Membranous glomerulonephritis associated with renal cell carcinoma: failure to detect a nephritogenic tumor antigen

A 57-year-old man with renal cell carcinoma associated with membranous glomerulonephropathy (MGN) developed a transient amelioration of the nephrotic syndrome after excision of the tumor. We tried to identify a nephritogenic tumor antigen using the immunoblotting technique in this patient with MGN, since previous studies examined the interaction between tumor antigens and IgG eluted from the kidney tissue using immunofluorescence or immunodiffusion techniques, and no studies have identified the specific tumor antigen with the immunoblotting method. In the present study, no significant immunoreactivity was noted between the IgG eluted from renal cortical tissues of the patient and renal cell carcinoma proteins. Further studies are necessary to establish the pathogenic mechanism of MGN associated with malignancy.     

Beta-2-microglobulins as a differentiation marker in bladder cancer

The transformation of a normal cell through dysplasia to the malignant state usually is associated with changes at the molecular level within the nucleus, cytoplasm and cell surface. These changes can be monitored by the loss of normal cell surface antigens, such as the blood group antigen ABO(H) and major histocompatibility complex antigens, which in the human correlate with the histocompatibility locus antigens. A group of patients with bladder biopsy and diagnosis ranging from normal through severe dysplasia to papillary transitional cell carcinoma, invasive transitional cell carcinoma and carcinoma in situ were evaluated for the presence or absence of beta-2-microglobulin. This 11,000 molecular weight protein was used as an indirect marker for the major histocompatibility complex antigens on the cell surface. With immunoperoxidase as a marker, the presence of beta-2-microglobulin was seen in all patients with normal epithelium as well as benign disease. However, with progression through dysplasia to carcinoma there was progressive deletion of the beta-2-microglobulin. Carcinoma in situ exhibited minimal expression of the beta-2-microglobulin. The use of beta-2-microglobulin as a marker for major histocompatibility complex antigens on the cell surface may prove to be useful for monitoring transitional cell carcinoma.     

Lack of antibody production against Hanganutziu-Deicher (H-D) antigens with N-glycolylneuraminic acid in patients with porcine exposure history

The significance of non-alphagalactosyl antigens remains unclear in pig-to-primate xenotransplantation. Hanganutziu-Deicher (H-D) antigens with terminal N-glycolylneuraminic acid (NeuGc) are widely expressed on endothelial cells of mammalian species, with the exception of humans. As baboons and monkeys also express H-D antigens, a pig-to-non-human primate experimental model cannot resolve the question of whether H-D antigens can elicit a potent humoral response in human recipients. The purpose of this study was to elucidate the clinical significance of H-D antigens by examining the sera from patients who have been previously exposed to porcine tissue. After the digestion of porcine aortic endothelial cells (PAEC) by neuraminidase, NeuGc and N-acetylneuraminic acid (NeuAc) were quantitated by HPLC. IgG and IgM antibody levels against H-D antigens were measured by NeuGc-GM3-coated ELISA plates in the sera of patients who had undergone ex vivo kidney perfusion 1 to 3 weeks and 2 years previously (n=2) or had been injected with fetal porcine islets 2 months previously (n= 10). HPLC determined that 9.7x 10(7) NeuAc and 6.3x 10(7) NeuGc residues per cell were released from PAEC by neuraminidase, while 25.7x 10(7) NeuAc and an undetectable level of NeuGc were released from human aortic endothelial cells (HAEC). No significant elevation of IgG or IgM antibody levels against NeuGc-GM3 was observed in sera from patients with a history of porcine exposure. Considering the active production of antibody against the foreign galactosyl antigens after pig-to-human xenotransplantation, some production of antibodies against the equally foreign H-D antigens would be expected, because large amounts of NeuGc terminated saccharides are present in the pig endothelial cell surface. However, no production of antibodies directed to H-D antigens could be found in patients exposed to porcine tissue. Further studies are warranted to explain why H-D antigens do not elicit a significant antibody production.     

