New Insights in Immunometabolism in Neonatal Monocytes and Macrophages in Health and Disease.

It is well established that the neonatal immune system is different from the adult immune system. A major task of the neonatal immune system is to bridge the achievement of tolerance towards harmless antigens and commensal bacteria while providing protection against pathogens. This is highly important because neonates are immunologically challenged directly after birth by a rigorous change from a semi-allogeneic sterile environment into a world rich with microbes. A so called disease tolerogenic state is typical for neonates and is anticipated to prevent immunopathological damage potentially at the cost of uncontrolled pathogen proliferation. As a consequence, neonates are more susceptible than adults to life-threatening infections. At the basis of a well-functioning immune response, both for adults and neonates, innate immune cells such as monocytes and monocyte-derived macrophages play an essential role. A well-responsive monocyte will alter its cellular metabolism to subsequently induce certain immune effector function, a process which is called immunometabolism. Immunometabolism has received extensive attention in the last decade; however, it has not been broadly studied in neonates. This review focuses on carbohydrate metabolism in monocytes and macrophages in neonates. We will exhibit pathways involving glycolysis, the tricarboxylic acid (TCA) cycle and oxidative phosphorylation and their role in shaping neonates' immune systems to a favorable tolerogenic state. More insight into these pathways will elucidate potential treatments targets in life-threatening conditions including neonatal sepsis or expose potential targets which can be used to induce tolerance in conditions where tolerance is harmfully impaired such as in autoimmune diseases.

Lung Epithelial CYP1 Activity Regulates Aryl Hydrocarbon Receptor Dependent Allergic Airway Inflammation.

The lung epithelial barrier serves as a guardian towards environmental insults and responds to allergen encounter with a cascade of immune reactions that can possibly lead to inflammation. Whether the environmental sensor aryl hydrocarbon receptor (AhR) together with its downstream targets cytochrome P450 (CYP1) family members contribute to the regulation of allergic airway inflammation remains unexplored. By employing knockout mice for AhR and for single CYP1 family members, we found that AhR-/- and CYP1B1-/- but not CYP1A1-/- or CYP1A2-/- animals display enhanced allergic airway inflammation compared to WT. Expression analysis, immunofluorescence staining of murine and human lung sections and bone marrow chimeras suggest an important role of CYP1B1 in non-hematopoietic lung epithelial cells to prevent exacerbation of allergic airway inflammation. Transcriptional analysis of murine and human lung epithelial cells indicates a functional link of AhR to barrier protection/inflammatory mediator signaling upon allergen challenge. In contrast, CYP1B1 deficiency leads to enhanced expression and activity of CYP1A1 in lung epithelial cells and to an increased availability of the AhR ligand kynurenic acid following allergen challenge. Thus, differential CYP1 family member expression and signaling via the AhR in epithelial cells represents an immunoregulatory layer protecting the lung from exacerbation of allergic airway inflammation.

RelB Deficiency in Dendritic Cells Protects from Autoimmune Inflammation Due to Spontaneous Accumulation of Tissue T Regulatory Cells.

Foxp3+ regulatory T cells are well-known immune suppressor cells in various settings. In this study, we provide evidence that knockout of the relB gene in dendritic cells (DCs) of C57BL/6 mice results in a spontaneous and systemic accumulation of Foxp3+ T regulatory T cells (Tregs) partially at the expense of microbiota-reactive Tregs. Deletion of nfkb2 does not fully recapitulate this phenotype, indicating that alternative NF-κB activation via the RelB/p52 complex is not solely responsible for Treg accumulation. Deletion of RelB in DCs further results in an impaired oral tolerance induction and a marked type 2 immune bias among accumulated Foxp3+ Tregs reminiscent of a tissue Treg signature. Tissue Tregs were fully functional, expanded independently of IL-33, and led to an almost complete Treg-dependent protection from experimental autoimmune encephalomyelitis. Thus, we provide clear evidence that RelB-dependent pathways regulate the capacity of DCs to quantitatively and qualitatively impact on Treg biology and constitute an attractive target for treatment of autoimmune diseases but may come at risk for reduced immune tolerance in the intestinal tract.

Pro-Angiogenic Macrophage Phenotype to Promote Myocardial Repair.

