Serum deoxyribonuclease and ribonuclease in pancreatic cancer and chronic pancreatitis

Serum ribonuclease (RNase) and deoxyribonuclease (DNase) were investigated in 18 control subjects, and in 22 patients with pancreatic cancer, 13 with chronic pancreatitis and 29 with extrapancreatic diseases in order to assess their clinical usefulness in pancreatic cancer diagnosis and to evaluate whether modifications were consensual and/or age-related. Increased DNase and RNase values were found not only in a notable proportion of pancreatic cancer, but also in chronic pancreatitis and extra-pancreatic diseases. Thus the clinical value of both enzymes in pancreatic cancer diagnosis is negligible. DNase does not seem to be strictly age-dependent, whereas serum RNase does. Elevated levels of the two enzymes, when present, were consensual, suggesting that factors involved in such an increase were partially common to both.     

TMSB4Y is a candidate tumor suppressor on the Y chromosome and is deleted in male breast cancer

Male breast cancer comprises less than 1% of breast cancer diagnoses. Although estrogen exposure has been causally linked to the development of female breast cancers, the etiology of male breast cancer is unclear. Here, we show via fluorescence in situ hybridization (FISH) and droplet digital PCR (ddPCR) that the Y chromosome was clonally lost at a frequency of ~16% (5/31) in two independent cohorts of male breast cancer patients. We also show somatic loss of the Y chromosome gene TMSB4Y in a male breast tumor, confirming prior reports of loss at this locus in male breast cancers. To further understand the function of TMSB4Y, we created inducible cell lines of TMSB4Y in the female human breast epithelial cell line MCF-10A. Expression of TMSB4Y resulted in aberrant cellular morphology and reduced cell proliferation, with a corresponding reduction in the fraction of metaphase cells. We further show that TMSB4Y interacts directly with β-actin, the main component of the actin cytoskeleton and a cell cycle modulator. Taken together, our results suggest that clonal loss of the Y chromosome may contribute to male breast carcinogenesis, and that the TMSB4Y gene has tumor suppressor properties.     

Loss of Y chromosome with retention of Y heterochromatin in a marker chromosome from a human melanoma

A single copy of a der 15 chromosome (m3) characterized by a C- and distamycin A-Dapi-positive region was observed in the -Y hyperploid karyotype of a primary human melanoma (Me 1402). The heterochromatic region was located pericentromerically, adjacent at one end to the NOR region of chromosome 15, and at the other to an unclassifiable chromosomal piece. We established that the C-positive block in the marker chromosome originated from Y heterochromatin by high-stringency in situ hybridization with a DNA probe for the 2.1 Hae III Y-specific repeat. Loss of the Y chromosome in tumors has been considered to be a secondary event associated with malignant evolution. It is significant that Me 1402 cells, which are highly malignant, lack the Y chromosome, but retain its heterochromatic portion in the rearranged m3 chromosome.     

Loss of miR-126 is crucial to pancreatic cancer progression

Evaluation of: Hamada S, Satoh K, Fujibuchi W et al. miR-126 acts as a tumor suppressor in pancreatic cancer cells via the regulation of ADAM9. Mol. Cancer Res. 10(1), 3-10 (2012). The tumor-suppressor miRNA 126 (miR-126) is downregulated in many tumors and has recently been placed at the heart of complex metastatic pathways. Hamada and colleagues have identified miR-126 as being downregulated in pancreatic ductal adenocarcinoma (PDAC) patient samples and cell lines. The protein ADAM9 has been implicated in the progression of various solid tumors including PDAC. ADAM9 is overexpressed in PDAC and also a direct target of miR-126. The miR-126/ADAM9 axis was subsequently established to control migration and invasion in PDAC, as well as reversal of epithelial-to-mesenchymal transition. miR-126 is also known to target other crucial oncogenes in PDAC such as KRAS and CRK. Replacing miR-126 in PDAC patients may be a novel strategy for preventing progression and metastasis.     

Loss of miR-126 is crucial to pancreatic cancer progression

Evaluation of: Hamada S, Satoh K, Fujibuchi W et al. miR-126 acts as a tumor suppressor in pancreatic cancer cells via the regulation of ADAM9. Mol. Cancer Res. 10(1), 3–10 (2012).

