[CANCER RESEARCH 41, 1133-1139, March 1981] 0008-5472/81 /0041-OOOOS02.00 Differential Effects of an Immunosuppressive Fraction from Ascites Fluid of Patients with Ovarian Cancer on Spontaneous and Antibody-dependent Cytotoxicity1
Alison M. Badger,2 Sue K. Oh, and Frederick R. Moolten
Department of Microbiology and the Cancer Research Center, Boston University School of Medicine, Boston, Massachusetts 02118
ABSTRACT In ADCC, effector (killer) cells bearing receptors for the Fc portion of antibody (FcR) bind to and lyse antibody-coated The ascites fluids from humans with cancer that has me- target cells (36, 45). This mechanism of cell injury has been tastasized to the peritoneum is suppressive of the immune implicated in the immune response to a wide variety of tumors, response both in vitro and in vivo. The active moiety is present allografts, and infectious agents in humans and experimental in a high-molecular-weight fraction which élûtesinthe void animals (5, 8, 11, 17,47). volume of a Sephadex G-200 column. This fraction (designated Our laboratory has carried out a number of studies on the Peak I) has been shown to inhibit a number of in vitro responses immunosuppressive effects of ascites fluids from patients with and will inhibit the antibody response to sheep red blood cells cancer metastatic to the peritoneum. These fluids are inhibitory in vivo. In this report, the Peak I protein fraction from the of a variety of immune responses both in vitro and in vivo. In ascites of patients with ovarian cancer is shown to inhibit this report, we have examined the effect of a high-molecular- spontaneous cytotoxicity of human mononuclear cells against weight fraction isolated from the ascitic fluids of 2 patients with the myeloid cell line K562 and against the T-lymphoid cell line ovarian cancer on the NK and ADCC activity of human periph Molt-4F. This fraction was active at concentrations of 1 to 3 eral mononuclear cells. mg/ml. In contrast to this finding, it was not possible to dem onstrate an inhibition of antibody-dependent cell cytotoxicity MATERIALS AND METHODS against chicken red blood cells. Preincubation of the effector cells with Peak I prior to addition to the chicken red blood cell Effector Cells. Mononuclear cells were isolated from hepa- targets or modification of the antibody concentration in the rinized human peripheral blood from normal healthy volunteers assay did not result in suppression; in fact, in many experi on Ficoll-Hypaque (Pharmacia Fine Chemicals, Inc., Piscata- ments, the Peak I potentiated cytotoxicity against chicken red way, N. J.) density gradients. The cells were washed 3 times blood cells. The Peak I proteins were also tested in an antibody- in HBSS and resuspended at the desired concentration in dependent cell cytotoxicity assay against a transformed cell Roswell Park Memorial Institute Tissue Culture Medium 1640 line (HeLa cells), and there was no significant suppression of (Flow Laboratories, Rockville, Md.) containing 10% heat-inac cytotoxicity. Peak I protein fractions prepared as controls from tivated (56°,30 min) fetal calf serum (Flow Laboratories), 2 HIM normal human serum and a congestive heart failure fluid were glutamine, penicillin (100 units/ml), and streptomycin (100 ¡ig/ not suppressive, whereas the Peak I fraction from a cirrhotic ml). This medium will be referred to as RPMI-10%. fluid was suppressive of natural killing activity. Target Cells. The myeloid cell line K562 and the T-lymphoid cell line Molt-4F were maintained in stationary suspension in INTRODUCTION 75 sq cm plastic flasks in RPMI-10%. The tumor cells were passaged every 3 days and on the day preceding an assay. Normal peripheral blood mononuclear cells are spontane HeLa cells were also maintained in RPMI-10% and passaged ously cytotoxic to some transformed and tumor cells in vitro (14, 46, 54). This cytotoxicity or NK3 is now a well-recognized by standard techniques every 3 to 4 days and on the day preceding an assay. CRBC were obtained from Grand Island phenomenon that is particularly effective with human lympho Biological Co. (Grand Island, N. Y.) and were washed 2 times cytes (41, 