Transient Warm Autoimmune Hemolytic Anemia and Cryoglobulinemia Associated with Seminoma

A patient with a pure seminoma presented with severe IgG-mediated warm autoimmune hemolytic anemia. Monoclonal IgM-kappa cryoglobulinemia and a biological false positive test for syphilis were also found. Treatment directed at both the seminoma and the hemolysis resulted in the complete disappearance of these antibodies. It is possible that these immunological phenomena occurred in response to the tumor. The occurrence of warm autoimmune hemolytic anemia and monoclonal paraproteinemia in association with solid tumors is reviewed.     

Dynamics of chemiluminescence response of Syrian hamster blood neutrophils to the growth of 10 different strains of tumor cells]

The production of H2O2 by blood neutrophils of Syrian hamsters, bearing 10 different transplanted tumors was studied at different stages of tumor growth by the luminol-dependent chemiluminescence (CL) test. It was demonstrated that during the tumors growth, two types of the blood neutrophils CL reaction can be registered. The first type of reaction represents very early and significant decrease of spontaneous CL of blood neutrophils, already evident before the appearance of palpable tumor nodules in animals, bearing in vivo selected cell strains. Subsequent subcutaneous tumor appearance in the animals was followed by increased CL of neutrophils. The second type of reaction was characteristic for in vitro transformed cells never selected in vivo. In this case the increase of CL of blood neutrophils at early stages of tumor growth was followed with the decrease of this activity at the period of active tumor growth. Possible relation of this reactions to the survival and growth of different tumor cells in vivo is discussed.     

Thy and Ly markers on lymphocytes initiating tumor rejection

Nonadherent spleen cells from mice specifically immune to the syngeneic methylcholanthrene-induced fibrosarcoma H-7 caused suppression of tumor growth in footpads of nonimmune mice. The cells were Thy-1 positive, largely Ly-1 positive, but Ly-2 and Ly-3 negative, and resembled T cells causing delayed-type hypersensitivity reactions rather than cytolytic T cells. It is suggested that some tumors fail to elicit the production of rapidly cytolytic T cells but can be rejected by a mechanism akin to that of delayed-type hypersensitivity, mediated by a different class of T cells working in collaboration with macrophages.     

Experimental erythrocyte autoantibodies. V. Induction and suppression of red blood cell autoantibodies in mice injected with rat bromelain-treated red blood cells

Mice injected with rat red blood cells (RBC), or rat bromelain-treated (brom) RBC, produce RBC autoantibodies and suppressor cells that specifically inhibit the autoimmune response without inhibiting the net production of antibodies against rat RBC. It has been investigated whether suppressor cells induced by injections of rat RBC are effective in preventing autoantibody production induced by rat brom RBC and vice versa. Autoantibodies were induced in C3H mice by weekly ip injections, each 0.2 ml, of a 6% suspension of rat RBC or rat brom RBC. Autoantibody production was assayed using Coombs' test. Suppressor cells were present in the spleens of mice positive in Coombs' tests and were shown by intravenous injections of 40 X 10(6) viable cells per mouse into untreated syngeneic mice 18 hr before the first injection of rat RBC or rat brom RBC. Autoantibodies eluted from mice positive in Coombs' tests after injections of rat RBC or brom RBC were absorbed by either type of rat RBC but not by RBC from sheep. This suggests that rat RBC and rat brom RBC display antigens that are similar, if not identical, to autoantigens on the mouse RBC. Spleen cells from mice injected with rat RBC suppressed autoantibodies induced by both rat RBC and rat brom RBC. In contrast, spleen cells from mice injected with rat brom RBC suppressed autoantibodies induced by rat brom RBC but not those induced by unmodified rat RBC. This differential suppression may be due to the removal from rat RBC, by bromelain, of a suppressor site and/or autoantigens of some specificities. Thus rat brom RBC may not induce the total range of specificities of autoantibodies, and of suppressor cells, induced by rat RBC.     

Autoantibodies to the thrombocytes and erythrocytes in HIV-infected patients

In 48 patients with HIV infection were tested for the presence of autologous and allogenic antibodies to red blood cells with the use of Coombs' direct and indirect tests. 18 HIV-infected patients had IgG antibodies to thrombocytes, circulating in the blood (detected by the method of EIA) and bound to thrombocytes (detected by the method of RIA). In 5 out of 48 patients Coombs' direct test yielded positive results with red blood cells. 6 out of 18 examined patients had an elevated content of thrombocyte-bound antibodies. The presence of cross reactions between gp120 of human immunodeficiency virus and gp3a thrombocytes led to the formation of antithrombocyte antibodies and, consequently, to a decrease in the number of thrombocytes.     