The tumor-suppressor miRNA 126 (miR-126) is downregulated in many tumors and has recently been placed at the heart of complex metastatic pathways. Hamada and colleagues have identified miR-126 as being downregulated in pancreatic ductal adenocarcinoma (PDAC) patient samples and cell lines. The protein ADAM9 has been implicated in the progression of various solid tumors including PDAC. ADAM9 is overexpressed in PDAC and also a direct target of miR-126. The miR-126/ADAM9 axis was subsequently established to control migration and invasion in PDAC, as well as reversal of epithelial-to-mesenchymal transition. miR-126 is also known to target other crucial oncogenes in PDAC such as KRAS and CRK. Replacing miR-126 in PDAC patients may be a novel strategy for preventing progression and metastasis.     

Loss of Fhit is frequent in stage I non-small cell lung cancer and in the lungs of chronic smokers.

Abnormalities of FHIT, a candidate tumor suppressor gene at 3p14.2, have been found frequently in multiple tumor types including non-small cell lung cancer (NSCLC). To investigate whether FHIT inactivation plays a role in early lung tumorigenesis, Fhit levels were determined by immunohistochemistry in tumors from 87 patients with stage I NSCLC and in 372 bronchial biopsy specimens from 86 chronic smokers without evidence of malignancy. We found that 49% of NSCLC specimens demonstrated significantly decreased staining or lack of staining for Fhit. However, Fhit expression status was not significantly associated with disease-free survival or overall survival. Analysis of a subset of 76 specimens on which microsatellite analysis at the FHIT locus was performed did not show a strong association between loss of heterozygosity at FHIT and Fhit expression, suggesting the presence of complex mechanisms of Fhit inactivation. Of 372 bronchial biopsies from chronic smokers, 86 biopsies (23%) exhibited decreased Fhit expression or lack of Fhit expression. In 37 of 86 (43%) subjects, decreased Fhit expression or lack of expression was observed in at least one biopsy site. Loss of Fhit expression was significantly higher in bronchial metaplastic lesions (23 of 49 lesions, 47%) than in histologically normal bronchial epithelium (63 of 323 specimens, 20%; P < 0.001). Smokers with a metaplasia index of > 15% had a higher frequency of loss of Fhit expression than those with a metaplasia index of < or = 15% (P = 0.015). Interestingly, current smokers had a higher rate of loss of Fhit expression than former smokers (P = 0.02). Our data indicate that Fhit expression is significantly reduced in a substantial number of early-stage NSCLC and preneoplastic lesions in chronic smokers. The association between cigarette smoking and Fhit expression suggests a role for FHIT in the initiation of smoking-related lung tumorigenesis.     

Cripto,a member of the epidermal growth factor family,is over-expressed in human pancreatic cancer and chronic pancreatitis

Cripto is a 188 amino-acid protein containing a central segment that shares amino-acid sequence homology with epidermal growth factor (EGF) and transforming growth factor alpha (TGF-α). The EGF receptor, EGF and TGF-α are expressed in the normal human pancreas, and are over-expressed in human pancreatic cancer. Therefore, in the present study we sought to determine whether cripto is found in the normal human pancreas and whether its expression is altered in pancreatic cancer. Because chronic pancreatitis (CP) is associated with interstitial fibrosis similar to that observed in pancreatic cancer, we also examined cripto expression in pancreatic tissues from patients with CP. In the normal pancreas, cripto immunoreactivity was found at moderate levels in most ductal cells and was present very faintly in a rare acinar cell. In 26 of 58 pancreatic cancers, cripto immunoreactivity was present in many cancer cells. Its presence was associated with advanced tumor stage, but not with shorter post-operative survival. Cripto was also present in acinar and ductal cells adjacent to the cancer cells, and in many ductal and atrophic acinar cells in the CP samples. Northern blot analysis revealed a marked increase in cripto mRNA levels in the cancer and CP samples. By densitometry, there was a 11 - and 4-fold increase in cripto mRNA levels in pancreatic cancer and CP respectively. Southern blot analysis did not reveal an increase in gene copies encoding cripto either in cancer or in CP. These findings indicate that cripto expression may contribute to disease progression in pancreatic cancer, and implicate cripto in the histopathological alterations that occur in the pancreas both in cancer and in CP.     