49) and has been described in mouse (15, 25) and in 0.15 N NaCI and 2 times in HBSS prior to use. rat (39) systems. NK takes place in the absence of any appar s1CrLabeling of Target Cells. Approximately 5 x 106 K562, ent antigenic stimulation. The possible significance of NK as a Molt-4F, or HeLa cells (in suspension) and 108 CRBC were mechanism for immunosurveillance has generated much inter resuspended in 1 ml RPMI-10%. One to 200 /iCi of Na251CrO4 est (22, 26, 27), and while the in vivo significance of NK in (specific activity, 200 to 500 Ci/g chromium; New England humans is not completely understood, there are several animal Nuclear, Boston, Mass.) were added to the target cells which models where tumor resistance has been correlated to levels were then incubated at 37° for 1 to 2 hr with occasional of NK (4, 13). A second in vitro cytotoxicity reaction is ADCC. shaking. Care was taken to keep the HeLa cells in suspension. ' This work was supported by Grant CA-15129 from the National Cancer Labeled target cells were washed 3 times in HBSS, resus Institute. pended in RPMI-10%, and adjusted to the desired concentra 2 To whom requests for reprints should be addressed, at Biological Research, tion. Smith, Kline & French Laboratories, 1500 Spring Garden St., Philadelphia, Pa. 19101. Preparation of Ascitic Proteins. Sterile ascites fluids were 3 The abbreviations used are: NK, natural killing; ADCC, antibody-dependent cell cytotoxicity; HBSS, Hanks' balanced salt solution; CRBC, chicken red blood obtained from 2 patients with ovarian cancer metastatic to the cells; E:T, effectortarget. peritoneal cavity. These patients had not been treated with Received November 28, 1979; accepted December 12, 1980. either chemotherapy or radiotherapy, and their ascites fluids
MARCH 1981 1133
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. A. M. Badger et al. have previously been shown to be immunosuppressive (1). These particular fluids were used for these experiments as they are currently being fractionated for purification of the immu nosuppressive moiety (40). The fluids had a protein concentra tion of approximately 50 mg/ml and were dialyzed against phosphate-buffered saline (pH 7.4) and applied to Sephadex G-200 columns. The elution profile of a typical column run is shown in Chart 1. The high-molecular-weight fraction eluting first from the column (designated Peak I) has been shown to be immunosuppressive in a number of in vitro systems (1, 2,
40). This fraction was dialyzed against water and lyophilized 200 250 and stored at -20°. Prior to testing, the Peak I was solubilized EFFLUENTVOLUME(ML) in RPMI-10% and sterilized by Millipore filtration (Ha, 0.45 /im). Chart 1. Sephadex G-200 filtration profile of an ascitic fluid obtained from a We have performed this fractionation procedure with a number patient with ovarian cancer. Proteins eluting from the column were pooled into 4 fractions (Peaks I to IV) and then dialyzed against distilled water and lyophilized. of different cancer ascites, and all Peak I fractions have been BSA. bovine serum albumin. immunosuppressive. The same purification procedure was used to prepare Peak I fractions from normal human serum, was for 18 or 40 hr. The supernatants were harvested and the fluid from a patient with congestive heart failure, and the counted as before, and the percentage of cytotoxicity was fluid from a patient with cirrhosis of the liver. The profile on calculated. Sephadex G-200 is similar to the cancer ascites in all cases, ADCC activity of the mononuclear cells against CRBC was with some variation in the volumes of the different peaks. Three also determined. 51Cr-CRBC were adjusted to a concentration additional control protein samples were included in several of of 107/ml, and 50 /¿Iwerepipetted into each well. Effector cells the experiments. These were freshly drawn normal human were added at 3 different concentrations giving E:T ratios of 4: serum, Peak II proteins, and Peak III proteins which we have 1,2:1, and 1:1. Anti-CRBC antibody was obtained from Cappel shown to be inactive as immunosuppressants in vitro (1, 2). Laboratories, Inc. (Cochranville, Pa.), and 25 /il of 1:1000 Assay for NK Activity. Effector cells were tested against dilution were added to the microtiter plates. 