The aim of the present study was to investigate within ovarian carcinoma and normal ovarian biopsies the gene expression of multiple secretoglobin family members relative to mammaglobin B, which we ...previously reported as a promising novel ovarian carcinoma prognostic marker.
Using quantitative real-time Reverse Transcription PCR we tested 53 ovarian carcinoma and 30 normal ovaries for the expression of 8 genes belonging to the secretoglobin family: mammaglobin A, lipophilin A, lipophilin B, uteroglobin, HIN-1, UGRP-1, RYD5 and IIS. Next, we decided to expand the LipB gene expression analysis to a further 48 ovarian carcinoma samples, for a total of 101 tumor tissues of various histologies and to study its protein expression by immunohistochemistry in formalin-fixed paraffin-embedded tumors and normal ovaries. Finally, we correlated lipophilin B gene and protein expression to conventional patient clinico-pathological features and outcome.
We found significant mammaglobin A, lipophilin A, lipophilin B and RYD5 gene overexpression in ovarian carcinomas compared to normal ovaries. Lipophilin B mRNA showed a higher presence in tumors (75.4%) compared to normal ovaries (16.6%) and the most significant correlation with mammaglobin B mRNA (rs =0.77, p < 0.001). By immunohistochemical analysis, we showed higher lipophilin B expression in the cytoplasm of tumor cells compared to normal ovaries (p < 0.001). Moreover, lipophilin B gene overexpression was significantly associated with serous histology (serous vs clear cell p = 0.027; serous vs undifferentiated p = 0.007) and lower tumor grade (p = 0.02). Lower LipB mRNA levels (low versus high tertiles) were associated to a shorter progression-free (p = 0.03, HR = 2.2) and disease-free survival (p = 0.02, HR = 2.5) by univariate survival analysis and, importantly, they remain an independent prognostic marker for decreased disease-free (p = 0.001, HR = 3.9) and progression-free survival (p = 0.004, HR = 2.8) in multivariate Cox regression analysis.
The present study represents the first quantitative evaluation of secretoglobin gene expression in normal and neoplastic ovarian tissues. Our results demonstrate lipophilin B gene and protein upregulation in ovarian carcinoma compared to normal ovary. Moreover, lipophilin B gene overexpression correlates with a less aggressive tumor phenotype and represents a novel ovarian carcinoma prognostic factor.
Objective: Retinoids are a class of compounds that are structurally related to vitamin A and have been found to be effective in the prevention and treatment of cervical cancer. To investigate whether ...enhanced immunogenicity might be responsible for such efficacy, we evaluated the effects of retinoic acid on the expression of major histocompatibility complex class I and class II and intercellular adhesion molecule ICAM-1 in human cervical carcinoma cell lines. Study Design: The expression of surface antigens (major histocompatibility complex class I and class II and ICAM-1) was evaluated by fluorescence-activated cell sorter analysis in 3 human cervical carcinoma cell lines after exposure to therapeutic doses of retinoic acid. In addition, the effects on human leukocyte antigen class I messenger ribonucleic acid expression were also evaluated by Northern blot analysis after such treatment. Results: CaSki, SiHa, and HT-3 cervical cancer cells expressed variable levels of major histocompatibility complex class I and ICAM-1 antigens, whereas class II surface antigens were not detectable. Exposure to therapeutic doses of retinoic acid were able to significantly increase the expression of major histocompatibility complex class I and ICAM-1 antigens in all the cell lines when compared with untreated tumor cells but were not able to induce the expression of class II surface human leukocyte antigens. Northern blot analysis showed that for major histocompatibility complex class I molecules such up-regulation was the result of an increased expression at the transcriptional level of major histocompatibility complex class I messenger ribonucleic acid. Conclusions: These data indicate that retinoic acid increases the expression of immunologically important surface antigens, suggesting that the efficacy of retinoic acid in the treatment of cervical cancer may be, at least in part, the result of immunologic modulation. Such findings support additional clinical research investigating the use of retinoids for the treatment of cervical cancer. (Am J Obstet Gynecol 1998;179:1020-5.)
