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Lung cancer is the leading cause of cancer deaths in the United States, surpassing breast cancer as the primary cause of cancer-related mortality in women. The goal of the present study was to identify early molecular changes in the lung induced by exposure to tobacco smoke and thus identify potential targets for chemoprevention. Female A/J mice were exposed to either tobacco smoke or HEPA-filtered air via a whole-body exposure chamber (6 h/d, 5 d/wk for 3, 8, and 20 weeks). Gene expression profiles of lung tissue from control and smoke-exposed animals were established using a 15K cDNA microarray. Cytochrome P450 1b1, a phase I enzyme involved in both the metabolism of xenobiotics and the 4-hydroxylation of 17β-estradiol (E2), was modulated to the greatest extent following smoke exposure. A panel of 10 genes were found to be differentially expressed in control and smoke-exposed lung tissues at 3, 8, and 20 weeks (P < 0.001). The interaction network of these differentially expressed genes revealed new pathways modulated by short-term smoke exposure, including estrogen metabolism. In addition, E2 was detected within murine lung tissue by gas chromatography-coupled mass spectrometry and immunohistochemistry. Identification of the early molecular events that contribute to lung tumor formation is anticipated to lead to the development of promising targeted chemopreventive therapies. In conclusion, the presence of E2 within lung tissue when combined with the modulation of cytochrome P450 1b1 and other estrogen metabolism genes by tobacco smoke provides novel insight into a possible role for estrogens in lung cancer. Cancer Prev Res; 3(6); 707–17. ©2010 AACR.

Perspective on this Article from Early Changes in Gene Expression Induced by Tobacco Smoke: Evidence for the Importance of Estrogen within Lung Tissue

Supplementary Figure 10 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

Supplementary Figure 7 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

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Supplementary Figure 5 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

Supplementary Figure 6 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

Supplementary Figure 9 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

Supplementary Figure 4 from Molecular Classifiers for Gastric Cancer and Nonmalignant Diseases of the Gastric Mucosa

High incidence of gastric cancer-related death is mainly due to diagnosis at an advanced stage in addition to the lack of adequate neoadjuvant therapy. Hence, new tools aimed at early diagnosis would have a positive impact in the outcome of the disease. Using cDNA arrays having 376 genes either identified previously as altered in gastric tumors or known to be altered in human cancer, we determined expression signature of 99 tissue fragments representing normal gastric mucosa, gastritis, intestinal metaplasia, and adenocarcinomas. We first validated the array by identifying molecular markers that are associated with intestinal metaplasia, considered as a transition stage of gastric adenocarcinomas of the intestinal type as well as markers that are associated with diffuse type of gastric adenocarcinomas. Next, we applied Fisher’s linear discriminant analysis in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Many classifiers could distinguish between normal and tumor samples, whereas, for the distinction of gastritis from tumor and for metaplasia from tumor, fewer classifiers were identified. Statistical validations showed that trios that discriminate between normal and tumor samples are powerful classifiers to distinguish between tumor and nontumor samples. More relevant, it was possible to identify samples of intestinal metaplasia that have expression signature resembling that of an adenocarcinoma and can now be used for follow-up of patients to determine their potential as a prognostic test for malignant transformation.

Background: Although Chemoradiation (CRT) is the curative treatment for SCCAC, many patients present primary resistance. Since it is a rare tumor, response predictors remain unknown. Methods: We performed a prospective cohort study to evaluate biomarkers associated with CRT response, progression-free survival (PFS), and overall survival (OS). The primary endpoint was response at 6 months (m). Tumor DNA and HPV were analyzed by next-generation sequencing, while KI-67 and PD-L1 by immunohistochemistry in tumor tissue. Results: Seventy-eight patients were recruited between October/2011 and December/2015, and 75 were response evaluable. The median age was 57 years, 65% (n=49) were stage III and 12% (n=9) were HIV positive (HIV+). At 6m, 62.7% (n=47) presented CR. On multivariate analyses, stage II patients were 4.7 more likely to achieve response than stage III (OR, 4.70; 95%CI, 1.36-16.30; p=0.015). HIV+ was associated with a worse response (OR, 5.72; 95%CI, 2.5-13.0; p