BACKGROUND: Heart failure following myocardial infarction (MI) remains one of the major causes of death worldwide, and its treatment is a crucial challenge of cardiovascular medicine. An attractive therapeutic strategy is to stimulate endogenous mechanisms of myocardial regeneration. OBJECTIVES: This study evaluates the potential therapeutic treatment with annexin A1 (AnxA1) to induce cardiac repair after MI. METHODS: AnxA1 knockout (AnxA1-/-) and wild-type mice underwent MI induced by ligation of the left anterior descending coronary artery. Cardiac functionality was assessed by longitudinal echocardiographic measurements. Histological, fluorescence-activated cell sorting, dot blot analysis, and in vitro/ex vivo studies were used to assess the myocardial neovascularization, macrophage content, and activity in response to AnxA1. RESULTS: AnxA1-/- mice showed a reduced cardiac functionality and an expansion of proinflammatory macrophages in the ischemic area. Cardiac macrophages from AnxA1-/- mice exhibited a dramatically reduced ability to release the proangiogenic mediator vascular endothelial growth factor (VEGF)-A. However, AnxA1 treatment enhanced VEGF-A release from cardiac macrophages, and its delivery in vivo markedly improved cardiac performance. The positive effect of AnxA1 treatment on cardiac performance was abolished in wild-type mice transplanted with bone marrow derived from Cx3cr1creERT2Vegfflox/flox or in mice depleted of macrophages. Similarly, cardioprotective effects of AnxA1 were obtained in pigs in which full-length AnxA1 was overexpressed by use of a cardiotropic adeno-associated virus. CONCLUSIONS: AnxA1 has a direct action on cardiac macrophage polarization toward a pro-angiogenic, reparative phenotype. AnxA1 stimulated cardiac macrophages to release high amounts of VEGF-A, thus inducing neovascularization and cardiac repair.

Hematopoietic ChemR23 (Chemerin Receptor 23) Fuels Atherosclerosis by Sustaining an M1 Macrophage-Phenotype and Guidance of Plasmacytoid Dendritic Cells to Murine Lesions-Brief Report.

Objective- Expression of the chemokine-like receptor ChemR23 (chemerin receptor 23) has been specifically attributed to plasmacytoid dendritic cells (pDCs) and macrophages and ChemR23 has been suggested to mediate an inflammatory immune response in these cells. Because chemokine receptors are important in perpetuating chronic inflammation, we aimed to establish the role of ChemR23-deficiency on macrophages and pDCs in atherosclerosis. Approach and Results- ChemR23-knockout/knockin mice expressing eGFP (enhanced green fluorescent protein) were generated and after crossing with apolipoprotein E-deficient ( Apoe-/- ChemR23 e/e) animals were fed a western-type diet for 4 and 12 weeks. Apoe-/- ChemR23 e/e mice displayed reduced lesion formation and reduced leukocyte adhesion to the vessel wall after 4 weeks, as well as diminished plaque growth, a decreased number of lesional macrophages with an increased proportion of M2 cells and a less inflammatory lesion composition after 12 weeks of western-type diet feeding. Hematopoietic ChemR23-deficiency similarly reduced atherosclerosis. Additional experiments revealed that ChemR23-deficiency induces an alternatively activated macrophage phenotype, an increased cholesterol efflux and a systemic reduction in pDC frequencies. Consequently, expression of the pDC marker SiglecH in atherosclerotic plaques of Apoe-/- ChemR23 e/e mice was declined. ChemR23-knockout pDCs also exhibited a reduced migratory capacity and decreased CCR (CC-type chemokine receptor)7 expression. Finally, adoptive transfer of sorted wild-type and knockout pDCs into Apoe-/- recipient mice revealed reduced accumulation of ChemR23-deficient pDCs in atherosclerotic lesions. Conclusions- Hematopoietic ChemR23-deficiency increases the proportion of alternatively activated M2 macrophages in atherosclerotic lesions and attenuates pDC homing to lymphatic organs and recruitment to atherosclerotic lesions, which synergistically restricts atherosclerotic plaque formation and progression.

Chrono-pharmacological Targeting of the CCL2-CCR2 Axis Ameliorates Atherosclerosis.

