Jason A. Wampfler

Correlation of IHC and FISH for ALK Gene Rearrangement in Non-small Cell Lung Carcinoma: IHC Score Algorithm for FISH

Introduction: Accurate, cost-effective methods for testing anaplastic lymphoma kinase gene rearrangement (ALK+) are needed to select patients with non-small cell lung carcinoma for ALK-inhibitor therapy. Fluorescent in situ hybridization (FISH) is used to detect ALK+, but it is expensive and not routinely available. We explored the potential of an immunohistochemistry (IHC) scoring system as an affordable, accessible approach. Methods: One hundred one samples were obtained from an enriched cohort of never-smokers with adenocarcinoma from the Mayo Clinic Lung Cancer Cohort. IHC was performed using the ALK1 monoclonal antibody with ADVANCE detection system (Dako) and FISH with dual-color, break-apart probe (Abbott Molecular) on formalin-fixed, paraffin-embedded tissue. Results: Cases were assessed as IHC score 0 (no staining; n = 69), 1+ (faint cytoplasmic staining, n = 21), 2+ (moderate, smooth cytoplasmic staining; n = 3), or 3+ (intense, granular cytoplasmic staining in ≥10% of tumor cells; n = 8). All IHC 3+ cases were FISH+, whereas 1 of 3 IHC 2+ and 1 of 21 IHC 1+ cases were FISH+. All 69 IHC 0 cases were FISH−. Considering FISH a gold-standard reference in this study, sensitivity and specificity of IHC were 90 and 97.8%, respectively, when 2+ and 3+ were regarded as IHC positive and 0 and 1+ as IHC negative. Conclusions: IHC scoring correlates with FISH and may be a useful algorithm in testing ALK+ by FISH in non-small cell lung carcinoma, similar to human epidermal growth factor-2 testing in breast cancer. Further study is needed to validate this approach.

Mutations in the Tyrosine Kinase Domain of the Epidermal Growth Factor Receptor in Non–Small Cell Lung Cancer

We evaluated somatic genetic alterations in the kinase domain of the EGFR gene in the tumors of 219 non–small cell lung cancer patients of primarily Caucasian and African American origins. We identified 26 patients (12%) whose tumors had a mutation in the EGFR gene, and 11 (5%) patients carried novel genomic variations consistent with germ-line polymorphisms. All but one mutation were identified in Caucasian patients affected with adenocarcinoma. EGFR mutations were more frequent in women and in nonsmokers, but a significant portion of the affected patients were men (12 of 26) and current or past smokers accounted for half of the patients affected (13 of 26). Screening subjects with EGFR mutations may identify patients whose tumors could respond to targeted therapy using tyrosine kinase inhibitors.

Tumor B7-H1 and B7-H3 expression in squamous cell carcinoma of the lung

BACKGROUND Pulmonary squamous cell carcinoma has a poor prognosis, and new therapeutic targets are needed. The aberrant expression of the immunomodulatory proteins B7-H1 and B7-H3 by malignant cells may contribute to tumoral immune evasion. Data about the expression of these proteins by squamous cell carcinoma of the lung are limited. MATERIALS AND METHODS Immunohistochemistry for B7-H1 and B7-H3 was performed on 214 resected pulmonary squamous cell carcinoma specimens. RESULTS At the last follow-up, 171 of 214 (80%) of patients were deceased (median survival time, 3.76 years). Forty-two (19.6%) of 214 cases showed positivity with B7-H1, with a range of 5% to 60% of cells that stained positively. A total of 189 (88.3%) of 214 cases showed positivity with B7-H3, with a range of 5% to 80% of cells staining positively. By using multivariate analysis, no degree of B7-H1 or B7-H3 positivity was significantly associated with patient outcome. CONCLUSIONS Although B7-H1 and B7-H3 are not of independent prognostic value, they are commonly expressed on a subset of tumor cells in pulmonary squamous cell carcinomas. Known interaction of the B7-H proteins with cytotoxic T-lymphocyte antigen-4 may make them attractive candidate biomarkers for response to immunomodulatory therapeutics, eg, ipilimumab, and warrants further study.

Primary salivary gland-type lung cancer: spectrum of clinical presentation, histopathologic and prognostic factors.

Primary salivary‐type lung cancers are rare tumors that include adenoid cystic carcinoma (ACC) and mucoepidermoid carcinoma (MEC). The clinicopathologic profiles, symptoms on presentation, and long‐term outcomes of patients with ACC and MEC as an overall group have not been defined recently.

Alpha1-antitrypsin deficiency carriers, tobacco smoke, chronic obstructive pulmonary disease, and lung cancer risk.