We describe a 65-year-old woman with a large surgically unresectable and chemoresistant liver metastasis of endometrial carcinoma who was treated by infusion with peripheral blood T cells stimulated ...with tumor lysate-pulsed autologous dendritic cells (DC). Extensive in vitro characterization of the DC-activated T cells included phenotypic analysis, cytotoxicity, and intracellular cytokine secretion. High cytotoxicity was observed against autologous tumor cells, but not against NK-sensitive K562 cells, autologous Con-A lymphoblasts, or autologous Epstein-Barr virus-transformed lymphoblastoid cells. Blocking studies demonstrated that lytic activity was HLA class I restricted. Two-color flow cytometric analysis revealed that a significant proportion of CD8+ T cells was also CD56+, and analysis of intracellular IFN-gamma and IL-4 expression suggested a type 1 cytokine bias. The patient was treated by three infusions of tumor-specific T cells at 3- to 4-week intervals, and in vivo distribution of the T cells was followed by (111)In oxine labeling and serial gamma camera imaging. Tumor localization and accumulation of labeled lymphocytes was consistently detected at serial time points following each injection. However, deep infiltration of the large tumor mass by activated T cells was minimal, as evaluated in 3 dimensions by single photon emission computerized tomography (SPECT) imaging. Transient serum increases of the tumor marker lactate dehydrogenase (LDH), were detectable after each injection. Similar posttreatment elevations were seen for serum uric acid and potassium. Clinically, stabilization of the large liver metastasis was obtained during treatment. Collectively, these results indicate that tumor-specific CD8+ cytotoxic T-cell responses can be generated in patients with endometrial cancer, and suggest that T-cell immunotherapy may be of therapeutic value in patients harboring metastatic disease.
Purpose: To compare the effects of concurrent administration of cisplatinum (40 mg/m
2/weekly) with radiation therapy (C-RT) to those induced by radiation therapy alone (RT) on the immune function of ...patients with locally advanced cervical cancer.
Methods and Materials: In 8 prospectively randomized patients (i.e., 4 receiving RT vs. 4 receiving C-RT), lymphocyte populations including CD3+, CD4+ and CD8+ T-cell subsets, B cells (CD19+) and natural killer cells (CD56+, CD16+, CD3−) were studied before, during, and after therapy. Expression of the activation marker CD25 on CD3+ T cells, intracellular levels of perforin in CD8+ and CD56+ cells, and interferon-γ (IFN-γ) and IL-2 in CD4+ and CD8+ T cells was also measured. Finally, lymphoblast transformation and natural killer (NK) cytotoxic activity were assessed.
Results: Both RT and C-RT significantly decreased the mean absolute number of all lymphocyte subsets compared to pretreatment levels (
p > 0.001). However, no differences were detected in the characteristics or the magnitude of the lymphopenia induced by the two treatments. Both RT and C-RT increased similarly the percentages of CD25-positive lymphocytes (
p > 0.001), and significantly decreased PHA-induced T-cell lymphoblast transformation (
p > 0.001) and NK cytotoxic activity against K562 cells (
p > 0.001). The percentage of perforin-positive and CD8+ T cells was not altered during either treatment, whereas the percentage of perforin-positive and CD56+ cells was significantly reduced during both treatments, and correlated with reduced cytotoxicity against K562 cells. The percentages of CD8+ IFN-γ+ and CD4+ IFN-γ+ T cells as well as that of CD8+ IL-2+ and CD4+ IL2+ T cells were not significantly altered by C-RT compared to RT alone. Finally, with both regimens, NK cells and B-cell numbers showed a more rapid recovery than T-cell numbers.
Conclusion: Administration of concurrent cisplatinum to radiation may synergistically increase cytotoxic effects of radiation on tumor cells but does not alter the magnitude and the characteristics of radiation-induced immunosuppression.
