Hepatic granulomas following liver transplantation: A retrospective survey, and analysis of possible microbiological etiology.

BACKGROUND: Liver granulomas have always been a diagnostic challenge for pathologists. They have been described in up to 15% of liver biopsies and can also be seen in liver allograft biopsy specimens, but there is a paucity of information regarding the prevalence and associated etiologic factors of granulomas in liver transplanted patients. The aim of this study is to shed light on the etiology of liver granulomas. METHODS: Liver biopsies from liver transplanted patients, in the period from 01.01.2011 - 01.05.2017, were examined. We registered the histo-morphological characteristics and clinicopathological data of all biopsies and performed next-generation sequencing (NGS) to detect possible pathogens (bacteria, fungi, and parasites) in the biopsies containing granulomas. RESULTS: We reviewed a total of 400 liver biopsies from 217 liver transplant patients. Of these, 131 liver biopsies (32.8%) from 98 patients (45.2%) revealed granulomas. Most were epithelioid granulomas located parenchymal and were detected in 115 (87.7%) biopsies. We also identified 10 cases (7.6%) with both lobular and portal granulomas and six biopsies (4.5%) with portal granulomas alone. In 54 biopsies (41.2%), granulomas were found in biopsies with acute cellular rejection (ACR). Fifty (51%) patients with granulomas underwent liver transplantation for autoimmune-related end-stage liver disease (AILD). The granulomas were found most frequently in the first six months after transplantation, where patients also more often were biopsied. NGS analysis did not reveal any potential infectious agent, and no significant differences were observed in the microbiological diversity (microbiome) between clinical- and granuloma characteristics concerning bacteria, fungi, and parasites. CONCLUSION: Our study confirmed that granulomas are frequently seen in liver allograft biopsy specimens, and most often localized in the parenchyma, occurring in the first post-transplant period in patients with AILD, and often seen simultaneously with episodes of ACR. Neither a specific microbiological etiological agent nor a consistent microbiome was detected in any case.

Extensive genomic analysis in patients with KRAS-mutated solid tumors shows high frequencies of concurrent alterations and potential targets but has limited clinical impact.

BACKGROUND: This study aimed to investigate the distribution and frequency of concurrent alterations in different cancers across KRAS subtypes and in different KRAS subtypes across cancers, and to identify potentially actionable targets and patients who received targeted treatment matched to their genomic profile (GP). MATERIALS AND METHODS: In this descriptive and single-center study, we included 188 patients with solid tumors harboring KRAS mutations in codon 12, 13, 61, 117, or 146, referred to the Phase 1 Unit, Rigshospitalet, Copenhagen, Denmark from mid-2016 to 2020. Genomic co-alterations were detected with whole-exome sequencing, RNA sequencing, SNP array, and mRNA expression array on fresh biopsies. The study is part of the Copenhagen Prospective Personalized Oncology study (NCT02290522). RESULTS: The majority of patients had colorectal cancer (60.1%), non-small cell lung cancer (11.2%), or pancreatic cancer (10.6%). Most tumors were KRAS-mutated in codon 12 or 13 (93.7%) including G12D (27.1%), G12V (26.6%), G12C (11.7%), and G13D (11.2%). A total of 175 different co-alterations were found, most frequently pathogenic APC and TP53 mutations (55.9% and 46.4%, respectively) and high expression of CEACAM5 (73.4%). Different cancers and KRAS subtypes showed different patterns of co-alterations, and 157 tumors (83.5%) had potentially actionable targets with varying evidence of targetability (assessed using ESMO Scale for Clinical Actionability of molecular Targets). Of the 188 patients included in the study, 15 (7.4%) received treatment matched to their GP (e.g., immunotherapy and synthetic lethality drugs), of whom one had objective partial response according to Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. CONCLUSION: Performing extensive genomic analysis in patients with known KRAS-mutated solid tumors may contribute with information to the genomic landscape of cancers and identify targets for immunotherapy or synthetic lethality drugs, but currently appears to have overall limited clinical impact, as few patients received targeted therapy matched to their GP.

Noninvasive Prediction of Advanced Fibrosis in Pediatric Liver Disease-Discriminatory Performance of 2D Shear Wave Elastography, Transient Elastography and Magnetic Resonance Elastography in Comparison to Histopathology.