Effect of erythrocytes isolated from the blood of animals with experimental tumors on the proliferative activity of cultured tumor cells

It has been established that erythrocytes taken from blood of mice with experimental tumors being in the active phase of growth, or of mice with a recidivation of the primary tumor intensify proliferative processes in the culture of tumor cells. Erythrocytes from animals with tumors being in the stationary phase of growth did not reveal such a capacity. The authors connect such an effect with the erythrocyte transport function due to which they can store up those factors excreted by tumors which stimulate proliferation. It is supposed that the test defining the peculiarities of the effect of erythrocytes on the proliferative capacity of tumor cells in the tissue culture can be used for the judgement about an individual activity of the growth of primary tumors as well as for the diagnostics of recidivation of tumor growth.     

Selected immunohistochemical markers in stromal neoplasms of the gastrointestinal tract.

We studied immunohistochemical reactions to vimentin, desmin and protein S-100 in 43 cases of stromal tumors of the gastrointestinal tract. The material studied included: 1 esophageal tumor, 18 gastric tumors, 19 small intestinal tumors and 5 colonic tumors, classified in 13 cases as benign and in 30 cases as malignant neoplasms of various degree of malignancy. Mean age of the patients was 58.9 years. A positive reaction to vimentin was found in 37 cases, a negative reaction concerned an esophageal tumor, two benign tumors (gastric and small intestinal) and three malignant tumors (gastric and two small intestinal). A positive reaction to desmin was detected in an esophageal tumor and in nine gastric tumors. Only one benign small intestinal tumor and three benign colonic tumors showed a positive reaction to desmin. Protein S-100 was found in an esophageal tumor and in 7 out of 18 gastric tumors and in 12 out of 24 intestinal tumors. Coexpression of vimentin and desmin was found in 8 gastric tumors, only in one small intestinal tumor and in three colonic tumors. Three gastric tumors showing both these reactions were all benign. Coexpression of desmin and protein S-100 was found in 7 out of 43 tumors of the alimentary tract. In six cases these tumors were benign. Basing on the results we may say that the presence of these antigens reflects the degree of differentiation of gastrointestinal stromal tumors of the gastrointestinal tract, though it does not allow to choose unequivocally conclusions as to their histogenesis.     

Immunoenzyme method in the diagnosis of human brucellosis

The optimum conditions for the determination of specific antibodies in the sera of brucellosis patients by means of enzyme immunoassay (EIA) have been selected. The comparative study of the specificity and sensitivity of EIA and other serological tests has demonstrated that EIA has high diagnostic effectiveness in the diagnosis of acute and chronic brucellosis. The presence of direct correlation between the results of EIA and Coombs' test is observed, which is indicative of the capacity of EIA for detecting both complete and incomplete specific antibodies. It should be pointed out that in all cases the titer of specific antibodies in EIA has been found to be 5-16 times higher than in Coombs' test, the passive hemagglutination test, and agglutination test.     

Interactions of sera from patients with autoimmune diseases with cDNA expressed fragment of topoisomerase I and monoclonal antibodies against enzyme

The interaction of sera from 34 patients with different autoimmune diseases with the expressed fusion protein cloned in lambda gt11 vector (topoisomerase I--beta galactosidase) and monoclonal antibodies against enzyme was studied. It was demonstrated that 100% of Scl cases possessed positive activity against fusion protein. It was shown that this test is not absolutely specific for Scl, i. e. 57.1% of Sle and 84.6% of RA demonstrated positive activity against "topoisomerase test". Autoimmune sera had the positive activity against monoclonal antibodies for topoisomerase I. This activity was shown to be due to the presence of antiidiotypic antibodies against topoisomerase in the autoimmune sera.     

Serological activity of incomplete antibodies]

Blood sera containing incomplete antibodies were studied in various serological tests, by the results of Coombs' and the inhibition of complement fixation tests. It was shown that division of antibodies into complete and incomplete by their serological activity was conditioned. Antibodies detected in Coombs' test could fix the complement. The blocking antibodies depressing the complement fixation test could fix the homologous complement. Proceeding from the latter the author suggests the test of the conglutinating complex fixation which proved to be effective in detection of antibodies inactive in the agglutination and complement fixation tests.     