Onset of cardiovascular complications as a consequence of atherosclerosis exhibits a circadian incidence with a peak in the morning hours. Although development of atherosclerosis extends for long periods of time through arterial leukocyte recruitment, we hypothesized that discrete diurnal invasion of the arterial wall could sustain atherogenic growth. Here, we show that myeloid cell recruitment to atherosclerotic lesions oscillates with a peak during the transition from the activity to the resting phase. This diurnal phenotype is regulated by rhythmic release of myeloid cell-derived CCL2, and blockade of its signaling abolished oscillatory leukocyte adhesion. In contrast, we show that myeloid cell adhesion to microvascular beds peaks during the early activity phase. Consequently, timed pharmacological CCR2 neutralization during the activity phase caused inhibition of atherosclerosis without disturbing microvascular recruitment. These findings demonstrate that chronic inflammation of large vessels feeds on rhythmic myeloid cell recruitment, and lay the foundation for chrono-pharmacology-based therapy.

Pulmonary Function Changes After Radiotherapy for Lung or Esophageal Cancer: A Systematic Review Focusing on Dose-Volume Parameters.

BACKGROUND: Despite technical developments in treatment delivery, radiation-induced lung toxicity (RILT) remains a crucial problem in thoracic radiotherapy. Clinically based RILT scores have their limitations, and more objective measures such as pulmonary functions tests (PFTs) might help to improve treatment strategies. PURPOSE: To summarize the available evidence about the effect of dose to the lung in thoracic radiotherapy on forced expiratory volume in one second (FEV1) and diffusion capacity (DLCO) in patients with lung and esophageal cancer treated with curative intent. MATERIAL AND METHODS: A systematic review following the PRISMA guidelines was performed, using MEDLINE and including clinical studies using (chemo)radiotherapy (CRT) or stereotactic ablative radiotherapy (SABR) for lung or CRT for esophageal cancer that reported both lung dose-volume histogram (DVH) parameters and changes in PFT results. Search terms included lung and esophageal neoplasms, respiratory function tests, and radiotherapy. RESULTS: Fifteen studies met the inclusion criteria. Seven out of 13 studies on lung cancer reported significant declines (defined as a p value < .05) in PFT results. Both esophageal studies reported significant DLCO declines. One SABR study found a correlation between low lung-dose parameters and FEV1 decline. Relations between decline of FEV1 (three studies) or decline of DLCO (five studies), respectively, and DVH parameters were found in eight studies analyzing CRT. Furthermore, a heterogeneous range of clinical risk factors for pulmonary function changes were reported in the selected studies. CONCLUSIONS: There is evidence that pulmonary function declines after RT in a dose-dependent manner, but solid data about lung DVH parameters predicting changes in PFT results are scarce. A major disadvantage was the wide variety of methods used, frequently lacking multivariable analyses. Studies using prospective high-quality data, analyzed with appropriate statistical methods, are needed. The Oncologist 2017;22:1257-1264 IMPLICATIONS FOR PRACTICE: Radiation-induced lung toxicity remains crucial in thoracic radiotherapy. To prevent this toxicity in the future and individualize patient treatment, objective measures of pulmonary toxicity are needed. Pulmonary function tests may provide such objective measures. This systematic review, included all available clinical studies using external beam radiotherapy for lung or esophageal cancer reporting pulmonary function combined with dose-volume histogram parameters. There is preliminary evidence that pulmonary function declines post radiotherapy in a dose-dependent manner. Data quality and analyses were generally limited. Analyses of high-quality data are therefore urgently needed to improve individualization of advanced radiation therapy.

Human Neutrophil Peptide 1 Limits Hypercholesterolemia-induced Atherosclerosis by Increasing Hepatic LDL Clearance.

Increases in plasma LDL-cholesterol have unequivocally been established as a causal risk factor for atherosclerosis. Hence, strategies for lowering of LDL-cholesterol may have immediate therapeutic relevance. Here we study the role of human neutrophil peptide 1 (HNP1) in a mouse model of atherosclerosis and identify its potent atheroprotective effect both upon transgenic overexpression and therapeutic delivery. The effect was found to be due to a reduction of plasma LDL-cholesterol. Mechanistically, HNP1 binds to apolipoproteins enriched in LDL. This interaction facilitates clearance of LDL particles in the liver via LDL receptor. Thus, we here identify a non-redundant mechanism by which HNP1 allows for reduction of LDL-cholesterol, a process that may be therapeutically instructed to lower cardiovascular risk.