Expression and modulation of major histocompatibility antigens on murine primary brain tumor in vitro

Lysis of tumor cells by activated cytotoxic lymphocytes requires their recognition of antigens associated with major histocompatibility complex molecules. The authors studied the constitutive expression of Class I and Class II major histocompatibility complex antigens on mouse brain-tumor cells and the capacity of different cytokines and cytokine combinations to alter this expression in vitro. Cells from the murine glioma 26 (GL26), glioma 261 (GL261), and ependymoblastoma A (EpA) cell lines were established in monolayer culture and treated for 48 hours with either alpha interferon, gamma interferon, tumor necrosis factor alpha, tumor necrosis factor alpha plus gamma interferon, or interleukin-2. They were then analyzed by flow cytometry for baseline and cytokine-altered major histocompatibility complex expression. All cell lines had a similar constitutive major histocompatibility complex pattern with low Class I antigen expression and no detectable Class II antigen expression. Alpha interferon substantially induced and up-regulated Class I antigen expression, but had no effect on Class II antigen expression. Gamma interferon also stimulated up-regulation of Class I antigen expression, generally doubling the anti-Class I antigen fluorescence of treated cells. Its effect on Class II antigen expression was more extensive. In the GL26 and GL261 cell lines the expression of Class II antigen determinants increased to 12 x and 14 x control values and as many as 75% of cells that had no detectable constitutive expression of Class II antigen expressed this antigen after priming with gamma interferon. The addition of tumor necrosis factor alpha to gamma interferon further increased Class II antigen expression on EpA tumor cells only. Interleukin-2 and tumor necrosis factor alpha alone had no effect on Class I or Class II antigen expression of any cell lines. It is concluded that Class I and Class II antigen expression in mouse glioma cell lines is induced and enhanced after treatment with certain cytokines in vitro. Use of these cell lines to create in situ primary brain tumors in C57BL/6 mice should provide an excellent animal system to study major histocompatibility complex modulation in brain tumor cells and to examine the potential impact of major histocompatibility complex up-regulation on the response of brain tumors to immunotherapy.     

Autoimmune responses against renal tissue proteins in long-term surviving allograft recipients

Major histocompatibility complex antigens (MHC) are classical targets of recipient responses to allotransplants. However, the role of an immune response directed against autologous graft tissue determinants is poorly defined. In this study, we investigated (i) whether autologous kidney tissue extract can induce an immune response to autologous kidney proteins in normal rats, and (ii) if a similar autologous response develops in the long-term surviving LEW.1A recipients of an MHC-mismatched LEW.1W kidney (RT1^u to RT1^a). LEW.1A rats immunized with allo- or syngeneic soluble kidney extracts developed a T-cell response to self antigens as shown by the frequency of specific IFN-γ-producing T cells from LEW.1A rats in the presence of extracts (ELISPOT). In contrast, they responded only marginally to dominant RT1^u determinants. The ELISPOT against fractions of soluble autologous kidney extracts separated by an FPLC gel-filtration system indicated a preferential response to megalin, a high molecular weight protein that has been shown to be involved in experimental Heymann nephritis. In a model of long-term kidney allograft survival by anti-CD28 administration, recipients also developed humoral but not cellular responses to megalin. Our data suggest that autoimmune processes develop in long-term surviving kidney allograft recipients.     

Fetal bone grafts do not elicit allograft rejection because of protecting anti-Ia alloantibodies. Implications to the immune survival of fetuses in allogeneic mothers.

In a previous study we showed that allografts of BN fetal bone, unlike allografts of adult bone, are not rejected by allogeneic recipients of the Lewis strain in spite of the existence of major histocompatibility complex (MHC) incompatibility between donors and hosts. In the present study, we analyzed the relationships existing between the host and fetal tissue that determine graft survival. We found that (1) the fetal BN graft, unlike adult grafts, induces in Lewis recipients a vigorous humoral response consisting mainly in the production of IgG antibody that seems to be directed against antigens of Ia-like specificities. (2) The BN rats are genetically defective in their capacity to respond to determinants and thus are not capable of producing anti-Ia antibodies; in accordance, Lewis fetal bone grafts are rejected by the BN recipients. (3) Chondrocytes isolated from fetal mouse bones do express Ia antigenic determinants. We suggest that the survival of an allogeneic fetal graft in an immunologically intact recipient depends on an active and selective immune response directed against the Ia components associated with the MHC on the embryonic and fetal cells. On the basis of these notions, we propose that the capacity of Ia determinants expressed on cells of the embryo, to elicit anti-Ia and IgG alloantibodies in the pregnant mother, determines the capacity of the embryo to escape rejection by the histoincompatible mother.     

