Expression In Biopsy Specimens Research Articles

Aromatase CYP19A1, progesterone receptor PGR and estrogen receptor ESR1 gene expression in biopsy specimens of endometrioid heterotopia and endometrial tissue by reverse transcription PCR

Hypothesis/aims of study. Endometriosis is one of the most pressing problems of gynecology. Clarifying the expression of the estrogen receptor (ESR1) and the progesterone receptor (PGR) genes and polymorphisms in the aromatase (CYP19A1) gene in endometriosis will expand the understanding of the pathogenesis of the disease and the causes of resistance to its therapy. The objective of this study was to conduct a comparative analysis of mRNA expression of PGR, ESR1 and CYP19A1 genes in paired samples of the eutopic endometrium and peritoneal endometrioid lesions in order to search for predictive markers of response to hormonal therapy. In the future, this may allow personalizing the selection of hormonal preparations for the treatment of endometriosis.
 Study design, materials and methods. Reverse transcription real-time PCR made it possible to evaluate CYP19A1, PGR and ESR1 gene expression levels in studied tissue samples from 22 patients with endometriosis and 9 women in the comparison group.
 Results. Quantitative analysis revealed a high heterogeneity in the expression level of the studied genes, in both the endometrium and endometrioid lesions from patients with endometriosis. In the endometrium of patients in the comparison group, the heterogeneity of the expression level was observed only for the ESR1 gene.
 Conclusion. Our findings suggest a high variability in CYP19A1, ESR1 and PGR gene expression levels in the endometrium and peritoneal foci in patients with endometriosis. This information indicates the need for an individual approach to prescribing targeted therapy, since it is obvious that the effect of treatment will depend primarily on the availability of a therapeutic target in a particular patient. The absence of a typical expression pattern for each of the genes in patients with endometriosis indicates the heterogeneity of the disease and the need to develop a molecular classification of this common pathology.

Prognostic significance of lactate dehydrogenase-5 expression in patients with prostate cancer treated with androgen deprivation therapy and androgen receptor axis-targeted agents.

300 Background: Evidence suggests that hypoxic zones are prevalent in prostate cancer (PC). Recent clinical data showed that the expression of lactate dehydrogenase-5 (LDH5), a tumor hypoxia marker, affected the outcome of PC treated with radiation therapy. However, the impact of its expression on hormone therapy remains unclear. We investigated the predictive value of LDH5 expression in biopsy specimens from patients with PC treated with hormone therapy. Methods: Baseline data were available from 50 patients with PC treated with both androgen deprivation therapy (ADT) and novel androgen receptor axis-targeted agents (ARATs). LDH5 expression in the biopsy specimens was analyzed using immunohistochemical staining. Patients were stratified according to LDH5 expression in needle biopsy specimens. The relationships between LDH5 expression and baseline data at diagnosis, time to emergence of castration resistance, prostate-specific antigen (PSA) progression, and overall survival from induction of ADT and ARATs were examined. Results: Biopsy specimens from 19 patients highly expressed LDH5. High LDH5 expression was significantly associated with high Gleason scores (≥8) (p = 0.03), high clinical T stage (p = 0.008), and the likelihood of having at least one lymph node (p = 0.019) and metastatic lesion (p = 0.024). In a multivariate logistic regression model, LDH5 expression was significantly associated with high clinical T stage (p = 0.038), the presence of lymph node metastasis (p = 0.013), and high neutrophil-to-lymphocyte ratio (p = 0.016). Multivariate Cox regression analysis showed that high LDH5 expression was a significant predictor of time to emergence of castration resistance (hazard ratio [HR] 2.45, 95% confidence interval [CI] 1.12–5.36, p = 0.025) and overall survival (HR 9.3, 95% CI 2.17–39.8, p = 0.003). No significant association was observed between LDH5 expression and time to PSA progression from the induction of ARATs in univariate Cox regression analysis (p = 0.28). Conclusions: The results of this study suggest that LDH5 expression is a predictor of treatment outcomes in patients with PC treated with ADT.

Abstract 914: COX-2 expression is associated with chemoresistance through cancer stemness property in head and neck squamous cell carcinoma