Protective Aptitude of Annexin A1 in Arterial Neointima Formation in Atherosclerosis-Prone Mice-Brief Report.

OBJECTIVE: Restenosis as a consequence of arterial injury is aggravated by inflammatory pathways. Here, we investigate the role of the proresolving protein annexin A1 (AnxA1) in healing after wire injury. APPROACH AND RESULTS: Apoe-/- and Apoe-/-Anxa1-/- mice were subjected to wire injury while fed a high-cholesterol diet. Subsequently, localization of AnxA1 and AnxA1 plasma levels were examined. AnxA1 was found to localize within endothelial cells and macrophages in the neointima. Levels of AnxA1 in the plasma and its lesional expression negatively correlated with neointima size, and in the absence of AnxA1, neointima formation was aggravated by the accumulation and proliferation of macrophages. In contrast, reendothelialization and smooth muscle cell infiltration were not affected in Apoe-/-Anxa1-/- mice. CONCLUSIONS: AnxA1 is protective in healing after wire injury and could, therefore, be an attractive therapeutic compound to prevent from restenosis after vascular damage.

The advantageous role of annexin A1 in cardiovascular disease.

The inflammatory response protects the human body against infection and injury. However, uncontrolled and unresolved inflammation can lead to tissue damage and chronic inflammatory diseases. Therefore, active resolution of inflammation is essential to restore tissue homeostasis. This review focuses on the pro-resolving molecule annexin A1 (ANXA1) and its derived peptides. Mechanisms instructed by ANXA1 are multidisciplinary and affect leukocytes as well as endothelial cells and tissue resident cells like macrophages and mast cells. ANXA1 has an outstanding role in limiting leukocyte recruitment and different aspects of ANXA1 as modulator of the leukocyte adhesion cascade are discussed here. Additionally, this review details the therapeutic relevance of ANXA1 and its derived peptides in cardiovascular diseases since atherosclerosis stands out as a chronic inflammatory disease with impaired resolution and continuous leukocyte recruitment.

Cathepsin G Controls Arterial But Not Venular Myeloid Cell Recruitment.

BACKGROUND: Therapeutic targeting of arterial leukocyte recruitment in the context of atherosclerosis has been disappointing in clinical studies. Reasons for such failures include the lack of knowledge of arterial-specific recruitment patterns. Here we establish the importance of the cathepsin G (CatG) in the context of arterial myeloid cell recruitment. METHODS: Intravital microscopy of the carotid artery, the jugular vein, and cremasteric arterioles and venules in Apoe-/-and CatG-deficient mice (Apoe-/-Ctsg-/-) was used to study site-specific myeloid cell behavior after high-fat diet feeding or tumor necrosis factor stimulation. Atherosclerosis development was assessed in aortic root sections after 4 weeks of high-fat diet, whereas lung inflammation was assessed after inhalation of lipopolysaccharide. Endothelial deposition of CatG and CCL5 was quantified in whole-mount preparations using 2-photon and confocal microscopy. RESULTS: Our observations elucidated a crucial role for CatG during arterial leukocyte adhesion, an effect not found during venular adhesion. Consequently, CatG deficiency attenuates atherosclerosis but not acute lung inflammation. Mechanistically, CatG is immobilized on arterial endothelium where it activates leukocytes to firmly adhere engaging integrin clustering, a process of crucial importance to achieve effective adherence under high-shear flow. Therapeutic neutralization of CatG specifically abrogated arterial leukocyte adhesion without affecting myeloid cell adhesion in the microcirculation. Repetitive application of CatG-neutralizing antibodies permitted inhibition of atherogenesis in mice. CONCLUSIONS: Taken together, these findings present evidence of an arterial-specific recruitment pattern centered on CatG-instructed adhesion strengthening. The inhibition of this process could provide a novel strategy for treatment of arterial inflammation with limited side effects.

Annexin A1 counteracts chemokine-induced arterial myeloid cell recruitment.