BACKGROUND Genetic susceptibility in lung cancer risk has long been recognized but remains ill defined, as does the role of tobacco smoke exposure and chronic obstructive pulmonary disease (COPD). METHODS Using a dual case-control design, we tested whether alpha(1)-antitrypsin deficiency (alpha(1)ATD) carriers are predisposed to a higher risk of lung cancer, adjusting for the effects of tobacco smoke exposure and COPD. A total of 1856 patients with incident lung cancer were included in the study; 1585 community residents served as controls. A second control group was composed of 902 full siblings of the patients. We first modeled 1585 case-control pairs without the alpha(1)ATD variable using multiple logistic regression analysis and then modeled the alpha(1)ATD allele type in the presence of other known risk factors of lung cancer. RESULTS We found a significantly increased lung cancer risk among alpha(1)ATD carriers from 2 parallel case-control comparisons: when patients were compared with unrelated controls, alpha(1)ATD carriers had a 70% higher risk of developing lung cancer than noncarriers (odds ratio, 1.7; 95% confidence interval, 1.2-2.4). In a further comparison of patients with their cancer-free siblings, we found a 2-fold increased lung cancer risk in alpha(1)ATD carriers (95% confidence interval, 1.4-2.7). Stratified analysis by tumor histologic subtypes showed a significant increase for adenocarcinoma and squamous cell carcinoma among alpha(1)ATD carriers. CONCLUSION Our results suggest that alpha(1)ATD carriers are at a 70% to 100% increased risk of lung cancer and may account for 11% to 12% of the patients with lung cancer in our study.

Relationship Between Deficits in Overall Quality of Life and Non–Small-Cell Lung Cancer Survival

PURPOSE Evidence has suggested a clinically meaningful relationship between self-reported quality of life (QOL) of a patient with cancer at the time of receiving a cancer diagnosis and overall survival (OS). This study evaluated the prognostic value of QOL assessments with regard to OS in a large cohort of patients with lung cancer. PATIENTS AND METHODS A total of 2,442 patients with non-small-cell lung cancer were observed between 1997 and 2007 and completed a single-item measure of overall QOL within the first 6 months of receiving a lung cancer diagnosis; these were dichotomized using an a priori definition of a clinically deficient score (CDS; ≤ 50 v > 50). Kaplan-Meier estimates and Cox models were used to evaluate the prognostic importance of QOL on OS alone and in the presence of covariates. Logistic regression modeling was used to identify which clinical and patient characteristics were related to a clinically meaningful deficit in QOL. RESULTS QOL deficits at time of lung cancer diagnosis were significantly associated with OS (hazard ratio [HR], 1.55; P < .001), as were performance status, older age, smoking history, male sex, treatment factors, and stage of disease. The median survival for patients with CDS QOL was 1.6 years versus 5.6 years for patients with non-CDS QOL. After controlling for all these covariates, the indication of a clinically deficient baseline QOL still contributed significantly to the prediction of patient survival (HR, 0.67; P < .001). CONCLUSION Overall QOL measured by a simple single item at the time of lung cancer diagnosis is a significant and independent prognostic factor for survival in patients with lung cancer.

Worse disease-free survival in never-smokers with ALK+ lung adenocarcinoma

Introduction: The EML4–anaplastic lymphoma kinase (ALK) translocation is a recognized oncogenic driver in non-small cell lung cancer. We investigated immunohistochemistry (IHC) screening with fluorescence in situ hybridization (FISH) confirmation for ALK detection and estimated the prevalence of ALK positivity in our patient cohort of never-smokers, together with differences in clinical outcomes and prognostic factors for patients with ALK-positive and ALK-negative tumors. Methods: We designed a three-phase study (training, validation, and testing) in 300 never-smokers with lung adenocarcinoma from the observational Mayo Clinic Lung Cancer Cohort. Tumor samples were tested using IHC and FISH, and concordance between the methods was assessed. Clinical outcomes were assessed via 5-year progression- or recurrence-free survival from diagnosis. Prognostic factors for ALK-positive tumors and metastases were also investigated. Results: ALK-positive patients were significantly (p < 0.05) younger and had higher grade tumors than ALK-negative patients. ALK positivity was 12.2% by IHC and confirmed at 8.2% of tumors by FISH, with complete concordance between IHC 3+/0 and FISH+/− assessments, respectively. Five-year risk of progression or recurrence was doubled for patients with ALK-positive compared with ALK-negative tumors; ALK-positive tumors also appeared to be associated with a higher risk of brain and liver metastases. Conclusions: Our findings suggest that ALK positivity is associated with a significantly poor outcome in nonsmoking-related adenocarcinoma and that ALK-positive tumors may be associated with an increased risk of brain and liver metastases compared with ALK-negative disease. Consequently, an unmet medical need exists in ALK-positive lung cancer patients, and effective ALK-specific therapies are needed.