Loss of heterozygosity at chromosome 9q22-31 is a frequent and early event in ovarian tumors

Loss of heterozygosity (LOH) studies in ovarian tumors, have highlighted the chromosomal regions at 9q22-31 and 9q32-34 as being potentially important in tumor development. We have investigated LOH at 9q22-31 in 85 patients with epithelial ovarian cancer, 15 with non-epithelial tumors and 16 with benign disease. Varying patterns of LOH were observed across the markers used between different tumors, the most common (71%) being interstitial discontinuous losses. LOH was frequent, and was detected at equally high levels in malignant (71%) and benign tumors (70%). LOH occurred in epithelial invasive tumors, borderline tumors, fibromas and dermoid tumors. In malignant epithelial tumors LOH at 9q22-31 was not significantly associated with patient clinical and pathological parameters; however, survival was 29 months at the 50th centile survival, in those women whose tumors displayed LOH compared with 60 months in women whose tumors retained heterozygosity. LOH at 9q22-31 was significantly associated with LOH at the p53 locus (p=0.02) and the ovarian suppressor locus at 3p21 (p=0.05). We conclude that the chromosome region at 9q22-31, flanked by the microsatellite markers D9S1796 and D9S53, is a frequent and early event in ovarian tumorigenesis. With the of extent of discontinuous LOH, high density deletion mapping of this region using LOH as a strategy to identify candidate genes may be problematic. However with the completion of the human genome sequencing project several candidate genes are identified.     

Differential diagnosis of chronic pancreatitis and pancreatic cancer in brush cytology specimens

Discrimination between chronic pancreatitis and pancreatic carcinoma can be complicated, particularly in brush cytology specimens. Previous studies have shown that the oxygen insensitivity of the histochemical reaction to detect glucose-6-phosphate dehydrogenase activity based on neotetrazolium reduction can be used for discriminating malignant cells from nonmalignant cells. In the present study, we investigated the value of the assay for differential diagnosis between the two pancreatic diseases. Oxygen insensitivity in ductal epithelial cells in normal human pancreas, chronic pancreatitis, and pancreatic carcinoma was determined by quantitative image analysis in sections of biopsies and in brush cytology preparations. In sections, the reaction in the absence of oxygen was a proper reflection of glucose-6-phosphate dehydrogenase activity, whereas in the presence of oxygen only malignant cells showed a significant reaction. Of 39 brush cytology specimens, diagnosis of all 11 cases of pancreatitis and 28 cases of cancer with the oxygen insensitivity test were in agreement with independent measures of chronic pancreatitis and cancer. The oxygen insensitivity test is a simple and valuable tool in addition to conventional pathology for differential diagnosis between pancreatitis and pancreatic cancer, both in biopsies and in brush cytology specimens.     

Presence of pancreatic intraepithelial neoplasia‐3 in a background of chronic pancreatitis in pancreatic cancer patients

The clinical significance of pancreatic intraepithelial neoplasia (PanIN) lesions in non‐neoplastic pancreata of pancreatic ductal adenocarcinoma (PDAC) patients remains controversial. As chronic inflammation has been recently demonstrated to promote dissemination of in situ precancerous lesions, we investigated the prognostic significance of PanINs associated with chronic pancreatitis (CP) in PDAC patients. This retrospective study analyzed 125 curatively resected PDAC specimens for the presence of PanIN and CP. Univariate and multivariate analyses were performed to identify significant predictive factors for poor disease‐free survival (DFS) and overall survival (OS). Immunohistochemical staining for E‐cadherin and S100A4, markers of epithelial‐mesenchymal transition, was performed on resected specimens containing PanIN‐3 lesions. CP was observed in 27.2% (34/125) and PanIN‐3 in 25.6% (32/125) of specimens. In the presence of CP, PanIN‐3 was significantly associated with decreased survival (DFS: 4.3 vs 15.5 months, P = 0.021; OS: 16.3 vs 30.9 months, P = 0.004). PanIN‐3 was not a prognostic factor in the absence of CP. The presence of both PanIN‐3 and CP was associated with a reduced survival compared to the other cases, in both univariate (DFS: P = 0.039; OS: P = 0.023) and multivariate (DFS: P = 0.020; OS: P = 0.076) analyses. Furthermore, E‐cadherin loss and S100A4 expression were more frequently observed in PanIN‐3 lesions of CP specimens than in those of non‐CP specimens, although not statistically significant. PanIN‐3 in association with CP is a significant prognostic factor for decreased survival in PDAC patients, suggesting that chronic inflammation may accelerate the progression of preinvasive high‐grade PanIN.     