Abstract Introduction: COX-2 is one of the two isoforms of cyclooxygenase, an enzyme that catalyzes the conversion of arachidonic acid to prostaglandins. COX-2 is considered to be associated with progression in various cancers, and its expression has been reported to have an impact on poor prognosis in head and neck squamous cell carcinomas (HNSCCs). Furthermore, COX-2 expression has been shown to be associated with resistance to anticancer drugs. However, the precise mechanism of COX-2 for chemoresistance in HNSCC is not fully elucidated. The aim of the present study was to investigate the impact of COX-2 on cancer stem cell property, and to reveal its effect on chemoresistance by in vitro and clinicopathological assays. Methods: We examined the chemoresistance to docetaxel with or without COX-2 inhibitor (celecoxib) in HNSCC cell lines (FaDu, Detroit512) by MTS assays. To evaluate the association of COX-2 expression with stemness property, we assessed alteration of expression levels of the genes related to cancer stem cell (CSC) property after exposure to celecoxib by quantitative real-time PCR. We also performed a sphere formation assay using ultra low attachment dishes and microscopic imaging. Moreover, we analyzed immunohistochemical expressions of COX-2 and clinicopathological factors by using matched samples of pre-treatment biopsy and surgical specimens from hypopharyngeal carcinoma patients who had undergone trans-oral tumor resection with preoperative induction chemotherapy including docetaxel. Results: Cell survival rate under exposure to docetaxel was decreased by addition of celecoxib. COX-2 inhibitor downregulated the expression of Oct3/4 and Nanog. Sphere formation was inhibited by co-culture with COX-2 inhibitor, especially in FaDu cells. Immunohistochemical study in biopsy specimens revealed a negative correlation between COX-2 expression in biopsy specimens and pathological effect of induction chemotherapy in surgical specimens. Conclusion: The present study suggests that COX-2 expression is associated with cancer stemness property, and inhibition of COX-2 has a possibility to enhance chemosensitivity in HNSCCs. Furthermore, initial COX-2 expression in biopsy specimens may predict the effect of chemotherapy in HNSCCs. These results indicate that COX-2 can be an attractive target for the treatment of HNSCC. Citation Format: Shin Saito, Hiroyuki Ozawa, Yorihisa Imanishi, Mariko Sekimizu, Yoshihiro Watanabe, Fumihiro Ito, Yuichi Ikari, Nana Nakahara, Kaori Kameyama, Kaoru Ogawa. COX-2 expression is associated with chemoresistance through cancer stemness property in head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 914.

Rogaratinib treatment of patients with advanced urothelial carcinomas prescreened for tumor FGFR mRNA expression.

494 Background: Activation of FGFR signaling is involved in a variety of malignancies including advanced urothelial cancer (UC). Rogaratinib is an oral pan-FGFR kinase inhibitor. We report here the results from a phase I expansion cohort in UC patients prescreened for FGFR1-3 mRNA expression levels and activating mutations. (NCT01976741) Methods: Patients with advanced urothelial carcinomas were selected based on high FGFR1-3 mRNA expression in biopsy specimens. Selected patients were treated with rogaratinib 800mg twice daily until tumour progression, untolerable toxicity, or withdrawal. Tumor response was assessed by RECIST, v1.1. Adverse events were reported using CTCAE v4.03 criteria. Results: A total of 219 UC patients were prescreened for FGFR1-3 mRNA expression levels and FGFR3 activating mutations, with 99 samples (45%) found to be FGFR-positive. Of those, 87% of samples were positive for FGFR3 mRNA, 5% for FGFR1 mRNA and 8% were double FGFR mRNA-positive (FGFR1/2, 1/3 or 2/3). Frequency of FGFR3 activating mutations in UC samples was 7%, all of which also had high FGFR3 mRNA. Fifty two patients (median prior line of treatment 2) started treatment and 51 were evaluable for response. Rogaratinib was generally well tolerated and AEs manageable with dose modification. The most common AEs were diarrhea (49%) and hyperphosphatemia (49%). Objective response rate (ORR) was 24% (12/51; all PRs) and disease control rate (DCR) was 73% (37/51). Eleven of 12 pts with a PR were positive for FGFR3 mRNA, 5 of whom also had FGFR3 mutations, and one patient was positive for FGFR1 mRNA. Ten FGFR-positive UC patients had prior immuno-oncology (I/O) treatment, 9 of whom had progressive disease as best response. For these 10 patients the ORR was 30% and the DCR 80%. Conclusions: Selection of pts for treatment with rogaratinib based on FGFR mRNA expression levels was feasible and identified drug-sensitive patients with and without underlying DNA alterations. Rogaratinib had a favorable safety profile and showed promising anti-tumor activity in UC patients. Responses and disease stabilization were observed with rogoratinib in UC pts refractory to prior I/O treatment. Clinical trial information: NCT01976741.