Transplantation immunology and ABO incompatibility

An allograft is tissue transplanted from another individual within the same species. Mechanical trauma to a graft and recipient transplant site along with graft-derived proinflammatory mediators stimulate a non-specific innate immune response. Dendritic cells and macrophages present foreign antigen to the adaptive immune system cells and thus initiate a specific and directed response. In order to respond to a specific pathogen, an individual must be able to recognize foreign cells as non-self. Major and minor histocompatibility antigens and the ABO blood group antigens are central to distinguishing one human from another and therefore in recognizing self from non-self. Genetic polymorphism describes genes encoded by varying alleles resulting in varied phenotypes within a species. The blood group and major histocompatibility complex (MHC) are polymorphic, with many different possible allelic combinations leading to differences between individuals and allowing an individual to recognize self from non-self. Rejection describes the graft injury and loss of function due to the recipient's non-acceptance of the graft as ‘self’ and the response which aims to remove it from the body. Rejection can be classified into hyperacute, acute and chronic phases. Both cell-mediated and antibody-mediated mechanisms lead to allograft tissue destruction. By minimizing MHC mismatch and using immunosuppression therapy, the immune response to a graft can be reduced. This involves familial grafting when possible, matching donors and recipients for similar human leucocyte antigen and identification of preformed recipient antibodies.     

Natural influenza infection produces a greater diversity of humoral responses than vaccination in immunosuppressed transplant recipients

The humoral immune response to influenza virus infection is complex and may be different compared to the antibody response elicited by vaccination. We analyzed the breadth of IgG and IgA responses in solid organ transplant (SOT) recipients to a diverse collection of 86 influenza antigens elicited by natural influenza A virus (IAV) infection or by vaccination. Antibody levels were quantified using a custom antigen microarray. A total of 120 patients were included: 80 IAV infected (40 A/H1N1 and 40 A/H3N2) and 40 vaccinated. Based on hierarchical clustering analysis, infection with either H1N1 or H3N2 virus showed a more diverse antibody response compared to vaccination. Similarly, H1N1-infected individuals showed a significant IgG response to 27.9% of array antigens and H3N2-infected patients to 43.0% of antigens, whereas vaccination elicited a less broad immune response (7.0% of antigens). Immune responses were not exclusively targeting influenza hemagglutinin (HA) proteins but were also directed against conserved influenza antigens. Serum IgA responses followed a similar profile. This study provides novel data on the breadth of antibody responses to influenza. We also found that the diversity of response is greater in influenza-infected rather than vaccinated patients, providing a potential mechanistic rationale for suboptimal vaccine efficacy in this population.     

Studies on the cellular immune response in patients with urinary bladder carcinoma. XI: Evaluation of specific ADCC activity

We previously reported that the activity of antibody dependent cell-mediated cytotoxicity (ADCC) of lymphocytes from the patients with urinary bladder carcinoma was lower than that of normal subjects. We performed the specific ADCC assay using SRBC as the target cells, coated with tumor extract from the urinary bladder carcinoma. At the same time, the antiserum was collected from the immunized rabbit by the same tumor extract. The addition of the antiserum from the immunized rabbit gave the highest value for the patient sub-group that produced a precipitin band (Ouchterlony gel diffusion assay). Therefore it is suggested that the reaction of lymphocytes to tumor associated antigens is maintained to a considerable extent, although ADCC activity of the patient group was decreased. However, the antibody producing phenomenon toward the tumor associated antigen might be restrained.     

Development of monoclonal antibody imaging of metastatic prostatic carcinoma

Indium-111-labeled monoclonal antibody directed against prostate-specific antigen was injected into ten patients with known prostatic carcinoma, nine with metastatic disease. The monoclonal antibody scan was compared to the standard technetium bone scan, and both were correlated to clinical status and to 2-year follow-up. The ratio of target-to-nontarget activity and pharmacokinetics of the radiolabeled antibody were determined. Based on these findings we are hopeful that modifications of this radiolabeled antibody approach may be used for staging and may be developed as a therapeutic adjuvant.     

