Comprehensive Molecular Portraits of Invasive Lobular Breast Cancer.

Invasive lobular carcinoma (ILC) is the second most prevalent histologic subtype of invasive breast cancer. Here, we comprehensively profiled 817 breast tumors, including 127 ILC, 490 ductal (IDC), and 88 mixed IDC/ILC. Besides E-cadherin loss, the best known ILC genetic hallmark, we identified mutations targeting PTEN, TBX3, and FOXA1 as ILC enriched features. PTEN loss associated with increased AKT phosphorylation, which was highest in ILC among all breast cancer subtypes. Spatially clustered FOXA1 mutations correlated with increased FOXA1 expression and activity. Conversely, GATA3 mutations and high expression characterized luminal A IDC, suggesting differential modulation of ER activity in ILC and IDC. Proliferation and immune-related signatures determined three ILC transcriptional subtypes associated with survival differences. Mixed IDC/ILC cases were molecularly classified as ILC-like and IDC-like revealing no true hybrid features. This multidimensional molecular atlas sheds new light on the genetic bases of ILC and provides potential clinical options.

SALL4 promotes tumor progression in breast cancer by targeting EMT.

Sal-like protein 4 (SALL4) is overexpressed in breast cancer and might contribute to breast cancer progression, but the molecular mechanism remains unknown. Here, we found that within a group of 371 ethnic Chinese breast cancer patients, SALL4 was associated with lower grade (P = .002) and progesterone receptor positivity (P = .004) for overall cases; lower Ki67 (P = .045) and high vimentin (P = .007) for luminal cases. Patients with high SALL4 expression in lymph node metastasis showed a significantly worse survival than those with low expression. Knockout of SALL4 in a triple-negative breast cancer cell line MDA-MB-231-Red-FLuc-GFP led to suppressed ability in proliferation, clonogenic formation, migration, and mammosphere formation in vitro, tumorigenicity and lung colonization in vivo. On the other hand, overexpression of SALL4 enhanced migration and mammosphere formation in vitro and tumorigenicity in vivo. Mechanistically, there was a positive correlation between SALL4 expression and mesenchymal markers including Zinc finger E-box binding homeobox 1 (ZEB1), vimentin, Slug, and Snail in vivo. Chromatin immunoprecipitation experiment indicated that SALL4 can bind to the promoter region of vimentin (-778 to -550 bp). Taken together, we hypothesize that SALL4 promotes tumor progression in breast cancer by inducing the mesenchymal markers like vimentin through directly binding to its promoter. Increased SALL4 level in metastatic lymph node relative to the primary site is an important poor survival marker in breast cancer.

Diagnostic upgrade of atypical ductal hyperplasia of the breast based on evaluation of histopathological features and calcification on core needle biopsy.

AIMS: Atypical ductal hyperplasia (ADH) of breast is increasingly diagnosed in core needle biopsy (CNB). As higher-grade lesions were found in the excision in a substantial proportion of ADH on CNB, factors predicting risk of subsequent upgrade are clinically significant. This study aims to investigate relevant histopathological factors in CNB that could predict diagnostic upgrade at excision. METHODS AND RESULTS: One hundred and forty-three cases of CNB with paired subsequent excision were evaluated for multiple clinicopathological parameters related to CNB sampling, ADH morphology, calcification and other co-existing histological features, and which of these parameters were associated with diagnostic upgrade at subsequent excisions were determined. Forty-eight cases (34.3%) were upgraded to malignancy, including 15 invasive cancers and 33 ductal carcinomas in situ (DCIS). An increased tissue area occupied by ADH (P = 0.026), a higher number of ADH foci (P = 0.004), the presence of solid pattern (P = 0.037) and older age (P = 0.012) were positively associated with upgrade, while negative associations were found with the presence of micropapillary pattern (P = 0.025), co-existing columnar cell lesions (CCL) (P = 0.001) and the presence of calcifications (P = 0.009). Multivariate logistic regression analysis showed that the number of ADH foci (HR = 2.810, P = 0.013) was an independent positive predictor, while co-existing CCL (HR = 0.391, P = 0.013) was an independent negative predictor for upgrade. CONCLUSIONS: Patients with ADH in CNB showing the presence of co-existing CCL and a lower number of ADH foci have a lower risk of disease upgrade at excision, and are potential candidates for observation-only management.