Differential Expression of RAGE, EGFR and Ki-67 in Primary Tumors and Lymph Node Deposits of Breast Carcinoma

Background:Breast cancer is a complex disease that results from the inheritance of a number of susceptible genes. Intensive search wok was conducted world-wide on molecular bases of breast cancer in order to achieve the best therapeutic modalities; however, breast cancer still remains a challengeable task. It is very important to determine if the biological parameters in metastatic regional lymph nodes are similar to that in the primary breast cancer because therapy is indicated for patients with synchronous metastatic regional lymph nodes of breast cancer. Difference in therapeutic response in cases of breast cancer may be assumed partially to variability in the biological behavior of tumor tissue in primary breast cancer and lymph node metastasis.Aim:Our aim is to evaluate any variability in the expression of three types of tissue markers in both the primary breast tumors and corresponding axillary lymph nodes in order to expect the targeted therapeutic effect on both sites.Material and Methods:Three markers from different categories; RAGE, EGFR and Ki-67 were immunohistochemicalyl studied for their expression in biopsy specimens from primary breast tumors and their corresponding axillary lymph nodes.Results:There was a statistically significant difference in the expression of these markers between benign and malignant breast lesions. Although we found some differences in the expression of the three studied markers between primary breast cancer and corresponding axillary lymph nodes, yet these variations were mostly not statistically significant.Conclusion:Our findings support the validity of anti-RAGE and anti-EGFR therapy for treatment of both primary and nodal metastatic breast cancer in immunopositive cases.

IMP3 expression in biopsy specimens as a diagnostic biomarker for colorectal cancer

No single biological marker is used in routine diagnosis of colorectal cancer (CRC) in endoscopic biopsies. IMP3 is a good independent prognostic biomarker for CRC. However, the expression of IMP3 in hyperplastic polyp (HP) and adenoma has not yet been studied. Moreover, no studies have established the diagnostic value of IMP3 in biopsies. This study aims to assess IMP3 expression in HP, adenoma, and CRC in resection specimens and to investigate its value in diagnosis of CRC in biopsies. A total of 1328 specimens (633 of polypectomy, 395 surgical resections, 300 biopsies) were retrospectively analyzed. IMP3 expression was observed in 0 of 197 (0%) normal tissues, 0 of 130 (0%) HPs, 14 of 504 (2.8%) adenomas, and 139 of 197 (70.6%) CRCs. IMP3 was found to be overexpressed in CRC compared with adenoma (P<.001). Among the 300 biopsies, 56 were diagnosed as adenoma, and 244 were CRCs. Of the 56 adenoma cases, 22 (39.3%) were confirmed, whereas 34 (60.7%) were diagnosed as CRC in resection specimens. All 244 CRC biopsies were confirmed by resection specimens. IMP3-positive expression was observed in 204 of 300 (68.0%) biopsies, including in 22 of 56 (39.3%) adenomas and 182 of 244 (74.6%) CRCs. All IMP3-positive expressions in the biopsies were finally diagnosed as CRC. Our findings demonstrated that IMP3 is a reliable marker for the diagnosis of CRC in endoscopic biopsies.

Calcium and 1,25-dihydroxyvitamin D3 modulate genes of immune and inflammatory pathways in the human colon: a human crossover trial1–3

A high dietary calcium intake with adequate vitamin D status has been linked to lower colorectal cancer risk, but the mechanisms of these effects are poorly understood. The objective of this study was to elucidate the effects of a Western-style diet (WD) and supplemental calcium and/or 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] on the colorectal mucosa. We conducted 2 crossover trials to define molecular pathways in the human colorectum altered by 1) a 4-wk WD supplemented with and without 2 g calcium carbonate/d and 2) a 4-wk WD supplemented with 1,25(OH)2D3 (0.5 μg/d) with or without 2 g calcium carbonate/d. The primary study endpoint was genome-wide gene expression in biopsy specimens of the rectosigmoid colonic mucosa. Serum and urinary calcium concentrations were also measured. Changes in urinary calcium accurately reflected calcium consumption. The WD induced modest upregulation of genes involved in inflammatory pathways, including interferon signaling, and calcium supplementation reversed these toward baseline. In contrast, supplementation of the WD with 1,25(OH)2D3 induced striking upregulation of genes involved in inflammation, immune response, extracellular matrix, and cell adhesion. Calcium supplementation largely abrogated these changes. Supplementing 1,25(OH)2D3 to a WD markedly upregulated genes in immune response and inflammation pathways, which were largely reversed by calcium supplementation. This study provides clinical trial evidence of global gene expression changes occurring in the human colorectum in response to calcium and 1,25(OH)2D3 intervention. One action of 1,25(OH)2D3 is to upregulate adaptive immunity. Calcium appears to modulate this effect, pointing to its biological interaction in the mucosa. This trial was registered at clinicaltrials.gov as NCT00298545 Trial protocol is available at http://clinicalstudies.rucares.org (protocol numbers PHO475 and PHO554).

Significance of MUC1 expression in biopsy specimens of submucosal invasive gastric carcinoma: The association with lymph node metastasis.