PURPOSE Li-Fraumeni syndrome (LFS) is rare in the worldwide population, but it is highly prevalent in the Brazilian population because of a founder mutation, TP53 p.R337H, accounting for 0.3% of south and southeastern population. Clinical criteria for LFS may not identify all individuals at risk of carrying the Brazilian founder mutation because of its lower penetrance and variable expressivity. This variant is rarely described in databases of somatic mutations. Somatic findings in tumor molecular profiling may give insight to identify individuals who might be carriers of LFS and allow the adoption of risk reduction strategies for cancer. MATERIALS AND METHODS We determined the frequency of the TP53 p.R337H variant in tumor genomic profiling from 755 consecutive Brazilian patients with pan-cancer. This is a retrospective cohort from January 2013 to March 2020 at a tertiary care center in Brazil. RESULTS The TP53 p.R337H variant was found in 2% (15 of 755) of the samples. The mutation allele frequency ranged from 30% to 91.7%. A total of seven patients were referred for genetic counseling and germline testing after tumor genomic profiling results were disclosed. All the patients who proceeded with germline testing (6 of 6) confirmed the diagnosis of LFS. Family history was available in 12 cases. Nine patients (9 of 12) did not meet LFS clinical criteria. CONCLUSION The identification of the TP53 p.R337H variant in tumor genomic profiling should be a predictive finding of LFS in the Brazilian population and should prompt testing for germline status confirmation.

// Jing Peng 1 , Sibele I. Meireles 1 , Xia Xu 2 , William E. Smith 3 , Michael J. Slifker 1 , Stacy L. Riel 1 , Shumenghui Zhai 4 , Guo Zhang 4 , Xiang Ma 4 , Mindy S. Kurzer 3 , Grace X. Ma 4 and Margie L. Clapper 1 1 Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, PA 19111, USA 2 Leidos Biomedical Research, Inc., Frederick, MD 21701, USA 3 Department of Food Science and Nutrition and Department of Medicine, University of Minnesota, St. Paul, MN 55108, USA 4 Center for Asian Health, Temple University, Philadelphia, PA 19140, USA Correspondence to: Margie L. Clapper, email: Margie.Clapper@fccc.edu Keywords: estrogen metabolism; 4-hydroxy estrogen; non-small cell lung cancer; tobacco smoke; never-smoking Chinese women Received: August 18, 2017 Accepted: September 30, 2017 Published: November 01, 2017 ABSTRACT Previous data from this group demonstrate that the murine lung metabolizes estrogen. Production of the putative carcinogen 4-hydroxyestrogen (4-OHE) is elevated within the lungs of female vs. male mice and accelerated by tobacco smoke. The goal of this study was to determine if the human lung metabolizes estrogen and evaluate the impact of tumor formation, smoke, sex and race/ethnicity on metabolism. Urine and lung tissue (normal, tumor) were obtained from 49 non-small cell lung cancer patients. Healthy postmenopausal Caucasian ( n = 19) and Chinese ( n = 20) American women (never-smokers) donated urine. Quantitative RT-PCR analyses indicate that multiple estrogen synthesis and metabolism genes are expressed in human bronchoalveolar cells. Estrogen and its metabolites were measured in lung tissue and urine using liquid chromatography/tandem mass spectrometry. Wilcoxon rank tests were used for statistical comparisons. E 1 , E 2 , E 3 and estrogen metabolites 2-OHE 1 , 2-OHE 2 , 4-OHE 1 , 4-OHE 2 , 2-OME 1 and 2-OME 2 were detected at higher levels in tumor vs. adjacent normal tissue and in women vs. men ( P < 0.05). The proportion of 4-OHEs was higher in tumors than in normal lung tissue ( P < 0.05), and elevated in normal tissue from current- vs. never-smoking women ( P = 0.006); similar trends were observed in urine. The proportion of 4-OHEs in the urine of postmenopausal Chinese American women was 1.8-fold higher than that of Caucasian women ( P = 0.015). These data indicate that estrogen metabolites are present in the human lung. A shift towards 4-hydroxylation during lung tumorigenesis may contribute to the risk conferred by smoking, sex or race/ethnicity.