Amyloid Precursor Protein Is Associated with Aggressive Behavior in Nonluminal Breast Cancers.

BACKGROUND: ß-amyloid precursor protein (APP), a potential target for Alzheimer's disease treatment, has recently been shown to take part in carcinogenesis. Increased APP promotes migration, survival, and proliferation in breast cancer cell lines. We examined the clinical value of APP in breast cancers. A comprehensive examination of clinicopathological features related to APP expression in a large cohort of breast cancers and the corresponding metastatic lymph nodes was performed. APP expression and its prognostic impact in different breast cancer subtypes were examined. RESULTS: APP was highly expressed in nonluminal breast cancers and correlated with features associated with nonluminal breast cancers (including higher grade, the presence of necrosis, and higher proliferative index, growth factor receptor, and basal marker expression). Multivariate Cox hazard analysis demonstrated that APP was an independent adverse prognostic factor of disease-free survival (DFS; hazard ratio [HR], 2.090; p = .013; 95% confidence interval [CI], 1.165-3.748) and breast cancer-specific survival (BCSS; HR, 2.631; p = .002; 95% CI, 1.408-4.915) in the nonluminal group. The independent prognostic impact was also seen in triple negative breast cancers. Interestingly, a higher expression of APP was found in nodal metastasis compared with primary tumor. Such APP upregulation was correlated with further distal metastasis and poorer outcome (DFS: log-rank, 12.848; p < .001; BCSS: log-rank, 13.947; p < .001). CONCLUSION: Our findings provided evidence of oncogenic roles of APP in clinical breast cancers. Patients with positive APP expression, particularly those with APP upregulation in lymph node metastases, may require vigilant monitoring of their disease and more aggressive therapy. IMPLICATIONS FOR PRACTICE: ß-amyloid precursor protein (APP), a potential target for Alzheimer's disease, has recently been implicated in oncogenesis. Here, evidence of its roles in clinical breast cancers is provided. Positive APP expression was found to be an independent prognostic factor in nonluminal cancers, particularly triple negative breast cancers (TNBCs). Interestingly, a higher APP in nodal metastases was associated with distal metastases. TNBCs are heterogeneous and currently have no available target therapy. APP could have therapeutic potential and be used to define the more aggressive cases in TNBCs. Current prognostic analysis is based on primary tumor. The present data suggest that investigation of nodal metastases could provide additional prognostic value.

CD117 expression in breast phyllodes tumors correlates with adverse pathologic parameters and reduced survival.

CD117 (c-kit) is a type III receptor tyrosine kinase encoded by the KIT gene. Deregulation of expression and mutations in the gene are implicated in various tumors. Reports of CD117 expression in phyllodes tumors have been controversial. We aim to investigate the protein expression of CD117 and mutations in the KIT gene in phyllodes tumors, and correlate the findings with pathological parameters and clinical outcome. A total of 272 cases were included in this study. CD117 expression was investigated by immunohistochemistry on tissue microarray sections. Toluidine blue staining was performed to indicate mast cells. Overall, 28 (10%) cases were CD117 positive. CD117 expression was significantly associated with tumor grade (P<0.001), increased stromal hypercellularity (P=0.003), stromal atypia (P=0.01), and stromal mitotic activity (P<0.001), permeative microscopic margins (P=0.002), and presence of hemorrhage (P=0.001). Expression was also associated with poorer overall survival (P=0.003). Nineteen cases were further selected for mutation screening through the Affymetrix OncoScan platform. No mutation of the KIT gene was found. Despite a lack of mutations in the KIT gene, CD117 protein expression is associated with unfavorable pathological parameters and poorer prognosis, suggesting an underlying role in the biology of phyllodes tumors.

