Reduced IQGAP2 Promotes Bladder Cancer through Regulation of MAPK/ERK Pathway and Cytokines.

The progression of non-muscle-invasive bladder cancer (NMIBC) to muscle-invasive bladder cancer (MIBC) is a major challenge in urologic oncology. However, understanding of the molecular processes remains limited. The dysregulation of IQGAP2 is becoming increasingly evident in most tumor entities, and it plays a role in multiple oncogenic pathways, so we evaluated the role of IQGAP2 in bladder cancer. IQGAP2 was downregulated in tumors compared with normal urothelium tissues and cells. IQGAP2 effectively attenuated bladder cancer cell growth independently from apoptosis. Reduced IQGAP2 promoted EMT in bladder cancer cells via activation of the MAPK/ERK pathway. In addition, IQGAP2 might influence key cellular processes, such as proliferation and metastasis, through the regulation of cytokines. In conclusion, we suggest that IQGAP2 plays a tumor-suppressing role in bladder cancer, possibly via inhibiting the MAPK/ERK pathway and reducing cytokines.

Wnt/ß-Catenin Signalling and Its Cofactor BCL9L Have an Oncogenic Effect in Bladder Cancer Cells.

Bladder cancer (BC) is characterised by a high recurrence and progression rate. However, the molecular mechanisms of BC progression remain poorly understood. BCL9L, a coactivator of ß-catenin was mutated in the 5' and 3' untranslated regions (UTRs). We assessed the influence of UTRs mutations on BCL9L, and the role of BCL9L and Wnt/ß-catenin signalling in BC cells. UTR mutations were analysed by a luciferase reporter. BCL9L protein was assessed by immunohistochemistry in BC tissues. Cell proliferation was examined by crystal violet staining and by the spheroid model. Moreover, migration and invasion were analysed in real-time using the xCelligence RTCA system. The A > T mutation at 3' UTR of BCL9L reduces the luciferase reporter mRNA expression and activity. BCL9L is predominantly increased in dysplastic urothelial cells and muscle-invasive BC. Knockdown of BCL9L and inhibition of Wnt/ß-catenin signalling significantly repress the proliferation, migration and invasion of Cal29 and T24. In addition, BCL9L knockdown reduces mRNA level of Wnt/ß-catenin target genes in Cal29 but not in T24 cells. BCL9L and Wnt/ß-catenin signalling play an oncogenic role in bladder cancer cells and seems to be associated with BC progression. Nevertheless, the involvement of BCL9L in Wnt/ß-catenin signalling is cell-line specific.

SLC35F2, a Transporter Sporadically Mutated in the Untranslated Region, Promotes Growth, Migration, and Invasion of Bladder Cancer Cells.

Bladder cancer is a very heterogeneous disease and the molecular mechanisms of carcinogenesis and progression are insufficiently investigated. From the DNA sequencing analysis of matched non-muscle-invasive bladder cancer (NMIBC) and muscle-invasive bladder cancer (MIBC) samples from eight patients, we identified the tumour-associated gene SLC35F2 to be mutated in the 5' and 3' untranslated region (UTR). One mutation in 3'UTR increased the luciferase activity reporter, suggesting its influence on the protein expression of SLC35F2. The mRNA level of SLC35F2 was increased in MIBC compared with NMIBC. Furthermore, in immunohistochemical staining, we observed a strong intensity of SLC35F2 in single tumour cells and in the border cells of solid tumour areas with an atypical accumulation around the nucleus, especially in the MIBC. This suggests that SLC35F2 might be highly expressed in aggressive and invasive tumour cells. Moreover, knockdown of SLC35F2 repressed the growth of bladder cancer cells in the monolayer and spheroid model and suppressed migration and invasion of bladder cancer cells. In conclusion, we suggest that SLC35F2 is involved in bladder cancer progression and might provide a new therapeutic approach, for example, by the anti-cancer drug YM155, a cargo of the SLC35F2 transporter.

CAMK2N1/RUNX3 methylation is an independent prognostic biomarker for progression-free and overall survival of platinum-sensitive epithelial ovarian cancer patients.