Background: Elastography can be measured with different imaging techniques and is increasingly used for noninvasive assessment of hepatic fibrosis. Little is known about the performance, and interrelation of different elastographic techniques, in prediction of hepatic fibrosis in pediatric liver disease. Objectives: We aimed to determine the discriminatory value for advanced fibrosis (Metavir F3-4) and evaluate the applicability of 2D shear wave ultrasound elastography (USe), Transient Elastography (TE) and Magnetic Resonance elastography (MRe) in pediatric liver disease. Methods: In patients with pediatric liver disease aged 0−19 years, USe, TE and MRe were compared with histopathological fibrosis stage. Multivariate logistic regression models for advanced fibrosis were considered. Discriminative performance was assessed by the area under the receiver operating characteristic curve and the Brier Score. Primary analyses included complete cases. Multiple imputation was used as sensitivity analysis. Results: In 93 histologically evaluated patients USe, TE and MRe were performed 89, 93 and 61 times respectively. With increased liver stiffness values, significantly increased odds for presenting F3-4 were seen in individual models for ALT < 470 U/L, whereas the effect for ALT > 470 U/L was non-significant. Area under the curve and Brier Score for discrimination of advanced fibrosis were 0.798 (0.661−0.935) and 0.115 (0.064−0.166); 0.862 (0.758−0.966) and 0.118 (0.065−0.171); 0.896 (0.798−0.994) and 0.098 (0.049−0.148) for USe, TE and MRe respectively. No significant increase in discriminatory ability was found when combining elastographic modalities. Conclusions: In pediatric liver disease, USe, TE and MRe had a good discriminatory ability for assessment of advanced liver fibrosis, although TE and MRe performed best. In most children with pediatric liver disease, TE is a reliable and easily applicable measure.

Differences in gene expression despite identical histomorphology in sinonasal intestinal-type adenocarcinoma and metastases from colorectal adenocarcinoma.

Sinonasal intestinal-type adenocarcinoma (sITAC) is histomorphologically indistinguishable from colorectal adenocarcinoma (CRC) leading to diagnostic challenges. Metastases from CRCs to the sinonasal tract have been reported. The aim of the study was to identify a biomarker making it possible to distinguish between sITAC and metastases of colorectal origin. Formalin-fixated paraffin-embedded (FFPE) tissue from 20 consecutive patients with sITAC treated at Rigshospitalet, Denmark from 2005 to 2017, 20 patients with CRC, and second patients with both sinonasal and colorectal carcinomas were included, and RNA-sequencing was performed on all samples. Moreover, a series of 26 samples from metastasizing CRC were included (in-house data). 3139 differentially expressed genes were identified, of these several were deemed as possible biomarkers, including CSDE1, for which immunohistochemical staining was performed. sITAC and CRC differ in genomic expression. CSDE1, previously found upregulated in CRC, was significantly differentially expressed. Using immunohistochemical staining, no sITACs displayed strong and diffuse staining for CSDE1, which represents a potential marker to use in distinguishing sITAC from a metastasis of colorectal origin. This knowledge could improve the diagnostic process and hopefully the outcome in patients with this rare tumor.

Increase of Ki-67 index and influence on mortality in patients with neuroendocrine neoplasms.

An increase in the Ki-67 index in neuroendocrine neoplasms over time in relation to prognosis has scarcely been investigated. We aimed to assess whether the Ki-67 index changed over time and also whether a change influenced prognosis. Second, we investigated the difference in the Ki-67 index between primary tumour and metastases. From 1 January 1995 to 31 December 2019, 108 consecutive patients with gastroenteropancreatic tumours were included. Patients were followed with regard to an increase in the Ki-67 index and all-cause mortality. Ki-67 determination of the primary tumour at diagnosis and at the time of radiological progression, including developed metastases, was performed. A significant increase in the Ki-67 index was defined as a doubling of the value at disease progression compared to the value at diagnosis. In addition, in 14 patients, the Ki-67 index of the primary tumour and present metastases at the time of diagnosis was investigated. At diagnosis, there were no differences in the Ki-67 index between primary tumours and metastases (P = .41). Sixty-five patients had a doubling of the Ki-67 index. The median Ki-67 index at the time of progression 17% (1%-90%) vs 5% (1%-60%) at the time of diagnosis (P = .006). A doubling of the Ki-67 index was independently associated with all-cause mortality (hazard ratio = 2.7 [1.3-6.3], P = 0.02), after adjustment for relevant co-variables including the Ki-67 index at baseline. Doubling of the Ki-67 index at the time of disease progression was associated with a significantly higher risk of all-cause mortality. We recommend that a Ki-67 index is obtained whenever disease progression is recorded by demonstrated progression because it may have impact on the choice of treatment.