Mucin 1, cell surface associated (MUC1) is a tumor-associated glycoprotein that has been reported to have an important role in lymphatic invasion and metastasis. The present study aimed to investigate the significance of MUC1 expression in endoscopic biopsy specimens of submucosal invasive gastric carcinomas and the association with lymph node metastasis. The clinicopathological features of 144 cases of surgically resected submucosal invasive gastric carcinomas and their paired endoscopic biopsy specimens were reviewed. Immunohistochemical staining for MUC1 was performed for the 144 endoscopic biopsy specimens. Positive MUC1 expression was identified in 70 (49%) cases. In addition, univariate analysis revealed that MUC1 expression was significantly associated with the presence of poorly-differentiated (P=0.001) and poorly-cohesive (P=0.015) carcinoma cells, undifferentiated type by Japanese classification (P<0.001), diffuse type of Lauren classification (P<0.001) and lymph node metastasis (P=0.024). By multivariate analysis, diffuse type of Lauren classification (P<0.001) and lymph node metastasis (P=0.035) were identified as independent factors for MUC1 expression. Furthermore, MUC1 expression (P=0.007), tumor size (P=0.018) and lymphatic invasion (P<0.001) were demonstrated to be independent factors for lymph node metastasis under multivariate analysis. In conclusion, the results of the present study indicated that positive MUC1 expression in endoscopic biopsy specimens may be a predictive factor of lymph node metastasis in submucosal invasive gastric carcinoma.

The Use of Transcriptional Profiling to Improve Personalized Diagnosis and Management of Cutaneous T-cell Lymphoma (CTCL).

Although many patients with mycosis fungoides presenting with stage I disease enjoy an indolent disease course and normal life expectancy, about 15% to 20% of them progress to higher stages and most ultimately succumb to their disease. Currently, it is not possible to predict which patients will progress and which patients will have a stable disease. Previously, we conducted microarray analyses with RT-PCR validation of gene expression in biopsy specimens from 60 patients with stage I-IV cutaneous T-cell lymphoma (CTCL), identified three distinct clusters based upon transcription profile, and correlated our molecular findings with 6 years of clinical follow-up. We test by RT-PCR within our prediction model the expression of about 240 genes that were previously reported to play an important role in CTCL carcinogenesis. We further extend the clinical follow-up of our patients to 11 years. We compare the expression of selected genes between mycosis fungoides/Sézary syndrome and benign inflammatory dermatoses that often mimic this cancer. Our findings demonstrate that 52 of the about 240 genes can be classified into cluster 1-3 expression patterns and such expression is consistent with their suggested biologic roles. Moreover, we determined that 17 genes (CCL18, CCL26, FYB, T3JAM, MMP12, LEF1, LCK, ITK, GNLY, IL2RA, IL26, IL22, CCR4, GTSF1, SYCP1, STAT5A, and TOX) are able to both identify patients who are at risk of progression and also distinguish mycosis fungoides/Sézary syndrome from benign mimickers. This study, combined with other gene expression analyses, prepares the foundation for the development of personalized molecular approach toward diagnosis and treatment of CTCL.

Correlation of Aurora-A expression with the effect of chemoradiation therapy on esophageal squamous cell carcinoma.

BackgroundChemoradiation therapy (CRT) is one of the most useful treatments for esophageal squamous cell carcinoma (ESCC). However, because some patients respond well to CRT and others do not, it is important to be able to predict response to CRT before beginning treatment by using markers. Aurora-A encodes a cell cycle regulated serine/threonine kinase that has essential functions in centrosome maturation and chromosome segregation. In this study, we investigated the relationship between the expression of Aurora-A and the response to CRT in patients with ESCC.MethodsWe immunohistochemically investigated the expression of Aurora-A in biopsy specimens of untreated primary tumors of 78 patients with ESCC and determined the relationship between Aurora-A levels and patient responses to CRT, which consisted of 5-fluorouracil plus cisplatin and 40 Gy of radiation.ResultsTumors were judged as Aurora-A positive when more than 10% of the cancer cells displayed a distinct positive nuclear anti-Aurora-A immunoreaction by immunohistochemical evaluation. The tumors of 46 of 78 patients (58.9%) displayed positive expression of Aurora-A. In terms of clinical response the percentage of patients showing complete response (CR), incomplete response/stable disease of primary lesion (IR/SD), and progressive disease (PD) was 19.2, 69.2, and 11.5%, respectively. In terms of histological response the tumor grade of the 41 patients who underwent surgery was as follows: grade 1, 48.8%; grade 2, 29.2%; grade 3, 22.0%. CRT was effective for patients who had Aurora-A (+) tumors (clinically: P = 0.0003, histologically: P = 0.036).ConclusionsOur results suggest that Aurora-A expression in biopsy specimens of primary tumors is associated with CRT efficacy in patients with ESCC. Assessment of Aurora-A expression in biopsy specimens maybe useful for regarding the potential utility of CRT therapy for patients with ESCC before treatment.

