Ovarian Cancer-Associated Ascites Have High Proportions of Cytokine-Responsive CD56bright NK Cells.

Ovarian cancer is the most lethal gynecological malignancy, with serous histotype as the most prevalent epithelial ovarian cancer (EOC). Peritoneal ascites is a frequent comorbidity in advanced EOC. EOC-associated ascites provide a reliable sampling source for studying lymphocytes directly from tumor environment. Herein, we carried out flow cytometry-based analysis to readdress issues on NK and T lymphocyte subsets in women with advanced EOC, additionally evaluating phenotypic modulation of their intracellular pathways involved in interleukin (IL)-2 and IL-15 signaling. Results depicted ascites as an inflammatory and immunosuppressive environment, presenting significantly (p < 0.0001) higher amounts of IL-6 and IL-10 than in the patients' blood, as well as significantly (p < 0.05) increased expression of checkpoint inhibitory receptors (programmed death protein-1, PD-1) and ectonucleotidase (CD39) on T lymphocytes. However, NK lymphocytes from EOC-associated ascites showed higher (p < 0.05) pS6 phosphorylation compared with NK from blood. Additionally, in vitro treatment of lymphocytes with IL-2 or IL-15 elicited significantly (p < 0.001) phosphorylation of the STAT5 protein in NK, CD3 and CD8 lymphocytes, both from blood and ascites. EOC-associated ascites had a significantly (p < 0.0001) higher proportion of NK CD56bright lymphocytes than blood, which, in addition, were more responsive (p < 0.05) to stimulation by IL-2 than CD56dim NK. EOC-associated ascites allow studies on lymphocyte phenotype modulation in the tumor environment, where inflammatory profile contrasts with the presence of immunosuppressive elements and development of cellular self-regulating mechanisms.

P-MAPA immunotherapy potentiates the effect of cisplatin on serous ovarian carcinoma through targeting TLR4 signaling.

BACKGROUND: Toll-like receptors (TLRs) are transmembrane proteins expressed on the surface of ovarian cancer (OC) and immune cells. Identifying the specific roles of the TLR-mediated signaling pathways in OC cells is important to guide new treatments. Because immunotherapies have emerged as the adjuvant treatment for patients with OC, we investigated the effect of a promising immunotherapeutic strategy based on protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride (P-MAPA) combined with cisplatin (CIS) on the TLR2 and TLR4 signaling pathways via myeloid differentiation factor 88 (MyD88) and TLR-associated activator of interferon (TRIF) in an in vivo model of OC. METHODS: Tumors were chemically induced by a single injection of 100 µg of 7,12-dimethylbenz(a)anthracene (DMBA) directly under the left ovarian bursa in Fischer 344 rats. After the rats developed serous papillary OC, they were given P-MAPA, CIS or the combination P-MAPA+CIS as therapies. To understand the effects of the treatments, we assessed the tumor size, histopathology, and the TLR2- and TLR4-mediated inflammatory responses. RESULTS: Although CIS therapy was more effective than P-MAPA in reducing the tumor size, P-MAPA immunotherapy significantly increased the expressions of TLR2 and TLR4. More importantly, the combination of P-MAPA with CIS showed a greater survival rate compared to CIS alone, and exhibited a significant reduction in tumor volume compared to P-MAPA alone. The combination therapy also promoted the increase in the levels of the following OC-related proteins: TLR4, MyD88, TRIF, inhibitor of phosphorylated NF-kB alpha (p-IkBα), and nuclear factor kappa B (NF-kB p65) in both cytoplasmic and nuclear sites. While P-MAPA had no apparent effect on tumor necrosis factor alpha (TNF-α) and interleukin (IL)-6, it seems to increase interferon-γ (IFN-γ), which may induce the Thelper (Th1)-mediated immune response. CONCLUSION: Collectively, our results suggest that P-MAPA immunotherapy combined with cisplatin could be considered an important therapeutic strategy against OC cells based on signaling pathways activated by TLR4.

Serum IL-6 and IL-8 Correlate with Prognostic Factors in Ovarian Cancer.

The aim of the study was to correlate serum levels of IL-2, IL-5, IL-6, IL-8, IL-10, and TNF-α with clinical, laboratory, and pathological prognostic factors in patients with primary ovarian malignancy. Patients treated at the Pelvic Mass Ambulatory of the Discipline of Gynecology and Obstetrics/Oncology Research Institute (IPON) of the UFTM with confirmed diagnosis of malignant ovarian neoplasia (n = 26) were evaluated. Serum collection was performed preoperatively for the determination of tumor markers. The cytokines IL-2, IL-5, IL-6, IL-8, IL-10, and TNF-α were assayed by enzyme-linked immunosorbent assay (ELISA). The prognostic factors were compared using the Mann-Whitney test, with significance level lower than 0.05. When evaluating IL6, it was observed that higher serum levels were associated with overall survival less than 60 months (p = 0.0382). In the evaluation of IL8, higher serum levels were associated with neutrophil-to-lymphocyte ratio (NLR) ≥ 4 and platelet-to-lymphocyte ratio (PLR) ≥ 200 (p = 0.0198 and p = 0.0072, respectively), altered values of serum CA125 (p = 0.0457), and stage IIIC (p = 0.0486). Therefore, increased levels of IL-6 and IL-8 are associated with factors of worse prognosis in ovarian cancer. Additional studies with a larger sample of patients are needed to confirm the role of cytokines as prognostic factors, in the definition of treatment, and in the development of future target therapies.

Influence of ovarian cancer type I and type II microenvironment on the phenotype and function of monocyte-derived dendritic cells.