RATIONALE: Chemokine-controlled arterial leukocyte recruitment is a crucial process in atherosclerosis. Formyl peptide receptor 2 (FPR2) is a chemoattractant receptor that recognizes proinflammatory and proresolving ligands. The contribution of FPR2 and its proresolving ligand annexin A1 to atherosclerotic lesion formation is largely undefined. OBJECTIVE: Because of the ambivalence of FPR2 ligands, we here investigate the role of FPR2 and its resolving ligand annexin A1 in atherogenesis. METHODS AND RESULTS: Deletion of FPR2 or its ligand annexin A1 enhances atherosclerotic lesion formation, arterial myeloid cell adhesion, and recruitment. Mechanistically, we identify annexin A1 as an endogenous inhibitor of integrin activation evoked by the chemokines CCL5, CCL2, and CXCL1. Specifically, the annexin A1 fragment Ac2-26 counteracts conformational activation and clustering of integrins on myeloid cells evoked by CCL5, CCL2, and CXCL1 through inhibiting activation of the small GTPase Rap1. In vivo administration of Ac2-26 largely diminishes arterial recruitment of myeloid cells in a FPR2-dependent fashion. This effect is also observed in the presence of selective antagonists to CCR5, CCR2, or CXCR2, whereas Ac2-26 was without effect when all 3 chemokine receptors were antagonized simultaneously. Finally, repeated treatment with Ac2-26 reduces atherosclerotic lesion sizes and lesional macrophage accumulation. CONCLUSIONS: Instructing the annexin A1-FPR2 axis harbors a novel approach to target arterial leukocyte recruitment. With the ability of Ac2-26 to counteract integrin activation exerted by various chemokines, delivery of Ac2-26 may be superior in inhibition of arterial leukocyte recruitment when compared with blocking individual chemokine receptors.

Less gastrointestinal toxicity after adjuvant radiotherapy on a small pelvic field compared to a standard pelvic field in patients with endometrial carcinoma.

OBJECTIVE: Radiotherapy is associated with short-term and long-term morbidity. This study compared toxicity rates among patients with endometrial carcinoma (EC) treated with adjuvant external beam radiation therapy (EBRT) on a small pelvic field (SmPF) in comparison with a standard pelvic field (StPF) or an extended field (EF). METHODS: Patients with EC preoperatively diagnosed with high-grade histological disease (grade 3 endometrioid, papillary serous, clear cell, and mixed tumor type) or cervical involvement were treated with total abdominal hysterectomy, bilateral salpingo-oophorectomy, and lymphadenectomy in the University Medical Center Groningen between 1999 and 2008. Patients who received adjuvant EBRT were included in this study. External beam radiation therapy on SmPF (includes only the central pelvis and proximal vagina) was applied in case of negative lymph nodes after adequate lymphadenectomy (≥10 lymph nodes removed at the bilateral obturator and external iliac nodal stations). In case of positive pelvic lymph nodes or inadequate lymphadenectomy, EBRT on StPF was given. External beam radiation therapy on EF was applied in case of common iliac and/or para-aortic lymph node metastases. Retrospectively, using the Common Terminology Criteria for Adverse Events v3.0, acute toxicity was scored during radiotherapy, whereas late toxicity was scored, from 3 months onward after treatment. RESULTS: Toxicity could be evaluated in 75 patients treated with SmPF (n = 33), StPF (n = 28), and EF EBRT (n = 14). Most patients with late adverse events had also reported toxicity during radiotherapy (71%). The most common late adverse events were gastrointestinal tract related, more frequently present in the StPF group (60.7%) compared to SmPF (33.3%; P = 0.032). In particular, nausea and anorexia were more frequent in the StPF group (32.1%) compared to the SmPF group (3.0%; P = 0.004), as well as ileus (14.3% vs 0%, P = 0.039, respectively). CONCLUSIONS: Treatment with adjuvant EBRT on SmPF results in less gastrointestinal late adverse events compared to treatment with EBRT on StPF in patients with surgically staged EC.

Prognostic role of indoleamine 2,3-dioxygenase in endometrial carcinoma.