We describe a 27-year-old woman with systemic chemoresistant and radioresistant metastatic disease secondary to a recurrence of human papillomavirus (HPV) 18 infected cervical adenocarcinoma of the ...uterine cervix who received adoptive transfer of peripheral blood T cells stimulated with HPV 18 E7-pulsed autologous dendritic cells (DC). Extensive in vitro characterization of the DC-activated T cells derived from peripheral blood mononuclear cells (PBMC) included phenotypic analysis, cytotoxicity and intracellular cytokine production. High cytotoxicity activity was observed by CD8+T cells against autologous tumor cells, but not against NK-sensitive K562 cells, autologous Con-A lymphoblasts, or autologous Epstein-Barr virus-transformed lymphoblastoid cells. Blocking studies demonstrated that lytic activity was significantly inhibited by pretreatment of tumor targets with MAb specific for HLA class I as well as that of effector cells with anti-CD8, anti-LFA-1, but not anti CD3 MAb. Two-color flow cytometric analysis of the cytotoxic T cells revealed that a significant proportion of CD8+ cells was also CD56+. These double positive CTLs were thymically derived, as shown by expression of heterodimeric CD8 molecules (alpha/beta CD8) and were endowed with high cytotoxic activity against tumor cells. Analysis of intracellular cytokine expression showed that the striking majority of E7-pulsed DC activated CD8+ T cells strongly expressed IFN-gamma, TNF-alpha and IL-2 but not IL-4. The patient received two infusions of cytotoxic tumor-specific T cells at 2 week intervals, and in vivo distribution of the T cells was followed by 111 oxine labeling and serial gamma camera imaging. Persistent accumulation of radioactivity in the lungs, which harbored extensive metastatic disease, was detected up to 120 hrs after the infusion. Taken together, these results illustrate the potential of E7-specific and tumor-specific CTL-based immunotherapy for the treatment of patients with invasive cervical cancer.
Little is known about the prevalence of serum hepatitis C virus (HCV) RNA in blood donors with HCV antibodies and persistently normal alanine aminotransferase (ALT) levels.
Thirty-nine ...anti-HCV-positive donors with normal ALT on four determinations at 3-month intervals were further tested monthly for 6 months, and they had normal ALT values. The presence of HCV RNA was determined in these 39 donors.
Serum HCV RNA was detected in 16 of 39 donors, 14 of 14 who reacted on second-generation recombinant immunoblot assay (RIBA-2) and 2 of 15 who were indeterminate. None of the 10 RIBA-2-nonreactive donors had evidence of viremia. The 15 RIBA-2-indeterminate samples were tested with third-generation RIBA (RIBA-3); the results showed reactivity in 5 (including the 2 HCV RNA positive), an indeterminate pattern in 7, and nonreactivity in 3 (all RNA negative). Among HCV RNA-positive subjects, mean age (p < 0.05), mean ALT (p < 0.001), signal-to-cutoff (S:CO) ratio on second-generation enzyme-linked immunosorbent assay (p < 0.001), and gamma globulin levels (p < 0.05) were higher than those among HCV RNA-negative subjects. During 6 additional months of ALT monitoring, completed by 36 of 39 donors, increased values were detected in 6 (5 HCV RNA positive). In 4 of those 6, however, ALT levels were less than 1.5-fold the upper normal limit. HCV RNA results were unchanged at the end of 1-year follow-up.
Forty-one percent of anti-HCV-positive donors with persistently normal ALT had active HCV infection. Long-term ALT monitoring allowed the detection of significantly increased enzyme values in only 2 of 16 viremic donors. Reactivity on RIBA-2 or -3, greater age, mean ALT levels in the upper range of normal, higher S:CO ratio on second-generation enzyme-linked immunosorbent assay, and higher gamma globulin levels were predictive of viremia.