4053 Background: Chemoradiation (CRT) is a curative treatment for SCCAC. However, some patients (pts) present primary CRT resistance. As a rare tumor, there is a lack of prospective studies of prognostic factors in this setting. Methods: This prospective cohort study was aimed to evaluate predictive biomarkers (Ki-67, PD-L1, Human papillomavirus (HPV), HIV status, and tumor DNA mutations) in SCCAC. We published the 6 months (m) response rate (RR) of this cohort showing that HIV- were 5.7 times more likely to achieve response 6m post CRT (OR 5.72, CI 95% 2.5-13.0, P < 0.001). Now we report the long-term follow-up results of 5-year progression-free survival (PFS) and overall survival (OS). Eligible pts had T2-4/N0-3/M0 disease and were candidates to standard CRT. DNA mutations were analyzed by next-generation sequencing (NGS). HPV positivity was tested by PapilloCheck Test. KI-67 and PD-L1 were evaluated by immunohistochemistry. Results: 78 pts were recruited from Jan/2011 to Dec/2015. 75 were evaluable for PFS and OS. The median age was 57 years; 49 (65%) were stage III, and 9 (12%) were HIV+. HPV was evaluated in 67 and found in 47 (70.1%); HPV16 was the most common. PD-L1 was tested in 61; 10 (16.4%) had positive expression > 1%. Ki-67 was performed in 65, with a median of 50% (range 1-90%). The median follow up is 66m. 5-year PFS and OS rates were 63.3% (95% CI 51.2-73.2%) and 76.4% (95% CI 64.8-84.6%), respectively. In a multivariate analysis, age (HR 1.06, P = 0.022, IC 95% 1.01-1.11) and absence of complete response at 6m (HR 3.36, P = 0.007, IC 95% 1.39-8.09) was associated with inferior OS. The OS rate was 62.5% in HIV+ group (95% CI 22.9-86%) in comparison with 78% (95% CI 65.7-86.3%) among HIV- pts, although this difference was not statistically significant (P = 0.400). A tendency to inferior OS was observed among pts with p53 codon 72 polymorphism (HR 2.83, P = 0.181, 95% CI 0.61-13.02). Other tumor mutations, HPV, Ki-67 expression, and PD-L1 expression, were not associated with PFS and OS. Conclusions: HIV- pts were 5.7 times more likely to achieve response 6m post CRT. The absence of complete response at 6m was the main factor associated with poor 5-year OS. New strategies of follow up and complementary treatment should be studied in late responders and HIV+ pts to ensure the success of curative treatment. Clinical trial information: 36211 .

The establishment of a reliable method for using RNA from formalin-fixed, paraffin-embedded (FFPE) tissue would provide an opportunity to obtain novel gene expression data from the vast amounts of archived tissue. A custom-designed 22,000 oligonucleotide array was used in the present study to compare the gene expression profile of colonic epithelial cells isolated by laser capture microdissection from FFPE-archived samples with that of the same cell population from matched frozen samples, the preferred source of RNA. Total RNA was extracted from FFPE tissues, amplified, and labeled using the Paradise Reagent System. The quality of the input RNA was assessed by the Bioanalyzer profile, reverse transcriptase-polymerase chain reaction, and agarose gel electrophoresis. The results demonstrate that it is possible to obtain reliable microarray data from FFPE samples using RNA acquired by laser capture microdissection. The concordance between matched FFPE and frozen samples was evaluated and expressed as a Pearson's correlation coefficient, with values ranging from 0.80 to 0.97. The presence of ribosomal RNA peaks in FFPE-derived RNA was reflected by a high correlation with paired frozen samples. A set of practical recommendations for evaluating the RNA integrity and quality in FFPE samples is reported.

Using cDNA fragments from the FAPESP/lICR Cancer Genome Project, we constructed a cDNA array having 4512 elements and determined gene expression in six normal and six tumor gastric tissues. Using t-statistics, we identified 80 cDNAs whose expression in normal and tumor samples differed more than 3.5 sample standard deviations. Using Self-Organizing Map, the expression profile of these cDNAs allowed perfect separation of malignant and non-malignant samples. Using the supervised learning procedure Support Vector Machine, we identified trios of cDNAs that could be used to classify samples as normal or tumor, based on single-array analysis. Finally, we identified genes with altered linear correlation when their expression in normal and tumor samples were compared. Further investigation concerning the function of these genes could contribute to the understanding of gastric carcinogenesis and may prove useful in molecular diagnostics.

e15059Background: Chemoradiation (CRT) is the standard curative treatment for Squamous Cell Carcinoma of the Anal Canal (SCCAC). However, some patients develop progressive disease and ultimately di...

Lung cancer is the leading cause of cancer mortality in women worldwide. Although the rise and growing epidemic status of this disease is overwhelmingly attributed to tobacco use, the rank of lung cancer in nonsmokers as the seventh most common cause of cancer worldwide suggests that other factors contribute to this disease. The majority of lung cancers among nonsmokers occur in women. Aside from geographic, cultural and genetic differences, hormonal and possibly infectious factors may also play etiologic roles. This review aims to discuss the epidemiology of lung cancer in women as well as the incidence of second primaries, and presents current opinions on the myriad of causes.