BACKGROUND: To date, no predictive or prognostic molecular biomarkers except BRCA mutations are clinically established for epithelial ovarian cancer (EOC) despite being the deadliest gynecological malignancy. Aim of this biomarker study was the analysis of DNA methylation biomarkers for their prognostic value independent from clinical variables in a heterogeneous cohort of 203 EOC patients from two university medical centers. RESULTS: The marker combination CAMK2N1/RUNX3 exhibited a significant prognostic value for progression-free (PFS) and overall survival (OS) of sporadic platinum-sensitive EOC (n = 188) both in univariate Kaplan-Meier (LogRank p < 0.05) and multivariate Cox regression analysis (p < 0.05; hazard ratio HR = 1.587). KRT86 methylation showed a prognostic value only in univariate analysis because of an association with FIGO staging (Fisher's exact test p < 0.01). Thus, it may represent a marker for EOC staging. Dichotomous prognostic values were observed for KATNAL2 methylation depending on BRCA aberrations. KATNAL2 methylation exhibited a negative prognostic value for PFS in sporadic EOC patients without BRCA1 methylation (HR 1.591, p = 0.012) but positive prognostic value in sporadic EOC with BRCA1 methylation (HR 0.332, p = 0.04) or BRCA-mutated EOC (HR 0.620, n.s.). CONCLUSION: The retrospective analysis of 188 sporadic platinum-sensitive EOC proved an independent prognostic value of the methylation marker combination CAMK2N1/RUNX3 for PFS and OS. If validated prospectively this combination may identify EOC patients with worse prognosis after standard therapy potentially benefiting from intensive follow-up, maintenance therapies or inclusion in therapeutic studies. The dichotomous prognostic value of KATNAL2 should be validated in larger sample sets of EOC.

Gain-of-function mutation in SCN11A causes itch and affects neurogenic inflammation and muscle function in Scn11a+/L799P mice.

Mutations in the genes encoding for voltage-gated sodium channels cause profound sensory disturbances and other symptoms dependent on the distribution of a particular channel subtype in different organs. Humans with the gain-of-function mutation p.Leu811Pro in SCN11A (encoding for the voltage-gated Nav1.9 channel) exhibit congenital insensitivity to pain, pruritus, self-inflicted injuries, slow healing wounds, muscle weakness, Charcot-like arthropathies, and intestinal dysmotility. As already shown, knock-in mice (Scn11a+/L799P) carrying the orthologous mutation p.Leu799Pro replicate reduced pain sensitivity and show frequent tissue lesions. In the present study we explored whether Scn11a+/L799P mice develop also pruritus, muscle weakness, and changes in gastrointestinal transit time. Furthermore, we analyzed morphological and functional differences in nerves, skeletal muscle, joints and small intestine from Scn11a+/L799P and Scn11a+/+ wild type mice. Compared to Scn11a+/+ mice, Scn11a+/L799P mice showed enhanced scratching bouts before skin lesions developed, indicating pruritus. Scn11a+/L799P mice exhibited reduced grip strength, but no disturbances in motor coordination. Skeletal muscle fiber types and joint architecture were unaltered in Scn11a+/L799P mice. Their gastrointestinal transit time was unaltered. The small intestine from Scn11a+/L799P showed a small shift towards less frequent peristaltic movements. Similar proportions of lumbar dorsal root ganglion neurons from Scn11a+/L799P and Scn11a+/+ mice were calcitonin gene-related peptide (CGRP-) positive, but isolated sciatic nerves from Scn11a+/L799P mice exhibited a significant reduction of the capsaicin-evoked release of CGRP indicating reduced neurogenic inflammation. These data indicate important Nav1.9 channel functions in several organs in both humans and mice. They support the pathophysiological relevance of increased basal activity of Nav1.9 channels for sensory abnormalities (pain and itch) and suggest resulting malfunctions of the motor system and of the gastrointestinal tract. Scn11a+/L799P mice are suitable to investigate the role of Nav1.9, and to explore the pathophysiological changes and mechanisms which develop as a consequence of Nav1.9 hyperactivity.

High Detection Rate for Non-Muscle-Invasive Bladder Cancer Using an Approved DNA Methylation Signature Test.