Predictors of an informative or actionable cytological diagnosis on repeat breast aspiration after an insufficient aspirate: a multicentre retrospective review.

An insufficient/inadequate diagnosis on fine-needle aspiration cytology (FNAC) of the breast is not an uncommon diagnostic dilemma. This study aims to review the rate and clinical features predicting an informative or actionable diagnosis on repeating breast aspiration after an insufficient aspirate. METHODS: Unsatisfactory/insufficient/inadequate or equivalent breast aspirates were retrieved from the involved institutions, and those with a repeat aspiration performed within 365 days were included. Clinical and radiological information were retrieved. Available cytological slides were reviewed. RESULTS: Totally 539 paired aspirates were retrieved, with 61.2% (n=330/539) and 10.9% (n=59/539) cytological diagnosis being informative (not insufficient) and actionable (not insufficient nor benign) on repeat aspiration. Younger age (p=0.005) was associated with an informative diagnosis and prior radiotherapy (p=0.097) and insufficient aspirates performed under free-hand (p=0.097) trended with an actionable diagnosis. Radiological findings of calcification (p=0.026) and hyperechogenicity (p=0.045), a small lesion size on initial (p=0.037) and repeat (p=0.059) radiological assessment and interval size increment (p=0.019) correlated with informative/actionable diagnoses. Cytomorphological parameters, except for a trend with crushing artefact (p=0.063), do not correlate with the cytologic diagnosis of the repeat aspirate. CONCLUSIONS: Repeating breast FNAC on patients after an insufficient diagnosis yields an informative ('sufficient') result in over 60% of cases. Small lesions with calcification, hyperechogenicity and/or interval size increment are more likely to yield diagnostic results on repeat aspiration and indicate select patients suitable for repeat FNAC over more invasive procedures. The lack of associations with cytomorphological parameters cautions against overinterpretation of insufficient breast aspirates.

The nerve cells in gastrointestinal cancers: from molecular mechanisms to clinical intervention.

Gastrointestinal (GI) cancer is a formidable malignancy with significant morbidity and mortality rates. Recent studies have shed light on the complex interplay between the nervous system and the GI system, influencing various aspects of GI tumorigenesis, such as the malignance of cancer cells, the conformation of tumor microenvironment (TME), and the resistance to chemotherapies. The discussion in this review first focused on exploring the intricate details of the biological function of the nervous system in the development of the GI tract and the progression of tumors within it. Meanwhile, the cancer cell-originated feedback regulation on the nervous system is revealed to play a crucial role in the growth and development of nerve cells within tumor tissues. This interaction is vital for understanding the complex relationship between the nervous system and GI oncogenesis. Additionally, the study identified various components within the TME that possess a significant influence on the occurrence and progression of GI cancer, including microbiota, immune cells, and fibroblasts. Moreover, we highlighted the transformation relationship between non-neuronal cells and neuronal cells during GI cancer progression, inspiring the development of strategies for nervous system-guided anti-tumor drugs. By further elucidating the deep mechanism of various neuroregulatory signals and neuronal intervention, we underlined the potential of these targeted drugs translating into effective therapies for GI cancer treatment. In summary, this review provides an overview of the mechanisms of neuromodulation and explores potential therapeutic opportunities, providing insights into the understanding and management of GI cancers.

Cellular zinc metabolism and zinc signaling: from biological functions to diseases and therapeutic targets.