Loss and reduction of FUS1 protein expression is a frequent phenomenon in the pathogenesis of lung cancer.

PURPOSE: FUS1, a novel tumor-suppressor gene located in the chromosome 3p21.3 region, may play an important role in lung cancer development. Currently, FUS1-expressing nanoparticles have been developed for treating patients with lung cancer. However, the expression of Fus1 protein has not been examined in a large series of lung cancers and their sequential preneoplastic lesions. EXPERIMENTAL DESIGN: Using tissue microarrays, we examined Fus1 immunohistochemical expression in 281 non-small cell lung carcinoma (NSCLC) and 22 small cell lung carcinoma tissue specimens and correlated the findings with patients' clinicopathologic features. To investigate the expression of Fus1 in the early sequential pathogenesis of NSCLC, we studied Fus1 expression in 211 histologically normal and mildly abnormal bronchial epithelia, and 118 bronchial and alveolar preneoplastic lesions obtained from patients with lung cancer. RESULTS: Loss and reduction of expression was detected in 82% of NSCLCs and 100% of small cell lung carcinomas. In NSCLCs, loss of Fus1 immunohistochemical expression was associated with significantly worse overall survival. Bronchial squamous metaplastic and dysplastic lesions expressed significantly lower levels of Fus1 compared with normal (P = 0.014 and 0.047, respectively) and hyperplastic (P = 0.013 and 0.028, respectively) epithelia. CONCLUSIONS: Our findings show a high frequency of Fus1 protein loss and reduction of expression in lung cancer, and suggests that this reduction may play an important role in the early pathogenesis of lung squamous cell carcinoma. These findings support the concept that FUS1 gene and Fus1 protein abnormalities could be used to develop new strategies for molecular cancer therapy for a significant subset of lung tumors.     

Evidence of chromosome instability in chronic pancreatitis

In the current study we analyzed chromosome instability on peripheral blood lymphocytes cultured from 7 untreated patients with chronic pancreatitis (CP) by assessing telomeric associations (TAS), chromosome aberrations (CA) and sister chromatid exchanges (SCE). Seven healthy individuals were also analyzed. Mean frequencies of TAS were significantly higher in CP patients (X +/- SE: 11.00 +/- 2.37) compared to controls (1.00 +/- 0.30) (p<0.001). Chromosomes preferentially involved in TAS were: 9, 20, 16 and 21, being the most affected arms: 9p, 20q, 16p, 9q and 21q. All these terminal bands were coincident with cancer breakpoints (p<0.03), two of them (40%) were specifically associated to pancreatic carcinoma rearrangements. Three bands (60%) were coincident with oncogene location. The mean frequency of CA was significantly higher in patients (3.88 +/- 0.80) compared to controls (0.63 +/- 0.49) (p<0.001). Chromosomes 1, 2 and 13 were the most damaged. No specifically affected breakpoints were found. SCE analysis showed higher levels in patients (8.33 +/- 0.70) than in controls (6.62 +/- 0.34) (p<0.025), but no differences were observed in cell cycle kinetics. Our results clearly indicate that CP patients exhibit chromosome instability, showing the presence of an unstable genome that could be related to the cancer development observed in this disease.     

基于CT影像组学对局灶性自身免 疫性胰腺炎与胰腺导管腺 癌鉴别诊断的价值北大核心

目的:探讨基于CT影像组学对局灶性自身免疫性胰腺炎(focal-autoimmune pancreatitis,f-AIP)与胰腺导管腺癌(pancreatic ductal adenocarcinoma,PDAC)鉴别诊断的价值。方法:回顾性收集f-AIP患者73例,PDAC患者74例的CT影像资料。首先,由两位放射科医生对所有f-AIP与PDAC患者的CT征象进行分析,确定病变的类型,得出人工诊断的准确性。其次,将显示病灶的所有层面的三期薄层图像以DICOM格式上传至达尔文医准科研平台,在病灶的中心层面及其上下层面勾画感兴趣区(ROI),提取组学特征,运用方差选择法、最小绝对收缩算法(LASSO)对组学特征进行筛选和降维,建立支持向量机(SVM)组学模型,并建立CT征象联合组学特征的联合模型,最后使用受试者工作特性曲线(ROC曲线)对各模型进行评价。结果:放射科医生诊断的准确性为0.83,在平扫、动脉期及静脉期中筛选出5、11、6个组学特征建立模型,联合CT征象组建联合模型,分别筛选出14、8、8个特征。在组学分析中,基于平扫、动脉期及静脉期所建组学模型,静脉期组学模型敏感度0.96、特异度0.90、准确度0.93及AUC值0.97均高于其他两期;在联合模型中,各期联合模型均较同期单纯组学模型的AUC值、敏感度及准确度高,仅平扫单纯模型的特异度高于联合组学模型。结论:在f-AIP与PDAC的鉴别中,人工诊断具有一定的诊断能力;构建多期相SVM单纯组学模型,动脉期及静脉期组学模型诊断效能均高于人工诊断,且静脉期诊断价值最高;采用CT征象联合组学特征建立联合模型,诊断效能明显优于单一模型,进一步提高了对f-AIP与PDAC的鉴别能力。     