Clinical assessment of tumor regression grade systems in gastroesophageal adenocarcinoma following neoadjuvant chemotherapy.

BACKGROUND: The standard treatment for gastroesophageal cancer is neoadjuvant chemotherapy, followed by surgery, which has been shown to increase survival compared with surgery alone. Evidence is mounting that characterization of the oncologically induced tumor regression is of prognostic importance. However, no consensus regarding the optimal system for describing tumor regression exists. Thus, this study aims to explore three validated/promising tumor regression systems with a focus on their interobserver reliability and usability. METHODS: We included 100 consecutive patients with gastroesophageal adenocarcinoma who had undergone neoadjuvant oncological treatment followed by surgery. The tumors underwent tumor regression grade (TRG) assessment according to the Standard Mandard-, Modified Mandard-, and Becker systems to assess the interobserver reliability between two consultant pathologists. The interobserver reliability was determined by both Fleiss kappa and weighted kappa metrics. Besides, a semi-quantitative usability questionary was completed and it was expanded with usability comments. RESULTS: The Fleiss kappa interobserver agreement was 0.67 [95% CI, 0.55-0.79], 0.88 [95% CI, 0.73-1.00], and 0.88 [95% CI, 0.73-1.00] for Standard Mandard-, Modified Mandard-, and the Becker systems, respectively. The weighted kappa (linear) was 0.80 [95% CI, 0.72-0.89], 0.91 [95% CI, 0.84-0.98], and 0.91 [95% CI, 0.84-0.98] for the Standard Mandard-, Modified Mandard-, and the Becker systems, respectively. The usability was scored on a scale of 8-24 by both raters. The systems were scored accordingly: 47 (Modified Mandard system), 43 (Becker system), and 37 (Standard Mandard system). CONCLUSION: The Modified Mandard- and Becker systems had excellent interobserver reliability and usability. However, the systems could be improved by a better characterization of the different tiers and tumor morphology.

IRDye800CW labeled uPAR-targeting peptide for fluorescence-guided glioblastoma surgery: Preclinical studies in orthotopic xenografts.

Glioblastoma (GBM) is a devastating cancer with basically no curative treatment. Even with aggressive treatment, the median survival is disappointing 14 months. Surgery remains the key treatment and the postoperative survival is determined by the extent of resection. Unfortunately, the invasive growth with irregular infiltrating margins complicates an optimal surgical resection. Precise intraoperative tumor visualization is therefore highly needed and molecular targeted near-infrared (NIR) fluorescence imaging potentially constitutes such a tool. The urokinase-type Plasminogen Activator Receptor (uPAR) is expressed in most solid cancers primarily at the invading front and the adjacent activated peritumoral stroma making it an attractive target for targeted fluorescence imaging. The purpose of this study was to develop and evaluate a new uPAR-targeted optical probe, IRDye800CW-AE344, for fluorescence guided surgery (FGS). Methods: In the present study we characterized the fluorescent probe with regard to binding affinity, optical properties, and plasma stability. Further, in vivo imaging characterization was performed in nude mice with orthotopic human patient derived glioblastoma xenografts, and we performed head-to-head comparison within FGS between our probe and the traditional procedure using 5-ALA. Finally, the blood-brain barrier (BBB) penetration was characterized in a 3D BBB spheroid model. Results: The probe effectively visualized GBM in vivo with a tumor-to-background ratio (TBR) above 4.5 between 1 to 12 h post injection and could be used for FGS of orthotopic human glioblastoma xenografts in mice where it was superior to 5-ALA. The probe showed a favorable safety profile with no evidence of any acute toxicity. Finally, the 3D BBB model showed uptake of the probe into the spheroids indicating that the probe crosses the BBB. Conclusion: IRDye800CW-AE344 is a promising uPAR-targeted optical probe for FGS and a candidate for translation into human use.

Neuroendocrine neoplasms of the appendix: Characterization of 335 patients referred to the Copenhagen NET Center of Excellence.

