cells in vitro. The addition of either ascitic fluid or serum to cultures of normal blood cells greatly enhanced the blastogenic response of cells to phytohemagglutinin P, and markedly depressed the responses to concanavalin A and succinyl-concanavalin A. The blastogenic response of the cells to pokeweed mitogen was unaffected by the addition of serum and depressed by the addition of ascitic fluid. Autologous normal serum also enhanced the response to phytohemagglutinin P but had no effect on the response to the other mitogens. These activities were concentration-dependent and heat-stable (56 degrees C, 60 min) and could be detected even if the ascitic fluid or serum was added as late as the second day of culture. Cells that had been preincubated with serum or ascitic fluid and washed well before culturing with the mitogens responded in the same manner as cells cultured in the presence of serum or ascitic fluid. The mitogen-induced blastogenic responses of mononuclear cells were not affected by the addition of autologous cells that had been preincubated with either serum or ascitic fluid, washed, and treated with mitomycin C. Indomethacin (2 X 10(-7)M) did not prevent the ascitic fluid-mediated depression of blastogenic responses of normal cells. The ascitic fluid and serum of these cancer patients appeared to contain a specific immunoinhibiting substance which exerted its effects by a direct action on the responding mononuclear cells and not by the induction of suppressor cells.

Depressed mononuclear cell function in advanced neoplastic disease.

The ascites fluids from 9 patients with invasive gynecologic neoplasms were examined to determine the immunocompetence of the mononuclear ascites cells and the immunoregulatory properties of the cell-free ascitic fluid. Blood mononuclear cells (from 5 patients) were also tested. The mononuclear cells from the cancer patients responded poorly to stimulation with polyclonal mitogens; only the blood mononuclear cell response to pokeweed mitogen was not significantly less than that of normal subjects. The addition of autologous serum or cell-free ascitic fluid to the cell cultures enhanced the response of the cells to phytohemagglutinin and pokeweed mitogen, had little effect on the response to concanavalin-A, and greatly depressed the response to succinyl-concanavalin-A. We found no evidence for the presence of suppressor cells in the ascites cell populations. The data are consistent with the thesis that the depressed immune responses are the result of malnutrition associated with advanced malignant disease.

Response of human adenocarcinoma to chemotherapy: as sole agents and in combination with sodium ibuprofen.

The sensitivity of 12 human tumors to various chemotherapeutic agents was measured in the 6-day subrenal capsule xenograft assay. All of the tumors were adenocarcinomas: 9 ovarian, 2 colon, and one endometrial. Doxorubicin, cisplatin, melphalan, 5- fluorouracil , methotrexate, and vinblastine sulfate were tested for antineoplastic activity on all tumors. In addition, a prostaglandin synthetase inhibitor, sodium ibuprofen, was assayed for cytostatic activity and for the ability to enhance the cytotoxic activity of melphalan and vinblastine sulfate. The response of the tumors to the cytotoxic agents were variable, but 5 of the 12 tumors showed a significant reduction of growth when the animals were treated with sodium ibuprofen alone. The addition of ibuprofen to the chemotherapeutic agents did not significantly alter the therapeutic activity of melphalan, but decreased the effectiveness of vinblastine in one case. When ibuprofen was given in combination with a cytotoxic drug, the primary cytoreductive effect was that of the cytotoxic agent.

Recovery of lymphocyte function after radiation therapy for cancer in relationship to prognosis.

The number and functional abilities of lymphocytes from patients with either breast cancer or cervical cancer were assessed before, during and after radiation therapy given to treat the disease and the patients were followed for several years. Radiation depleted both T and B cells and depressed the responses to PHA and Con-A in all patients groups. The response to mitogens in the weeks immediately following radiation therapy was greater in patients whose disease did not recur and was depressed for a much longer time in patients whose disease recurred within the next few years. This difference was most clearly reflected in the ratio of mitogen response to the number of T-cells. The relationship of responsive to non-responsive or suppressor lymphocytes in the weeks following radiation therapy may be a clue to prognosis.

The immunoinhibitory activities of the lung lavage materials and sera from patients with pulmonary alveolar proteinosis (PAP).