To assess expression of the immunosuppressive cytokines IL-10 and TGF-beta in the ascitic fluid and plasma of advanced ovarian cancer patients.
A prospective study.
The Department of Obstetrics and ...Gynaecology at the University of Arkansas for Medical Sciences.
Twenty-eight women diagnosed with advanced ovarian cancer and ten normal female controls.
Plasma and ascitic samples were collected at the time of surgery and analysed for the presence of IL-10 and TGF-beta using a sensitive enzyme-linked immunosorbent assay.
Elevated levels of IL-10 were detected in the plasma mean (SD) = 12 (5) pg/mL; range 8 to 23 pg/mL and in the peritoneal fluid mean (SD) = 165 (137) pg/mL; range 50 to 556 pg/mL of ovarian cancer patients, while no detectable IL-10 was found in any of the normal control plasma samples tested. Similarly, plasma levels of TGF-beta in ovarian cancer patients were significantly higher mean (SD) = 1,506 (246) pg/mL; range 1,020 to 2,070 pg/mL compared with controls mean (SD) = 937 (187) pg/mL; range 770 to 1,140 pg/mL(P < 0.001). Surprisingly, however, although elevated TGF-beta levels were also detected in the peritoneal fluid of all ovarian cancer patients mean (SD) = 407 (158) pg/mL; range 140 to 770 pg/mL, these levels were significantly lower than those seen in matched plasma samples (P < 0.001).
Local and systemic secretion of immunosuppressive cytokines may play an important role in the impaired anti-tumour immune function commonly observed in advanced ovarian cancer. However, the observation that plasma levels of TGF-beta are significantly higher than those detected in the ascitic fluid raises the possibility that cells other than tumour cells are responsible for TGF-beta release in the bloodstream of these patients.
Objective: The purpose of this study was to evaluate the potential of dendritic cells pulsed with whole-tumor extracts derived from autologous ovarian cancer cells in eliciting a tumor-specific ...cytotoxic T-cell response in vitro from patients with advanced ovarian cancer. Study Design: CD8+ T lymphocytes stimulated in vitro with autologous ovarian tumor lysate–pulsed dendritic cells were tested for their ability to induce a human leukocyte antigen class I–restricted cytotoxic T-lymphocyte response able to specifically kill autologous tumor cells in standard 6-hour chromium 51 cytotoxicity assays. In addition, to correlate cytotoxic activity by cytotoxic T-lymphocytes with a particular lymphoid subset, 2-color flow cytometric analysis of intracellular cytokine expression (interferon γ and interleukin 4) at the single-cell level was performed. Results: Cytotoxic T lymphocytes specific for autologous ovarian tumor cells were elicited from 3 patients with advanced ovarian cancer. Although cytotoxic T-lymphocyte populations expressed strong cytolytic activity against autologous tumor cells, they did not lyse concanavalin A–stimulated autologous lymphocytes or autologous Epstein-Barr virus–transformed lymphoblastoid cell lines and showed negligible cytotoxicity against the natural killer cell–sensitive cell line K-562. Cytotoxic effect against the autologous tumor cells was inhibited by an anti–human leukocyte antigen class I monoclonal antibody (W6/32). It is interesting that CD8+ cytotoxic T lymphocytes expressed variable levels of CD56, a marker that may be associated with high cytotoxic activity. Finally, most of the tumor-specific CD8+ T cells exhibited a TH1 cytokine bias, and a high percentage of interferon γ expressors among cytotoxic T lymphocytes was correlated with higher cytotoxic activity. Conclusion: These data show that tumor lysate–pulsed dendritic cells can consistently induce in vitro specific CD8+ cytotoxic T lymphocytes able to kill autologous tumor cells from patients with advanced stage ovarian cancer. This novel approach may have important implications for the treatment of residual or resistant disease with active or adoptive immunotherapy after standard surgical and cytotoxic treatment. (Am J Obstet Gynecol 2000;183:601-9.)
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