51Cr-labeled K562 and Molt-4F target cells. Triplicate cultures In all ADCC experiments, the background spontaneous cy were established in round-bottomed microtiter plates in a total totoxicity of the cells against CRBC and HeLa cell targets was volume of 200 /il. Effector cells were added in 100 /tl medium examined in the absence of antibody. In addition, the Peak I to give E:T cell ratios of 100:1, 50:1, 25:1, and 12.5:1. A and control proteins were incubated with target cells alone and constant number of 51Cr-labeled target cells (2 x 10") were with target cells and antibody. There was no additional spon added to each well in 50 /il medium. The volume in each well taneous release of 51Cr label from the target cells due to was brought to 200 ¿dwith RPMI-10%, Peak I, or control antibody or sample addition. protein. The cultures were incubated at 37°in 5% CO2 incu bator for 4 or 16 hr, and the supernatants were harvested using RESULTS a Titertek supernatant collection system (Flow Laboratories). The amount of chromium released into the supernatant was The immunosuppressive activity of Peak I protein from an determined in a Beckman 300 system gamma counter. Cyto- ovarian ascites fluid on NK against K562 targets in a 4-hr toxicity was calculated as follows: assay is shown in Chart 2. There was a dose-dependent experimental 5'Cr release suppression of NK activity at all 3 E:T cell ratios that were - spontaneous 51Cr release tested. The Peak I material was not cytotoxic to either the % of cytotoxicity = total 5'Cr counts x 100 effector or the target cells. In the experiment shown in Table 1, - spontaneous 5'Cr release the Peak II proteins were included as controls, and it can be seen that there was no suppression of the control response. In all experiments, spontaneous 51Cr release was measured The Peak I fraction (3 mg/ml) was suppressive at all E:T ratios with the target K562 and Molt-4F cells alone and target cells tested. The Peak III proteins were included in many of the incubated with Peak I or control proteins. In no case did the experiments and never showed any activity. Normal human added samples cause toxicity to the target cells. This absence serum and human serum albumin have also been used as of toxicity was shown by trypan blue exclusion and the lack of controls in these experiments without demonstrating any im increased spontaneous release of radioactivity of the target munosuppressive activity. cells. The immunosuppressive activity from a second Peak I frac Assay for ADCC. The ADCC assay against HeLa targets was tion of ovarian ascites fluid is shown in Chart 3. This fraction established in a manner similar to that for NK against K562 in was highly suppressive with 100% suppression being obtained that similar numbers of effector and target cells were utilized. with 1.25 mg protein per ml. In this experiment, we also tested Antibody against HeLa cells was prepared in rabbits by re the Peak I fractions from normal human serum, the fluid from peated injections of 107 to 108 live HeLa cells i.p. Rabbits were a patient with congestive heart failure, and the fluid from a bled by cardiac puncture, and the y-globulin fraction of serum patient with cirrhosis of the liver. Various concentrations of the was prepared. Lyophilized y-globulin was suspended in HBSS Peak I fractions were tested on NK activity at a 50:1 E:T ratio. at a concentration of 4 mg/ml and titered to determine the High concentrations of the Peak I fractions from normal human concentration required for maximum killing (1:1000). Twenty- serum and from the congestive heart failure fluid were required five /il of antisera were added to the cultures, and incubation to give suppression of the response (approximately 5 mg
1134 CANCER RESEARCH VOL. 41
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. Modulation of NK and ADCC by Ascites Proteins
was no longer apparent at 40 hr, the effect of Peak I was examined in an 18-hr ADCC assay against CRBC. The results