Toll-like receptor 3 signaling contributes to the expression of a neutrophil chemoattractant, CXCL1 in human mesangial cells.

Mesangial proinflammatory chemokine/cytokine expressions via innate immunity play a pivotal role in the pathogenesis of glomerulonephritis. CXCL1/GROα is a strong neutrophil chemoattractant cytokine and reportedly plays an important role in regional inflammatory reactions. However, detailed signaling of mesangial CXCL1 expression induced by viral or "pseudoviral" immunity remains to be determined. We treated normal human mesangial cells (MCs) in culture with polyinosinic-polycytidylic acid (poly IC), an authentic double-stranded RNA, and analyzed the expression of CXCL1 by reverse transcription-polymerase chain reaction (RT-PCR), real-time quantitative RT-PCR and enzyme-linked immunosorbent assay. To elucidate the poly IC-induced signaling pathway for CXCL1 expression, we subjected the cells to RNA interference against Toll-like receptor (TLR) 3, retinoic acid-inducible gene-I (RIG-I), melanoma differentiation-associated gene 5 (MDA5), interferon (IFN)-β, nuclear factor (NF)-κB p65 and IFN regulatory factor (IRF) 3. We also conducted an immunofluorescence study to examine mesangial CXCL1 expression in biopsy specimens from patients with lupus nephritis (LN) and IgA nephropathy (IgAN). We found that activation of TLR3 signaling could induce the expression of CXCL1 in MCs. NF-κB, IRF3 and IFN-β, but neither RIG-I nor MDA5, were found to be involved in mesangial CXCL1 expression in this setting. Induction of CXCL1 by poly IC was inhibited by pretreatment of cells with dexamethasone. Intense glomerular CXCL1 expression was observed in biopsy specimens from patients with LN, whereas only a trace staining occurred in specimens from patients with IgAN. TLR3 signaling also contributes to the CXCL1 expression in MCs. These observations further support the implication of viral and "pseudoviral" immunity in the pathogenesis of inflammatory renal diseases, especially in LN.

Multidrug resistance-associated protein 3 confers resistance to chemoradiotherapy for rectal cancer by regulating reactive oxygen species and caspase-3-dependent apoptotic pathway

This study aimed to clarify the role of multidrug resistance-associated protein 3 (MRP3) in resistance to neoadjuvant chemoradiotherapy and long-term prognosis of advanced rectal cancer. Immunohistochemistry was used to measure MRP3 expression in biopsy specimens of 144 stage II–III rectal cancer patients who received preoperative chemoradiotherapy. The effect of MRP3 expression on short-term pathological response and postoperative long-term prognosis were assessed using the Cox proportional hazards model. Short interfering RNAs targeting MRP3 were synthesized and used to transfect human colorectal carcinoma cell lines. The effect of MRP3 down-regulation on cell proliferation and apoptosis in response to 5-fluorouracil and/or irradiation were examined in vitro and in xenograft mouse models, respectively. The content of intracellular reactive oxygen species and the activity of caspase-3-dependent apoptotic pathway in response to irradiation were further evaluated. High expression (immunoreactive score > 6) of MRP3 significantly predicted poor pathological response to chemoradiotherapy (tumor regression grade ≤ 2 vs. ≥3, p = 0.002) in univariate analysis and unfavorable long-term prognosis (5-year overall survival: HR = 1.612, 95% CI, 1.094–2.375, p = 0.016; 5-year disease-free survival: HR = 1.513, 95% CI, 1.041–2.200, p = 0.030) in multivariate Cox analysis. MRP3 down-regulation significantly increased 5-fluorouracil or irradiation-induced cell apoptosis and attenuated tumor growth following irradiation in animal models. MRP3 inhibition significantly reduced intracellular reactive oxygen species exporting from cells following irradiation, and increased expression of cleaved poly ADP-ribose polymerase and caspase-3. Aberrant expression of MRP3 in rectal cancer confers chemo-radioresistance. MRP3 might be a predictive factor and an attractive target in treating advanced rectal cancer.

ERCC1 as a biological marker guiding management in malignant pleural mesothelioma.

