Kidney damage relates to agonal bacteremia: a single-center retrospective study.

BACKGROUND: Agonal bacteremia, diagnosed with postmortem positive blood culture results, is considered a possible contributing factor to death. We hypothesized that some premortem organ damage, such as kidney damage, can enhance agonal bacteremia. METHODS: We performed a postmortem blood and alveolar fluid culture study in 30 cadavers and evaluated the relationship between blood culture results and clinical parameters, including organ damage (brain, heart, lung, kidney, liver and gastrointestinal tract). RESULTS: A total of 23 cases (76.7%) were positive for blood culture; the number of cultured species was one in 12 cases, two in 7 cases, and three in 4 cases. The ratio of agonal bacteremia was significantly higher in patients with heart damage (100%, n = 13) and those with kidney damage (end-stage kidney damage, acute kidney injury, obstructive kidney failure, or metastatic kidney tumours) (100%, n = 13). The mean number of cultured species was 0.67 ± 0.98 in heart or kidney damage, 1.40 ± 0.55 in heart damage only, 1.40 ± 0.55 in kidney damage only, and 2.00 ± 0.93 in heart and kidney damage. As the number of damaged organs increased (0 organs, no heart/kidney damage; 1 organ, heart or kidney damage; and 2 organs, heart and kidney damage), the mean number of cultured species increased significantly (p for trend = 0.001964). CONCLUSION: Premortem kidney damage relates to agonal bacteremia.

Automated assessment of Ki-67 in breast cancer: the utility of digital image analysis using virtual triple staining and whole slide imaging.

AIMS: Precise evaluation of proliferative activity is essential for the stratified treatment of luminal-type breast cancer (BC). Immunohistochemical staining of Ki-67 has been widely used to determine proliferative activity and is recognised to be a useful prognostic marker. However, there remains discussion concerning the methodology. We aimed to develop an automated and reliable Ki-67 assessment approach for invasive BC. MATERIALS AND RESULTS: A retrospective study was designed to include two cohorts consisting of 152 and 261 consecutive patients with luminal-type BC. Representative tissue blocks following surgery were collected, and three serial sections were stained automatically with Ki-67, pan-cytokeratin and p63. The whole slides were scanned digitally and aligned using VirtualTripleStaining - an extension to the VirtualDoubleStaining™ technique provided by Visiopharm software. The aligned files underwent automated invasive cancer detection, hot-spot identification and Ki-67 counting. The automated scores showed a significant positive correlation with the pathologists' scores (r = 0.82, P < 0.0001). Among selected patients with curative surgery and standard adjuvant therapies (n = 130), the digitally assessed low Ki-67 group (<20%) demonstrated a significantly better prognosis (breast cancer-specific survival, P = 0.030; hazard ratio = 0.038) than the high Ki-67 group. CONCLUSIONS: Digital image analysis yielded similar results to the scores determined by experienced pathologists. The prognostic utility was verified in our cohort, and an automated process is expected to have high reproducibility. Although some pitfalls were confirmed and thus need to be monitored by laboratory staff, the application could be utilised for the assessment of BC.

Targeted next-generation sequencing assays using triplet samples of normal breast tissue, primary breast cancer, and recurrent/metastatic lesions.

BACKGROUND: Next-generation sequencing (NGS) has shown that recurrent/metastatic breast cancer lesions may have additional genetic changes compared with the primary tumor. These additional changes may be related to tumor progression and/or drug resistance. However, breast cancer-targeted NGS is not still widely used in clinical practice to compare the genomic profiles of primary breast cancer and recurrent/metastatic lesions. METHODS: Triplet samples of genomic DNA were extracted from each patient's normal breast tissue, primary breast cancer, and recurrent/metastatic lesion(s). A DNA library was constructed using the QIAseq Human Breast Cancer Panel (93 genes, Qiagen) and then sequenced using MiSeq (Illumina). The Qiagen web portal was utilized for data analysis. RESULTS: Successful results for three or four samples (normal breast tissue, primary tumor, and at least one metastatic/recurrent lesion) were obtained for 11 of 35 breast cancer patients with recurrence/metastases (36 samples). We detected shared somatic mutations in all but one patient, who had a germline mutation in TP53. Additional mutations that were detected in recurrent/metastatic lesions compared with primary tumor were in genes including TP53 (three patients) and one case each of ATR, BLM, CBFB, EP300, ERBB2, MUC16, PBRM1, and PIK3CA. Actionable mutations and/or copy number variations (CNVs) were detected in 73% (8/11) of recurrent/metastatic breast cancer lesions. CONCLUSIONS: The QIAseq Human Breast Cancer Panel assay showed that recurrent/metastatic breast cancers sometimes acquired additional mutations and CNV. Such additional genomic changes could provide therapeutic target.