Mutational Profiling Can Establish Clonal or Independent Origin in Synchronous Bilateral Breast and Other Tumors

Background

Synchronous tumors can be independent primary tumors or a primary-metastatic (clonal) pair, which may have clinical implications. Mutational profiling of tumor DNA is increasingly common in the clinic. We investigated whether mutational profiling can distinguish independent from clonal tumors in breast and other cancers, using a carefully defined test based on the Clonal Likelihood Score (CLS = 100 x # shared high confidence (HC) mutations/ # total HC mutations).

Methods

Statistical properties of a formal test using the CLS were investigated. A high CLS is evidence in favor of clonality; the test is implemented as a one-sided binomial test of proportions. Test parameters were empirically determined using 16,422 independent breast tumor pairs and 15 primary-metastatic tumor pairs from 10 cancer types using The Cancer Genome Atlas.

Results

We validated performance of the test with its established parameters, using five published data sets comprising 15,758 known independent tumor pairs (maximum CLS = 4.1%, minimum p-value = 0.48) and 283 known tumor clonal pairs (minimum CLS 13%, maximum p-value <0.01), across renal cell, testicular, and colorectal cancer. The CLS test correctly classified all validation samples but one, which it appears may have been incorrectly classified in the published data. As proof-of-concept we then applied the CLS test to two new cases of invasive synchronous bilateral breast cancer at our institution, each with one hormone receptor positive (ER+/PR+/HER2-) lobular and one triple negative ductal carcinoma. High confidence mutations were identified by exome sequencing and results were validated using deep targeted sequencing. The first tumor pair had CLS of 81% (p-value < 10–15), supporting clonality. In the second pair, no common mutations of 184 variants were validated (p-value >0.99), supporting independence. A plausible molecular mechanism for the shift from hormone receptor positive to triple negative was identified in the clonal pair.

Conclusion

We have developed the statistical properties of a carefully defined Clonal Likelihood Score test from mutational profiling of tumor DNA. Under identified conditions, the test appears to reliably distinguish between synchronous tumors of clonal and of independent origin in several cancer types. This approach may have scientific and clinical utility.     

Assessment of syngeneic cell-mediated immunity to mouse mammary tumors by three different assays

Summary The purpose of this study was to compare the MCT, CRT, and Winn assays for assessment of CMI to syngeneic tumors. Spontaneous and MMTV-induced mammary tumors in Balb/c and Balb/cfC₃H mice were used. LNC were obtained at various times relative to S.C. injection of sensitizing tumor cells and surgical removal of the outgrowth. In direct comparison to Winn assays, CRT often gave false negatives (i.e., LNC could inhibit tumor growth in the Winn assay without being cytolytic), but when LNC were cytolytic in the CRT, they inhibited in Winn assays. Thus, a positive CRT was associated with a positive Winn assay. In contrast, results from MCT tests were generally not associated with those obtained by CRT or Winn assays. This divergence in assays was independent of differences in status of the LNC donor. With all three assays, stimulation of tumor growth or increased survival was occasionally observed. For the CRT this was manifested in the release of smaller amounts of ⁵¹Cr in the presence of immune LNC than in the presence of normal LNC. For MCT and Winn assays it was seen in better tumor growth after treatment with LNC than after treatment with medium alone. Detection of this phenomenon was independent among the three assays.Data from the CRT and Winn assays were analyzed to see whether they correlated with in vivo behavior of the sensitizing tumors. Detection of CMI by these tests was associated with tumors growing out after relatively long latency periods, but was independent of tumor growth rate. This is in contrast to our previously reported analysis of MCT data, in which detection of CMI was associated with slower growth rate but was independent of the latency period (Hager et al., 1978).Abbreviations CMI cell-mediated immunity - CRT chromium-release test - MCT microcytotoxicity test - MMTV murine mammary tumor virus - LDA lymphocyte-dependent antibody - LNC lymph node cells - SBF serum-blocking factors - SC subcutaneous - TCM tissue culture medium     

Influence of T-lymphocyte deprivation on the antileukemic effect of bacillus calmette-guérin in vivo

Summary The effect in AKR mice of T-lymphocyte deprivation in vivo, obtained by adult thymectomy plus/minus whole-body irradiation and bone-marrow reconstitution, was studied in the growth of grafted leukemia cells obtained from spontaneous AKR leukemia. Both thymectomized mice and mice subjected to thymectomy, whole-body irradiation, and bone-marrow reconstitution (B) had a lower take-frequency of graft leukemia than conventional mice. Growth of graft leukemia was inhibited by BCG treatment both in thymectomized mice and in B mice. Concomitant with the increased inhibition of leukemia growth, an increased incidence of wasting-like disease was observed. In vitro cytotoxicity studies revealed that spleen lymphoid cells from nonleukemic mice were cytotoxic to AKR leukemia cells, to nonmalignant AKR fibroblasts, and to other nonmalignant cells with H-2^k haplotype. The activity of this self-directed cytotoxicity was most marked in AKR mice with wasting-like disease. The presence of autocytotoxic cells was frequently associated with a positive direct Coombs' test. Immunofluorescence studies showed, further, that the cytotoxic activity was independent of retrovirus antigens as tested by indirect immunofluorescence with anti-MuLV antibodies. Adult thymectomy of AKR mice confers an increased antitumoral immune potential, but also an increased risk of development of serious autoimmune disease.     