Pulmonary alveolar proteinosis (PAP) is a diffuse pulmonary disease of unknown etiology, characterized by the accumulation of large amounts of amorphous phospholipid containing material in the alveolar spaces. Lavaging the lungs with large amounts of saline to remove the accumulated material results in marked clinical improvement and often complete remission. We have examined theperiperal blood and lung lavage material from 6 PAP patients in an attempt to detect abnormalities of the normal immune response. The complete blood counts and the numbers of T and B lymphocytes and monocytes were normal: the lavage fluids contained large numbers of macrophages, most of which were engorged with the amorphous material, and varying numbers of lymphocytes and polymorphonuclear leukocytes. The lavage material inhibited the blastogenic response of mononuclear cells to mitogens: 3HTdR incorporation in response to stimulation with concanavalin-A and Pokeweed mitogen was inhibited in both allogeneic and autologous cells, while response to phytohemagglutinin stimulation was inhibited only in autologous cells. The sera of the PAP patients inhibited 3HTdR incorporation in response to all three mitogens. Neither the lavage material nor the sera inhibited E-rosette formation, nor were they cytotoxic to allogeneic mononuclear cells. Thus while the peripheral blood monoculear cells from patients with PAP appeared to respond normally in vitro, the sera and lavage material contained substances which inhibited the in vitro response of mononuclear cells toi mitogens by interfering with normal monocyte-lymphocyte interactions. The inhibiting material does not appear to be immunoglobulin.

In vivo antineoplastic activity of various biological response modifiers for tumors of the ovary and breast.

Fourteen pharmacologic agents reported to have biological activities which are directly or indirectly antineoplastic, were assayed for their ability to inhibit the growth of a mouse neoplasia (M5076) and a rat mammary adenocarcinoma (13762) implanted beneath the renal capsule of the host. Ascorbic acid, cimetidine hydrochloride. Corynebacterium parvum, dimethylsulfoxide, naloxone hydrochloride, indomethacin, muramyl-dipeptide, Protein A from Staphylococcus aureus, theophylline, tilorone (analog R11, 877DA), tuftsin diacetate and sodium ibuprofen were completely inactive as antineoplastic agents for these 2 tumors. In fact, theophylline and dimethylsulfoxide seemed to enhance the formation of 13762 metastases. Blue tongue virus and polyinocinic-polycytidylic acid were marginally effective antineoplastic agents for 13762. Polyinocinic-polycytidylic acid was an excellent antineoplastic agent for M5076; this agent not only prevented the growth of M5076, it was oncolytic.

Breast milk lymphocyte response to K1 antigen of Escherichia coli.

Comparison milk and blood lymphocyte blastogenic responses to the K1 antigen of Escherichia coli and lipopolysaccharide (LPS) from E. coli O127,B8 were examined in 16 postpartum women by [3H]thymidine uptake. Rabbit hemolysincoated sheep erythrocyte monolayers were used to deplete macrophages from milk lymphocyte preparations and to enrich for T lymphocytes in order to make milk preparations more comparable to blood preparations. Response was defined as a stimulation index of greater than or equal to 2.0. There was no evidence of selective response to K1 antigen by milk lymphocytes, since both blood and milk lymphocytes responded in four women and neither blood nor milk lymphocytes responded in nine. Milk lymphocytes alone responded to K1 in one woman, whereas blood lymphocytes alone responded in two women. Additional nonpaired milk or blood cultures were available from three women. None of these responded to K1 antigen. Corresponding lymphocyte cultures were stimulated with LPS. A positive K1 response was always accompanied by an LPS response, and the LPS response correlated with the K1 response in 17 of 19 women. Stool cultures examined with an antiserum agar showed no correlation between the presence of K1 E. coli in the stool and milk or blood lymphocyte response to K1 antigen. In the system used here, no selectivity of response of breast milk lymphocytes to K1 antigen was noted.

Accurate predictions of tumor growth in vivo: the subrenal capsule implant site.

An assay system with exquisite predictive qualities is described for four animal tumors: two rat mammary adenocarcinomas, R3230 Ac and 13762; a mouse papillary mammary carcinoma, MXT; and the mouse reticulum cell sarcoma M5076. The tumors were implanted beneath the renal capsule of syngeneic hosts (BDF1 mice or Fischer 344 rats). This site is readily accessible, well-protected, and provides a rich vascular bed, thus controlling many of the variables involved in neoplastic growth. The growth of each implant was recorded as a function of time and/or the initial tumor size. The absolute growth of each of the tumors could be predicted accurately from a simple linear regression analysis of time (i.e., number of days post-implant). Even more accurate predictions of tumor growth were obtained when both variables (days post-implant and initial implant size) were evaluated by multiple regression analysis. The subrenal capsule tumor implant site provides a superior model system to investigate the effects of biological response modifiers and other immunological manipulations on tumor growth in intact normal immunocompetent animals.