Multiplex PCR analysis of apocrine lesions shows frequent PI3K-AKT pathway mutations in both benign and malignant apocrine breast tumors.

Pathological diagnosis of intraductal apocrine lesions can be challenging, because even benign apocrine lesions often show atypical cytology, and immunohistochemistry is of little assistance. A new diagnostic method for apocrine lesions is desirable. The mutations present in apocrine lesions have not been well studied. We performed a MassARRAY multiplex polymerase chain reaction (PCR) study of benign and malignant apocrine lesions, which included 152 mutations of 18 genes. We found that four of 11 benign lesions showed AKT1 or PIK3CA mutations, one of four noninvasive apocrine carcinomas showed a FBX4 mutation, two of 15 invasive apocrine carcinomas showed a PIK3CA mutation, and one invasive apocrine carcinoma showed both PIK3CA and TP53 mutations. The mutation frequency did not differ significantly between benign and malignant lesions (p = 0.683). We demonstrated that both benign and malignant apocrine lesions may contain mutations of genes in the PI3K-AKT pathway, this pathway could be a good therapeutic target of these diseases.

CD1d- and PJA2-related immune microenvironment differs between invasive breast carcinomas with and without a micropapillary feature.

BACKGROUND: Invasive micropapillary carcinoma (IMPC) of the breast is characterized by its unique morphology and frequent nodal metastasis. However, the mechanism for development of this unique subtype has not been clearly elucidated. The aim of this study was to obtain a better understanding of IMPC. METHODS: Using representative cases of mixed IMPC, mRNA expression in the micropapillary area and usual invasive area was compared. Then, immunohistochemical analyses for 294 cases (76 invasive carcinomas with a micropapillary feature [ICMF] and 218 invasive carcinomas without a micropapillary feature [ICNMF]) were conducted. Clinicopathological analyses were also studied. RESULTS: DNA microarray analyses for mixed IMPC showed that BC-1514 (C21orf118) was commonly upregulated in the micropapillary area. CAMK2N1, CD1d, PJA2, RPL5, SAMD13, TCF4, and TXNIP were commonly downregulated in the micropapillary area. Immunohistochemically, we confirmed that BC-1514 was more upregulated in ICMF than in ICNMF. CD1d and PJA2 were more downregulated in ICMF than ICNMF. All patients with cases of PJA2 overexpression survived without cancer recurrence during the follow-up period, although the differences for disease-free (p = 0.153) or overall survival (p = 0.272) were not significant. CONCLUSIONS: The CD1d- and PJA2-related tumour microenvironment might be crucial for IMPC. Further study of the immune microenvironment and micropapillary features is warranted.

Phosphorylated HER3 and FITC-labeled trastuzumab immunohistochemistry in patients with HER2-positive breast cancer treated with adjuvant trastuzumab.

The development of trastuzumab has significantly improved the prognosis of HER2-positive breast cancer. However, disease recurs in some patients with HER2-positive breast cancer. A new strategy for treating HER2-positive breast cancer is necessary. Although several studies have reported that HER3 is a prognostic factor for HER2-positive breast cancers, phosphorylated HER3 (pHER3) has not been well studied. There has been no survival analysis including immunohistochemistry with trastuzumab as the primary antibody. We analyzed immunohistochemistry using anti-pHER3 antibody and FITC-labeled trastuzumab (FITC-tra). Of 78 patients enrolled in the study, we could evaluate the immunohistochemistry for pHER3 in 71 cases and that for FITC-tra in 72 cases. Sixteen cases were positive for pHER3 (16/71, 22.5%), and 19 positive for FITC-tra (19/72, 26.4%). Kaplan-Meier analysis showed a significant association of pHER3 positivity (p = 0.011) but not HER3 positivity or FITC-tra positivity with disease-free survival. Therefore, immunohistochemical evaluation of pHER3 in HER2-positive breast cancer may provide a useful biomarker. An expanded study of pHER3 involving standardization of the pHER3 test to be encouraged.