Zinc metabolism at the cellular level is critical for many biological processes in the body. A key observation is the disruption of cellular homeostasis, often coinciding with disease progression. As an essential factor in maintaining cellular equilibrium, cellular zinc has been increasingly spotlighted in the context of disease development. Extensive research suggests zinc's involvement in promoting malignancy and invasion in cancer cells, despite its low tissue concentration. This has led to a growing body of literature investigating zinc's cellular metabolism, particularly the functions of zinc transporters and storage mechanisms during cancer progression. Zinc transportation is under the control of two major transporter families: SLC30 (ZnT) for the excretion of zinc and SLC39 (ZIP) for the zinc intake. Additionally, the storage of this essential element is predominantly mediated by metallothioneins (MTs). This review consolidates knowledge on the critical functions of cellular zinc signaling and underscores potential molecular pathways linking zinc metabolism to disease progression, with a special focus on cancer. We also compile a summary of clinical trials involving zinc ions. Given the main localization of zinc transporters at the cell membrane, the potential for targeted therapies, including small molecules and monoclonal antibodies, offers promising avenues for future exploration.

Phyllodes tumours of the breast: the role of CD34, vascular endothelial growth factor and ß-catenin in histological grading and clinical outcome.

AIMS: The grading and prognostication of breast phyllodes tumours remain challenging, and the value of biological markers continues to be elusive. The aim of this study was to evaluate CD34, vascular endothelial growth factor (VEGF) and ß-catenin in a series of 185 breast phyllodes tumours comprising 120 benign, 48 borderline and 17 malignant lesions. METHODS AND RESULTS: Immunohistochemistry on tissue microarrays of phyllodes tumours was performed. CD34, VEGF and ß-catenin in stromal cells were expressed, respectively, in: 38.3%, 29.2% and 27.5% of benign phyllodes tumours; 33.3%, 58.3% and 54.2% of borderline phyllodes tumours; and 5.9%, 64.7% and 76.5% of malignant phyllodes tumours; these associations with histological grade were statistically significant. There was a statistically significant inverse association of CD34 stromal expression with adverse histological features, and a positive correlation of VEGF and cytoplasmic ß-catenin stromal staining with unfavourable microscopic parameters. At a median follow-up duration of 42 months, 11 women suffered recurrences, with three succumbing from phyllodes tumour. Patients whose tumours expressed VEGF had poorer overall survival (P = 0.033), and there was a trend for worse overall survival in patients with tumour ß-catenin cytoplasmic expression. CONCLUSIONS: CD34, VEGF and ß-catenin play biological roles in breast phyllodes tumours, and may provide insights into tumour progression, with differential expression accompanying higher grades.

Exploring the Microbiome in Gastric Cancer: Assessing Potential Implications and Contextualizing Microorganisms beyond H. pylori and Epstein-Barr Virus.

While previous research has primarily focused on the impact of H. pylori and Epstein-Barr virus (EBV), emerging evidence suggests that other microbial influences, including viral and fungal infections, may also contribute to gastric cancer (GC) development. The intricate interactions between these microbes and the host's immune response provide a more comprehensive understanding of gastric cancer pathogenesis, diagnosis, and treatment. The review highlights the roles of established players such as H. pylori and EBV and the potential impacts of gut bacteria, mainly Lactobacillus, Streptococcus, hepatitis B virus, hepatitis C virus, and fungi such as Candida albicans. Advanced sequencing technologies offer unprecedented insights into the complexities of the gastric microbiome, from microbial diversity to potential diagnostic applications. Furthermore, the review highlights the potential for advanced GC diagnosis and therapies through a better understanding of the gut microbiome.

H. pylori-induced NF-κB-PIEZO1-YAP1-CTGF axis drives gastric cancer progression and cancer-associated fibroblast-mediated tumour microenvironment remodelling.