To determine prognostic value of excision repair cross-complementation 1 (ERCC1) in patients with malignant pleural mesothelioma (MPM). The study included 60 patients with MPM who were diagnosed and treated in the Radiation Oncology Department of Kayseri Teaching Hospital and Medical Oncology Department of Erciyes University, Medicine School between 2005 and 2013. By using immunohistochemical methods, ERCC1 expression in biopsy specimens was evaluated. We retrospectively assessed whether there is a correlation between ERCC1 and response to anti-neoplastic therapy or survival. There were 50 men and 10 women with median age of 62 years (range: 39-83). Histological type was epithelial mesothelioma in the majority of the cases (85%), most commonly presenting in stage four. Of the cases, 20 (33%) received radiotherapy, 60 (%100) received first-line chemotherapy and 15 (%25) received second-line chemotherapy. In the assessment after therapy, it was found that there was partial response in 12 cases (20%), stable disease in 19 cases (31.4%) and progression in 25 cases (41.7%). ERCC1 was positive in 43% of the cases. Mean OS was 11.7 months and mean DFS was 9.5 months in ERCC1-positive cases regardless of therapy, while they were 19.2 months and 17.1 months in ERCC1-negative cases, respectively. The difference was found to be significant (p<0.05). In univariate analysis, stage, comorbidity, response to treatment and ERCC1 expression were found to be significantly associated with OS (p=0.083; p=0.043; p=0.041; p=0.050). In multivariate analysis, response to treatment remained to be significant for OS (p=0.005). In univariate and multivariate analyses, response to treatment and ERCC1 were found to be significantly associated with DFS (p=0.049; p=0.041). ERCC1 was identified as poor prognostic factor in patients with MPM.

Glomerular expression of myxovirus resistance protein 1 in human mesangial cells: Possible activation of innate immunity in the pathogenesis of lupus nephritis

Since viral infections activate type I interferon (IFN) pathways and cause subsequent release of IFN-dependent proinflammatory chemokines and cytokines, the innate immune system plays an important role in the pathogenesis of lupus nephritis (LN). It has been reported that human myxovirus resistance protein 1 (Mx1), a type I IFN-dependent transcript, acts against a wide range of RNA viruses. Although the expression of Mx1 in biopsy specimens obtained from patients with dermatomyositis and cutaneous lupus has been described, the expression of Mx1 in human mesangial cells (MCs) has remained largely unknown. We treated normal human MCs in culture with polyinosinic-polycytidylic acid (poly IC), an authentic double-stranded RNA, and analyzed the expression of Mx1 by reverse transcription-polymerase chain reaction and western blotting. To elucidate the poly IC-signalling pathway, we subjected the cells to RNA interference against IFN-β. We also conducted an immunofluorescence study to examine mesangial Mx1 expression in biopsy specimens from patients with LN. Poly IC-induced Mx1 expression in MCs are shown both time- and dose-dependently, and RNA interference against IFN-β inhibited poly IC-induced Mx1 expression. Intense glomerular Mx1 expression was observed in biopsy specimens from patients with LN, whereas negative staining occurred in specimens from patients with IgA nephropathy or purpura nephritis. These preliminary observations support, at least in part, the theory of innate immune system activation in the pathogenesis of LN.

CD44 Expression in Biopsy Specimens of Patients with Nasopharyngeal Carcinoma Is Associated with Prognosis

Objectives: CD44 is a major cancer stem cell marker, and it has been reported to be associated with biological malignancy in several cancers. However, the potential oncogenic role of CD44 and its clinical/prognostic significance in nasopharyngeal carcinoma (NPC) are unclear. In this study, we evaluated the association of CD44, especially its variant isoforms V3 and V6, with prognostic factors in NPC patients. We examined CD44 expression by immunohistochemical analysis of biopsy specimens obtained from 42 NPC patients who underwent treatment. Methods: The subjects were 42 patients (35 men and 7 women; average age, 56.3 years) who received concurrent chemoradiotherapy (CCRT) consisting of chemotherapy (CBDCA or CDGP or DOC) and radiation therapy (60-66 Gy) as an initial treatment. Thereafter, the biopsy specimens from these patients were examined immunohistochemically to study the association between CD44 expression (V3, V6) and prognosis of NPC. Results: High CD44 (V3, V6) expression was significantly correlated with increased clinical incomplete response to CCRT at the primary site ( P = 0.008 in V3 and V6 by χ2-test) and poor disease-specific survival ( P = 0.0003 in V3 and P = 0.0029 in V6 by log-rank test). However, distant metastasis showed no association with CD44 expression. Conclusions: High expression of CD44 (V3, V6) is associated with poor prognosis in patients with NPC. Evaluation of CD44, especially the variant isoforms V3 and V6, may be useful for predicting tumor response to CCRT and prognosis of patients with NPC.