Anti-cancer stem cell activity of a hedgehog inhibitor GANT61 in estrogen receptor-positive breast cancer cells.

Estradiol (E2) increases not only the cell growth but also the cancer stem cell (CSC) proportion in estrogen receptor (ER)-positive breast cancer cells. It has been suggested that the non-canonical hedgehog (Hh) pathway activated by E2 plays an important role in the regulation of CSC proportion in ER-positive breast cancer cells. We studied anti-CSC activity of a non-canonical Hh inhibitor GANT61 in ER-positive breast cancer cells. Effects of GANT61 on the cell growth, cell cycle progression, apoptosis and CSC proportion were investigated in four ER-positive breast cancer cell lines. CSC proportion was measured using either the mammosphere assay or CD44/CD24 assay. Expression levels of pivotal molecules in the Hh pathway were measured. Combined effects of GANT61 with antiestrogens on the anti-cell growth and anti-CSC activities were investigated. E2 significantly increased the cell growth and CSC proportion in all ER-positive cell lines. E2 increased the expression levels of glioma-associated oncogene (GLI) 1 and/or GLI2. GANT61 decreased the cell growth in association with a G1-S cell cycle retardation and increased apoptosis. GANT61 decreased the E2-induced CSC proportion measured by the mammosphere assay in all cell lines. Antiestrogens also decreased the E2-induced cell growth and CSC proportion. Combined treatments of GANT61 with antiestrogens additively enhanced anti-cell growth and/or anti-CSC activities in some ER-positive cell lines. In conclusion, the non-canonical Hh inhibitor GANT61 inhibited not only the cell growth but also the CSC proportion increased by E2 in ER-positive breast cancer cells. GANT61 enhanced anti-cell growth and/or anti-CSC activities of antiestrogens in ER-positive cell lines.

Vasohibin-1 Is a Poor Prognostic Factor of Ovarian Carcinoma.

Vasohibin-1 (VASH1) is an identified negative feedback inhibitor of angiogenesis induced by vascular endothelial growth factor (VEGF) in vascular endothelial cells (ECs). Expression of VASH1 has been reported not only in ECs of normal tissue, but also in ECs surrounding malignant tumors. In malignant tumors, VASH1 is also gaining attention as a prognosis prediction marker. The aim of this study is to investigate the correlation between VASH1 expression and vascular-related factors and various clinicopathological outcomes in clinical cases of ovarian carcinoma. We retrospectively analyzed clinical records of 58 patients with ovarian carcinoma. The expression patterns of VASH1 and other vascular-related factors (CD31 as markers of microvessel density (MVD), VEGF receptor type 2 (VEGFR2), D2-40 as markers of lymphovessel density), and Ki67 (as proliferation markers of cancer cells) were examined immunohistochemically. We studied the correlation between immunohistochemical expression and overall survival. VASH1 expression pattern significantly differed between Federation of Obstetrics and Gynecology (FIGO) Stages. Numbers of VASH1-positive vessels had a significant positive correlation with MVD (Speaman's correlation coefficient (ρ) was 0.51, p < 0.001), VEGFR2-positive vessels (ρ = 0.61, p < 0.001), and percentage of Ki67 (ρ = 0.28, p = 0.034). The Cox univariable analyses revealed that the group of high VASH1 expression (> 14.6 vessels per mm2) at Stages I-III is a prognostic factor (HR = 3.3, 95%CI = 0.4-8.4; p = 0.013). Our results indicate that VASH1 expression in ovarian carcinoma is significantly associated with vascular-related factors and Ki67 expression. We propose that VASH1 is a prognostic marker in ovarian carcinoma.

Immunohistochemistry of Vasohibin-2 in Human Kidney Disease : Implications in Impaired Glucose Tolerance and Reduced Renal Function.