Depressed natural killer cell activity in tumor-bearing rats: effect of immunotherapy and cytoreductive chemotherapy.

A single-cell cytotoxicity assay was used to determine the number of effector cells in the peripheral blood of normal and tumor-bearing Fischer 344 female rats which bound to and lysed K562 target cells. Of the blood leukocytes of normal rats 18.4 +/- 2.6% bound to K562 target cells; one-third of the conjugated target cells were killed as evidenced by trypan blue uptake. Cells isolated from the blood of rats bearing mammary adenocarcinoma R3230 or 13762 bound fewer target cells, 6.3 +/- 1.1 and 7.1 +/- 1.8%, respectively. The proportion of dead conjugated target cells was not different from that of normal rat leukocytes if the leukocytes were from rats bearing 13762 but was reduced by one-half if the leukocytes came from rats bearing R3230. Treatment of the tumor-bearing rats with cytoreductive chemotherapy returned the percentage conjugation to target cells to normal levels if the tumor growth was inhibited more than 70%. Treatment of the tumor-bearing rats with immunomodulating agents did not inhibit the growth of the tumors, but treatment with sodium ibuprofen or muramyl dipeptide increased the percentage conjugation significantly.

Structural aspects of immune recognition of lysozymes. III. T cell specificity restriction and its consequences for antibody specificity.

We have studied the relationship between the determinants encountered by T cells on an antigenic molecule and the specificities of the antibodies eventually produced by the B cells with which these T cells cooperate. The number of epitopes on the hen lysozyme (HEL) molecule available to T cell receptors was functionally limited by inducing T cell tolerance to HEL in rabbits. Highly cross-reactive lysozymes were then used to challenge the HEL-unresponsive rabbits. Only T cells which recognize new epitopes on the challenge lysozymes could act as helpers in generating an anti-lysozyme response. Amino acid differences between Japanese quail lysozyme (JEL) and HEL are segregated within a single quadrant of this small antigen molecule. HEL-tolerant rabbits challenged with JEL produced antibodies which were totally cross-reactive with the tolerogen HEL. This result is in contrast to the result obtained in nontolerant rabbits which produced antibodies to JEL which were only 50-70% cross-reactive with HEL. We conclude that T cells restricted to the JEL-unique epitopes were only capable of cooperating with B cells specific for common epitopes shared between JEL and the tolerogen HEL. Turkey lysozyme (TEL), on the other hand, bears different amino acids which are distributed over several regions on the surface of the molecule. Any one HEL-tolerant rabbit developed a restricted response to TEL; in some rabbits the anti-TEL was highly HEL cross-reactive, while in others little cross-reactivity with HEL was observed. Each of four HEL-tolerant rabbits injected with the minimally altered bob-white quail lysozyme possessed the reactive T cells necessary to mount a limited response to this challenge lysozyme, suggesting a diverse library of T cell specificities. Recognition of the small differences between the challenge lysozymes and the T cells of these tolerant rabbits to make a fine discrimination between minimally changed epitopes.

Concordance of combination and single agent chemosensitivity prediction in ovarian carcinoma using the subrenal capsule xenograft assay (SRCA).

The tumors from 62 patients with advanced ovarian adenocarcinoma were assayed by the subrenal capsule xenograft assay (SRCA) for sensitivity to doxorubicin (A), cis-platinum (P), and cyclophosphamide (C), individually and in combination. In some instances only one or two of the individual drugs were assayed; however, the combination, CAP, was always tested. All patients received an optimal surgical debulking (absence of any residual tumor masses greater than or equal to 2 cm) followed by chemotherapy with CAP. Forty-two tumors were predicted to be sensitive to CAP by the SRCA; 51 of 71 (72%) individually tested drugs agreed with this determination. Twenty-one tumors were predicted to be resistant to CAP and 32 of 36 (89%) individually tested drugs agreed with this determination. In this preliminary study, 11 patients had surgically documented partial responses to CAP chemotherapy. All of these patients had tumors which prospectively tested as sensitive to CAP in the SRCA: 13 of 18 (72%) of separately tested drugs were in concordance with this sensitivity. Fourteen patients failed CAP therapy and three of these failures were predicted prospectively by the SRCA: 9 of 9 (100%) of separately tested drugs were in concordance. Thus, there is an overall concordance of 82% (22/27) between the individual components of a combination chemotherapy and the combination therapy itself. It would seem that extrapolations of sensitivity or resistance can be made from the individual components.