Loss of BNIP3 expression is a late event in pancreatic cancer contributing to chemoresistance and worsened prognosis

Altered expression of apoptosis-regulating genes plays an important role in the aggressive growth behavior and chemoresistance of pancreatic ductal adenocarcinoma. In the present study, the hypoxia-inducible proapoptotic gene, BNIP3, was analysed in terms of expression, effect on patient survival, and chemo-responsiveness in pancreatic cancer cell lines. cDNA microarray, real-time light cycler quantitative polymerase chain reaction, laser-capture microdissection, and immunohistochemistry analyses were used to evaluate BNIP3 expression in normal and diseased pancreatic specimens. Modulation of BNIP3 expression was achieved using specific siRNA molecules. The effect of chemotherapeutic agents on pancreatic cancer cells was assessed utilizing 3-(4,5-methylthiazol-2-yl)-2,5-diphenyl-tetrazolium-bromide assays. BNIP3 mRNA levels were 3.0- and 6.3-fold lower in chronic pancreatitis and pancreatic cancer compared to the normal pancreas, respectively. Microdissection analysis confirmed the reduction of BNIP3 expression in pancreatic cancer cells compared to normal duct cells. By immunohistochemistry, BNIP3 was predominantly expressed in the acinar cells of the normal and diseased pancreas. Interestingly, while BNIP3 was undetectable in the cancer cells of 59% of the cases, 75-100% of PanIN2/3 lesions displayed BNIP3 immunoreactivity. Loss of BNIP3 expression correlated with poorer survival of patients (8 vs 14 months for BNIP3 negative vs positive tumors). Hypoxia induced BNIP3 expression in four out of eight pancreatic cancer cell lines, while it was absent under normoxic and hypoxic conditions in the remaining four. Downregulation of BNIP3 resulted in increased resistance to 5-fluoro-uracil and gemcitabine. In conclusion, loss of BNIP3 expression occurs late in pancreatic cancer, contributes to resistance to chemotherapy, and correlates with a worsened prognosis.     

Loss of Betaig-h3 protein is frequent in primary lung carcinoma and related to tumorigenic phenotype in lung cancer cells

Betaig-h3 as a secreted protein induced by transforming growth factor-beta has been suggested to modulate cell adhesion and tumor formation. Although we have previously shown that downregulation of Betaig-h3 gene is involved in the cellular transformation of human bronchial epithelial cells induced by radiation, its regulation in primary human lung cancers is not clearly understood. In this study, Betaig-h3 expression was studied in 130 primary human lung carcinomas by immunohistochemistry. Betaig-h3 protein was absent or reduced by more than two-fold in 45 of 130 primary lung carcinomas relative to normal lung tissues examined. Recovery of Betaig-h3 expression in H522 lung cancer cells lacking endogenous Betaig-h3 protein significantly suppressed their in vitro cellular growth and in vivo tumorigenicity. In addition, parental H522 cancer cells are resistant to the etoposide induced apoptosis compared with normal human bronchial epithelial cells. However, recovery of Betaig-h3 expression in H522 cancer cells results in significantly higher sensitivity to apoptotic induction than parental tumor cells. IGFBP3 is upregulated in Betaigh3-transfected H522 cells that may mediate the apoptotic sensitivity and antitumor function of Betaig-h3 gene. These observations demonstrate that downregulation of Betaig-h3 gene is a frequent event and related to the tumor progression in human lung cancer.