OBJECTIVE: Indoleamine-2,3-dioxygenase (IDO) suppresses the function of T-lymphocytes and is an important immune escape mechanism for cancer. Therefore, it is to be expected that IDO influences prognosis of cancer patients. This study aimed to investigate the prognostic role of IDO expression in a large cohort of endometrial carcinoma (EC) patients. METHODS: A tissue microarray containing primary EC tissue of 355 patients treated in a single institution was used to evaluate IDO expression. Expression of IDO was associated with clinicopathological characteristics, survival and previously determined numbers of CD8(+) and Foxp3(+) T-lymphocytes. RESULTS: IDO(high) expression was associated with lower numbers of intratumoral CD8(+) T-lymphocytes (p=0.031). Next to well-known prognostic parameters, IDO(high) expression was independently associated with poor disease specific survival in the general cohort of EC patients (HR 2.62, 95% C.I. 1.48-4.66, p=0.001) and among patients with early stage EC (HR 3.06, 95% C.I. 1.10-8.54, p=0.032). CONCLUSION: Our results show that IDO expression is associated with poor survival. This provides evidence that further research into the use of IDO blocking agents in cancer treatment is valid where it might be a promising new therapeutic strategy.

Loss of HLA class I and mismatch repair protein expression in sporadic endometrioid endometrial carcinomas.

Tumor cells can escape from cytotoxic T-cell responses by downregulation of human leukocyte antigen (HLA) class I molecules expressed at the cell surface which has been associated with a deficient mismatch repair (MMR) system in colorectal carcinomas. Our study investigated the association between expression of MMR proteins and HLA class I in sporadic endometrioid endometrial carcinomas (EC). In a consecutively selected cohort of 486 EC patients, MMR proteins (MLH1, MSH2 and MSH6) and HLA class I (HLA-A, -B, -C or ß(2) m) were investigated by immunohistochemistry. Expression levels of MMR proteins and HLA class I were compared between low-grade and high-grade ECs. HLA class I expression was compared between tumors with loss (negative immunostaining of ≥1 MMR protein) and expression of MMR proteins. Associations between previously determined numbers of intratumoral CD8(+) T-lymphocytes and expression of MMR proteins and HLA class I and the influence on survival was determined. ECs with loss of MMR protein expression (33.5%) more frequently have loss of HLA-B/C (37.3%), compared to ECs with MMR protein expression (25.5%, p = 0.007). Patients with loss of MMR proteins have a worse disease-specific survival compared to patients with expression (p = 0.039). CD8(+) T-lymphocytes have a positive influence on disease-free and disease-specific survival in the total EC cohort but not in patients with loss of MMR protein expression. In conclusion, our results indicate that loss of MMR protein expression is related to selective downregulation of HLA class I which contributes to immune escape in EC with an abnormal MMR system.

Serum tryptophan and kynurenine concentrations as parameters for indoleamine 2,3-dioxygenase activity in patients with endometrial, ovarian, and vulvar cancer.

OBJECTIVE: Indoleamine 2,3-dioxygenase (IDO) suppresses the function of T-lymphocytes and is involved in immune escape of cancers. Indoleamine 2,3-dioxygenase catalyzes the initial rate-limiting step in the degradation of the essential amino acid tryptophan. In this study, we investigated cancer-induced IDO activity in sera of endometrial, ovarian, and vulvar cancer patients. METHODS: Concentrations of tryptophan and kynurenine were determined in pretreatment serum samples of patients with endometrial (n = 41), ovarian (n = 28), and vulvar cancer (n = 40) and compared to 19 healthy female controls. In serum of a subgroup of endometrial (n = 22), ovarian (n = 21), and vulvar (n = 21) cancer patients, tryptophan, kynurenine, and the kynurenine-to-tryptophan ratio (kyn/trp) were determined at different time points: preoperative, at clinical remission, and at the time of diagnosis of recurrent disease. Analyses were performed by an automated online solid-phase extraction-liquid chromatographic-tandem mass spectrometric method. Indoleamine 2,3-dioxygenase activity was estimated by calculating the kyn/trp ratio. RESULTS: Kynurenine concentrations and the kyn/trp ratio were higher in preoperative serum of endometrial, ovarian, and vulvar cancer patients compared to controls (all: P < 0.001). Preoperative serum of ovarian cancer patients contained higher kynurenine concentrations (median, 2.53 µM; interquartile range [IQR], 1.72-4.29 µM) and a higher kyn/trp ratio (median, 39.3 µmol/mmol; IQR, 26.5-61.7 µmol/mmol) compared to serum collected at clinical remission (median, 2.02 µM; IQR, 1.68-2.72 µM, P = 0.035; and median, 29.9 µmol/mmol; IQR, 23.4-38.9 µmol/mmol, P = 0.005, respectively). CONCLUSIONS: Patients with endometrial, ovarian, and vulvar cancer have increased tryptophan degradation compared to controls resulting in higher serum kynurenine concentrations and a higher kyn/trp ratio. Our results suggest that IDO-induced immune escape may play an important role in these gynecologic cancers.