INTRODUCTION: Neuroendocrine neoplasms (NEN) of the appendix are often incidentally discovered after appendectomy. Appropriate management is debated. The purpose was to characterize a cohort of 335 appendix NEN and evaluate the risk of recurrence. METHODS: Retrospective collection of data from 335 patients referred to the Neuroendocrine Tumor Center at Rigshospitalet 2000-2019. Appendix goblet cell carcinoids and mixed neuroendocrine non-neuroendocrine neoplasms were excluded. Patients were followed until December 31st, 2019. No patients were lost to follow-up. RESULTS: Sixty-three percent of the patients were female. The median (range) age at diagnosis was 34 (9-92) years. Median follow-up was 66 (1-250) months. Median tumor size was 7 (1-45) mm with 10 (3%) tumors >20 mm. In 18 specimens (5%) resection margins were positive. Mesoappendiceal invasion was found in 113 (35%). Sixty-three (19%) patients underwent right-sided completion hemicolectomy (RHC) after appendectomy according to ENETS guidelines. Among these, 11 (17%) had lymph node metastases in the resected tissue. Further, one patient who underwent initial RHC due to colonic adenocarcinoma had lymph node metastases. All lymph node metastases were detected in patients with serotonin positive tumors. No patients with glucagon positive tumors (n = 85) had lymph node metastases. Mesoappendiceal invasion >3 mm and positive resection margins were associated with presence of lymph node metastases. No recurrences were recorded. CONCLUSION: Following ENETS guidelines may lead to overtreatment of patients with respect to completion RHC. The risk of over- and undertreatment needs to be further evaluated.

Major driver mutations are shared between sinonasal intestinal-type adenocarcinoma and the morphologically identical colorectal adenocarcinoma.

PURPOSE: The purpose of our study was to compare genomic changes in sinonasal intestinal-type adenocarcinoma (sITAC) and colorectal adenocarcinoma (CRC), as they are histomorphologically indistinguishable. This can cause diagnostic difficulties as sinonasal tumours initially diagnosed as sITAC may represent metastasis from CRC, a frequent cancer. Previous studies have not uncovered the underlying mechanism behind the histomorphological resemblance. METHODS/PATIENTS: Tissue samples from all consecutive patients with sITAC at our facility (20 patients) were compared to samples from 20 patients with CRC as well as samples from 2 patients with both CRC and sinonasal tumours. DNA sequencing was performed using Illumina TruSight Oncology 500 panel consisting of 523 cancer-associated genes. Frequent mutations were inspected manually using the Integrative Genomics Viewer. RESULTS: Several well-known cancer-associated genes were mutated in the CRC group, but also in the sinonasal ITAC group. These genes included APC mutated in 65% of the CRC group and 37% of the sinonasal ITAC group, and TP53 mutated in 65% of CRC samples and 58% of ITAC samples. These shared mutations may explain the histomorphological similarities. Successful DNA sequencing was performed on the colorectal sample from one of the two patients with both CRC and sinonasal tumour. Comparing mutations in these samples from one patient we have shown that the sinonasal tumour in all probability was a CRC metastasis. CONCLUSION: We have identified several genetic similarities between sITAC and CRC. This discovery brings us closer to understanding mechanisms behind the development of sITAC-and hopefully in the future targeted therapy.

The macrophage activation marker soluble CD163 is elevated and associated with liver disease phenotype in patients with Wilson's disease.