BACKGROUND: Gastric cancer (GC) is one of the most common tumours in East Asia countries and is associated with Helicobacter pylori infection. H. pylori utilizes virulence factors, CagA and VacA, to up-regulate pro-inflammatory cytokines and activate NF-κB signaling. Meanwhile, the PIEZO1 upregulation and cancer-associated fibroblast (CAF) enrichment were found in GC progression. However, the mechanisms of PIEZO1 upregulation and its involvement in GC progression have not been fully elucidated. METHODS: The CAF enrichment and clinical significance were investigated in animal models and primary samples. The expression of NF-κB and PIEZO1 in GC was confirmed by immunohistochemistry staining, and expression correlation was analysed in multiple GC datasets. GSEA and Western blot analysis revealed the YAP1-CTGF axis regulation by PIEZO1. The stimulatory effects of CTGF on CAFs were validated by the co-culture system and animal studies. Patient-derived organoid and peritoneal dissemination models were employed to confirm the role of the PIEZO1-YAP1-CTGF cascade in GC. RESULTS: Both CAF signature and PIEZO1 were positively correlated with H. pylori infection. PIEZO1, a mechanosensor, was confirmed as a direct downstream of NF-κB to promote the transformation from intestinal metaplasia to GC. Mechanistic studies revealed that PIEZO1 transduced the oncogenic signal from NF-κB into YAP1 signaling, a well-documented oncogenic pathway in GC progression. PIEZO1 expression was positively correlated with the YAP1 signature (CTGF, CYR61, and c-Myc, etc.) in primary samples. The secreted CTGF by cancer cells stimulated the CAF infiltration to form a stiffened collagen-enrichment microenvironment, thus activating PIEZO1 to form a positive feedback loop. Both PIEZO1 depletion by shRNA and CTGF inhibition by Procyanidin C1 enhanced the efficacy of 5-FU in suppressing the GC cell peritoneal metastasis. CONCLUSION: This study elucidates a novel driving PIEZO1-YAP1-CTGF force, which opens a novel therapeutic avenue to block the transformation from precancerous lesions to GC. H. pylori-NF-κB activates the PIEZO1-YAP1-CTGF axis to remodel the GC microenvironment by promoting CAF infiltration. Targeting PIEZO1-YAP1-CTGF plus chemotherapy might serve as a potential therapeutic option to block GC progression and peritoneal metastasis.

The molecular classification of cancer-associated fibroblasts on a pan-cancer single-cell transcriptional atlas.

BACKGROUND: Cancer-associated fibroblasts (CAFs), integral to the tumour microenvironment, are pivotal in cancer progression, exhibiting either pro-tumourigenic or anti-tumourigenic functions. Their inherent phenotypic and functional diversity allows for the subdivision of CAFs into various subpopulations. While several classification systems have been suggested for different cancer types, a unified molecular classification of CAFs on a single-cell pan-cancer scale has yet to be established. METHODS: We employed a comprehensive single-cell transcriptomic atlas encompassing 12 solid tumour types. Our objective was to establish a novel molecular classification and to elucidate the evolutionary trajectories of CAFs. We investigated the functional profiles of each CAF subtype using Single-Cell Regulatory Network Inference and Clustering and single-cell gene set enrichment analysis. The clinical relevance of these subtypes was assessed through survival curve analysis. Concurrently, we employed multiplex immunofluorescence staining on tumour tissues to determine the dynamic changes of CAF subtypes across different tumour stages. Additionally, we identified the small molecule procyanidin C1 (PCC1) as a target for matrix-producing CAF (matCAF) using molecular docking techniques and further validated these findings through in vitro and in vivo experiments. RESULTS: In our investigation of solid tumours, we identified four molecular clusters of CAFs: progenitor CAF (proCAF), inflammatory CAF (iCAF), myofibroblastic CAF (myCAF) and matCAF, each characterised by distinct molecular traits. This classification was consistently applicable across all nine studied solid tumour types. These CAF subtypes displayed unique evolutionary pathways, functional roles and clinical relevance in various solid tumours. Notably, the matCAF subtype was associated with poorer prognoses in several cancer types. The targeting of matCAF using the identified small molecule, PCC1, demonstrated promising antitumour activity. CONCLUSIONS: Collectively, the various subtypes of CAFs, particularly matCAF, are crucial in the initiation and progression of cancer. Focusing therapeutic strategies on targeting matCAF in solid tumours holds significant potential for cancer treatment.

Primary nasopharyngeal carcinoma: diagnostic accuracy of MR imaging versus that of endoscopy and endoscopic biopsy.