PURPOSE: The aim of this study was to evaluate the influence of ovarian cancer cell lysates isolated from type I or type II ovarian cancer (OC) on the phenotype of monocyte-derived dendritic cells (Mo-DCs) and the cytokine profile. We also determined whether the Mo-DCs and tumor microenvironment, reflected by peritoneal fluid (PF) from type I or II ovarian cancer, could promote regulatory T cell (Tregs) differentiation from naive CD4+ lymphocytes in vitro. RESULTS: Our results show a significant role of the ovarian cancer microenvironment reflected by PF from type I or II OC in the inhibition of the DC differentiation process. Interestingly, the percentage of cells co-expressing CD45 and CD14 antigens in the cultures stimulated with PF from both type I and type II OC was higher than in the control. Furthermore, the percentage of cells expressing CD1a, i.e., a marker of immature DCs, was significantly reduced in the cultures stimulated with PF from type I and type II OC. The results obtained show that ovarian cancer type II lysates induce differentiation of monocytes into macrophage-like cells with a CD1a+/HLA-DR+/CD83- phenotype and significantly higher CD86/HLA-DR expression. We show that ovarian cancer type II Mo-DCs are able to prevent an immune response by release of IL-10, whereas OC type I Mo-DCs can promote the generation of Tregs. CONCLUSIONS: We demonstrate that each type of ovarian cancer can induce a unique phenotype of DCs and differentiation of Tregs, both associated with immune-suppressive function, which may be an obstacle while developing effective anticancer dendritic cell vaccination.

Up-regulation of DNAM-1 and NKp30, associated with improvement of NK cells activation after long-term culture of mononuclear cells from patients with ovarian neoplasia.

This study aimed at evaluating the functional activation and activating receptors expression on resting, short- and long-term NK and NK-like T cells from blood of ovarian neoplasia patients. Blood from patients with adnexal benign alterations (n = 10) and ovarian cancer (grade I-IV n = 14) were collected after signed consent. Effector cells activation was evaluated by the expression of the CD107a molecule. Short-term culture was conducted overnight with IL-2 and long-term culture for 21 days, by a method designed to expand CD56(+) lymphocytes. Short-term culture significantly increased NK cells activation compared to resting NK cells (p<0.05), however, the long-term procedure supported an even higher increase (p<0.001). Resting NK-like T cells showed poor activation, which was not altered by the culture procedures. The long-term culture effectively increased the expression of the activating receptors on NK and NK-like T cells, either by increasing the number of cells expressing a given receptor and/or by up-regulating their expression intensity. As a conclusion, the long-term culture system employed, resulted in a high number of functional NK cells. The culture system was particularly efficient on the up-regulation of NKp30 and DNAM-1 receptors on NK cells.

Carcinoma subseroso de la vesícula biliar: expresión del complejo caderina-catenina / Subserous gallbladder carcinoma: expression of cadherine-catenine complex

BACKGROUND: Subserous gallbladder carcinoma is difficult to diagnose and treat. There are no tissue markers with prognostic value in this type of tumor. AIM: To study the immunohistochemical expression of E-cadherin alpha and beta catenin in subserous gallbladder carcinoma. PATIENTS AND METHODS: One hundred seventeen subjects (103 women and 14 men aged 62 and 69 years as a mean, respectively), were studied. Thirty five gallbladder samples without evidence of cancer were used as controls. Expression of markers was studied with standard immunohistochemical techniques for formalin fixed and paraffin embedded tissue. RESULTS: Ninety seven percent of tumors were adenocarcinoma. A lower or absent expression of E-cadherin, alpha catenin and beta catenin was observed in 26, 33 and 29 per cent, respectively. Actuarial five years survival was 37 per cent. No association between macroscopic features of the tumor and survival was observed. Well differentiated tumors had a 73 per cent survival, whereas less differentiated tumors had a 30 per cent survival. Tumors with a normal expression of the markers had a slightly better survival, although not significant (p = 0.06). CONCLUSIONS: Approximately 30 per cent of subserous gallbladder carcinoma have an abnormal expression of E-cadherin, alpha catenin and beta catenin. This abnormal expression has no relationship with prognosis and is probably secondary to the aberrant genic expression of the tumor.

[Subserous gallbladder carcinoma: expression of cadherine-catenine complex]. / Carcinoma subseroso de la vesícula biliar: expresión del complejo caderina-catenina.

BACKGROUND: Subserous gallbladder carcinoma is difficult to diagnose and treat. There are no tissue markers with prognostic value in this type of tumor. AIM: To study the immunohistochemical expression of E-cadherin alpha and beta catenin in subserous gallbladder carcinoma. PATIENTS AND METHODS: One hundred seventeen subjects (103 women and 14 men aged 62 and 69 years as a mean, respectively), were studied. Thirty five gallbladder samples without evidence of cancer were used as controls. Expression of markers was studied with standard immunohistochemical techniques for formalin fixed and paraffin embedded tissue. RESULTS: Ninety seven percent of tumors were adenocarcinoma. A lower or absent expression of E-cadherin, alpha catenin and beta catenin was observed in 26, 33 and 29% of tumors, respectively. Actuarial five years survival was 37%. No association between macroscopic features of the tumor and survival was observed. Well differentiated tumors had a 73% survival, whereas less differentiated tumors had a 30% survival. Tumors with a normal expression of the markers had a slightly better survival, although not significant (p = 0.06). CONCLUSIONS: Approximately 30% of subserous gallbladder carcinoma have an abnormal expression of E-cadherin, alpha catenin and beta catenin. This abnormal expression has no relationship with prognosis and is probably secondary to the aberrant genic expression of the tumor.