70 of this experiment are shown in Chart 4. It can be seen that the Peak I was still unable to suppress the ADCC; in fact, as noticed previously, these proteins enhanced the cytotoxicity 60 for CRBC. This increase in killing was not due to toxic effects of Peak I on the target cells because the spontaneous release 50 of 51Cr from labeled CRBC incubated with Peak I was no «G/ML different to the release from CRBC incubated alone. 5 10 The absence of inhibition by Peak I in this assay could be MG/ML due to some form of competition between the antibody and the 30 immunosuppressive moiety for a site on the lymphocyte sur face. Effector cells were therefore incubated with Peak I (3
20 mg/ml) in microtiter plates for 3 hr prior to the addition of antibody and CRBC. The cultures were then incubated for an additional 18 hr. As seen in Chart 5, despite the 3-hr preincu- 10 bation of the effector cells with inhibitory material, there was still no evidence that the Peak I could inhibit the ADCC in an 25-1 50-1 100-1 18-hr assay. Cordier et al. (7) have demonstrated that high concentrations of a2-macroglobulin (a protein which is present EFFECTOR:TARGETRATIO in the Peak I fraction) can inhibit ADCC against CRBC and that Chart 2. Reduction in NK activity of human mononuclear cells against K562 cells in a 4-hr assay by 3 different concentrations of Peak I proteins. this inhibition is markedly increased upon dilution of the anti- CRBC antibody. In order to test this in our system, the ADCC Table 1 Effect of Peak I and Peak II protein fractions on NK activity of human peripheral blood mononuclear cells ion cytotoxicityE:T % of
sup 80 II I pres sion360 ratio100:1 (3mg/ml)56.9 (3mg/ml)21.0 ±5.16 ±2.9° ±3.7 50:1 52.7 ±1.7 46.7 ±3.0 22.3 ±2.5 58 25:1aControl51.9 43.5 ±5.4Peak56/cytotoxicity43.0 ±3.5Peak 19.0 ±3.4%of « 60
of Peak l-treated cellsN i<*.<-.inn-7T1nn I I y 1nn \ cytotoxicity of control cells / 40 Mean ±S.D Treated cultures were significantly different from untreated control cultures protein per ml was required to suppress the response by 50%). 20 The Peak I fraction from the cirrhosis fluid, on the other hand, gave approximately 75% suppression with 2.5 mg protein per ml. We have demonstrated this immunosuppressive activity in cirrhosis fluid previously (1). The high NK of the control cells 0.625 1.25 2.5 5.0 (80% at 50:1) may reflect a change in the susceptibility of the PROTEINCONCENTRATION(MG/HL) K562 target cells while being maintained in culture. We have Chart 3. Effect of 4 Peak I fractions on the NK activity of human mononuclear also tested the ascites Peak I proteins on NK against the T- cells against K562 target cells in a 4-hr assay. Four protein concentrations lymphoid cell line Molt-4F. These results are shown in Table 2. (0.625 to 5.0 mg/ml) were tested on a 50:1 E:T ratio. O, congestive heart failure; Again, the Peak I proteins were suppressive of NK activity, A. normal human serum; •cirrhosis; •,ovarian cancer. Bars. S.D. whereas the Peak II and the Peak III protein fractions were Table 2 inactive. Effect of Peak I, II. and III proteins on NK activity of human mononuclear cells The effect of the immunosuppressive fraction on ADCC ac against the Molt-4F cell line tivity of human mononuclear cells against CRBC and HeLa cell cytotoxicityE:T % of targets was then examined. Table 3 shows the results of a 40- I II III hr ADCC assay against CRBC. Using similar concentrations of ratio1 (3mg/ml)20.7 (3mg/ml)56.8 (3mg/ml)51.2 ±0.9a ±1.66 Peak I proteins as in the NK experiments, we found that there 00: 1 ±7.8 ±4.2 was no immunosuppression at any of the E:T ratios tested or 50:1 52.7 ±4.9 17.9 ±2.0 55.1 ±7.0 44.6 ±4.4 NDC at any Peak I concentration tested. There was, in fact, an 25:1Control51.046.7 ±1.0Peak 13.9 ±5.0Peak 54.2 ±3.7Peak enhancement of cytotoxicity at the highest E:T ratios, although Mean ±S.D. All Peak l-treated cultures were significantly different from control cultures this was not always significant. In order to negate the possibility (p< 0.001). that an early suppression of ADCC may have taken place which c ND. not determined.
MARCH 1981 1135
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. A. M. Badger et al.