PURPOSE: To compare the accuracy of magnetic resonance (MR) imaging with that of the current clinical standard of endoscopy and endoscopic biopsy, to determine whether MR imaging depicts subclinical cancers missed at endoscopy and endoscopic biopsy, and to determine whether MR imaging can identify patients without nasopharyngeal carcinoma (NPC) who do not need to undergo invasive sampling biopsy. MATERIALS AND METHODS: The study protocol was approved by the institutional review board; written informed consent was obtained from all patients. Patients suspected of having NPC underwent MR imaging, endoscopy, and endoscopic biopsy. Endoscopic biopsy targeted the suspected tumor or sampled the endoscopically normal nasopharynx. The final diagnosis was based on results of the endoscopic biopsy or on results of a repeat biopsy directed at the lesion detected at MR imaging. The sensitivity and specificity of the three investigations were compared by using the Fisher exact test. RESULTS: NPC was present in 77 (31%) of 246 patients and absent in 169 (69%) patients. The combined sensitivity, specificity, and accuracy, respectively, were 100%, 93%, and 95% for MR imaging, 90%, 93%, and 92% for endoscopy, and 95%, 100%, and 98% for endoscopic biopsy. Benign disease was mistaken for NPC in 12 (7%) of 169 patients at MR imaging and in 11 (6%) patients at endoscopy. The sensitivity of MR imaging was significantly higher than that of endoscopy (P = .006) and was similar to that of endoscopic biopsy (P = .120). The specificity of MR imaging was similar to that of endoscopy (P = .120) and was significantly lower than that of endoscopic biopsy (P < .001). CONCLUSION: MR imaging is an accurate test for the diagnosis of NPC. MR imaging depicts subclinical cancers missed at endoscopy and endoscopic biopsy and helps identify the majority of patients who do not have NPC and who therefore do not need to undergo invasive sampling biopsies.

Pretreatment and early intratreatment prediction of clinicopathologic response of head and neck cancer to chemoradiotherapy using 1H-MRS.

PURPOSE: To determine if choline (cho) identified by proton magnetic resonance spectroscopy ((1)H-MRS) performed pretreatment and early in the course of treatment predicts clinicopathologic response of head and neck squamous cell carcinoma (HNSCC). MATERIALS AND METHODS: In all, 60 patients with HNSCC scheduled to undergo concurrent chemoradiotherapy or radiotherapy alone were recruited. (1)H-MRS was performed pretreatment and early intratreatment (2 weeks after start of treatment). Cho:creatine and cho:water ratios at each timepoint and change in the ratios between the two timepoints were correlated with locoregional failure, distant metastases, overall survival, and cancer-related death. Statistical analysis was performed using logistic regression and chi-square and a P-value of < 0.05 was considered statistically significant. RESULTS: Cho was identified in 47/49 successful pretreatment spectra and 42 of these 47 underwent successful (1)H-MRS early intratreatment, of which 21 showed persistent cho. Locoregional failure occurred in 15, distant metastases in 6, and death in 15 patients; the follow-up period in survivors ranged from 13-64 months (mean, 39 months). No statistically significant correlation was found between (1)H-MRS parameters and clinical endpoints. CONCLUSION: The pretreatment cho and change in cho early during a course of treatment did not predict clinical outcome.

Squamous cell carcinoma of the head and neck: diffusion-weighted MR imaging for prediction and monitoring of treatment response.