Circadian disruption accelerates tumor growth and angio/stromagenesis through a Wnt signaling pathway

Epidemiologic studies show a high incidence of cancer in shift workers, suggesting a possible relationship between circadian rhythms and tumorigenesis. However, the precise molecular mechanism played by circadian rhythms in tumor progression is not known. To identify the possible mechanisms underlying tumor progression related to circadian rhythms, we set up nude mouse xenograft models. HeLa cells were injected in nude mice and nude mice were moved to two different cases, one case is exposed to a 24-hour light cycle (L/L), the other is a more "normal" 12-hour light/dark cycle (L/D). We found a significant increase in tumor volume in the L/L group compared with the L/D group. In addition, tumor microvessels and stroma were strongly increased in L/L mice. Although there was a hypervascularization in L/L tumors, there was no associated increase in the production of vascular endothelial cell growth factor (VEGF). DNA microarray analysis showed enhanced expression of WNT10A, and our subsequent study revealed that WNT10A stimulates the growth of both microvascular endothelial cells and fibroblasts in tumors from light-stressed mice, along with marked increases in angio/stromagenesis. Only the tumor stroma stained positive for WNT10A and WNT10A is also highly expressed in keloid dermal fibroblasts but not in normal dermal fibroblasts indicated that WNT10A may be a novel angio/stromagenic growth factor. These findings suggest that circadian disruption induces the progression of malignant tumors via a Wnt signaling pathway.     

Immune-deficient animals to study "hormone-dependent" breast and endometrial cancer

Athymic (nu/nu) mice are T cell deficient and can accept xenografts of human tumor material. Hormone-dependent tumor growth can be demonstrated in ovariectomized athymic mice by estrogen administration. Estrogen receptor (ER) positive MCF-7 breast cancer cells implanted into the axillary mammary fat do not grow into palpable tumors unless sustained release preparations of estrogen are administered. The non-steroidal antiestrogen tamoxifen, though it exhibits estrogenic properties in the mouse, does not facilitate MCF-7 tumor growth (during short term, i.e. 8 weeks of therapy) and can prevent estradiol-stimulated growth. In contrast, ER negative MDA-MB-231 cells grow with or without estrogen administration and tamoxifen does not control tumor growth. These statements reflect current dogma concerning the value of athymic mice to confirm the hormone dependent growth of cancer cells in vivo. Our aim has been to define the limits of this dogma and to investigate the growth relationship of hormone-dependent and independent cells with their host environment. The potential endocrine or paracine effect of ER negative tumors on the growth of ER positive tumors was evaluated by transplantation on opposite sides of athymic mice or by the inoculation of different ratios of ER positive/negative cells (MCF-7:MDA-MB-231 9:1, 99:1, 999:1). MCF-7 cells could not be encouraged to grow by a rapidly growing MDA-MB-231 tumor on the opposite side of the animal. Similarly ER negative tumors grew out of the mixed tumor inoculates suggesting that ER positive tumors could not be encouraged to grow preferentially by the paracrine influences of ER negative cells. However, estrogen facilitates the growth of an ER positive tumor following inoculation of mixed cell populations. Antiestrogen treatment can blunt estrogen-stimulated growth but cannot control the growth of ER positive/negative containing tumors. ER positive endometrial tumors grow in response to estrogen treatment and some (EnCa101) have been shown to grow in response to tamoxifen or a combination of tamoxifen and estrogen. More unusual though is our recent observation that an ER negative primary endometrial tumor (BR) and its metastasis (BR-MET) grow more rapidly in estrogen-treated athymic mice. This finding seems to have far-ranging consequences for our view of hormone-dependent growth. Either our view of estrogen-stimulated growth needs to be modified or the host is specifically altered during estrogen treatment. We have taken the position that since natural killer cells (present in athymic mice) can be lowered by estrogen this may result in an increased tumor cell survival in the heterotransplant model.(ABSTRACT TRUNCATED AT 400 WORDS)     

Genetic and biochemical studies on the suppression of and a recovery from the tumorous state in higher plants