INTRODUCTION: Cystoscopy and transurethral resection are the current reference standard tests to diagnose and histologically confirm non-muscle-invasive bladder cancer (NMIBC). In other tumor entities (ie, colon carcinoma, cervical cancer), DNA methylation markers have been approved as diagnostic tests with high diagnostic power. In our case-control study, we used an approved molecular cervical cancer diagnostics test that includes 6 DNA methylation markers (GynTect) for the detection of bladder cancer. PATIENTS AND METHODS: We included samples from 40 patients with bladder cancer and 34 control subjects. In a pilot study, we analyzed DNA methylation in 38 tumor tissues and 4 healthy ureters using methylation-specific polymerase chain reaction. Subsequently, we determined the sensitivity and specificity of the GynTect for the detection of bladder cancer in urine sediments from 40 patients with bladder cancer and 30 control subjects with benign prostatic hyperplasia or urolithiasis. RESULTS: The markers showed very different methylation rates in the NMIBC tissues, ranging from 2.6% to 78.9%. No methylation of any of the markers was detectable in the healthy ureters. Using the urine sediments from the patients with cancer and control subjects, we found surprisingly high sensitivity and specificity for the GynTect assay (60% and 96.7%, respectively). The application of different algorithms for evaluation of the markers included in GynTect resulted in a sensitivity of ≤ 90% and specificity of ≤ 100%. CONCLUSION: The GynTect assay, originally designed for cervical cancer diagnostics, showed unexpectedly high diagnostic accuracy for bladder cancer detection. The inclusion of additional methylation markers might allow for the development of a suitable diagnostic marker set based on the GynTect test for NMIBC diagnostics.

Combined assessment of 3q26 amplification and promoter methylation in patients with high grade cervical lesions show age specific differences.

A considerable proportion of high grade cervical intraepithelial lesions (CIN2/3) are known to resolve on their own especially among young women. However, since reliable prognostic markers are still lacking, the diagnosis "CIN3" is still an indication for surgery which may result in overtreatment. It is conceivable that a combination of different, ideally independent molecular markers may provide more reliable results. In the present cross-sectional study two established triage markers, 3q26 amplification and a methylation signature, were evaluated in an age-dependent manner. The patient cohort comprised 60 patients with histologically confirmed CIN2/3 in two equally sized age groups (<30 years, ≥30 years). Cervical scrapes were analyzed by interphase fluorescence in situ hybridization for 3q26 amplification and methylation specific PCR (GynTect®) for six different genome regions. Both assays showed a significantly different pattern of test outcome independent of age (P = .001). Moreover, the combination of both assays differed significantly for double positive and double negative cases when comparing the two age groups: In patients <30 years there were clearly less cases with positive methylation signature and amplification of 3q26 as in women ≥30 years (23% vs 63%, Bonferroni adjusted P = .016). Of particular interest is the finding that double negative results were exclusive for the young age group (0% vs 27%, Bonferroni adjusted P = .020). Since regression of CIN2/3 characteristically occurs among young women it is tempting to speculate that a double negative test result could be prognostic for regression of CIN2/3. This will have to be investigated further in a prospective longitudinal intervention study.

4-miRNA Score Predicts the Individual Metastatic Risk of Renal Cell Carcinoma Patients.

BACKGROUND: In order to improve individual prognostication as well as stratification for adjuvant therapy in patients with clinically localized clear cell renal cell carcinoma (ccRCC), reliable prognostic biomarkers are urgently needed. In this study, microRNAs (miRNAs) have emerged as promising candidates. We investigated whether a combination of differently expressed miRNAs in primary tumors can predict the individual metastatic risk. METHODS: Using two prospectively collected biobanks of academic centers, 108 ccRCCs were selected, including 57 from patients with metastatic disease at diagnosis or during follow-up and 51 without evidence of metastases. Fourteen previously identified candidate miRNAs were tested in 20 representative formalin-fixed and paraffin embedded samples in order to select the best discriminators between metastatic and nonmetastatic ccRCC. These miRNAs were approved in 108 tumor samples. We evaluated the association of altered miRNA expression with the metastatic potential of tumors using quantitative polymerase chain reaction. A prognostic 4-miRNA model has been established using a random forest classifier. Cox regression analyses were performed for correlation of the miRNA model and clinicopathological parameters to metastasis-free and overall survival. RESULTS: Nine miRNAs indicated significant expression alterations in the small cohort. These miRNAs were validated in the whole cohort. The established 4-miRNA score (miR-30a-3p/-30c-5p/-139-5p/-144-5p) has been identified as a superior predictor for metastasis-free survival (hazard ratio 12.402; p = 7.0E-05) and overall survival (p = 1.1E-04) compared with clinicopathological parameters, and likewise in the Leibovich score subgroups. CONCLUSIONS: We identified a 4-miRNA model that was found to be superior to clinicopathological parameters in accurately predicting individual metastatic risk and can support patient selection for risk-stratified follow-up and adjuvant therapy studies.

