Nirmal K. Veeramachaneni

Differential Pathogenesis of Lung Adenocarcinoma Subtypes Involving Sequence Mutations, Copy Number, Chromosomal Instability, and Methylation

Background Lung adenocarcinoma (LAD) has extreme genetic variation among patients, which is currently not well understood, limiting progress in therapy development and research. LAD intrinsic molecular subtypes are a validated stratification of naturally-occurring gene expression patterns and encompass different functional pathways and patient outcomes. Patients may have incurred different mutations and alterations that led to the different subtypes. We hypothesized that the LAD molecular subtypes co-occur with distinct mutations and alterations in patient tumors. Methodology/Principal Findings The LAD molecular subtypes (Bronchioid, Magnoid, and Squamoid) were tested for association with gene mutations and DNA copy number alterations using statistical methods and published cohorts (n = 504). A novel validation (n = 116) cohort was assayed and interrogated to confirm subtype-alteration associations. Gene mutation rates (EGFR, KRAS, STK11, TP53), chromosomal instability, regional copy number, and genomewide DNA methylation were significantly different among tumors of the molecular subtypes. Secondary analyses compared subtypes by integrated alterations and patient outcomes. Tumors having integrated alterations in the same gene associated with the subtypes, e.g. mutation, deletion and underexpression of STK11 with Magnoid, and mutation, amplification, and overexpression of EGFR with Bronchioid. The subtypes also associated with tumors having concurrent mutant genes, such as KRAS-STK11 with Magnoid. Patient overall survival, cisplatin plus vinorelbine therapy response and predicted gefitinib sensitivity were significantly different among the subtypes. Conclusions/ Significance The lung adenocarcinoma intrinsic molecular subtypes co-occur with grossly distinct genomic alterations and with patient therapy response. These results advance the understanding of lung adenocarcinoma etiology and nominate patient subgroups for future evaluation of treatment response.

Management of Stage IIIA Non-Small Cell Lung Cancer by Thoracic Surgeons in North America

BACKGROUND Stage IIIA(N2) non-small cell lung cancer is a heterogeneous spectrum ranging from microscopic lymph node metastases to bulky multistation nodal disease. While some favor surgical resection after neoadjuvant therapy, others favor definitive chemoradiation for treatment. Our aim was to determine practice patterns of thoracic surgeons. METHODS We invited 2,539 active surgeons identified on the Cardiothoracic Surgery Network as expressing interest in general thoracic surgery to participate in an anonymous Web-based survey. The participants evaluated clinical vignettes of a patient with single station N2 disease. RESULTS In all, 513 surgeons (20%) responded, with 222 (43%) in academic practice. For microscopic N2 disease, 84% (n=430) preferred neoadjuvant therapy followed by surgery. For grossly involved N2 disease, 62% (n=318) favored neoadjuvant therapy followed by surgery if N2 disease was downstaged. In patients with normal pulmonary function tests, requiring pneumonectomy, in the presence of bulky, single station N2 disease, there was less consensus: 32% (n=163) favored neoadjuvant therapy followed by lobectomy (less radical surgery than initially predicted) if feasible and N2 disease had downstaged, 30% (n=159) favored neoadjuvant therapy followed by pneumonectomy if N2 disease downstaged, 12% (n=60) would favor surgery regardless of N2 disease downstaging, and 22% (n=114) favored definitive chemoradiation. If the patient did not have adequate pulmonary function for pneumonectomy but could tolerate lobectomy, 50% favored neoadjuvant therapy followed by reassessment for lobectomy and 41% favored definitive chemoradiation. CONCLUSIONS There is no clear consensus on management of patients with stage IIIA lung cancer in the United States. Diversity of opinion is greatest in patients with more advanced lung cancer, and limited pulmonary function.