Expression of pAkt affects p53 codon 72 polymorphism-based prediction of response to radiotherapy in nasopharyngeal carcinoma

BackgroundCodon 72 (Arg/Pro), the most frequently studied single nucleotide polymorphism (SNP) of p53 to date, is associated with the ability of the gene to induce cell apoptosis. The PI3K/Akt pathway plays an essential role in the transcriptional activation function of p53, and is an important factor in radiotherapy resistance. The present study was designed to evaluate the prediction of response to radiotherapy based on p53 codon 72 SNP and pAkt expression in biopsy specimens of locoregional nasopharyngeal carcinoma (NPC) before treatment.Materials and methodsIn total, 75 consecutive patients with locoregional NPC were enrolled. The p53 codon 72 SNP was identified from retrospectively collected paraffin-embedded biopsy specimens using Sanger sequencing. Expression patterns of p53, p21, 14-3-3σ, and pAkt proteins were investigated using immunohistochemical analyses. The effects of genetic polymorphisms and protein expression on progression-free survival (PFS) were evaluated using the Cox proportional hazards model, Kaplan–Meier method, and log-rank test.ResultsThe p53 codon 72 Pro/Pro carriers showed lower risk of disease progression (local recurrence and distant metastases) (HR: 0.300; 95% CI: 0.092–0.983; p=0.047). However, this association between the p53 codon 72 polymorphism and PFS was not significant in the pAkt-positive subgroup. No association was observed between protein expression of p53, p21 or 14-3-3σ and p53 codon72 polymorphisms. Notably, positive expression of p53 protein appeared to be correlated with poorer PFS among patients diagnosed as local regional lymph node metastasis (N+) before treatment (p=0.032).ConclusionsThe p53 codon 72 Pro/Pro genotype may be an effective independent prognostic marker for better outcome in patients with locoregional NPC. Based on the current findings, we hypothesize that pAkt weakens the predictive value of p53 codon 72 SNP in NPC. A combination of positive p53 protein expression and local regional lymph node metastasis may additionally be predictive of high risk of disease progression.

Impaired regeneration in LGMD2A supported by increased PAX7‐positive satellite cell content and muscle‐specific microrna dysregulation

Recent in vitro studies suggest that CAPN3 deficiency leads initially to accelerated myofiber formation followed by depletion of satellite cells (SC). In normal muscle, up-regulation of miR-1 and miR-206 facilitates transition from proliferating SCs to differentiating myogenic progenitors. We examined the histopathological stages, Pax7 SC content, and muscle-specific microRNA expression in biopsy specimens from well-characterized LGMD 2A patients to gain insight into disease pathogenesis. Three distinct stages of pathological changes were identified that represented the continuum of the dystrophic process from prominent inflammation with necrosis and regeneration to prominent fibrosis, which correlated with age and disease duration. Pax7-positive SCs were highest in the fibrotic group and correlated with down-regulation of miR-1, miR-133a, and miR-206. These observations, and other published reports, are consistent with microRNA dysregulation leading to inability of Pax7-positive SCs to transit from proliferation to differentiation. This results in impaired regeneration and fibrosis.

TOX is a specific marker for cutaneous T cell lymphomas

Cutaneous T cell lymphoma (CTCL) is characterized by proliferation of clonally expanded T lymphocytes in skin, but without detectable systemic involvement at first diagnosis. The most common subtypes of CTCL are mycosis fungoides (MF) and Sézary syndrome (SS). It is sometimes very difficult to differentiate CTCL from benign inflammatory skin diseases such as atopic dermatitis (AD). Thymocyte selection-associated HMG box factor (TOX) is a DNA-binding factor required for development of CD4 T cells, natural killer T cells, and T regulatory cells. It has been recently reported that TOX serves as a molecular marker for histological diagnosis of early-stage MF. In this study, we evaluated TOX expression in biopsy specimens from various types of cutaneous lymphoma. The followings were enrolled; MF (2 patch, 2 plaque, and 2 tumor samples), SS (2 samples), anaplastic large cell lymphoma (ALCL; 5 samples), adult T cell lymphoma (ATL; 5 samples), diffuse large B dell lymphoma (DLBCL; 5 samples), lymphomatoid papulosis (Lyp; 5 samples), and peripheral T cell lymphoma not otherwise specified (PTL, NOS; 5 samples). We also evaluated TOX expression using skin specimen from healthy controls (5 samples) and AD (2 samples). Dermal atypical lymphocytes showed a high specific nuclear staining in MF/SS, which was consistent with the previous report. Infiltrating lymphocytes in Lyp, and PTL, NOS also showed positive-staining. Interestingly, almost all anaplastic large cells in Lyp were positive for TOX. Tumor cells in ALCL or ATL showed a slightly dim nuclear staining of TOX. In contrast, there was no TOX staining in infiltrating cells of DLBCL, AD, or healthy skin. In summary, TOX may be useful as a specific marker for histological diagnosis of CTCL. JSID AbstractsJournal of Dermatological ScienceVol. 69Issue 2Preview Full-Text PDF

Glomerular expression of fractalkine is induced by polyinosinic-polycytidylic acid in human mesangial cells: possible involvement of fractalkine after viral infection