Table 3 DISCUSSION Effect of Peak I proteins on antibody-dependent cellular cytotoxicity of human tnononuclear cells against 5'Cr-CR6C Human mononuclear cells are naturally cytotoxic for the Cultures were incubated for 40 hr prior to harvesting. Statistical evaluations myeloid cell line K562 and the T-lymphoid cell line Molt-4F. have not been included as there was clearly no ¡mmunosuppressive effect exerted by the Peak I proteins. We have demonstrated that a high-molecular-weight fraction of % of cytotoxicity at following E:T ratios human ovarian cancer ascites fluid (designated Peak I) which has previously been shown to be ¡mmunosuppressive (1, 2, 40) Treatment 4:1 2:1 1:1 will inhibit this activity in a dose-dependent fashion. In direct ControlculturesPeak untreated contrast, the Peak I proteins are unable to inhibit ADCC activity 13 mg/ml1.5 of the same mononuclear cell fraction against CRBC or HeLa mg/ml0.75 cell targets. In fact, in most of the experiments, we have found mg/ml526159595352505143443840 that these proteins will enhance cytotoxicity against CRBC in
100 100r
80 PEAKl CONTROL
LLS PREINCUBATED KITHPEAK 1 60 £ 60
10
20 20
25-1 50-1 100-1 12.5-1 25-1 50-1 100-1 EFFECTOR:TARGETRATIO EFFECTORiTARGETRATIO Chart 4. Inability of Peak I proteins from ascites fluid (3 mg/ml) to inhibit ADCC of human mononuclear cells against antibody-coated CRBC target cells in Chart 5. ADCC activity of human mononuclear cells against CRBC targets an 18-hr assay. •,control untreated cultures; O, cultures containing Peak I following preincubation with Peak I proteins. •control untreated cultures: O, cultures preincubated with Peak I proteins (3 mg/ml) for 3 hr prior to addition of proteins (3 mg/ml). CRBC and antibody. Bars, S.D. assay was established with 3 different concentrations of anti 70 r body: 1:1,000 (optimum); 1:10,000; and 1:50,000 (subopti mum). The Peak I proteins (3 mg/ml) were then tested for 60 1-1,000 inhibitory activity at each antibody dilution. As can be seen in Chart 6, increasing dilutions of the antisera resulted in decreas 50 ing cytotoxicity against CRBC. There was, however, no evi dence that Peak I proteins could inhibit the ADCC activity even >- 5 to at the lowest dilution of antibody. x ADCC activity of human peripheral mononuclear cells against g 1-10,000 an erythrocyte target (CRBC in this case) may not be identical «-> 30 to ADCC activity against a transformed cell line. There is M evidence to suggest that different cells are involved. We have _g 1-50,000 therefore established an ADCC assay against HeLa cells and 20 have examined the effect of Peak I proteins in this assay. The results of one of these experiments is shown in Chart 7. Human 10 mononuclear cells rarely display any natural cytotoxicity against HeLa cell targets. In one or 2 experiments, some activity could be detected, but it was minimal. In this experi 25-1 50-1 100-1 ment, no NK activity could be detected, but we were able to demonstrate good ADCC activity. As with the experiments EFFECTOR:TARGETRATIO utilizing CRBC as targets, we were unable to demonstrate any Chart 6. Effect of Peak I proteins on ADCC activity of human mononuclear cells against CRBC targets in an 18-hr assay at 3 different antibody concentra inhibition of the ADCC against HeLa cells with the Peak I tions. •.control untreated cultures; O, cultures treated with Peak I proteins (3 proteins. mg/ml).
1136 CANCER RESEARCH VOL. 41
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. Modulation of NK and ADCC by Ascites Proteins
ouabain (32), enzymes (24), and corticosteroids (44). There are also studies which demonstrate differences in these 2 cytotoxic activities in various pathological states (12, 28, 29, 35,51). ADCC In addition to the controversy concerning the NK and ADCC cells, there is also controversy as to whether the cells mediating ADCC against CRBC are the same as the cells mediating cytotoxicity against transformed cell lines. Some researchers have demonstrated that monocytes, lymphocytes, and granu- locytes are actively cytotoxic for CRBC (6, 38, 53), whereas a lymphocyte (K-cell) is active against transformed cells. In ad dition, Catalona ef al. (6) have shown that, in cancer patients, ADCC against Chang cell targets is impaired, whereas the cytotoxicity against CRBC was normal. Due to these reports, we felt that it was necessary to examine the effect of the immunosuppressive fraction on a target other than CRBC. We established an assay with HeLa cells and found that mononu clear cells are spontaneously cytotoxic (NK activity) for HeLa 12.5l 25-1 50l 100-1 cells, but poorly so. This NK activity, which ranged from 0 to EFFECTOR;TARGtTRATIO 10%, was completely suppressed by the Peak I proteins at all E:T ratios. The Peak I proteins were not, however, able to Chart 7. NK and ADCC activity of human