Laparoscopic radical hysterectomy with transvaginal closure of vaginal cuff - a multicenter analysis.

OBJECTIVE: Laparoscopic/robotic radical hysterectomy has been historically considered oncologically equivalent to open radical hysterectomy for patients with early cervical cancer. However, a recent prospective randomized trial (Laparoscopic Approach to Cervical Cancer, LACC) has demonstrated significant inferiority of the minimally invasive approach. The aim of this study is to evaluate the oncologic outcomes of combined laparoscopic-vaginal radical hysterectomy. METHODS: Between August 1994 and December 2018, patients with invasive cervical cancer were treated using minimally-invasive surgery at the Universities of Jena, Charité Berlin (Campus CCM and CBF) and Cologne and Asklepios Clinic Hamburg. 389 patients with inclusion criteria identical to the LACC trial were identified. In contrast to the laparoscopic/robotic technique used in the LACC trial, all patients in our cohort underwent a combined transvaginal-laparoscopic approach without the use of any uterine manipulator. RESULTS: A total of 1952 consecutive patients with cervical cancer were included in the analysis. Initial International Federation of Gynecology and Obstetrics (FIGO) stage was IA1 lymphovascular space invasion (LVSI+), IA2 and IB1/IIA1 in 32 (8%), 43 (11%), and 314 (81%) patients, respectively, and histology was squamous cell in 263 (68%), adenocarcinoma in 117 (30%), and adenosquamous in 9 (2%) patients. Lymphovascular invasion was confirmed in 106 (27%) patients. The median number of lymph nodes was 24 (range 2-86). Lymph nodes were tumor-free in 379 (97%) patients. Following radical hysterectomy, 71 (18%) patients underwent adjuvant chemoradiation or radiation. After a median follow-up of 99 (range 1-288) months, the 3-, 4.5-, and 10-year disease-free survival rates were 96.8%, 95.8%, and 93.1 %, and the 3-, 4.5-, and 10-year overall survival rates were 98.5%, 97.8%, and 95.8%, respectively. Recurrence location was loco-regional in 50% of cases with recurrence (n=10). Interestingly, 9/20 recurrences occurred more than 39 months after surgery. CONCLUSION: The combined laparoscopic-vaginal technique for radical hysterectomy with avoidance of spillage and manipulation of tumor cells provides excellent oncologic outcome for patients with early cervical cancer. Our retrospective data suggest that laparoscopic-vaginal surgery may be oncologically safe and should be validated in further randomized trials.

A three-gene methylation marker panel for the nodal metastatic risk assessment of muscle-invasive bladder cancer.

PURPOSE: In this study, we aimed to identify a DNA methylation pattern suitable for prognosis assessment of muscle-invasive bladder cancer and to investigate metastasis-associated processes regulated by DNA methylation. METHODS: Genome-wide methylation analysis was performed on 23 muscle-invasive bladder tumors by microarray analysis. Validation was performed by the qAMP technique in two different patient cohorts (n = 32 and n = 100). mRNA expression was analyzed in 12 samples. Protein expression was determined using tissue microarrays of 291 patients. Bladder cancer cell lines T24 and 253JB-V were used for functional analyses. RESULTS: Microarray analyses revealed KISS1R, SEPT9 and CSAD as putative biomarkers with hypermethylation in node-positive tumors. The combination of the three genes predicted the metastatic risk with sensitivity of 73% and specificity of 71% in cohort 1, and sensitivity of 82% and specificity of 54% in cohort 2. mRNA expression differences were detected for KISS1R (p = 0.04). Protein expression of KISS1R was significantly reduced (p < 0.001). Knockdown of SEPT9v3 resulted in increased cell migration by 28% (p = 0.04) and increased invasion by 22% (p = 0.004). KISS1R overexpression resulted in decreased cell migration (25%, p = 0.1). CONCLUSIONS: We identified a methylation marker panel suitable to differentiate between patients with positive and negative lymph nodes at time of cystectomy. This enables a risk assessment for patients who potentially benefit from extended lymph node resection as well as from neoadjuvant chemotherapy and could improve the survival rates. Furthermore, we examined the impact of putative markers on tumor behavior. Hence, KISS1R and SEPT9 could represent a starting point for the development of novel therapy approaches.