Local failure after complete resection of N0-1 non-small cell lung cancer

PURPOSE To estimate the risk of local-regional failure (LRF) after surgery for operable NSCLC, and the effect of clinical/pathologic factors on this risk. METHODS Records of 335 patients undergoing complete resection (lobectomy, pneumonectomy) for pathological T1-4 N0-1 NSCLC (without post-operative radiation) from 1996 to 2006 were reviewed. Crude and actuarial estimated failure rates were computed; local-regional sites included ipsilateral lung, surgical stump, hilar, mediastinal, or supraclavicular nodes. Failure times in sub-groups were calculated with the Kaplan-Meier method and compared via log-rank test. Independent factors adversely affecting LRF were determined with Cox regression. RESULTS The median follow-up duration for event-free surviving patients was 40 months (range: 1-150). The crude and actuarial 5-year probability of any failure (LR or distant) were 33% and 43%, respectively. Of all failures; 37% were LR only, 35% LR and distant and 28% distant only. The 5-year crude and actuarial probability of LRF were 24% and 35% (95% CI: 29-42%). Five-year crude LRF rates for T1-2N0, T1-2N1, T3-4N0 and T3-4N1 disease were 19% (41/216), 27% (16/59), 37.5% (15/40) and 40% (8/20), respectively. The corresponding actuarial estimates were T1-2N0 28%, T1-2N1 39%, T3-4N0 50% and T3-4N1 67%. In Cox multiple regression analysis, lymphovascular space invasion (p=0.03, HR: 1.7) and tumor size (p=0.01, HR: 1.67 for 5 cm increment) were associated with an increased risk of LRF. CONCLUSION Five-year LRF rates are ≥19% in essentially all patient subsets.

Rapid On-Site Pathologic Evaluation Does Not Increase the Efficacy of Endobronchial Ultrasonographic Biopsy for Mediastinal Staging

BACKGROUND Endobronchial ultrasonography with transbronchial needle aspiration (EBUS-TBNA) has been shown to be equivalent to mediastinoscopy in lung cancer staging for mediastinal node involvement. Rapid on-site evaluation (ROSE) to determine the adequacy of nodal sampling has been claimed to be beneficial. METHODS A retrospective evaluation was performed in 170 patients who underwent EBUS-TBNA from July 2008 to May 2011. The patients were classified as having either high or low pretest probability for mediastinal disease based on history and radiographic imaging. ROSE was compared with the final pathology reports based on slides and cell blocks. RESULTS One hundred thirty-one (77%) patients were classified as being in the high pretest cohort based on clinical staging. Of these, 101 (77%) patients had adequate tissue sampling based on ROSE, with 70 (69%) patients having positive mediastinal disease. In the 30 (23%) patients who had inadequate tissue by ROSE, the final analysis of all the prepared slides and cell blocks allowed for a diagnosis in all but 8 patients. The sensitivity and specificity of ROSE in the high pretest probability cohort were 89.5% and 96.4%, respectively, whereas the overall sensitivity and specificity of EBUS-TBNA was 92.1% and 100%, respectively. Despite having inadequate tissue on ROSE in 30 of 131 patients, sufficient tissue was available on final analysis for diagnosis in 22 of 30 patients. CONCLUSIONS ROSE does not impact clinical decision making if a thorough mediastinal staging using EBUS is performed. Despite inadequate tissue sampling assessment by ROSE, a final diagnosis was made in most patients, potentially avoiding an additional surgical procedure to prove mediastinal disease.

Effectiveness and Risks Associated With Intrapleural Alteplase by Means of Tube Thoracostomy

BACKGROUND The use of fibrinolytics has been described for the treatment of complex pleural processes. This has evolved from streptokinase to urokinase to alteplase. Intrapleural fibrinolysis has added an alternative to surgical intervention in patients with complex pleural processes. This study describes the use of alteplase as an alternative to surgical intervention for these processes. METHODS From December 2004 to March 2009, 118 patients required alteplase for complex pleural processes. The type of tube thoracostomy, pleural process, antithrombotic type, international normalized ratio, prothrombin time, partial thromboplastin time, platelets, doses, and outcomes were reviewed for each patient. Complications and the need for additional interventions were evaluated. RESULTS Patients received one to eight doses of intrapleural alteplase through a tube thoracostomy. Indications for intrapleural alteplase were empyema (n = 32; 27.1%), loculated pleural effusion (n = 44; 37.3%), hemothorax (n = 13; 11.0%), parapneumonic effusion (n = 25; 21.2%), and malignant effusion (n = 6; 5.1%). The success rate was 86.4% (102 of 118 patients). The incidence of bleeding was 8.5% (n = 10). Binary analysis did not demonstrate an increase in bleeding with abnormal coagulation variables. Of the patients with a bleeding complication, 7 required operative interventions. Twenty (16.9%) required a second tube thoracostomy for incomplete evacuation of the pleural process. Nine (7.6%) required an operative intervention for incomplete evacuation of the pleural process. CONCLUSIONS Intrapleural alteplase appears to be effective in treating complex parapneumonic processes. Systemic anticoagulation, prothrombin time, partial thromboplastin time, international normalized ratio, and platelet count do not appear to be risk factors for bleeding complications. One or two doses of alteplase appear most successful.