Overcoming immunologic incompatibility: transplanting the difficult to transplant patient

Immunologic incompatibilities between donor and recipient have limited the access to renal transplantation for many patients. Previously the presence of donor-specific alloantibodies directed against donor major histocompatibility complex (MHC) antigens or natural antibodies directed against donor ABO blood group antigens was considered an absolute contraindication to renal transplantation. However, with the current understanding of humoral immune responses, superior immunosuppressive agents, and improved diagnosis and treatment of antibody-mediated rejection, renal transplantation can be safely performed with outstanding results despite the presence of donor-specific antibody. In this review we discuss the biology of antibody-mediated rejection and sensitization. We discuss the diagnostic tests necessary to characterize the type, affinity, and avidity of the donor-directed antibodies. Current methods for performing renal transplants across ABO and human leukocyte antigen (HLA)-sensitized barriers are covered, including the potential morbidities. The rest of the review focuses on advances in managing these antibodies to increase the likelihood of receiving a deceased donor kidney or allow transplantation from a living donor against whom one has a prohibitive antibody.     

Primary prostatic carcinoid tumor with intracytoplasmic prostatic acid phosphatase and prostate-specific antigen

A case of prostatic carcinoid tumor with lymph node metastases is reported. The patient was a 78-year-old male who died in ventricular fibrillation. At autopsy, a 2 X 2 cm, white, irregular tumor was found in the prostate and there were several enlarged para-aortic lymph nodes. Both specimens contained a characteristic carcinoid tumor. Argyrophil stains revealed strong positivity in the primary as well as in the metastatic tumors. Electron micrographs prepared from formalin-fixed tissue demonstrated numerous membrane-bound dense-core granules. Immunoperoxidase-labeled antibodies against both prostatic acid phosphatase and prostate-specific antigen localized in the tumor cells. The ultrastructural and immunohistochemical results support differentiation of the tumor cells toward both prostatic epithelial cells and endocrine cells. We believe that this is the first reported case of a prostatic carcinoid tumor in which specific prostatic tissue markers have been demonstrated in the tumor cells.     

Inhibition of leukocyte migration by extracts of malignant prostatic tissue and correlation of degree of in vitro sensitization to clinical responsiveness in prostatic cancer patients

In an attempt to evaluate the degree of in vitro cellular sensitization to tumor and its relationship to clinical responsiveness, direct leukocyte migration tests were carried out in patients with varying degrees of adenocarcinoma of the prostate employing pooled allogeneic extracts of normal, benign, and malignant prostatic tissue as a source of antigen. Cell-mediated immunity to presumably common prostatic tumor associated antigens was observed. The degree of sensitization of clinically significant specific reactivity of the patients' leukocytes to malignant prostatic tissue was greatest in patients with localized disease, low-grade tumor, and clinically inactive disease than in patients with advanced disease, high-grade tumor, and clinically active disease. Evaluation of the possible correlation of specific reactivity to malignant prostatic tissue as a prognostic index of clinical responsiveness revealed a positive correlation with the degree of sensitization in 3 (43 per cent) of 7 patients. Correlation in 4 patients was questionable because of observations of "stimulation" of migration rather than inhibition, suggested by some to be reflective of weak sensitization to tumor. Evaluation of a larger patient population as well as a prospective study of the relationship of the degree of sensitization and clinical responsiveness will be necessary before any definitive conclusions may be drawn regarding the present observations.     

Intratumoral immunotherapy with anti-PD-1 and TLR9 agonist induces systemic antitumor immunity without accelerating rejection of cardiac allografts