BACKGROUND: Macrophages play a significant role in liver disease development and progression. The macrophage activation marker soluble (s)CD163 is associated with severity and prognosis in a number of different acute and chronic liver diseases but has been only sparsely examined in Wilson's disease (WD). We investigated sCD163 levels in patients with acute and chronic WD and hypothesized associations with liver disease phenotype and biochemical markers of liver injury. METHODS: We investigated sCD163 in two independent cohorts of WD patients: 28 patients with fulminant WD from the US Acute Liver Failure (ALF) Study Group registry and 147 patients with chronic disease from a German WD registry. We included a control group of 19 healthy individuals. Serum sCD163 levels were measured by ELISA. Liver CD163 expression was determined by immunohistochemistry. RESULTS: In the ALF cohort, median sCD163 was 10-fold higher than in healthy controls (14.6(2.5-30.9) vs. 1.5(1.0-2.7) mg/L, p < 0.001). In the chronic cohort, median sCD163 was 2.6(0.9-24.9) mg/L. There was no difference in sCD163 according to subgroups based on initial clinical presentation, i.e. asymptomatic, neurologic, hepatic, or mixed. Patients with cirrhosis at the time of diagnosis had higher sCD163 compared with those without cirrhosis (3.0(1.2-24.9) vs. 2.3(0.9-8.0) mg/L, p < 0.001); and both cohorts significantly lower than the ALF patients. Further, sCD163 correlated positively with ALT, AST, GGT and INR (rho = 0.27-0.53); and negatively with albumin (rho = - 0.37), (p ≤ 0.001, all). We observed immunohistochemical CD163 expression in liver tissue from ALF patients. CONCLUSIONS: Although sCD163 is not specific for WD, it was elevated in WD patients, especially in those with ALF. Further, sCD163 was higher in patients with cirrhosis compared to patients without cirrhosis and associated with biochemical markers of liver injury and hepatocellular function. Thus, macrophage activation is evident in WD and associates with liver disease phenotype and biochemical parameters of liver disease. Our findings suggest that sCD163 may be used as a marker of liver disease severity in WD patients.

A rare missense variant in APC interrupts splicing and causes AFAP in two Danish families.

BACKGROUND: We report the first case of a missense variant in the APC gene that interrupts splicing by creating a new cryptic acceptor site. The variant, c.289G>A, p.(Gly97Arg), is located in exon 3, and qualitative and semi-quantitative RNA splicing analysis reveal that the variant results in skipping of the last 70 nucleotides of the exon, which leads to the introduction of a frameshift and a premature stop codon. CASE PRESENTATION: The variant was detected in two, apparently unrelated, Danish families with an accumulation of colorectal cancers, colonic adenomas and other cancers. The families both have an attenuated familial adenomatous polyposis phenotype, which is consistent with the association of pathogenic variants in the 5' end of the gene.One variant-carrier also had Caroli Disease and a Caroli Disease associated hepatic mucinous cystadenocarcinoma. This is the first description of a person with both Caroli Disease and a pathogenic APC variant, and although the APC variant is not known to be connected to the development of the hepatic malformations in Caroli Disease, it remains unclear whether the variant could have contributed to the carcinogenesis of the liver tumour. CONCLUSIONS: Based on functional and co-segregation data we classify the APC c.289G>A, p.(Gly97Arg) variant as pathogenic (class 5). Our findings emphasize the importance of a functional evaluation of missense variants although located far from the exon-intron boundaries.

Hepatic epitheloid haemangioendothelioma: a rare malignant tumour.

Our case concerns a 66-year-old man. After experiencing recurrent episodes of abdominal pain, an initial CT scan, ultrasound and gastroscopy was carried out. All of which showed normal findings.As a consequence of persisting symptoms, another CT scan was performed. This scan revealed a hypodense area in the right lobe of the liver. This was interpreted as a possible haemangioma. Subsequent MRI scans indicated an intrahepatic cholangiocarcinoma. A final ultrasound-guided liver biopsy was performed and histology demonstrated epitheloid haemangioendothelioma, which was locally advanced and inoperable.

Quantitative perfusion assessment of intestinal anastomoses in pigs treated with glucagon-like peptide 2.

PURPOSE: Despite exhaustive research and improvement of techniques, anastomotic leakage remains a frequent complication in gastrointestinal surgery. As leakage is associated with poor perfusion, reliable objective methods to assess anastomotic perfusion are highly demanded. In addition, such methods enable evaluation of interventions that may improve anastomotic perfusion. Glucagon-like peptide 2 (GLP-2) is an enteroendocrine hormone that regulates mid-gut perfusion. In the present study, we aimed to explore if quantitative perfusion assessment with indocyanine green (q-ICG) could detect an increase in porcine anastomotic perfusion after treatment with GLP-2. METHODS: Nineteen pigs had two small bowel resections followed by anastomosis. Blinded to all investigators, animals were randomized to receive GLP-2 or placebo. Anastomotic perfusion was assessed at baseline, 30 min after injection of GLP-2/placebo, and after 5 days of treatment. Anastomotic strength and healing were evaluated by bursting pressure and histology. RESULTS: Q-ICG detected a significantly higher increase in anastomotic perfusion (p < 0.05) in animals treated with GLP-2, compared with placebo. No significant differences in anastomotic strength or healing were found. CONCLUSIONS: Q-ICG is a promising tool for perfusion assessment in gastrointestinal surgery and opens new opportunities in research of factors that may influence anastomotic healing, but further research is warranted to evaluate the effects of GLP-2 on anastomotic healing.

