Michael Z. Gilcrease

Smoke exposure, histologic type and geography-related differences in the methylation profiles of non-small cell lung cancer.

Aberrant methylation of several known or putative tumor suppressor genes occurs frequently during the pathogenesis of lung cancers. There are major smoke exposure, histology, geography and gender‐related changes in non‐small cell lung cancer (NSCLC). We investigated smoking‐related, histologic, geographic and gender differences in the methylation profiles of resected NSCLCs. We examined 514 cases of NSCLC and 84 corresponding nonmalignant lung tissues from 4 countries (USA, Australia, Japan and Taiwan) for the methylation status of 7 genes known to be frequently methylated in lung cancers [p16, RASSF1A (RAS association domain family 1), APC, RARβ, CDH13, MGMT and GSTP1]. Multivariate analyses were used for data analysis. Adenocarcinoma was the major histologic type in women and never smokers; analyses that involved smoke exposure and gender were limited to this histology. Our major findings are a) methylation status of any single gene was largely independent of methylation status of other genes; b) the rates of methylation of p16 and APC and the mean Methylation Index (MI), a reflection of the overall methylation status, were significantly higher in ever smokers than in never smokers; c) the mean MI of tumors arising in former smokers was significantly lower than the mean of current smokers; d) the methylation rates of APC, CDH13 and RARβ were significantly higher in adenocarcinomas than in squamous cell carcinomas; e) methylation rates of MGMT and GSTP1 were significantly higher in the USA and Australian cases than in those from Japan and Taiwan; and (f) no significant gender‐related differences in methylation patterns were noted. Our findings demonstrate important smoke exposure, histologic type and geography‐related differences in the methylation profiles of NSCLC tumors.

Synchronous Overexpression of Epidermal Growth Factor Receptor and HER2-neu Protein Is a Predictor of Poor Outcome in Patients with Stage I Non-Small Cell Lung Cancer

Purpose: Despite maximal therapy, surgically treated patients with stage I non-small cell lung cancer (NSCLC) are at risk for developing metastatic disease. Histopathologic findings cannot adequately predict disease progression, so there is a need to identify molecular factors that serve this purpose. Because the ErbB receptors play an important role in lung cancer progression, we analyzed the expression of epidermal growth factor receptor (EGFR), phosphorylated EGFR, transforming growth factor α (TGFα), and HER2-neu as potential prognostic factors in stage I NSCLC. Experimental Design: Using immunohistochemical techniques, we retrospectively analyzed formalin-fixed, paraffin-embedded samples from 111 patients with resected pathological stage I NSCLC. Then we correlated these data with patient clinical outcome. Results: Median follow-up was 69.3 months. EGFR overexpression (defined as >10% membranous staining) was found in 66 tumors (59.5%). It was significantly more common in T2 tumors than in T1 tumors (P = 0.001), and in more squamous cell carcinomas than in adenocarcinomas (P = 0.07). HER2-neu overexpression was found in 19 tumors (17.1%) and was significantly more common in adenocarcinomas than in squamous cell carcinomas (P = 0.035). Synchronous overexpression of EGFR and HER2-neu was found in 11 tumors (9.9%). Patients with these tumors had a significantly shorter time to recurrence (P = 0.006) and a trend toward shorter overall survival (P = 0.093). Phosphorylated EGFR and transforming growth factor α were detected but were not related to prognosis. Conclusions: Synchronous overexpression of EGFR and HER2-neu at the protein level predicts increased recurrence risk and may predict decreased survival in patients with stage I NSCLC. This suggests that important interactions take place among the different members of the ErbB family during tumor development and suggests a method for choosing targeted therapy. A prospective study is planned.

Feasibility and Accuracy of Sentinel Lymph Node Biopsy After Preoperative Chemotherapy in Breast Cancer Patients With Documented Axillary Metastases

The feasibility and accuracy of sentinel lymph node (SLN) biopsy in patients with breast cancer after preoperative chemotherapy has been demonstrated in a number of large, single‐institution studies. However, a relative contraindication to SLN biopsy after preoperative chemotherapy is the presence of axillary metastases at initial diagnosis. The objective of this study was to determine the feasibility and accuracy of SLN biopsy after preoperative chemotherapy in patients with documented axillary metastases at presentation.