Quality of Life and Symptom Burden among Long-Term Lung Cancer Survivors

Introduction: Information is limited regarding health-related quality of life (QOL) status of long-term (greater than 5 years) lung cancer survivors (LTLCS). Obtaining knowledge about their QOL changes over time is a critical step toward improving poor and maintaining good QOL. The primary aim of this study was to conduct a 7-year longitudinal study in survivors of primary lung cancer which identified factors associated with either decline or improvement in QOL over time. Methods: Between 1997 and 2003, 447 LTLCS were identified and followed through 2007 using validated questionnaires; data on overall QOL and specific symptoms were at two periods: short-term (less than 3 years) and long-term postdiagnosis. The main analyses were of clinically significant changes (greater than 10%) and factors associated with overall QOL and symptom burden for each period and for changes over time. Results: Three hundred two (68%) underwent surgical resection only and 122 (27%) received surgical resection and radiation/chemotherapy. Recurrent or new lung malignancies were observed in 84 (19%) survivors. Significant decline or improvement in overall QOL over time were reported in 155 (35%) and 67 (15%) of 447 survivors, respectively. Among the 155 whose QOL declined, significantly worsened symptoms were fatigue (69%), pain (59%), dyspnea (58%), depressed appetite (49%), and coughing (42%). The symptom burden did not lessen among the 67 who reported improvement in overall QOL, suggesting that survivors had adapted to their compromised physical condition. Conclusions: LTLCS suffered substantial symptom burden that significantly impaired their QOL, indicating a need for targeted interventions to alleviate their symptoms.

MET and EGFR Mutations Identified in ALK-Rearranged Pulmonary Adenocarcinoma: Molecular Analysis of 25 ALK-Positive Cases

Introduction: Oncogenic ALK kinase activity associated with ALK gene rearrangement is the target of crizotinib, an ALK inhibitor recently approved by the Food and Drug Administration for the treatment of ALK-rearranged (ALK+) non–small cell lung cancers. ALK+ status is generally thought to be mutually exclusive of epidermal growth factor receptor (EGFR) and KRAS mutations. However, the mutation status of other genes is not widely known in ALK+ tumors. The aim of this study is to survey for mutations involving other genes in 25 ALK+ cases confirmed by fluorescent in situ hybridization. Methods: Using the DNA extracted from formalin-fixed paraffin-embedded tumor samples, a MassArray-based Lung Cancer Mutations Screening Panel was performed to test for 179 individual mutations in 10 genes, including EGFR, KRAS, BRAF, ERBB2, JAK2, AKT1, AKT2, KIT, MET and PIK3CA, which have been implicated in lung carcinogenesis and/or considered as potential therapeutic targets. Results: Five of 25 ALK+ cases showed additional genetic abnormalities, which were verified by gene sequencing. One patient had EGFR del L747-S752. The remaining four mutations were in the MET gene: MET N375S (n = 2) and MET R988C (n = 2). No MET amplification was found by fluorescent in situ hybridization in the four cases with MET mutation. No mutations were detected in the other genes tested. Conclusions: In summary, additional mutations were found in 20% of ALK+ cases involving two of the 10 genes tested. Our study highlights that EGFR mutation can be present in ALK+ tumors, though uncommon. Clinical implication of MET mutation in our cases is uncertain and further study is needed.

CHRNA5 Risk Variant Predicts Delayed Smoking Cessation and Earlier Lung Cancer Diagnosis—A Meta-Analysis

BACKGROUND Recent meta-analyses show strong evidence of associations among genetic variants in CHRNA5 on chromosome 15q25, smoking quantity, and lung cancer. This meta-analysis tests whether the CHRNA5 variant rs16969968 predicts age of smoking cessation and age of lung cancer diagnosis. METHODS Meta-analyses examined associations between rs16969968, age of quitting smoking, and age of lung cancer diagnosis in 24 studies of European ancestry (n = 29 072). In each dataset, we used Cox regression models to evaluate the association between rs16969968 and the two primary phenotypes (age of smoking cessation among ever smokers and age of lung cancer diagnosis among lung cancer case patients) and the secondary phenotype of smoking duration. Heterogeneity across studies was assessed with the Cochran Q test. All statistical tests were two-sided. RESULTS The rs16969968 allele (A) was associated with a lower likelihood of smoking cessation (hazard ratio [HR] = 0.95, 95% confidence interval [CI] = 0.91 to 0.98, P = .0042), and the AA genotype was associated with a four-year delay in median age of quitting compared with the GG genotype. Among smokers with lung cancer diagnoses, the rs16969968 genotype (AA) was associated with a four-year earlier median age of diagnosis compared with the low-risk genotype (GG) (HR = 1.08, 95% CI = 1.04 to 1.12, P = 1.1*10(-5)). CONCLUSION These data support the clinical significance of the CHRNA5 variant rs16969968. It predicts delayed smoking cessation and an earlier age of lung cancer diagnosis in this meta-analysis. Given the existing evidence that this CHRNA5 variant predicts favorable response to cessation pharmacotherapy, these findings underscore the potential clinical and public health importance of rs16969968 in CHRNA5 in relation to smoking cessation success and lung cancer risk.