Molecular markers and clinical behavior of uterine carcinosarcomas: focus on the epithelial tumor component.

Carcinosarcomas (malignant mixed Müllerian tumors) of the uterus are rare and aggressive malignancies consisting of an epithelial (carcinoma) and a mesenchymal (sarcoma) tumor component and are considered as metaplastic endometrial carcinomas. This study evaluated molecular characteristics and clinical behavior of uterine carcinosarcomas to improve treatment regimens in the future. Immunohistochemical expression of estrogen receptor-α and -ß, progesterone receptor-A and -B, MLH1, MSH2, MSH6, PTEN (phosphatase and tensin homolog deleted on chromosome 10), p53, ß-catenin and cyclin D1 was determined in 40 uterine carcinosarcomas. Immunostaining was compared between epithelial and mesenchymal tumor components. To determine the prognostic role of the epithelial component, clinicopathological data and survival were compared between patients with endometrioid and non-endometrioid epithelial tumor components. To determine prognosis of carcinosarcomas compared with high-risk endometrial carcinomas, clinicopathological characteristics and survival were compared between these patients. Hormone receptor expression occurred infrequently: estrogen receptor-α (8%) and -ß (32%), progesterone receptor-A (0%) and -B (23%), next to ß-catenin (4%) and cyclin D1 (7%). PTEN, MLH1, MSH2 and MSH6 mutations occurred in 39%, 33%, 22% and 21%, respectively (based on absent immunostaining). Overexpression of p53 was observed in 38%. Expression patterns of p53, MSH2 and MSH6 corresponded between epithelial and mesenchymal tumor components. In our cohort, the epithelial component caused the majority of metastases (72%) and vascular invasion (70%). Survival tended to be worse for patients with a non-endometrioid epithelial component compared with an endometrioid epithelial component (5-year survival: 26% and 55%, respectively). Survival was worse for patients with uterine carcinosarcomas compared with high-risk endometrial carcinomas (grade 3 endometrioid and non-endometrioid); 5-year survival rates: 42%, 77% and 57%, respectively. Our results support the monoclonal origin of uterine carcinosarcomas. The epithelial component determines prognosis by causing the majority of metastases and vascular invasion. To improve prognosis, treatment should focus on the epithelial tumor component of uterine carcinosarcomas.

The prognostic role of classical and nonclassical MHC class I expression in endometrial cancer.

The aim of this study was to investigate classical MHC class I and nonclassical MHC (human leukocyte antigen-G [HLA-G]) expression in a large cohort of patients with endometrial cancer, to determine the prognostic value of these cell surface markers and their relation with clinicopathological variables. Tissue microarrays containing epithelial endometrial carcinoma tissue from 554 patients were stained for classical and nonclassical MHC class I using the following monoclonal antibodies: 4H84 (anti-HLA-G), beta2-m (anti-beta-2-microglobulin) and HC-10 (MHC class I antigen heavy chain). Expression data were linked to known clinicopathological characteristics and survival. HLA-G upregulation and MHC class I downregulation in neoplastic cells was observed in 40% and 48%, respectively. Nonendometrioid tumor type, advanced stage disease (FIGO stage > or = II) and poorly or undifferentiated tumors were associated with MHC class I downregulation. Absence of HLA-G expression was independently associated with MHC class I downregulation. In univariate analysis, MHC class I downregulation was a predictor of worse disease-specific survival. Prognostic unfavorable tumor characteristics were correlated with downregulation of MHC class I expression in endometrial cancer cells. Furthermore, downregulated MHC class I has a negative impact on disease-specific survival, observed in a large cohort of patients with endometrial cancer. As there seems to be a relation between classical and nonclassical MHC class I molecules (HLA-G), further research is warranted to unravel this regulatory mechanism.