Improved Axillary Evaluation Following Neoadjuvant Therapy for Patients With Node-Positive Breast Cancer Using Selective Evaluation of Clipped Nodes: Implementation of Targeted Axillary Dissection

PURPOSE Placing clips in nodes with biopsy-confirmed metastasis before initiating neoadjuvant therapy allows for evaluation of response in breast cancer. Our goal was to determine if pathologic changes in clipped nodes reflect the status of the nodal basin and if targeted axillary dissection (TAD), which includes sentinel lymph node dissection (SLND) and selective localization and removal of clipped nodes, improves the false-negative rate (FNR) compared with SLND alone. METHODS A prospective study of patients with biopsy-confirmed nodal metastases with a clip placed in the sampled node was performed. After neoadjuvant therapy, patients underwent axillary surgery and the pathology of the clipped node was compared with other nodes. Patients undergoing TAD had SLND and selective removal of the clipped node using iodine-125 seed localization. The FNR was determined in patients undergoing complete axillary lymphadenectomy (ALND). RESULTS Of 208 patients enrolled in this study, 191 underwent ALND, with residual disease identified in 120 (63%). The clipped node revealed metastases in 115 patients, resulting in an FNR of 4.2% (95% CI, 1.4 to 9.5) for the clipped node. In patients undergoing SLND and ALND (n = 118), the FNR was 10.1% (95% CI, 4.2 to 19.8), which included seven false-negative events in 69 patients with residual disease. Adding evaluation of the clipped node reduced the FNR to 1.4% (95% CI, 0.03 to 7.3; P = .03). The clipped node was not retrieved as an SLN in 23% (31 of 134) of patients, including six with negative SLNs but metastasis in the clipped node. TAD followed by ALND was performed in 85 patients, with an FNR of 2.0% (1 of 50; 95% CI, 0.05 to 10.7). CONCLUSION Marking nodes with biopsy-confirmed metastatic disease allows for selective removal and improves pathologic evaluation for residual nodal disease after chemotherapy.

Multimodality Imaging of Triple Receptor-Negative Tumors With Mammography, Ultrasound, and MRI

OBJECTIVE We retrospectively reviewed imaging findings for 44 patients with triple receptor-negative breast carcinomas on mammography, sonography, and MRI to determine the imaging characteristics of triple receptor-negative cancers that may improve diagnosis at the time of presentation. CONCLUSION Despite their large size at presentation, triple receptor-negative cancers may be occult on mammography or sonography and frequently have benign or indeterminate features. MRI identified all triple receptor-negative cancers and showed features that had a high positive predictive value for malignancy.

CD44 Expression Is Associated with Increased Survival in Node-Negative Invasive Breast Carcinoma

Purpose: CD44 is a multifunctional cell surface receptor with many known splice variants, some of which have been reported to play a role in tumor progression. The purpose of this study was to evaluate the prognostic significance of CD44 isoforms in early-stage, lymph node–negative invasive breast carcinoma. Experimental Design: Immunohistochemical staining for CD44 isoforms was done on archival paraffin tissue sections of invasive breast carcinoma from a cohort of lymph node–negative patients who received no adjuvant tamoxifen or chemotherapy and who had a mean clinical follow-up period of 15 years. Immunohistochemical staining was done with antibodies to CD44s, the standard isoform of CD44, and to isoforms containing variant exon 6 (CD44v6); levels of staining were correlated with clinical outcome data. Results: There was a trend towards increased disease-free survival for patients whose tumors had high anti-CD44s positivity (P = 0.05), and a significant association was observed between anti-CD44s positivity and disease-related survival (P = 0.04). Expression of CD44v6 isoforms did not correlate with clinical outcome. Conclusion: CD44 expression, as assessed by immunohistochemical staining with anti-CD44s, may be a favorable prognostic factor in patients with node-negative invasive breast carcinoma.

Clear cell adenocarcinoma and nephrogenic adenoma of the urethra and urinary bladder: A histopathologic and immunohistochemical comparison

Because of histological similarities between nephrogenic adenomas and clear cell adenocarcinomas of the urinary tract, there is the potential for diagnostic confusion between these two entities. The histopathologic features of 13 nephrogenic adenomas and five clear cell adenocarcinomas of the urethra and urinary bladder are compared in this report, and detailed immunohistochemical staining profiles are provided for these tumors. Only 2 of the 13 nephrogenic adenomas contained clear cells, and these constituted less than 10% of the lesions. In contrast, four of the five clear cell adenocarcinomas contained prominent areas with clear cells. Nephrogenic adenomas generally showed only mild cytologic atypia, whereas four of the five clear cell adenocarcinomas showed severe atypia. A single mitotic figure was identified in only two of the nephrogenic adenomas, whereas the mitotic rate in the clear cell adenocarcinomas ranged from 2 to 14 per 10 high-power fields. None of the nephrogenic adenomas showed evidence of necrosis, but focal necrosis was noted in four of the five clear cell adenocarcinomas. In general, the nephrogenic adenomas and clear cell adenocarcinomas showed negative to weak staining with CK903 but strong staining with AE1, AE3, and Cam 5.2. Variable staining was observed with Brst-3 and antibodies to S-100, CEA (monoclonal and polyclonal), LeuM-1, and CA19.9. Nephrogenic adenomas and clear cell adenocarcinomas were all negative for prostate-specific acid phosphatase (PSAP), prostate-specific antigen (PSA), and estrogen and progesterone receptors (except for two nephrogenic adenomas, which showed only focal weak staining for estrogen receptor). Neither bcl-2 nor c-erbB-2 staining was able to discriminate between the tumors. However, strong staining for p53 was noted in each clear cell adenocarcinoma and in none of the nephrogenic adenomas. MIB-1 positivity in nephrogenic adenomas ranged from 0 to 13 (average of 5.5) per 200 cells, whereas the positive range for clear cell adenocarcinomas was 33 to 70 (average of 47) per 200 cells. In summary, histopathologic features that favor clear cell adenocarcinoma over nephrogenic adenoma include a predominance of clear cells, severe cytological atypia, high mitotic rate, necrosis, high MIB-1 positivity, and strong staining for p53.