Viral infections often trigger the onset or worsening of glomerular diseases, but the details of this mechanism are unclear. Fractalkine/CX3CL1 (Fkn) is a chemokine that induces the chemotaxis and activation of cells expressing its receptor, CX3CR1. To examine the involvement of glomerular Fkn expression in the development of glomerulonephritis after viral infection, we conducted experimental studies using human mesangial cells (MCs) in culture. We examined the effect of polyinosinic-polycytidylic acid (poly IC), an authentic viral double-stranded RNA, on Fkn expression in MCs to investigate the involvement of Fkn in the antiviral reaction of MCs. Fkn mRNA and protein were analyzed using real-time PCR and enzyme-linked immunosorbent assay. Also, an immunofluorescent study to examine mesangial Fkn expression in biopsy specimens obtained from patients with glomerulonephritis was conducted. Poly IC-induced Fkn expression in MCs in both a time- and dose-dependent manner, and RNA interference (RNAi) against Toll-like receptor 3 (TLR3) or interferon regulatory factor 3 (IRF3) inhibited poly IC-induced Fkn expression. Significant glomerular Fkn expression was observed in biopsy specimens from patients with immunoglobulin A nephropathy and purpura nephritis, with increasing severity of glomerular inflammation. The TLR3/IRF3/Fkn signaling pathway may, at least in part, mediate immune and inflammatory responses against viral infection in MCs.

Clinical Features of Newly Diagnosed CD25-Positive Follicular Lymphoma

AbstractAbstract 1543 Background:CD25 (the alpha chain of the IL-2 receptor) has been shown to be expressed in hairy cell leukemia cells and a small subset of normal B cells. Recently, we reported that CD25 is expressed in a subset of diffuse large B-cell lymphoma (DLBCL) (ASH 2011 118:2666). However, its expression in follicular lymphoma (FL) has yet to be fully examined. The aim of this study was to clarify the clinical features of newly diagnosed CD25+ FL. Patients and Methods:Eighty-four patients with newly diagnosed CD25+ FL who were admitted to our hospital between 2002 and 2012 were retrospectively evaluated. Biopsy specimens from lymph nodes or related tissues were analyzed by flow cytometry (FCM) using a combined method of single- and two-color staining. CD25 expression was defined as positivity of >20% of clonal B cells with CD19 or 20 expression >70% in a gated region. Results were compared with patients with reactive lymphadenopathy (n=27), DLBCL at initial presentation (n=91), and FL at relapse (n=13). Results:CD25 expression in untreated patients with FL (mean±SD, 10.6±9.9%; n=84) was almost equivalent to that in patients with reactive lymphadenopathy (mean±SD, 9.1±4.4%; n=27, P =0.3), and lower than that in untreated patients with DLBCL (mean±SD, 28.5±31.1%; n=91, P <0.001). There was a trend of a higher CD25 expression in biopsy specimens from relapsed FL than in those from newly diagnosed FL (mean±SD, 18.6±16.3 vs. 10.6±9.9%, respectively, P=0.077). With the sum of treated and untreated CD25+ FL, two-color FCM showed that the mean±SD of CD25/CD19 and CD25/CD20 dual expression was 28.5±26.5% (n=4) and 28±24.3% (n=4), respectively. The patients with newly diagnosed CD25+ (more than 20%) FL (n=10) showed some clinical features such as B symptoms (P =0.09), soluble IL-2 receptor titers of more than 5,000 U/ml (P =0.013), and advanced ages over 60 (P =0.16), compared with CD25- FL patients (n=74). Besides, there were some differences between CD25+ and CD25- FL in terms of histology grade 3 (50 vs. 32%, respectively, P=0.29) and the presence of t(14;18) (33 vs. 56%, respectively, P=0.27).Patients with FL at the initial presentation underwent rituximab-containing chemotherapy (n=68), radiotherapy (n=4), and observation (n=12). Kaplan–Meier analysis showed poorer progression-free survival (PFS) and overall survival (OS) in patients with CD25+ FL than that in patients with CD25- FL (2-year PFS, 36 vs. 82%, respectively, P <0.001; 6-year OS, 47 vs. 88%, respectively, P <0.01). Multivariate analysis revealed that CD25 expression is a significant prognostic factor for PFS (HR 6.4, 95%CI 2.4–17.4, P <0.001) and OS (HR 6.6, 95%CI 1.2–36.9, P= 0.027), which is independent of FLIPI score variables. Conclusion:Newly diagnosed CD25+ FL may constitute a distinct subgroup associated with aggressive clinical features and an inferior prognosis. CD25 may be a new prognostic marker, and could be a therapeutic target in FL patients. Disclosures:No relevant conflicts of interest to declare.