mononuclear cells against HeLa cell targets. •.ADCCcontrol; O, ADCC with Peak I proteins (3 mg/ml); A. NK control. significantly suppress the ADCC against the HeLa cell targets. Bars, S.D. In those experiments where some NK activity was evident, there was a minimal inhibition of the ADCC, usually not signif the absence of toxicity. We do not have an explanation for this icant, which we hypothesize was a reduction of NK activity latter finding at this time, although it may be due to the presence against the target cells which is taking place concomitantly of complement factors in the Peak I fraction. Ghebrehiwet and with the ADCC. Müller-Eberhard (10) have demonstrated that C1 q can mediate There are some recent studies describing the immunosup the lysis of CRBC by human lymphoblastoid cell lines, and this pressive activity of serum and ascitic proteins on spontaneous factor is present in the Peak I fraction. cell cytotoxicity and ADCC. Cordier ef al. (7) have described The differential sensitivity of these 2 cytotoxicity assays to the inhibition of mitogen-induced cytotoxicity and ADCC the immunosuppressive activity of these Peak I proteins lends against CRBC by a human a2-macroglobulin fraction. This support to the hypothesis that there may be 2 different effector inhibition apparently increases with dilution of the antisera in cells or 2 different effector mechanisms mediating cytotoxicity. the assay. As there is an a2-macroglobulin in our Peak I There is currently a great deal of interest and controversy preparation, we have tried to duplicate these findings with our concerning the cells affecting NK and ADCC. NK cells have ¡mmunosuppressive fraction without success. Kuo (31) has been shown to be Fc receptor (FcR)-positive, largely comple shown that EL4/C57BI/6 ascitic fluid will block antibody com ment receptor-negative cells which may possess low-density plement-dependent cytolysis of murine spleen cells. This block or low-affinity receptors for sheep RBC (21, 57). They do not ing, however, was reduced when excess complement was used appear to have any surface immunoglobulin (52). It is therefore in the assay and therefore does not appear to be similar to the generally agreed that surface immunoglobulin-positive B-cells system that we are concerned with. Nair ef al. (37) have found and macrophages are not effectors in NK. Several authors that the sera from both normal and tumor-bearing animals have identified the most active NK cell as a non-T-cell (20, 48); inhibit the NK against RL5 1 and ADCC against CRBC, the however, other workers claim that a T-cell may contribute to cancer serum being the more inhibitory. We have found previ NK (3, 23), and others say that it is the main effector cell (18, ously (1 ) that not only cancerous ascites are immunosuppres 57). Koren and Williams (30) believe that NK and K-cells are 2 sive but that fluids from patients with liver disease and cirrhosis separate lymphoid populations, with NK cells bearing receptor suppress lymphocyte proliferation in response to phytohemag- sites for NK determinants and FcR, whereas K-cells bear only glutinin. We prepared Peak I fractions from a cirrhosis fluid and FcR. Insofar as the lymphocytes responsible for NK have in found that it suppressed NK activity of mononuclear cells most instances been shown to have surface receptors for the against the K562 cell line. Peak I fractions from normal human Fc fragment of IgG, they are indistinguishable from the effector serum and congestive heart failure were suppressive only at lymphocytes that mediate ADCC (24, 42, 43, 46). Many work high concentrations (5 mg/ml). We did not test these fractions ers therefore consider that a significant fraction of both K- and on ADCC function. Immunosuppressive activity in nonmalignant NK cells are Fc receptor-bearing T-cells (43, 50, 56). ascites has also been demonstrated by Hess ef al. (19) and Le It would certainly appear from the experiments presented in Bien ef al. (33), and they suggest that these factors may be a this study that 2 different mechanisms are responsible for the natural host-mediated response to inflammation. It remains to cytotoxicity. It must be remembered, however, that in vitro be determined whether the suppressive moieties in these fluids experiments are always suspect of artifactual results. There are identical. Our current purification studies have centered on are other studies which show that NK and ADCC have a the ovarian ascites fluids and must be repeated with the cirrho differential sensitivity to certain substances such as interferon sis and other inflammatory fluids. (9, 16, 34, 55), Protein A from Staphylococcus aureus (24), Although we feel that the Peak I proteins have a direct
MARCH 1981 1137
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. A. M. Badger et al.