Several angiogenesis-related factors are known to play important roles in the pathogenesis of kidney disease. Vasohibin-2 (VASH-2) was recently reported as a novel proangiogenic factor. Although VASH-2 was demonstrated to accelerate tumor angiogenesis, its roles in non-tumor processes including renal disease have not been well elucidated yet. Here, we performed a retrospective study including an immunohistochemical analysis of human kidney biopsy specimens from 82 Japanese patients with a variety of kidney diseases, and we evaluated the correlations between the immunoreactivity of VASH-2 and the patients' clinicopathological parameters. VASH-2 immunoreactivity was detected in varying degrees in renal tubules as well as in peritubular capillaries and vasa recta. The cortical and medullary tubule VASH-2+ scores were correlated with the presence of hypertension, and the medullary tubule VASH-2+ score was significantly correlated with the blood glucose (p=0.029, r=0.35) and hemoglobin A1c levels (p=0.0066, r=0.39). Moreover, decreased VASH-2+ scores in the vasa recta were associated with reduced renal function (p=0.0003). These results suggest that VASH-2 could play an important role in the pathogenesis of renal diseases, and that VASH-2 is closely associated with hypertension and impaired glucose tolerance.

Incremental value of high b value diffusion-weighted magnetic resonance imaging at 3-T for prediction of extracapsular extension in patients with prostate cancer: preliminary experience.

PURPOSE: To investigate whether high b value diffusion-weighted imaging (DWI) contributes to the improvement of diagnostic ability of extracapsular extension (ECE) in prostate cancer (PC). MATERIALS AND METHODS: Forty-three patients with PC underwent multiparametric MRI including DWI (b values: 0, 2000 s/mm2) at 3-T. Two radiologists assessed the presence of ECE and the diagnostic certainty degree using conventional diagnostic method by consensus. Tumor apparent diffusion coefficient (ADC, ×10-3 mm2/s) was also measured. Independent predictors of ECE were identified among PSA, tumor ADC, Gleason score, and conventional MRI. ECE in patients with low diagnostic certainty by conventional MRI was further reevaluated using ADC cutoff value, and the results were combined with those of patients with high diagnostic certainty by conventional MRI (MRI + ADC method). RESULTS: Tumor ADC was an independent predictor of ECE, and the ADC cutoff value was 0.72. The sensitivity, specificity, and accuracy of conventional MRI and MRI + ADC method in the diagnosis of ECE were 44, 92, and 72%, and 78, 96, and 88%, respectively. Among MRI findings leading to the judgement of low diagnostic certainty, broad tumor contact was most common (72% of the patients). CONCLUSIONS: The addition of ADC obtained with high b value DWI at 3-T to conventional MRI improved the diagnostic ability of ECE.

Value of preoperative 3T multiparametric MRI for surgical margin status in patients with prostate cancer.

PURPOSE: To examine the value of preoperative multiparametric magnetic resonance imaging (MRI) as a predictor of surgical margin (SM) status in patients with prostate cancer (PC). MATERIALS AND METHODS: The Institutional Review Board approved this retrospective study; the requirement for informed consent was waived. Fifty-six male patients with histologically proven PC underwent preoperative 3T multiparametric MRI including high b value (0, 2000 s/mm(2) ) diffusion-weighted imaging. In each patient, clinical data, such as biopsy Gleason score and D'Amico clinical risk score, and multiparametric MRI findings, such as tumor location, tumor size, tumor extension in the apical or proximal region, tumor apparent diffusion coefficient (ADC), and the presence or absence of MRI findings of extracapsular extension (ECE) were evaluated. Statistical evaluations included the Fisher's exact test, χ(2) test, Mann-Whitney U-test, and logistic regression analysis. RESULTS: On histopathological evaluation, 15 patients (27%) were SM-positive (SMP group), and 41 (73%) were SM-negative (SMN group). The tumor ADC was significantly lower in the SMP group than in the SMN group (P = 0.001). The frequency of tumor extension in the apex or base and suspected ECE on MRI were significantly higher in the SMP group than in the SMN group (P = 0.037 and 0.011, respectively). On multivariate analysis, tumor ADC was the only predictor of SM status in PC (P = 0.003). CONCLUSION: PC with positive SM was characterized by tumor extension in the apical and proximal regions, lower tumor ADC, and tumors with positive MRI findings of ECE, compared to tumors with negative SM. J. Magn. Reson. Imaging 2016;44:584-593.