OBJECTIVE: To investigate the role of diffusion-weighted imaging (DWI) in predicting and monitoring chemoradiotherapy response in head and neck squamous cell carcinoma (HNSCC). METHODS: Diffusion-weighted imaging was performed pre-treatment (n = 50), intra-treatment (n = 41) and post-treatment (n = 20). Apparent diffusion coefficient (ADC) values were correlated with locoregional failure (LF). RESULTS: Locoregional failure occurred in 20/50 (40%) patients. A significant correlation was found between LF and post-treatment ADC (p = 0.02) but not pre- or intra-treatment ADC. Serial change in ADC was even more significant (p = 0.00001), using a fall in ADC early (pre- to intra-treatment) or late (intra- to post-treatment) to indicate LF, achieved 100% specificity, 80% sensitivity and 90% accuracy. CONCLUSIONS: Single ADC measurements pre- or intra-treatment did not predict response, but ADC post-treatment was a marker for LF. Serial change in ADC was an even stronger marker, when using an early or late treatment fall in ADC to identify LF.

Monitoring of treatment response after chemoradiotherapy for head and neck cancer using in vivo 1H MR spectroscopy.

Elevated choline (Cho) level has been documented on proton magnetic resonance spectroscopy ((1)H MRS) in head and neck squamous cell carcinoma and therefore percentage changes in Cho levels after chemoradiotherapy may serve as a marker of residual cancer in a post-treatment mass (PTM). Forty-six patients underwent (1)H MRS before treatment and the 30 patients with a PTM underwent repeat (1)H MRS at 6 weeks post-treatment. The percentage change in Cho/creatine and Cho/water ratios were correlated with residual cancer. The mean pretreatment Cho/creatine and Cho/water ratios were 2.24 and 1.20 x 10(-3), respectively. Cho persisted in four out of nine PTMs with residual cancer. Cho was absent in five out of nine PTMs with residual cancer and 21/21 PTMs without cancer. The number of PTMs with persistent Cho was too small to allow analysis of percentage change in ratios but the presence of Cho in a PTM showed significant correlation with residual cancer (p = 0.0046), producing a sensitivity, specificity, positive predictive value and negative predictive value of 44%, 100%, 100% and 81%, respectively. Therefore, the presence of Cho in a PTM may serve as a marker of residual cancer. Furthermore since so few PTMs contain Cho, a percentage change in Cho ratios may not be a useful method for monitoring treatment response.

Apoptotic and necrotic effects of tumour necrosis factor-alpha potentiated with hyperthermia on L929 and tumour necrosis factor-alpha-resistant L929.

PURPOSE: The cytotoxic effect of the combination treatment of TNF-alpha and hyperthermia on L929 and TNF-alpha-resistant L929 (rL929) cells was investigated. MATERIALS AND METHODS: L929 cells were treated with TNF-alpha (5 ng/mL), heating at 43 degrees C or the combination of TNF-alpha and heating. The cells were harvested at different time within the 24-hour period. The viability and the type of cell death of the harvested cells were examined. RESULTS: When L929 cells were treated with a combination of TNF-alpha and heating the cells died quickly and apoptosis increased to an overwhelming extent, especially in the group pre-treated with TNF-alpha for 1 h prior to heating. Although rL929 cells were resistant to TNF-alpha alone, the cells became sensitive to TNF-alpha treatment when combined with heating. Similar to the L929 cell, the cells also died rapidly and exhibited apoptosis to a higher extent. Using an Annexin-V-FITC kit and flow cytometer, we found that both necrosis and apoptosis occurred. Agarose gel electrophoresis of DNA extracted from treated cells showed that the DNA fragments were multiples of approximately 200 bp. Furthermore, by studying the kinetics of cell death and apoptosis, we found that the loss of cell membrane integrity preceded the DNA fragmentation in both L929 and rL929 cells. CONCLUSION: The results suggested that hyperthermia may enhance the necrotic and apoptotic effects of TNF-alpha on some tumour cells and overcome the resistance of some tumour cells to TNF-alpha.

Increased expression of centrosomal alpha, gamma-tubulin in atypical ductal hyperplasia and carcinoma of the breast.