High incidence of gastric cancer-related death is mainly due to diagnosis at an advanced stage in addition to the lack of adequate neoadjuvant therapy. Hence, new tools aimed at early diagnosis would have a positive impact in the outcome of the disease. Using cDNA arrays having 376 genes either identified previously as altered in gastric tumors or known to be altered in human cancer, we determined expression signature of 99 tissue fragments representing normal gastric mucosa, gastritis, intestinal metaplasia, and adenocarcinomas. We first validated the array by identifying molecular markers that are associated with intestinal metaplasia, considered as a transition stage of gastric adenocarcinomas of the intestinal type as well as markers that are associated with diffuse type of gastric adenocarcinomas. Next, we applied Fisher’s linear discriminant analysis in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Many classifiers could distinguish between normal and tumor samples, whereas, for the distinction of gastritis from tumor and for metaplasia from tumor, fewer classifiers were identified. Statistical validations showed that trios that discriminate between normal and tumor samples are powerful classifiers to distinguish between tumor and nontumor samples. More relevant, it was possible to identify samples of intestinal metaplasia that have expression signature resembling that of an adenocarcinoma and can now be used for follow-up of patients to determine their potential as a prognostic test for malignant transformation.

We examined whether mechanical ventilation with low tidal volume induces polymorphonuclear infiltration and proinflammatory and profibrogenic responses in rat lungs compared dependent and nondependent lung region to expression of interleukin-1beta (IL-1beta) and alpha-1 procollagen III (PC III) mRNA.An experimental, randomized and controlled protocol with previously normal rats.Three groups of ten animals were studied. Two groups were ventilated (FIO2=0.3) in supine position for 1 h without positive end expiratory pressure, one group with a low tidal volume (6 ml/kg), and the other with a high tidal volume (24 ml/kg). In the third group animals were kept in spontaneous ventilation for 1 h.After ventilation the right lung was used to quantify polymorphonuclear infiltration. The left lung was divided into dependent and nondependent regions, and expression of IL-1beta and PC III mRNA was quantified by northern blot analysis. The group ventilated with low tidal volume had greater polymorphonuclear infiltration IL-1beta and PC III mRNA expression than the nonventilated group. Similar results were observed with high tidal volumes. There was no difference between low and high tidal volume ventilation. Expression levels of IL-1beta and PC III mRNA were higher in the nondependent region of ventilated groups and equal in the nonventilated group.Even a low tidal volume mode of mechanical ventilation induces proinflammatory and profibrogenic response, with a nondependent predominance for IL-1beta and PC III mRNA expression in supine, ventilated, previously normal rats.

Although results from recent studies suggest that the female hormone estrogen promotes lung cancer development, the molecular mechanisms underlying this oncogenic activity remain to be elucidated. Previous data from this group demonstrate that estrogen is metabolized extensively within the murine lung. Exposure to tobacco smoke accelerates the production of 4-hydroxyestrogen, a putative carcinogenic estrogen metabolite that is generated primarily by CYP1B1. While these data suggest that mutagenic estrogen metabolites may contribute to lung tumorigenesis in mice, their relevance to humans remains unknown. The purpose of the present study was to determine the capacity of the human lung to synthesize and metabolize estrogen and to compare the profile of estrogen metabolites in tumor tissue from non-small cell lung cancer (NSCLC) patients with that of adjacent normal tissue. Quantitative RT-PCR analyses revealed expression of estrogen synthesis (CYP17A1, CYP21, HSD17B3 and HSD17B7) and metabolism (CYP1B1, GSTA4, GSTT1, NQO1 and COMT) genes in 50-100% of both normal lung specimens and tumors. Transcripts for the estrogen synthesis genes CYP19 and HSD3B1 were present only in tumors, while expression of GSTM1 and CYP1A1 was detected in less than 30% of both normal and tumor tissues. Expression of ERα and ERβ was comparable in both tissue types. However, tumors exhibited decreased levels of HSD17B3 (converts androstenedione to testosterone) and progesterone receptor transcripts as compared to normal tissue. In order to determine if the detected genes are functional, the estrogen metabolite profile of human lung tissue was established using liquid chromatography/tandem mass spectrometry (LC-MS2) technology. Examination of paired normal and tumor specimens from 9 female patients with NSCLC indicated the presence of 3 estrogens (E1, E2 and E3) and 6 estrogen metabolites (2-OHE1, 2-OHE2, 4-OHE1, 4-OHE2, 2-OMeE1 and 2-OMeE2). With the exception of 2-OMeEs, putative protective estrogen metabolites, the levels of all estrogen species examined were elevated 1.5-2 fold in tumor tissue as compared to adjacent normal tissue (P Citation Format: Jing Peng, Xia Xu, Sibele Meireles, Stacy Mosier, Michael Slifker, Karthik Devarajan, Margie Clapper. Estrogen synthesis and metabolism within the lungs of non-small cell lung cancer patients. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr A05.