Pancreatic adenocarcinomas without secondary signs on multiphasic multidetector CT: association with clinical and histopathologic features.

PURPOSE: To determine the clinical, histopathologic and imaging features of pancreatic adenocarcinomas without secondary signs on dynamic CT. MATERIALS AND METHODS: Seventy patients (mean age 70 years) with histologically proven pancreatic adenocarcinoma underwent preoperative contrast material-enhanced multiphasic multidetector CT before pancreatic resection. In each patient, clinical data including carbohydrate antigen 19-9, frequency of isoattenuating tumours, and presence of secondary signs and histopathologic findings such as tumour location, tumour stage, and microscopic infiltrative growth grade were evaluated. RESULTS: Ten tumours (14%) were without secondary signs, and 60 (86%) were with secondary signs. Tumours without and with secondary signs were located in the uncinate process in 5 (50%) and 3 (5%), head in 3 (30%) and 29 (48%), body in 2 (20%) and 22 (37%), and tail in 0 (0%) and 6 (10%), respectively (p = .001). The frequency of isoattenuating pancreatic adenocarcinomas without secondary signs was significantly higher than those with secondary signs (p = 0.034). The tumour stage of pancreatic adenocarcinomas without secondary signs was earlier than that in tumours with secondary signs (p = 0.041). CONCLUSIONS: Pancreatic adenocarcinomas without secondary signs is characterized by the presence of uncinate and isoattenuating tumours and earlier tumour stage compared to tumours with secondary signs. KEY POINTS: Frequency of pancreatic adenocarcinomas without secondary signs on multiphasic CT is 14 . Pancreatic adenocarcinomas without secondary signs are common in the uncinate process. Pancreatic adenocarcinomas without secondary signs are common in isoattenuating tumours. Pancreatic adenocarcinomas without secondary signs are characterized by earlier-stage tumours.

[Genome Abnormality and Histological Findings in Breast Carcinoma].

Breast cancers contain variable histologies as well as biology. Gene expression profiling and cluster analyses have been performed since the beginning of the 21st century. The use of intrinsic subtype classification has replaced histological classification of breast carcinomas, as it frequently yields the same results. For examples, around 80% of triple negative (estrogen receptor-, progesterone receptor-, and HER2-negative) cancers are of the basal-like subtype. In daily practice, adjuvant therapy is selected based on histological features, but the results of ordinal cluster analyses and histological intrinsic subtypes are not always the same for individual cases. With advanced genetic analysis, new concepts have been elucidated, ie, the molecular identification of claudin-low breast cancer. Proposals of a new classification system and a new therapeutic approach are expected in the future.

Management of breast papillary lesions diagnosed in ultrasound-guided vacuum-assisted and core needle biopsies.

AIMS: To assess the outcome of breast papillary lesions diagnosed by ultrasound-guided core needle biopsy (CB) or vacuum-assisted 'mammotome' biopsy (MT), the accuracy of these diagnoses, and whether it is justified not to undertake surgical excision of non-malignant papillary lesions so diagnosed. METHODS AND RESULTS: Among 3219 (MT, 2195; CB, 1024) breast biopsies spanning 5 years, 185 (5.7%) papillary lesions [MT, 162 (88%); CB, 23 (12%)] were identified. Of these, 142 cases (77%; MT/CB, 125/17) were benign, 24 (13%, 23/1) were atypical, and 19 (10%; 14/5) were malignant. Of the 142 benign cases, 114 had imaging follow-up (FU) (FU period 2-81 months); 17 of 114 cases were excised, and four were malignant (3.5%) (FU period 4-57 months). Of the 24 atypical cases (23 had FU), 19 were excised: six were benign (32%) and 13 malignant (68%). The remaining four cases were considered to be non-malignant (FU period 7-54 months). CONCLUSIONS: Benign papillary lesions diagnosed by MT or CB might not require immediate excision, but should receive imaging FU for at least 5 years. Excision should be performed in cases showing changes in imaging features, as the possibilities of carcinoma coexisting with papilloma or carcinoma developing from papilloma cannot be excluded, as illustrated by the 4% upgrade rate at excision in this study.