immunosuppressive effect on the mononuclear effector cells, 21. Jondal, M., and Pross, H. Surface markers on human B and T lymphocytes. VI. Cytotoxicity against cell lines as a functional marker for lymphocyte sub- we have not ruled out the possibility that this fraction may be populations. Int. J. Cancer, 75 596-605, 1975. activating a suppressor cell population. This suppressor cell 22. Jondal, M., Spina, C., and Targen, S. Human spontaneous killer cells population would have to be specific for the NK activity and selective for tumor derived target cells. Nature (Lond.), 272. 62-64, 1978. 23. Kaplan, J., and Callewaert, D. M. Expression of human T-lymphocyte anti not affect the ADCC activity. In addition, these suppressor cells gens by natural killer cells. J. Nati. Cancer Inst., 60 961-964. 1978. would have to be activated very rapidly as the entire assay only 24. Kay, H. D., Bonnard, G. D.. West, W. H., and Herberman, R. B. A functional comparison of human Fc-receptor-bearing lymphocytes active in natural takes 4 hr. cytotoxicity and antibody-dependent cellular cytotoxicity. J. Immunol., 7 78: These studies clearly demonstrate that the spontaneous 2058-2066, 1977. cytotoxicity of normal mononuclear cells is inhibited by ascitic 25. Kiessling, R., Klein, E , and Wigzell, H. "Natural" killer cells in the mouse. I. Cytotoxic cells with specificity for mouse leukemia cells. Specificity and fluid proteins, whereas their ADCC activity may not be compro distribution according to genotypes. Eur J. Immunol., 5: 112-117, 1975. mised. This inhibition of NK activity may be a reflection of an in 26. Kiessling, R., Petranyi, G., Klein, G., and Wigzell, H. Genetic variations of in vivo reduction of the cancer patients' capability for immune vitro cytolytic activity and in vivo rejection potential of non-immunized semisyngeneic mice against a mouse lymphoma line. Int. J. Cancer, 75. surveillance. 933-940. 1975. 27. Kiessling, R., Petranyi, G., Klein, G., and Wigzell, H. Non-T-cell resistance REFERENCES against a mouse Moloney lymphoma. Int. J. Cancer, 77: 1-7, 1976. 28. Klein, M., Roder, J., Haliotis, T., Korec, S., Jett, J. R., Herberman, R. B., 1. Badger. A. M., Buehler, R. J.. and Cooperband, S. R. Immunosuppressive Katz, P., and Fauci, A. S. Chédiak-Higashi gene in humans. II. The selectivity activity and tissue polypeptide antigen content of human ascitic fluids. of the defect in natural killer and antibody dependent cell-mediated cytotox Cancer Res., 38 3365-3370, 1978. icity function. J. Exp. Med., 757. 1049-1058, 1980. 2. Badger, A. M., and Cooperband. S. R. An immunostimulatory substance in 29 Koren. H. S.. Amos, D. B., and Buckley, R. H. Natural killing in immunode- the ascites fluid of patients with cancer metastatic to the peritoneum. Cell. ficient patients. J. Immunol., 720. 796-799, 1978. Immunol., 45: 15-25. 1979. 30. Koren, H. S.. and Williams, M. S. Natural killing and antibody-dependent 3. Bakacs. T., Gergely, P., and Klein. E. Characterization of cytotoxic human cellular cytotoxicity are mediated by different mechanisms and by different lymphocyte subpopulations. The role of Fc-receptor carrying cells. Cell. cells. J. Immunol., 727. 1956-1960, 1978. Immunol.,32. 317-328, 1977. 31. Kuo, C. Y. In vitro blocking of antibody mediated tumor cell destruction by 4. Becker, S., and Klein, E. Decreased natural killer effect in tumor-bearing ascitic protein of mouse tumor. Fed. Proc.. 38: 920. 1979. mice and its relation to the immunity against oncorna virus-determined cell 32. Lawrence, E. C., Muchmore, A. V., Decker, J. M., Dooley, N. J., and Blaese, surface antigens. Eur. J. Immunol., 6. 892-898, 1976. R. M. Differential effects of oubain on human cell-mediated cytotoxicity. II. 5. Butterworth. A. E., Sturrock, R. F.. and Houba, V. Antibody-dependent cell- Stimulation of monocyte-mediated. spontaneous cytotoxicity against mediated damage to Schistosomu/a in vitro. Nature (Lond.,), 252. 