BACKGROUND: Sebaceous lesions comprises: hyperplasias, adenomas, sebaceomas and carcinomas, which can be sporadic or related to hereditary syndrome. Keratoacanthomas, as sebaceous lesions, can be associated with hereditary cancer, known as Muir-Torre syndrome (MTS), accepted as the presence of those lesions with visceral malignancies, mainly colorectal cancer (CRC). Besides, a subset of MTS shares clinical, pathological and molecular characteristics with hereditary nonpolyposis colorectal cancer (HNPCC). BRAF mutation has already been described in sebaceous hyperplasia but not in CRC in an onset of HNPCC. BRAF oncogene is part of the MAPKinase pathway, as KRAS gene, and both have an important role in cellular proliferation, differentiation and apoptosis. OBJECTIVE: The aim of this study was to evaluate the role of KRAS and BRAF genes in sebaceous lesions and keratoacanthomas in a sporadic setting or related with hereditary syndrome. METHODS: KRAS (codons 12 and 13) and BRAF (V600E) mutations of 52 sebaceous lesions and 42 keratoacanthomas samples was evaluated using Sanger sequencing. 24 (46%) patients had at least one internal malignancy and thus were diagnosed as having MTS. 7 (14%) patients presented clinical or molecular diagnosis of HNPCC. RESULTS: Mutations in KRAS was identified in 7 of 52 cases (14%) of sebaceous lesions and 10 of 42 cases (24%) of keratoacanthomas. BRAF mutation was observed in 2 of 52 cases (4%) of sebaceous lesions and 2 of 42 cases (5%) of Keratoacanthomas. Mutation in both genes occurred in just one sample. There was no statistical significance observed with clinical variables. BRAF mutation was not observed in patients with diagnosis of MTS or HNPCC. CONCLUSION: The results indicate that KRAS and BRAF mutations play a distinct role in carcinogenesis of sebaceous lesions and KRAS a more important event in keratoacanthomas. KRAS mutation was observed in sporadic lesions and also related to hereditary syndrome. However, BRAF mutation was not observed in patients with HNPCC, concordant with observed in CRC with the same origin. 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 335. doi:10.1158/1538-7445.AM2011-335

Lung cancer is the leading cause of cancer deaths in the U.S., surpassing breast cancer as the primary cause of cancer-related mortality in women. The goal of the present study was to identify early molecular changes in the lung induced by exposure to tobacco smoke and thus identify potential targets for chemoprevention. Female A/J mice were exposed to either tobacco smoke or HEPA-filtered air via a whole-body exposure chamber (6 h/day; 5 days/wk for 3, 8 and 20 wk). Gene expression profiles of lung tissue from control and smoke-exposed animals were established using a 15 K cDNA microarray. Cytochrome P450 1b1 (Cyp1b1), a Phase I enzyme involved in both the metabolism of xenobiotics and the 4-hydroxylation of 17β estradiol, was modulated to the greatest extent following smoke exposure. A panel of 10 genes were found to be differentially expressed between control and smoke-exposed animals at 3, 8 and 20 wk (P < 0.001). The interaction network of these differentially expressed genes revealed new pathways modulated by short-term smoke exposure including estrogen metabolism. In addition, 17β-estradiol was detected within murine lung tissue by gas chromatography coupled mass spectroscopy and immunohistochemistry. Identification of the early molecular events that contribute to lung tumor formation is anticipated to lead to the development of promising targeted chemopreventive therapies. In conclusion, the presence of 17β-estradiol within lung tissue when combined with the modulation of Cyp1b1 and other estrogen metabolism genes by tobacco smoke provides novel insight into a possible role for estrogens in lung cancer. (Supported by fellowships from the AACR-Pennsylvania Department of Health and the ASPO/CRPF, the Estate of Jane Villon, the Jerome M. Spencer and Arnold Zaslow Family Foundation and NIH CA-96310). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2459.