A Promising New Approach for the Treatment of Inflammatory Pain: Transfer of Stem Cell-Derived Tyrosine Hydroxylase-Positive Cells.

OBJECTIVES: The appearance of endogenous tyrosine hydroxylase-positive cells (TH+ cells) in collagen-induced arthritis was associated with an anti-inflammatory effect. Here we investigated putative anti-inflammatory and antinociceptive effects of the transfer of induced, bone marrow stem cell-derived TH+ cells (iTH+ cells) on murine antigen-induced arthritis (AIA). METHODS: Bone marrow-derived stem cells were differentiated into iTH+ cells. These cells were transferred to mice immunized against methylated bovine serum albumin (mBSA) 2 days before AIA was induced by injection of mBSA into one knee joint. In AIA control mice and iTH+-treated mice the severity of AIA, pain-related behavior, humoral and cellular responses, and the invasion of macrophages into the dorsal root ganglia were assessed. RESULTS: The intravenous transfer of iTH+ cells before AIA induction did not cause a sustained suppression of AIA severity but significantly reduced inflammation-evoked pain-related behavior. The iTH+ cells used for transfer exhibited enormous production of interleukin-4. A major difference between AIA control mice and iTH+-treated AIA mice was a massive invasion of the dorsal root ganglia by iNOS-negative, arginine 1-positive macrophages corresponding to an M2 phenotype. The differences in other cellular and humoral immune parameters such as release of cytokines from stimulated lymphocytes between AIA control mice and iTH+-treated mice were small. CONCLUSIONS: The transfer of iTH+ cells may cause a long-lasting reduction of arthritis-induced pain even if it does not ameliorate inflammation. The invasion of M2 macrophages into the dorsal root ganglia is likely to be an important mechanism of antinociception.

Induction of chronic destructive arthritis in SCID mice by arthritogenic fibroblast-like synoviocytes derived from mice with antigen-induced arthritis.

BACKGROUND: Fibroblast-like synoviocytes (FLSs) from patients with rheumatoid arthritis (RA) are autonomously activated to maintain inflammation and joint destruction in co-transplantation models. To elucidate inducing mechanisms involved in this altered behavior, the arthritogenic potential of FLSs from murine antigen-induced arthritis (AIA) were investigated in a transfer model. METHODS: FLSs were isolated, expanded in vitro, and transferred into knee joint cavities of severe combined immunodeficient (SCID) mice. Their arthritogenic capacity was assessed by monitoring joint swelling and evaluation of histological parameters 70 to 100 days after transfer. RESULTS: FLSs from AIA mice were able to transfer arthritis into recipient SCID mice. FLS transfer induced a chronic arthritis with recruitment of inflammatory cells and marked cartilage destruction. Long-lasting inflammation was not required for imprinting of arthritogenicity in FLSs since cells isolated from acute arthritic joints were fully competent to transfer arthritis. We also observed arthritogenic potential in FLSs isolated from contralateral non-arthritic joints in our monoarticular arthritis model. CONCLUSIONS: We show that the transformation of FLSs into arthritogenic cells occurs early in arthritis development. This challenges current hypotheses on the role of these cells in arthritis pathogenesis and opens up the way for further mechanistic studies.

Interleukin-17A is involved in mechanical hyperalgesia but not in the severity of murine antigen-induced arthritis.