Immune checkpoint inhibitors, such as programmed cell death 1 (PD-1) blockades, have revolutionized the field of cancer immunotherapy. However, there is a growing concern whether PD-1 inhibitors can be administered safely to transplant recipients with advanced cancer, as the T cells activated by checkpoint inhibitors may become reactive not only toward tumor antigens but also toward donor alloantigen, thereby resulting in allograft rejection. Here, immunotherapy with anti-PD-1/toll like receptor 9 agonist was administered to C57BL/6 mice bearing a cardiac allograft that were receiving maintenance immunosuppression or a PI4KIIIβ inhibitor-based tolerogenic regimen. Intratumoral (i.t.), but not systemic, immunotherapy promoted potent anti-tumor responses, but did not accelerate allograft rejection. This effect was associated with a pro-immunogenic effect induced by i.t. immunotherapy resulting in systemic cellular and humoral immune anti-tumor responses. Furthermore, when the tumor and cardiac allograft shared major histocompatibility complex (MHC) antigens, i.t. immunotherapy promoted immune responses directed against tumor and the cardiac allograft resulting in allograft rejection. The anti-tumor effect was compromised by maintenance immunosuppression with cyclosporin A, indicating that an optimal balance between enhanced anti-tumor immunity and decreased transplant immunoreactivity is critical. A clinically relevant approach could be to temporarily withdraw maintenance immunosuppression and/or replace it with a PI4KIIIβ inhibitor–based tolerance-inducing regimen to allow for effective immunotherapy to take place.     

Elution of in vivo bound antiprostatic epithelial antibodies following multiple cryotherapy of carcinoma of prostate

Antibodies reactive with the cytoplasmic membrane or intercellular areas of autologous human and monkey prostatic secretory epithelial cells have been demonstrated by immunofluorescence in an eluate prepared from prostatic tissue obtained from a patient with metastatic adenocarcinoma of the prostate following multiple cryotherapy of his primary prostatic tumor. Elution of antiprostatic antibodies provides preliminary evidence that prostatic tissue (tumor?)-specific or tumor-associated antigens are liberated into the circulation following cryotherapy of the prostate in man.     

Inflammatory cell infiltrates vary in experimental primary and metastatic brain tumors.

We have studied the cellular immune response that accompanies primary and metastatic brain cancers induced experimentally in rats by inoculation of RG-2 glioma and Walker 256 (W256) carcinoma cells, respectively. The inflammatory cell infiltrates were characterized with lectin histochemistry to visualize microglial cells and macrophages and with immunohistochemistry, using a panel of monoclonal antibodies, to detect major histocompatibility complex (MHC), lymphocytic, and macrophage antigens. The metastatic tumor was composed of a loose stroma with multiple, often large, necrotic areas, whereas the RG-2 glioma was composed of a dense collection of tumor cells showing only rare necrotic foci. Both tumor types were heavily infiltrated with microglia and/or macrophages, and these were positive for MHC Class II (Ia) antigens. Expression of MHC Class I antigens was absent from RG-2 glioma cells, but it was present in W256 metastatic carcinoma cells. The metastatic tumor was also characterized by a much heavier infiltrate of lymphocytes, as shown by the presence of cells positive for CD4, CD8, and leukocyte common antigens. These lymphocytic markers were absent from reactive microglia in the W256 carcinoma, whereas they were present in the RG-2 glioma. Polymorphonuclear leukocytes were seen only in the metastatic tumor. Our study delineates differences between the inflammatory cell infiltrates found in metastatic brain tumors and those found in primary brain tumors. The differences in cell composition and immunophenotype may indicate a more effective antitumor response in the metastatic tumor that could account for the observed tissue destruction.     

Evidence for cytotoxic T lymphocyte response against human lung cancer: reconstitution of antigenic epitope with peptide eluted from lung adenocarcinoma MHC class I

BACKGROUND: Cancer-associated, major histocompatibility complex (MHC)-restricted peptide antigens have been elucidated in human melanomas and ovarian, breast, and renal carcinomas; but relatively little is known about lung cancer antigens. METHODS: To work toward delineation of lung cancer-associated antigens, we developed tumor infiltrating lymphocytes (TILs), peripheral blood mononuclear cell-derived cytolytic T cell lines (CTL), autologous lung cancer cell lines, and normal lung cell lines from 17 patients undergoing lung cancer resections. The TILs and CTL lines were subsequently evaluated for markers of activation and specific lysis of autologous or allogeneic lung cancer cell lines or both. RESULTS: Freshly isolated TILs contained a more activated T cell population compared with the patients' peripheral blood T cells as evidenced by an increased expression of HLA-DR, CD25, and CD45RO. TILs isolated from 15 patients lysed allogeneic lung cancer lines. TILs lysed autologous lung cancer but not autologous normal lung or Epstein-Barr virus transformed B cell lines (B-LCL) in 4 of 8 cases tested, suggesting tumor specificity. A CTL line (RHPBL57.1) was generated from peripheral blood mononuclear cells of an HLA-A24(+) patient by stimulation against an established HLA-A24(+) allogeneic lung cancer cell line. RHPBL57.1 lysed the lung cancer cell line in an HLA-A24-restricted manner. Moreover, RHPBL57.1 specifically lysed autologous B-LCL pulsed with peptides, eluted from MHC class I and isolated from the HLA-A24(+) lung cancer cell line. CONCLUSIONS: TILs isolated from patients with lung cancer are predominantly an activated population of T cells with evidence of tumor and MHC class I-restricted lysis. Furthermore, we provide evidence for a lung cancer-associated, MHC class I-bound peptide antigen(s) that reconstitutes the epitope recognized by a lung cancer specific CD8(+) T cell line derived from a patient with lung cancer.     