Medullary carcinoma of the colon: can the undifferentiated be differentiated?

Medullary carcinoma of the colon is a rare variant of colorectal cancer claimed to have a more favorable prognosis than conventional adenocarcinomas. The histopathologic appearance may be difficult to distinguish from poorly differentiated adenocarcinoma. The study aimed to evaluate the diagnostic interobserver agreement and to characterize the immunohistochemical and molecular differences between these two subgroups. Fifteen cases initially classified as medullary carcinoma and 30 cases of poorly differentiated adenocarcinomas were included. Two pathologists reviewed the slides independently without knowledge of the original diagnosis and subgrouped the tumors into the two entities. Agreement was reached in 31 of 45 cases (69 %) with kappa = 0.32. An extensive immunohistochemical panel was performed, and KRAS, NRAS, and BRAF mutational status was assessed. Of the 31 cases with diagnostic agreement, the expression of only MLH-1 along with corresponding expression of PMS-2 differed significantly (p = 0.04). A high rate of BRAF mutations was detected in both subgroups without significant differences. Expression of MLH-1 was superior in dividing the tumors into two separate entities with significant differences in CK20 (p = 0.005) expression and in the rate of BRAF mutations (p = 0.0035). In conclusion, medullary carcinomas of the colon are difficult to discriminate from poorly differentiated adenocarcinoma even with the help of immunohistochemical and molecular analyses. This raises the question whether these morphological subtypes should be maintained or whether an alternative classification of poorly differentiated colorectal adenocarcinomas based on MLH-1 status rather than morphology should be suggested.

AMACR is not applicable as a diagnostic tool in hepatocellular carcinoma.

Alpha-methylacyl coenzyme A racemase (AMACR or P504S) is a mitochondrial and peroxisomal protein present in a variety of human cells. Demonstration of increased expression is used diagnostically in prostatic adenocarcinoma. AMACR is also produced by normal hepatocytes and it has been postulated that the demonstration of AMACR expression or its pattern of distribution is useful in the diagnosis of hepatocellular carcinoma (HCC) (Jiang et al., Hum Pathol 2003;34, Guzman et al., Appl Immunohistochem Mol Morphol 2006;14, Li et al., J Exp Clin Cancer Res 2008;27). The aim of the present study was to evaluate whether immunohistochemical staining for AMACR can be used in a routine histopathologic setting. Immunohistochemical staining for AMACR was performed on paraffin-embedded tissue from livers resected for HCC during 1980-2006 at Rigshospitalet, Copenhagen, Denmark (n = 44). Tumor sections as well as surrounding non-neoplastic tissues were studied. In both tumor and non-tumor tissues, intracellular localization and staining pattern were assessed and the staining intensity of AMACR was graded. The fraction of stained tumor cells was not significantly different from that of stained non-tumor cells in the same patients (p = 0.97). A significantly lower staining intensity was observed in clear cell areas (p = 0.005), but the AMACR expression did not correlate with the HCC type and could not distinguish neoplastic from non-neoplastic liver cells. AMACR is not applicable as a tool in the histopathologic diagnosis of HCC.

Intraoperative immunohistochemistry staining of sentinel nodes in breast cancer: clinical and economical implications.

The study aimed to evaluate intraoperative immunohistochemistry (IHC) staining of sentinel nodes in primary breast cancer surgery. We analysed retrospectively 1209 consecutive sentinel node procedures and compared the rate of late positive metastases in sentinel node biopsy (SNB) and the duration of the surgical procedures before (n=706) and after (n=503) introducing intraoperative IHC on frozen section. We also did a cost analysis. Intraoperative IHC staining led to a lowering of the late positive SNB rate. Introducing IHC gave a decrease in the late positive rate from 93 to 52% (p<0.0001) for isolated tumour cell metastasis, from 56 to 36.4% (p<0.02) for micrometastasis, and from 16 to 5% (p<0.01) for macrometastasis. The surgical procedures were slightly prolonged for lumpectomies but not for mastectomies after introducing intraoperative IHC staining. The cost analysis showed an overall cost saving of approximately 40%. In conclusion, intraoperative IHC staining of the SNB lowered the late positive rate and gave an overall cost saving.