Interleukin-17A (IL-17A) is considered an important pro-inflammatory cytokine but its importance in joint diseases such as rheumatoid arthritis (RA) is unclear. It has also been reported that IL-17A may induce pain but it is unclear whether pro-inflammatory and pro-nociceptive effects are linked. Here we studied in wild type (WT) and IL-17A knockout (IL-17AKO) mice inflammation and hyperalgesia in antigen-induced arthritis (AIA). We found that the severity and time course of AIA were indistinguishable in WT and IL-17AKO mice. Furthermore, the reduction of inflammation by sympathectomy, usually observed in WT mice, was preserved in IL-17AKO mice. Both findings suggest that IL-17A is redundant in AIA pathology. However, in the course of AIA IL-17AKO mice showed less mechanical hyperalgesia than WT mice indicating that IL-17A contributes to pain even if it is not crucial for arthritis pathology. In support for a role of IL-17A and other members of the IL-17 family in the generation of pain we found that sensory neurones in the dorsal root ganglia (DRG) express all IL-17 receptor subtypes. Furthermore, in isolated DRG neurones most IL-17 isoforms increased tetrodotoxin- (TTX-) resistant sodium currents which indicate a role of IL-17 members in inflammation-evoked sensitization of sensory nociceptive neurones.

Tribbles 2 mediates cisplatin sensitivity and DNA damage response in epithelial ovarian cancer.

Aim was to identify methylated genes with functional involvement in cisplatin-resistance development of epithelial ovarian cancer (EOC). Genome-wide analyses of hypermethylated CpG-islands in resistant cell lines in combination with qRT-PCR analyses were used to identify epigenetically silenced genes. EOC-Type-II tumors were analyzed for gene methylation and expression and TCGA data were interrogated in-silico. Experiments revealed 37 commonly hypermethylated genes in resistant cells of which Tribbles 2 (TRIB2) showed the most pronounced downregulation on mRNA level and was characterized further. TRIB2 showed a reactivation after 5'-Aza-Cytidine treatment in resistant cells but a cisplatin-dependent, prominent upregulation on mRNA level in sensitive cells, only. Re-expression in resistant A2780 cells increased the sensitivity to cisplatin and other DNA-damaging agents, but not taxanes. Contrary, knockdown of TRIB2 increased resistance to cisplatin in sensitive cells. TRIB2 was involved in the induction of a cisplatin-dependent cell cycle arrest and apoptosis by influencing p21 and survivin expression. An increased Pt-DNA-adduct formation in TRIB2 re-expressing cells did not translate in higher levels of dsDNA damage (yH2AX-foci). Thus, TRIB2 is potentially involved in the signal transduction from nucleotide excision repair of intrastrand cross links. Importantly, patient stratification of two homogenous cohorts of EOC-Type-II patients from Jena (n = 38) and the TCGA (n = 149) by TRIB2 mRNA expression consistently revealed a significantly decreased PFS for patients with low TRIB2 levels (log-rank p < 0.05). Tumors from resistant patients expressed the lowest levels of TRIB2. Downregulation of TRIB2 contributes to platin-resistance and TRIB2 expression should be validated as prognostic and predictive marker for EOC.

In situ foamable, degradable polyurethane as biomaterial for soft tissue repair.

Degradable foams which can be inserted endoscopically as liquid or pasty mixtures into soft tissue defects possess a promising potential for the surgical treatment of such defects. The defects can be sealed under in situ foaming and simultaneous material expansion. We developed an in situ foamable (l-lactide-co-ε-caprolactone)-based, star-shaped prepolymer by ring opening polymerization of l-lactide and ε-caprolactone in the presence of meso-erythritol as starter. By conversion of the terminal hydroxyl groups of the formed oligoester with lysine diisocyanate ethyl ester (LDI) an isocyanate-endcapped, reactive prepolymer has been received. Foaming can be initiated by addition of 1,4-diazabicyclo[2,2,2]octane (DABCO), water, LDI and DMSO. By varying the composition of these additives, the foaming and curing time could be varied within a clinically acceptable range. A porosity of approximately 90%, and an average tensile strength of 0.3MPa with elongations of 90% were determined for the foams. In vitro cytotoxicity on cured foams was assayed on 3T3 fibroblasts and demonstrated an excellent cytocompatibility. This was also confirmed in an in vivo study using an established rat model, where prefabricated foams and in situ hardening material were inserted into subdermal skin incisions in parallel. The feature of chronic inflammation was only weakly developed in both groups and slightly more pronounced and persisted for longer time in the group of in situ foamed material. In both groups the foreign materials were vascularized, degraded and substituted by connective tissue. The results encourage to proceed with trials where the materials are used to fill more heavily loaded defects.

Prognostic Value of "Prepectoral Edema" in MR-mammography.