Prognostic value of host immunocompetence in urologic cancer patients.

To evaluate the prognostic significance of host immunocompetence in urologic cancer patients, the subsequent clinical course of 95 patients was determined a year after skin testing with dinitrochlorobenzene. A close correlation was demonstrated between dinitrochlorobenzene reactivity and prognosis among 38 transitional carcinoma patients. Of 19 patients with impaired reactivity 13 had tumor recurrences and 11 of these died of cancer within 1 year. Only 5 of 19 patients with normal dinitrochlorobenzene reactivity had recurrences and none died during the same interval. Although not statistically significant, similar results were observed among 10 renal cell carcinoma patients of whom 3 of 5 with impaired dinitrochlorobenzene reactivity had tumor recurrences, while 4 of 5 with normal reactivity remained free of tumor. One testis tumor patient with impaired dinitrochlorobenzene reactivity died of cancer, while 3 of 4 with normal reactivity remained free of tumor. Similarly, 1 patient with carcinoma of the penis with impaired dinitrochlorobenzene reactivity died of cancer, while 2 of 3 with normal reactivity remained free of tumor. In contrast, reactivity to dinitrochlorobenzene did not correlate with the clinical course of 38 prostatic carcinoma patients. Ten of 19 patients with normal dinitrochlorobenzene reactivity and 9 of 19 with impaired reactivity were dead or had symptomatic recurrences within 1 year, while 9 of 19 with normal reactivity and 10 of 19 with impaired reactivity were either free of tumor or asymptomatic. However, a trend toward a correlation between dinitrochlorobenzene reactivity and tumor progression was observed among patients not receiving endocrine therapy. The differences with respect to the prognostic significance of host immunocompetence between transitional carcinoma patients and those with prostatic carcinoma may be explained by fundamental differences in the biologic properties of these tumors, especially the endocrine sensitivity of prostatic carcinoma.     

Comparison of 15 monoclonal antibodies against tumor-associated antigens of transitional cell carcinoma of the human bladder.

Quantitative urinary immunocytology with our monoclonal antibody (mab) 486p 3/12 proved to be valuable for diagnostic use in bladder-cancer patients' urine, especially in the followup of patients with superficial bladder carcinoma. To evaluate the use of other monoclonal antibodies in bladder cancer, we compared 15 mabs directed against bladder-tumor-associated antigens from seven research groups in a broad panel of cellular and tissue specimens (bladder tumor, prostatic adenoma, and kidney stone). Quantitative evaluation was done in cytocentrifuged preparations and tissue specimens. None of the 15 mabs was bladder-tumor-specific. All 15 stained normal urothelium to some extent and six stained granulocytes. Each of the 15 seemed to identify a different cellular antigen, as can be clearly demonstrated by the staining pattern of different regions in the normal kidney. The sensitivity of quantitative urinary immunocytology in bladder-tumor patients can be improved by using a panel, rather than one mab in bladder-tumor patients, but specificity decreases simultaneously. A main reason for the poor specificity of quantitative urinary immunocytology with all 15 mabs is that false-positive results are obtained with all mabs in kidney-stone patients. Our quantitative urinary immunocytology method is a general tool for the diagnostic use of all mabs in bladder-tumor patients. Mabs that have a high sensitivity might be useful in the followup of patients with superficial bladder carcinoma. None of the 15 mabs (because of their poor specificity) seems to be helpful in quantitative urinary immunocytology for screening a population for bladder carcinoma.