Renal distribution of Vasohibin-1 in patients with chronic kidney disease.

Experimental studies have demonstrated the involvement of angiogenesis-related factors in the progression of chronic kidney disease (CKD). There have so far been no reports investigating the distribution and clinical roles of Vasohibin-1 (VASH-1), a negative feedback regulator of angiogenesis, in CKD. We recruited 54 Japanese CKD patients and 6 patients who had normal renal tissues excised due to localized renal cell carcinoma. We evaluated the correlations between the renal expression level of VASH-1 and the clinical/histological parameters. VASH-1 was observed in renal endothelial/mesangial cells, crescentic lesions and interstitial inflammatory cells. Significant positive correlations were observed between 1) crescent formation and the number of VASH-1+ cells in the glomerulus (r＝0.48, p＝0.001) or cortex (r＝0.64, p＜0.0001), 2) interstitial cell infiltration and the number of VASH-1+ cells in the cortex (r＝0.34, p＝0.02), 3) the glomerular VEGFR-2+ area and the number of VASH-1+ cells in the glomerulus (r＝0.44, p＝0.01) or medulla (r＝0.63, p＝0.01). These results suggest that the renal levels of VASH-1 may be affected by local inflammation, crescentic lesions and VEGFR-2.

[Up-to-date pathological diagnosis for breast cancer].

The number of breast cancer patients among Japanese women has been increasing, and one in fourteen women may develop breast cancer during their lifetime. Most cases of breast cancer occur in the terminal duct-lobular units, but their pathological features are quite heterogeneous. Thus, it is necessary to examine them pathologically, to establish appropriate adjuvant therapy for individual patients. In addition to determining the histological type, tumor size, tumor grade, lymphovascular invasion, node metastases, and surgical margin status (for breast-conserving surgery), we should clarify the biomarker status. Analyses of estrogen receptor (ER), progesterone receptor (PgR), and human epidermal growth factor 2 (HER2) are routinely performed currently, but strict quality control is strongly recommended, including pre-analytical issues. In addition, the Ki-67 labeling index has been employed as a marker to determine the indication for adjuvant chemotherapy for hormone receptor (ER/PgR)-positive, HER2-negative invasive carcinoma patients. The significance of PgR may alter, too. The threshold for the therapeutic indication may alter with the progress of pharmacology. The evidence-based data and expert consensus (i.e., St. Gallen international meeting consensus) may influence our daily practice. In this issue, we introduce the newest histological classifications (by the WHO and the Japanese Society of Breast Cancer) and the Japanese Breast Cancer Treatment Guideline. Several issues to be included in the pathology reports are examined, and the current status as well as future perspectives of biomarker analysis will be discussed.

Using the DNA Integrity Number to Analyze DNA Quality in Specimens Collected from Liquid-Based Cytology after Fine-Needle Aspiration of Breast Tumors and Lesions.

INTRODUCTION: Cancer genome analysis using next-generation sequencing requires adequate and high-quality DNA samples. Genomic analyses were conventionally performed using formalin-fixed paraffin-embedded sections rather than cytology samples such as cell block or smear specimens. Specimens collected from liquid-based cytology (LBC) have the potential to be sources of high-quality DNA suitable for genetic analysis even after long-term storage. METHODS: We collected breast tumor/lesion fractions from 92 residual LBC specimens using fine-needle aspiration (FNA) biopsy, including breast carcinoma (1 invasive carcinoma and 4 ductal carcinomas in situ), papillomatous lesion (5 intraductal papillomas), and fibroepithelial lesion (19 phyllodes tumors and 53 fibroadenomas) samples, and others (1 ductal adenoma, 1 hamartoma, 1 fibrocystic disease, and 7 unknown). DNA was extracted from all samples and subjected to DNA integrity number (DIN) score analysis. RESULTS: Average DIN score collected from 92 LBC specimens was significantly higher score. In addition, high-quality DNA with high DIN values (7.39 ± 0.80) was successfully extracted more than 12 months after storage of residual LBC specimens. CONCLUSION: Residual LBC specimens collected from FNA of the breast were verified to carry high-quality DNA and could serve as an alternate source for genetic analysis.