503-505, chicken red cell targets. Cell. Immunol.. 46: 110-118, 1979 1974. 33. Le Bien, T. W.. Gerber, J. D., and McCarthy. R. E. Suppression of 2-ME 6. Catalona, W. J.. Ratcliff, T. L, and McCod, R. E. Discordance among cell- induced lymphocyte proliferation by murine ascitic fluids. Fed. Proc., 37: mediated cytolytic mechanisms in cancer patients: importance of the assay 1354. 1978. system. J. Immunol., 722 1009-1014, 1979. 34. Lindahl. P., Leary, P., and Gresser, I. Enhancement by interferon of the 7. Cordier, G., Latour, M., Bonneau, M., and Revillard. J P. Modulation of specific cytotoxicity of sensitized lymphocytes. Proc. Nati. Acad. Sei. U. S. antibody-dependent or phytohemagglutinin-induced cell-mediated cytotox A., 69: 721-725, 1972. icity by human
1138 CANCER RESEARCH VOL. 41
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. Modulation of NK and ADCC by Ascites Proteins
hosts immune to tumor antigens. Int. J. Cancer, 9. 316-323, 1972. lymphocyte. J. Immunol., 121: 526-531, 1978. 48. Press, H. F., and Jondal, M. Cytotoxic lymphocytes from normal donors. A 53. Shaw. G., Levy, P.. Balcerzak, S.. and LoBuglio. A. Human lymphocytes, functional marker of human non-T lymphocytes. Clin. Exp. Immunol., 27. monocytes and neutrophils possess similar potential for antibody dependent 226-235. 1975. cytotoxicity (ADCC). F9d. Proc., 38. 925, 1979. 49. Rosenberg, E. B., McCoy, J. L, Green, S. S., Donnelly, F. C., Siwarski, D. 54. Takasugi, M., Mickey, M. R.. and Terasaki, P I. Reactivity of lymphocytes F., Levine, P. H., and Herberman, R. B. Destruction of human lymphoid from normal persons on cultured tumor cells. Cancer Res.. 33 2898-2902. tissue culture cell lines by human peripheral lymphocytes in 51Cr-release 1973. cellular cytotoxicity assays. J. Nati. Cancer Inst, 52 345-352, 1974. 55. Trinchieri. G.. Santoli, D., and Koprowski. H. Spontaneous cell-mediated 50. Saal, J. G., Rieber, E. P., Hadam. M., and Riethmuller, G. Lymphocytes with cytotoxicity in humans: role of Interferon and immunoglobulins. J. Immunol., T-cell markers cooperate with IgG antibodies in the lysis of human tumor 120:1849-1855,1978. cells. Nature (Lond.). 265 158-160, 1977. 56. Wahlin. B.. Perlmann. H., and Perlmann, P. Analysis by a plaque assay of 51. Sanai. S. O.. and Buckley, R. H. Antibody-dependent cellular cytotoxicity in IgG or IgM-dependent cytolytic lymphocytes in human blood. J Exp. Med , primary immunodeficiency diseases and with normal leukocyte subpopula- 144: 1375-1380. 1976. tions. J. Clin. Invest.. 61: 1-10. 1978. 57. West. W. H.. Cannon, G. B.. Kay, H. D., Bonnard, G. B., and Herberman. R 52. Santoli. D.. Trinchieri, G., and Lief, F. S. Cell-mediated cytotoxicity against B. Natural cytotoxic reactivity of human lymphocytes against a myeloid cell virus-infected target cells in humans. I. Characterization of the effector line. Characterization of effector cells. J. Immunol.. ! Õ8.355-361, 1977.
MARCH 1981 1139
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research. Differential Effects of an Immunosuppressive Fraction from Ascites Fluid of Patients with Ovarian Cancer on Spontaneous and Antibody-dependent Cytotoxicity
Alison M. Badger, Sue K. Oh and Frederick R. Moolten
Cancer Res 1981;41:1133-1139.
Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/41/3/1133
E-mail alerts Sign up to receive free email-alerts related to this article or journal.
Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].
Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/41/3/1133. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.
Downloaded from cancerres.aacrjournals.org on October 7, 2021. © 1981 American Association for Cancer Research.