Aromatase, cyclooxygenase 2, HER-2/neu, and p53 as prognostic factors in endometrioid endometrial cancer.

The prognostic value of aromatase, cyclooxygenase 2 (COX-2), HER-2/neu, and p53 expression was determined in endometrioid endometrial cancer. Tissue microarrays were constructed comprising samples from 315 endometrioid endometrial cancer patients. Expression of aromatase, COX-2, HER-2/neu, and p53 was determined by immunostaining and related to classical clinicohistopathologic parameters, in addition to recurrence of disease and survival. Median follow-up time for all patients was 5.0 years. Patients were classified as Fédération Internationale de Gynécologie Obstétrique stage I (59.0%), stage II (17.1%), stage III (19.4%), and stage IV (4.1%). Sixty-five patients (20.6%) developed recurrent disease, and 38 (12.1%) died because of endometrial cancer. Aromatase, COX-2, HER-2/neu, and p53 expression was observed in 133 (42.2%), 107 (34.0%), 17 (5.4%), and 21 (6.7%) tumor cases, respectively. Aromatase expression in tumor cells was related to aromatase expression in stromal cells (P < 0.0001) and to HER-2/neu expression in tumor cells (P = 0.019). Aromatase expression in tumor as well as stromal cells was related to a low stage of disease (P = 0.02 and P = 0.001, respectively), whereas aromatase expression in stromal cells was also related to a low tumor grade (P = 0.021). P53 expression was related to a high stage and a high grade (P = 0.006 and P < 0.0001, respectively). In multivariate analysis, p53 overexpression was independently related to death because of the disease (P = 0.043; odds ratio 3.0; 95% confidence interval, 1.0-8.7). For COX-2, HER-2/neu, and aromatase, no relation with any other histopathologic parameter or survival was found. In conclusion, aromatase and p53 expression are related to tumor grade and stage of disease, whereas p53 is an independent prognostic factor in endometrioid endometrial cancer.

Prognostic significance of tumor-infiltrating T-lymphocytes in primary and metastatic lesions of advanced stage ovarian cancer.

PURPOSE: Ovarian cancer patients with intra-tumoral CD3(+) T-lymphocytes in primary tumor tissue have a better prognosis. This study aims to analyze the presence and relative influence of three important T-lymphocyte subsets, tumor-infiltrating CD8(+) cytotoxic T-lymphocytes (CTL), CD45R0(+) memory T-lymphocytes, and FoxP3(+) regulatory T-lymphocytes (Treg), in primary tumor tissue and omental metastases of patients with ovarian cancer. EXPERIMENTAL DESIGN: The number of CD8(+), CD45R0(+), and FoxP3(+) T-lymphocytes was determined by immunohistochemistry on a tissue micro array containing ovarian tumor tissue and/or omental metastases obtained at primary debulking surgery from 306 FIGO stage I-IV ovarian cancer patients. Immunohistochemistry data were correlated to clinicopathological parameters and survival data. RESULTS: High number of CD8(+) CTL and a high CD8(+)/FoxP3(+) ratio in ovarian-derived tumor tissue were associated with increased disease-specific survival and proved to be independent prognostic factors in multivariate analyses. In advanced stage patients, the presence of CD8(+) CTL, CD45R0(+) memory T-lymphocytes, FoxP3(+) Treg or a high CD8(+)/FoxP3(+) ratio in ovarian-derived tumor tissue was associated with an increased disease specific survival in univariate analysis, as was the presence of CD45R0(+) memory T-lymphocytes and FoxP3(+) Treg in omental metastases. Furthermore, in advanced stage patients CD8(+) cytotoxic and FoxP3(+) regulatory T-lymphocytes infiltrating ovarian-derived tumor tissue were independent predictors of increased prognosis. CONCLUSIONS: T-lymphocytes infiltrating primary and metastatic ovarian cancer sites are associated with improved prognosis. These associations are especially distinct in advanced stage patients, underlining the potential for immunotherapy as a broadly applicable therapeutic strategy.