Point mutations in codons 12, 13 and 61 of the KRAS gene and in codon 600 of the BRAF gene can be identified in neoplastic tissue from many different organs. KRAS mutations are present in approximately 40% of sporadic colorectal adenocarcinoma and this analysis is important to determine response to anti-EGFR targeted therapies. BRAF mutations are present in approximately 40% of the papillary thyroid carcinoma and 10% sporadic colorectal adenocarcinoma. BRAF mutation V600E is associated with more aggressive thyroid tumors and its detection is important for thyroid cancer prognosis. Detection of BRAF mutation in colorectal cancer is also useful for characterizing sporadic tumors. A method developed in-house to identify mutations in KRAS (codos 12 and 13) was validated using formalin-fixed paraffin-embeded tissue (FFPE) from 31 surgical specimens diagnosed with metastatic colorrectal cancer. Similarly, a method for BRAF mutation analysis (codon 600) was developed and validaded using 39 smears from fine needle aspiration biopsies (FNAB) of the thyroid. Cytology analysis was performed to confirm the presence of neoplastic or suspected lesions. DNA was extract from 1-4 slides containing the FFPE tissue sections or smears from the FNAB. Mutation analyses were performed by Sanger sequencing and results were compared with pyrosequencing. Automatic analysis of nucleotide sequences using the SeqScape software (Applied Biosystems) was also implemented. There was 85.7% and 100% concordance between Sanger sequencing and pyrosequencing for KRAS and BRAF mutation analysis, respectively. Precision intra- and inter-assay analysis were concordant, with the exception of one sample with KRAS mutation. Sensitivity was evaluated using pools containing different proportions of mutated and non-mutated DNA from cell lines. Data analysis by visual inspection resulted in the detection of the mutant allele in samples with at least 10% of mutated DNA. However, KRAS and BRAF mutations analysis using SeqScape software detected the mutant allele in pools containing at least 30% and 25% of mutated DNA, respectively. This is in accordance with the overall sensitivity observed by standard sequencing analysis. In conclusion, KRAS and BRAF mutation analysis can be successfully performed using small amount of tissue sections from FFPE specimens and smears from FNAB. This material has the advantage of requiring small amounts of sample and also allows cytology review of the tissue content prior to molecular analysis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 822.

In acute lung injury (ALI), elevation of procollagen type III (PC III) occurs early and has an adverse impact on outcome. We examined whether different high-inflation strategies of mechanical ventilation (MV) in oleic acid (OA) ALI alter regional expression of PC III. We designed an experimental, randomized, and controlled protocol in which rats were allocated to two control groups (no injury, recruited [alveolar recruitment maneuver after tracheotomy without MV; n = 4 rats] and control [n = 5 rats]) or four injured groups (one exposed to OA only [n = 10 rats] and three OA-injured and ventilated). The three OA-injured groups were ventilated for 1 hour according to the following strategies: LVHP-S (low volume-high positive end-expiratory pressure [PEEP], supine; n = 10 rats, tidal volume [VT] = 8 ml/kg, PEEP = 12 cm H2O), HVLP-S (high volume-low PEEP, supine; n = 10 rats, VT = 20 ml/kg, PEEP = 5 cm H2O), and HVLP-P (high volume-low PEEP, prone; n = 10 rats). Northern blot analysis for PC III and interleukin-1-beta (IL-1β) and polymorphonuclear infiltration index (PMI) counting were performed in nondependent and dependent regions. Regional differences between groups were assessed by two-way analysis of variance after logarithmic transformation and post hoc tests. A significant interaction for group and region effects was observed for PC III (p = 0.012) with higher expression in the nondependent region for HVLP-S and LVHP-S, intermediate for OA and HVLP-P, and lower for control (group effect, p < 0.00001, partial η2 = 0.767; region effect, p = 0.0007, partial η2 = 0.091). We found high expression of IL-1β (group effect, p < 0.00001, partial η2 = 0.944) in the OA, HVLP-S, and HVLP-P groups without regional differences (p = 0.16). PMI behaved similarly (group effect, p < 0.00001, partial η2 = 0.832). PC III expression is higher in nondependent regions and in ventilatory strategies that caused overdistension. This response was partially attenuated by prone positioning.