Four neoplastic diseases of plants: crown gall, which is caused by Ti plasmid DNA; Black's wound tumor disease by an RNA virus; the Kostoff genetic tumors by chromosomal imbalance; and habituation, which results from a spontaneous activation of select biosynthetic systems, have been analyzed and compared. It has been found that both the development of a capacity for autonomous growth and the nature of the heritable cellular change that underlies tumorigenesis are similar in the four instances. All develop a capacity for autonomous growth as a result of the persistent activation of select biosynthetic systems, the products of which are concerned with cell growth and division. That the persistent activation of these biosynthetic systems does not involve heritable changes of an irreversible type is indicated by the finding that a reversal of the neoplastic state occurred in three of the test systems. Since the tumor cells in these instances were found to remain totipotent the results suggest that whether the normal or tumor phenotype is expressed is determined by how the genetic information is regulated in a cell. Regulation appears to be accomplished in part through positive feedback control mechanisms. Foreign genetic information could act either in a regulatory manner to persistently activate normal biosynthetic systems or it could code for one or more essential but normally limiting substance(s) and thus replace a substance(s) that in the case of the Kostoff tumors or habituation is specified by host cell genes, or it could do both. In either case, the foreign genetic information can be regulated in much the same manner as are the host cell genes to give rise to either the normal or tumor phenotype.     

Immunotherapy of tumors with xenogeneic endothelial cells as a vaccine

The breaking of immune tolerance against autologous angiogenic endothelial cells should be a useful approach for cancer therapy. Here we show that immunotherapy of tumors using fixed xenogeneic whole endothelial cells as a vaccine was effective in affording protection from tumor growth, inducing regression of established tumors and prolonging survival of tumor-bearing mice. Furthermore, autoreactive immunity targeting to microvessels in solid tumors was induced and was probably responsible for the anti-tumor activity. These observations may provide a new vaccine strategy for cancer therapy through the induction of an autoimmune response against the tumor endothelium in a cross-reaction.     

Pathological cell aggregates in bone marrow cultures from patients with various hematological diseases.

Non-clonal growth of macroscopic cell aggregates in methylcellulose cultures of abnormal marrow is described. They were seen in all patients with Graft-versus-Host Disease, graft rejection, and autoimmune disease presumably directed against hemopoietic cells, we found them in 35% of patients with primary hematological neoplasias and rarely in patients with solid tumors. They were never encountered in 80 healthy controls. The aggregates originated from small cell clumps which sedimented with the "buffy coat" in contrast to normal bone marrow particles. They contained tumor cells, grafted myeloid cells, or target cells of autoimmune disease in association with a widely varying amount of macrophages. Preliminary results suggest that the frequency of macrophages within the aggregates correlates inversely with the aggressiveness of the clinical condition. We propose that appearance of such aggregates in an indicator of immune activation; we expect that further quantitation of the phenomenon will reveal important clinical correlations and provide a model for the study of host defense to "foreign" cells.     

Effect of melatonin and pineal extracts on human ovarian and mammary tumor cells in a chemosensitivity assay

Pinealectomy enhances tumor growth and metastatic spread in experimental animals. This effect is only in part due to melatonin since melatonin-free pineal extracts containing yet unidentified pineal substances have also shown tumor inhibiting activity. Despite numerous reports suggesting melatonin as a potential anti-cancer agent there have not been sufficient clinical trials to define the actual therapeutic potential of melatonin for the treatment of human cancers. To help fill this gap, we used a chemosensitivity assay designed to test the sensitivity of tumors from individual patients towards chemotherapeutic drugs for assessing the effect of melatonin and pineal extracts on primary human tumor cells. Primary cell cultures from seven ovarian and six mammary tumors were incubated with melatonin, the pineal extract YC05R (containing substances between 500 and 1000 daltons) and chemotherapeutic drugs. The pineal extract YC05R inhibited growth of all tumors in a dose-dependent manner. Physiological concentrations of melatonin (10(-8)-10(-10) M) inhibited the growth of one out of six mammary carcinomas in a dose-dependent manner. Primary cell cultures from three ovarian tumors were affected by melatonin in different ways, i.e., two were inhibited and one was slightly stimulated. There was no correlation between sensitivity towards melatonin and sex steroid receptor status, stage or grade of the tumor. It is concluded that, 1), melatonin may be an inhibitor of human mammary and ovarian carcinoma in individual cases and, 2), the pineal gland contains very active anti-tumor substances inhibiting both, the mammary and ovarian tumors, tested. These substances require chemical and biological identification.