Non–Small Cell Lung Cancer: Prognostic Importance of Positive FDG PET Findings in the Mediastinum for Patients with N0–N1 Disease at Pathologic Analysis

PURPOSE To assess the prognostic implications of mediastinal positron emission tomographic (PET) findings in patients undergoing curative resection of non-small cell lung cancer (NSCLC) who have histologically negative mediastinal lymph nodes (LNs), with the hypothesis that positive findings at PET are prognostic even in patients with negative histologic findings in the LNs. MATERIALS AND METHODS Records of patients with a preoperative PET undergoing curative surgery, without adjuvant radiation, for pathologic T1-3N0-1 NSCLC at the University of North Carolina between 2000 and 2006 were reviewed as an institutional review board-approved HIPAA-compliant retrospective study. Ninety patients were evaluable (all histologically negative in mediastinum; 44 with both mediastinoscopy and surgery); 13 patients had positive mediastinal PET findings, and 77 had negative mediastinal PET findings. Local-regional and distant failure rates in patients with and those without mediastinal abnormalities at preoperative PET were compared by using logistic regression and log-rank tests. RESULTS Median follow-up was 54.3 months (range, 1-99 months). There were higher rates of local-regional (P = .001) and distant (P < .001) failure as well as death (P = .001) in patients with postive PET findings than in patients with negative findings. In multivariable analysis (adjusting for other prognostic factors), positive PET findings in the mediastinum remained prognostic for distant failure (P < .001, hazard ratio = 6.9) and were marginally prognostic for local-regional failure (P = .093, hazard ratio = 1.9). CONCLUSION Positive findings at preoperative PET in the mediastinum appear to have prognostic implications despite the mediastinal LNs being histologically negative. The high rate of local-regional and distant failure suggests that postoperative radiation therapy and/or chemotherapy may be particularly helpful in patients with positive mediastinal findings at preoperative PET.

Annual review of advances in non-small cell lung cancer research: A report for the year 2010

Lung cancer is the leading cause of cancer mortality in the United States and worldwide,1,2 and the prognosis remains poor with an estimated 5-year overall survival (OS) rate of 15%.2 Approximately 85% of the cases of lung cancer are non-small cell lung cancer (NSCLC), and the majority of patients have advanced stage disease at the time of diagnosis.3 Significant investigations into the etiology, molecular characteristics, and the development of novel therapies for NSCLC have been performed and are ongoing. Consequently, NSCLC research and care have evolved into a multidisciplinary approach. As a variety of disciplines perform important research and the results are published in multiple journals, it has become increasingly difficult to remain current on the recent publications. We sought to provide a concise review of the major publications related to lung cancer in the fields of molecularly biology, surgery, radiation oncology, and medical oncology. We restricted the review to clinical trials that were published in peer-reviewed journals to the calendar year of 2010; given the rapid developments within the field of molecular biology, abstracts and presentations from 2010 and early 2011 were included in this review. Unfortunately, no appreciable progress was made in small cell lung cancer in the year 2010; therefore, the focus on the review will be NSCLC.

Variability in Defining T1N0 Non-Small Cell Lung Cancer Impacts Locoregional Failure and Survival

BACKGROUND Locoregional recurrence can occur despite complete anatomic resection of T1N0 non-small cell lung cancer. That may be the result of incomplete resection or inaccurate staging. We assessed the impact of extent of nodal staging on the rate of locoregional failure and patient survival. METHODS The records of 742 patients undergoing lobectomy, bilobectomy, or pneumonectomy for non-small cell lung cancer from 1996 to 2006 were reviewed. Operative reports and pathology reports were reviewed for the number of lymph nodes and the anatomic nodal stations examined. The Kaplan-Meier method was applied to analyze recurrence-free survival. RESULTS A total of 119 patients with pathologically staged Ia lung cancer were identified. Histology type included 61% (n = 73) adenocarcinoma, 27% (n = 32) squamous cell cancer, and 12% (n = 14) other. Median age was 65 years (range, 34 to 88). Mean follow-up duration was 40 months (median 47; range, 1 to 121). Locoregional recurrence occurred in 20% (n = 18). The N2 nodal stations were examined in 94% (n = 112). At least one defined N1 nodal station was examined in 70% (n = 83). Station undefined N1 nodes were examined in 27% (n = 32), and no N1 nodes were examined in 3% (n = 4). Median number of N1 lymph nodes analyzed was 5 (range, 0 to 18). The locoregional recurrence rate was 14% (12 of 83) for patients with a defined N1 station node versus 31% (11 of 36) for patients in whom there were undefined N1 nodes (p = 0.03). Similar differences were seen in disease-free survival, 78.2% versus 62.6%, respectively (p = 0.06). CONCLUSIONS Despite anatomic resection of stage Ia lung cancer and uniform analysis of N2 nodal stations, a high rate of locoregional recurrence occurs. Imprecise staging of N1 lymph nodes may contribute to the understaging and undertreatment of patients with early stage lung cancer.