BACKGROUND/AIM: Previous studies have indicated a highly significant correlation between invasive tumors and accompanying prepectoral edema (PE) in MR-mammography (MRM). The aim of the present study was to identify prognostic factors associated with PE as a diagnostic sign. MATERIALS AND METHODS: A total of 1,109 consecutive MRM exams were included in this study. Exclusion criteria were previous operation, biopsy, intervention, chemotherapy, hormone replacement therapy (HRT) or previous mastitis. One hundred and sixty-two patients with 180 lesions were evaluated and histologically correlated. Diagnostic evaluations were performed by four experienced radiologists in consensus. RESULTS: One hundred and eighty lesions included 104 malignant lesions (93 invasive and 11 non-invasive) and 76 benign lesions. PE was detected significantly more frequently in presence of lymphangiosis carcinomatosa ((53.8%; 14/26) vs. (9.8%; 4/41)) (p<0,000). PE significantly correlates with positive axillary nodal status ((19.4% (12/62) vs. 44.4% (12/27)) (p=0.020), as well as pathologic enhancement of the pectoral muscle (5.4% (5/93) vs. 22.7% (20/88)) (p=0,015). PE significantly correlates with higher tumor grading (G3) (33.9% vs. 13.9%) (p<0.05). There was no significant difference in a positive vs. negative estrogen (p=0.681) and progesterone (p=0.751) and/or human epidermal growth factor receptor-2 (HER-2) (p=0.726) receptor status accompanied by PE. CONCLUSION: The presence of PE may be a strong prognostic indicator for lymphatic spread and the cancerous infiltration of lymph nodes. It is also associated with the infiltration of the pectoral muscle, as well as high tumor grading. There is no correlation between prepectoral edema and positive tumor receptor status.

Gene expression analysis combined with functional genomics approach identifies ITIH5 as tumor suppressor gene in cervical carcinogenesis.

Progression from human papillomavirus-induced premalignant cervical intraepithelial neoplasia (CIN) to cervical cancer (CC) is driven by genetic and epigenetic events. Our microarray-based expression study has previously shown that inter-α-trypsin-inhibitor heavy chain 5 (ITIH5) mRNA levels in CCs were significantly lower than in high-grade precursor lesions (CIN3s). Therefore, we aimed to analyze in depth ITIH5 expression during cervical carcinogenesis in biopsy material and cell culture. Moreover, functional analyses were performed by ectopic expression of ITIH5 in different cell lines. We were able to confirm the validity of our microarray differential expression data by qPCR, demonstrating a clear ITIH5 downregulation in CC as compared with CIN2/3 or normal cervix. ITIH5 protein loss, evaluated by immunohistochemistry, was evident in 81% of CCs, whereas ITIH5 showed weak to moderate cytoplasmic staining in 91% of CIN2/3 cases. In addition, ITIH5 was strongly reduced or absent in seven CC cell lines and in three immortalized keratinocyte cell lines. Moreover, ITIH5 mRNA loss was associated with ITIH5 promoter methylation. ITIH5 expression could be restored in CC cell lines by pharmacological induction of DNA demethylation and histone acetylation. Functionally, ITIH5 overexpression significantly suppressed proliferation of SW756 cells and further resulted in a significant reduction of colony formation and cell migration in both CaSki and SW756 tumor models, but had no effect on invasion. Remarkably, ITIH5 overexpression did not influence the phenotype of HeLa cells. Taken together, ITIH5 gene silencing is a frequent event during disease progression, thereby providing evidence for a tumor suppressive role in cervical carcinogenesis.

Interleukin-6-dependent influence of nociceptive sensory neurons on antigen-induced arthritis.