Centrosomal abnormalities have been found in various cancer types. We sought to determine whether centrosomal dysfunctions occur in the atypical ductal hyperplasia (ADH)-carcinoma sequence of breast cancer. As alpha and gamma-tubulins are the structural components of centrosomes, we performed real time quantitative polymerase chain reaction (qPCR), in situ hybridization (ISH) and immunnohistochemistry (IHC) to determine the DNA copy levels, messenger RNA (mRNA) expression, and protein expression of alpha and gamma-tubulins respectively. Gamma-tubulin staining was used for the localization and quantification of centrosomes. We found that alpha-tubulin or gamma-tubulin mRNA was increasingly expressed from normal breast tissue (NBT) to ADH, ductal carcinoma in situ (DCIS), and infiltrative ductal carcinoma (IDC), respectively, with the highest expressions being found in DCIS. The expression profiles of alpha, gamma-tubulin proteins were concordant with that of mRNA, except that the highest expression was found in IDC. Similarly, DNA copies of alpha, gamma-tubulins showed a rising tendency, with the highest level for gamma-tubulin attained in IDC and that for alpha-tubulin was found in DCIS. However, there was no significant difference of alpha, gamma-tubulin DNA copy levels, mRNA expression, and protein expression between DCIS and IDC. Our results demonstrate that centrosomal aberrations may play key roles in the early stage of breast tumorogenesis. The malignant transformation sequence is probably attributable to the amplification of centrosomal DNA leading to mRNA and protein over-expression of these centrosomal proteins. Furthermore, determination of alpha, gamma-tubulins using combined qPCR with ISH may be useful in assisting the diagnosis of premalignant lesions of the breast.

Increased epidermal growth factor receptor (EGFR) expression in malignant mammary phyllodes tumors.

Mammary phyllodes tumors are uncommon stromal-epithelial neoplasms, and are divided into benign, borderline malignant and frankly malignant groups on the basis of their histological features. Accumulating evidence shows that epidermal growth factor receptor (EGFR) is involved in the pathogenesis and progression of many malignancies. This study investigated 453 phyllodes tumors (296 benign, 98 borderline, 59 malignant) for EGFR expression using immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) for gene amplification. The staining was correlated to tumor margin status, degree of malignancy, stromal cellularity, mitotic activity, nuclear pleomorphism and stromal overgrowth. Cases with strong positive IHC staining were selected for FISH. The overall positive rate for EGFR was 16.2% (48/296), 30.6% (30/98) and 56% (33/59) for benign, borderline malignant and frankly malignant phyllodes tumors, respectively. FISH demonstrated egfr gene amplification in 8% of immunohistochemically positive cases. The results of this study provide strong evidence that EGFR overexpression is involved in the pathogenesis of phyllodes tumors, although gene amplification may not be the major underlying mechanism for overexpression.

Mucin extravasation in breast core biopsies--clinical significance and outcome correlation.

AIMS: To document the spectrum of lesions associated with mucin extravasation (ME) in breast core biopsy specimens, and to correlate with open surgical excisions. METHODS AND RESULTS: Thirty-nine lesions in 37 women with ME on core biopsies constituted the study group. Fibrocystic change (FC), atypical ductal hyperplasia (ADH) and ductal carcinoma in situ (DCIS) were found in 21 (53.8%), 13 (33.3%) and four (10.3%) core biopsy specimens, respectively, with one (2.6%) consisting only of mucin pools. Except for the latter, all disclosed mucocoele-like lesions (MLL) accompanying ME. Columnar cell lesions (CCL) were frequently observed (84.6%). On open biopsy, three cases underdiagnosed on core biopsy included FC that later disclosed ADH; one ADH lesion on core later upgraded to DCIS; and a case of mucin pools that revealed mucinous carcinoma on excision. The extent of CCL on core biopsy appeared to predict sinister lesions on open excision. For calcified lesions that were completely removed on core biopsy, there were no malignant lesions discovered on open excision that had not already been diagnosed preoperatively. CONCLUSIONS: ME and MLL on core biopsy warrant close radiological-pathological correlation. When the entire radiological abnormality has been removed with large core mammotome biopsy specimens, surgery may potentially be avoided in histologically benign lesions, although such an approach requires further validation.