Radiofrequency ablation for nondysplastic Barrett's esophagus: Should we do it, because we can?

8. Gundrie JJ, PouwRE, Sondermeijer CM, Peters FP, CurversWL, RosmolenWD, et al. Stepwise circumferential and focal ablation of Barrett’s esophagus with high-grade dysplasia: results of the first prospective series of 11 patients. Endoscopy. 2008;40:359-69. 9. Ganz RA, Overholt BF, Sharma VK, Fleischer DE, Shaheen NJ, Lightdale CJ, et al. Circumferential ablation of Barrett’s esophagus that contains high-grade dysplasia: a U.S. multicenter registry. Gastrointest Endosc. 2008;68:35-40. 10. Velanovich V. Endoscopic endoluminal radiofrequency ablation of Barrett’s esophagus: initial results and lessons learned. Surg Endosc. 2009;23: 2175-80. 11. Lyday WD, Corbett FS, Kuperman DA, Kalvaria I, Mavrelis PG, Shughoury AB, et al. Radiofrequency ablation of Barrett’s esophagus: outcomes of 429 patients from a multicenter community practice registry. Endoscopy. 2010;42:272-8.

Abstract 4839: Lung cancer patients exhibit a genomewide chromosomal instability and DNA methylation correlation which varies by expression subtype

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Introduction: Lung cancer is a deadly disease, which manifests gross DNA alterations. Chromosome instability (CIN) and DNA methylation (DM) instability are important DNA alteration processes that contribute to the initiation and progression of lung cancer. Considering that DNA replication and methylation maintenance are coordinated, we hypothesized that CIN and DM may be coordinated in the etiology of lung cancer patients. Methods: A cohort of 63 patients with surgically-resected lung tumors was collected. DNA copy number was measured by Affymetrix 250K_StyI microarrays. DNA methylation was measured by the MSNP assay, which involves comparing the copy number of HpaII (methylation sensitive) digested DNA to undigested DNA (Yuan, 2006). The sample genomewide CIN score was the median of chromosome arm absolute copy numbers. The sample genomewide DNA methylation score was the median of the percent methylation across all probes. mRNA expression was measured by Agilent 44K microarrays. Results: CIN and DM varied substantially over lung cancers. Histological classes did not differ in CIN or DM. Lung cancer mRNA expression subtypes (described in Hayes et al., J Clin Oncol, 2006 and Wilkerson et al., Clin Cancer Res 2010) were significantly different in CIN (P<0.05) with the magnoid adenocarcinoma, classical squamous cell carcinoma (SCC), and basal SCC subtypes having the largest CIN scores. DM was also significantly different between expression subtypes (P<0.05), with the magnoid adenocarcinoma and the classical SCC having the largest DM scores. CIN and DM were significantly positively correlated (0.37; P<0.01), indicating that the most copy number disrupted genomes were also the most DNA hypermethylated. To evaluate the combined effect of these alterations, a combined CIN and DM (CIN+DM) standardized score was evaluated. Histological classes did not differ in CIN+DM. Expression subtypes were significantly different in CIN+DM (P<0.01) with the magnoid adenocarcinoma subtype and the classical SCC subtype exhibiting the largest CIN+DM scores. Patients with distant recurrences had a trend of larger CIN+DM scores than patients with local recurrences or no observed recurrences (P<0.21). An mRNA expression signature based on the CIN+DM score was positively correlated (GSEA, FDR < 0.05) with overexpression of DNA repair pathways, providing corroborating evidence for this DNA alteration phenotype. Conclusions: Genomewide chromosomal instability and DNA methylation are significantly correlated in lung cancer. These co-occurring DNA alterations are the most extreme in the magnoid adenocarcinoma and classical squamous cell carcinoma subtypes, suggesting the subtypes have evolved differently. These results elucidate new lung cancer etiology and future therapies could be focused on exploiting these DNA alterations. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4839. doi:10.1158/1538-7445.AM2011-4839