A lymph node metastatic mouse model reveals alterations of metastasis-related gene expression in metastatic human oral carcinoma sublines selected from a poorly metastatic parental cell line.

Greater than 40% of patients with squamous cell carcinoma (SCC) of the oral cavity have lymph node metastasis at the time of diagnosis and a 5‐year survival rate of less than 50%. Changes in gene expression that regulate metastasis of SCC to lymph nodes have not been identified.

Reliability of chromogenic in situ hybridization for detecting HER-2 gene status in breast cancer: comparison with fluorescence in situ hybridization and assessment of interobserver reproducibility

Accurate determination of HER-2 status is important in the management of patients with breast cancer, especially in determining their eligibility for trastuzumab therapy. Fluorescence in situ hybridization (FISH) has been regarded as the gold standard method for detecting HER-2 gene amplification. Recently, chromogenic in situ hybridization (CISH), in which HER-2 is detected by a peroxidase reaction and the gene copies are determined by regular bright-field microscopy, has emerged as a potential alternative to FISH. However, this method requires validation before it can be adopted into clinical practice. In this study, we evaluated 80 cases of invasive breast carcinoma by CISH, compared the results with those obtained by FISH, and assessed interobserver reproducibility among three observers. We found that agreement among the three pathologists on the CISH-determined HER-2 status was achieved in 73 cases (91%), all of which had results matching the corresponding FISH results: 54 nonamplified and 19 amplified. Of the 19 amplified cases, 13 were scored unanimously as high-level amplification; six had a minor scoring discrepancy (ie, low-level vs high-level amplification). A major scoring discrepancy (ie, nonamplification vs amplification) was found in the remaining seven cases, three of which were amplified and four of which were nonamplified by FISH. Two of the latter cases had a polysomy of chromosome 17. The cases that caused scoring difficulty were those with an equivocal or borderline signal number against a high background. Overall, there was nearly perfect agreement between the CISH and corresponding FISH results, and interpretation of CISH results were highly reproducible among the three pathologists. We conclude that, in general, HER-2 status can be reliably assessed by CISH. Confirmatory FISH is recommended in cases with equivocal or borderline CISH copy numbers.

Metaplastic sarcomatoid carcinoma of the breast with absent or minimal overt invasive carcinomatous component : A misnomer

Metaplastic carcinomas of the breast are a heterogeneous group of neoplasms ranging from tumors with a predominant component of overt carcinoma and focal mesenchymal differentiation to keratin-positive tumors with pure sarcomatoid morphology. We examined the clinicopathologic features of 22 patients previously diagnosed at M.D. Anderson Cancer Center with metaplastic carcinoma of the breast with pure or almost pure sarcomatoid morphology. Patients were included in the study if their tumors had sarcomatoid morphology and: 1) an invasive carcinomatous component identifiable on hematoxylin and eosin stains comprising less than 5% of the invasive tumor; or 2) associated ductal carcinoma in situ; or 3) immunohistochemical expression of keratin in the sarcomatoid areas. Patients with low-grade fibromatosis-like metaplastic tumors and those who received neoadjuvant chemotherapy were excluded. Axillary lymph node dissection or limited axillary node excision was performed in 17 patients, including 1 patient who had a sentinel lymph node biopsy. Lymph node involvement occurred in only 1 patient and consisted of a single 3.5-mm metastasis. Clinical follow-up was available for 21 patients and ranged from 4 months to 155 months (median follow-up, 35 months). Ten patients experienced local relapse, including 7 of 11 patients treated with breast-conserving surgery, and 9 developed distant metastases, most frequently to the lungs. These findings suggest that metaplastic sarcomatoid carcinomas that lack or have only a minimal overt invasive carcinomatous component have a biologic behavior similar to that of sarcomas. In addition to systemic treatment, early aggressive local therapy is recommended, as these patients have a high rate of local relapse.