INTRODUCTION: Interleukin-6 (IL-6) is an important mediator of inflammation. In addition to cells involved in inflammation, sensory nociceptive neurons express the IL-6 signal-transducer glycoprotein 130 (gp130). These neurons are not only involved in pain generation but also produce neurogenic inflammation by release of neuropeptides such as calcitonin gene-related peptide (CGRP). Whether IL-6 activation of sensory neurons contributes to the induction of inflammation is unknown. This study explored whether the action of IL-6 on sensory neurons plays a role in the generation of neurogenic inflammation and arthritis induction. METHODS: In SNS-gp130(-/-) mice lacking gp130 selectively in sensory neurons and appropriate control littermates (SNS-gp130(flox/flox)), we induced antigen-induced arthritis (AIA), and assessed swelling, histopathological arthritis scores, pain scores, expression of CGRP in sensory neurons, serum concentrations of CGRP and cytokines, and the cytokine release from single cell suspensions from lymph nodes and spleens. In wild-type mice CGRP release was determined during development of AIA and, in cultured sensory neurons, upon IL-6 stimulation. RESULTS: Compared to SNS-gp130(flox/flox) mice SNS-gp130(-/-) mice showed significantly weaker initial swelling, reduced serum concentrations of CGRP, IL-6, and IL-2, no inflammation-evoked upregulation of CGRP in sensory neurons, but similar histopathological arthritis scores during AIA. During the initial swelling phase of AIA, CGRP was significantly increased in the serum, knee and spleen. In vitro, IL-6 augmented the release of CGRP from cultured sensory neurons. Upon antigen-specific restimulation lymphocytes from SNS-gp130(-/-) mice released more interleukin-17 and interferon-γ than lymphocytes from SNS-gp130(flox/flox) mice. In naive lymphocytes from SNS-gp130(flox/flox) and SNS-gp130(-/-) mice CGRP reduced the release of IL-2 (a cytokine which inhibits the release of interleukin-17 and interferon-γ). CONCLUSIONS: IL-6 signaling in sensory neurons plays a role in the expression of arthritis. Selective deletion of gp130 signaling in sensory neurons reduces the swelling of the joint (most likely by reducing neurogenic inflammation) but increases some proinflammatory systemic cellular responses such as the release of interleukin-17 and interferon-γ from lymphocytes upon antigen-specific restimulation. Thus IL-6 signaling in sensory neurons is not only involved in pain generation but also in the coordination of the inflammatory response.

Means to increase the therapeutic efficiency of magnetic heating of tumors.

The treatment of tumors via hyperthermia has gained increased attention in the last years. Among the different modalities available so far, magnetic hyperthermia has the particular advantage of offering the possibility of depositing the heating source directly into the tumor. In this study, we summarized the present knowledge we gained on how to improve the therapeutic efficiency of magnetic hyperthermia using magnetic nanoparticles (MNPs), with particular consideration of the intratumoral infiltration of the magnetic material. We found that (1) MNPs will be mainly immobilized at the tumor area and that this aspect has to be considered when estimating the heating potential of MNPs, (2) the intratumoral distribution patterns via slow infiltration might well be modulated by specific MNP coating and magnetic targeting, (3) imaging of the nanoparticle depositions within the tumor might allow to correct the distribution pattern via multiple applications, (4) multiple therapeutic sessions are feasible because MNPs are not delivered from the tumor site during the heating process, (5) the utilization of MNPs that internalize into cells will favor the production of intracellular heating spots rather than extracellular ones, (6) utilization of MNPs functionalized with chemotherapeutic agents will allow us to exploit the additive effects of both therapeutic modalities, and (7) distinct cytopathological and histopathological alterations in target tissues are induced as a result of magnetic hyperthermia. However, the accumulation at the tumor via intravenous application remains a matter of challenge.

Stress-resistant Translation of Cathepsin L mRNA in Breast Cancer Progression.

The cysteine protease cathepsin L (CTSL) is often thought to act as a tumor promoter by enhancing tumor progression and metastasis. This goes along with increased CTSL activity in various tumor entities; however, the mechanisms leading to high CTSL levels are incompletely understood. With the help of the polyoma middle T oncogene driven breast cancer mouse model expressing a human CTSL genomic transgene, we show that CTSL indeed promotes breast cancer metastasis to the lung. During tumor formation and progression high expression levels of CTSL are maintained by enduring translation of CTSL mRNA. Interestingly, human breast cancer specimens expressed the same pattern of 5' untranslated region (UTR) splice variants as the transgenic mice and the human cancer cell line MDA-MB 321. By polyribosome profiling of tumor tissues and human breast cancer cells, we observe an intrinsic resistance of CTSL to stress-induced shutdown of translation. This ability can be attributed to all 5' UTR variants of CTSL and is not dependent on a previously described internal ribosomal entry site motif. In conclusion, we provide in vivo functional evidence for overexpressed CTSL as a promoter of lung metastasis, whereas high CTSL levels are maintained during tumor progression due to stress-resistant mRNA translation.