Jelena Stojsic

Identification of genes associated with non-small-cell lung cancer promotion and progression

Lung cancer is the most common cause of neoplasia-related death worldwide. One of the crucial early events in carcinogenesis is the induction of genomic instability and mutator phenotype. We investigated genomic instability in 30 patients with non-small-cell lung cancer (NSCLC) by comparing DNA fingerprints of paired tumor and normal tissues using arbitrarily primed polymerase chain reaction (AP-PCR). Selected 21 DNA bands with altered mobility were isolated from polyacrylamide gels, cloned and sequenced. Obtained sequences were submitted to homology search in GenBank database which revealed the following genes: TSPAN14, CDH12, RDH10, CYP4Z1, KIR, E2F4, PHACTR3, PHF20, PRAME family member and SLC2A13. Following the identification of these genes we examined their relation to the clinicopathological parameters and survival of the patients. Our study revealed that genetic alterations of TSPAN14, SLC2A13 and PHF20 appeared prevalently in tumors of grade 1, stage I suggesting that structural changes of these genes could play a role in NSCLC promotion. Contrary to this CYP4Z1, KIR and RDH10 were prevalently mutated in tumors of grade 3, stage III suggesting that they could play a role in NSCLC progression. E2F4, PHACTR3, PRAME family member and CDH12 most probably play important role in NSCLC geneses. In conclusion, our study revealed altered genes previously not described in regard to this type of cancer.

Aberrant Promoter Methylation of CDH13 and MGMT Genes is Associated With Clinicopathologic Characteristics of Primary Non–Small-Cell Lung Carcinoma

UNLABELLED Non–small-cell lung carcinoma (NSCLC) (n = 65) were analyzed for promoter methylation of RASSF1A, CDH13, MGMT, ESR1, and DAPK genes in matching lung tumors, normal lung tissue, and blood samples. Aberrant methylation in CDH13 and MGMT was associated with clinicopathologic features of NSCLC. Hypermethylation detected in primary tumors was not observed in corresponding blood samples, which rendered this an unsuitable blood-based test for NSCLC detection. INTRODUCTION Systemic methylation changes may be a diagnostic marker for tumor development or prognosis. Here, we investigate the relationship between gene methylation in lung tumors relative to normal lung tissue and whether DNA methylation changes can be detected in paired blood samples. MATERIAL AND METHODS Sixty-five patients were enrolled in a surgical case series of non-small-cell lung carcinoma at a single institution. By using bisulfite pyrosequencing, CpG methylation was quantified at 5 genes (RASSF1A, CDH13, MGMT, ESR1, and DAPK) in lung tumor, pathologically normal lung tissue, and circulating blood from enrolled cases. RESULTS The analyses of methylation in tumors compared with normal lung tissue identified higher methylation of CDH13, RASSF1A, and DAPK genes, whereas ESR1 and MGMT methylation did not differ significantly between these tissue types. We then examined whether the 3 aberrantly methylated genes could be detected in blood. The difference in methylation observed in tumors was not reflected in methylation status of matching blood samples, which indicated a low feasibility of detecting lung cancer by analyzing these genes in a blood-based test. Lastly, we probed whether tumor methylation was associated with clinical and demographic characteristics. Histology and sex were associated with methylation at the CDH13 gene, whereas, stage was associated with methylation at MGMT. CONCLUSION Our results showed higher methylation of RASSF1A, CDH13, and DAPK genes in lung tumors compared with normal lung. The lack of reflection of these methylation changes in blood samples from patients with non-small-cell lung carcinoma indicates their poor suitability for a screening test.

Preoperative pulmonary rehabilitation in patients with non-small cell lung cancer and chronic obstructive pulmonary disease

Introduction The aim of this study was to assess the effects of preoperative pulmonary rehabilitation (PPR) on preoperative clinical status changes in patients with chronic obstructive pulmonary disease (COPD) and non-small cell lung cancer (NSCLC), and net effects of PPR and cancer resection on residual pulmonary function and functional capacity. Material and methods This prospective single group study included 83 COPD patients (62 ±8 years, 85% males, FEV1 = 1844 ±618 ml, Tiffeneau index = 54 ±9%) with NSCLC, on 2–4-week PPR, before resection. Pulmonary function, and functional and symptom status were evaluated by spirometry, 6-minute walking distance (6MWD) and Borg scale, on admission, after PPR and after surgery. Results Following PPR significant improvement was registered in the majority of spirometry parameters (FEV1 by 374 ml, p < 0.001; VLC by 407 ml, p < 0.001; FEF50 by 3%, p = 0.003), 6MWD (for 56 m, p < 0.001) and dyspnoeal symptoms (by 1.0 Borg unit, p < 0.001). A positive correlation was identified between preoperative increments of FEV1 and 6MWD (r s = 0.503, p = 0.001). Negative correlations were found between basal FEV1 and its percentage increment (r s = –0.479, p = 0.001) and between basal 6MWD and its percentage change (r s = –0.603, p < 0.001) during PPR. Compared to basal values, after resection a significant reduction of most spirometry parameters and 6MWD were recorded, while Tiffeneau index, FEF25 and dyspnoea severity remained stable (p = NS). Conclusions Preoperative pulmonary rehabilitation significantly enhances clinical status of COPD patients before NSCLC resection. Preoperative increase of exercise tolerance was the result of pulmonary function improvement during PPR. The beneficial effects of PPR were most emphasized in patients with initially the worst pulmonary function and the weakest functional capacity.

Relapse in resected lung cancer revisited: does intensified follow up really matter? A prospective study

Backgroundbeside the well known predominance of distant vs. loco-regional relapse, several aspects of the relapse pattern still have not been fully elucidated.Methodsprospective, controlled study on 88 patients operated for non-small cell lung cancer (NSCLC) in a 15 months period. Stage IIIA existed in 35(39.8%) patients, whilst stages IB, IIA and IIB existed in 10.2%, 4.5% and 45.5% patients respectively. Inclusion criteria: stage I-IIIA, complete resection, systematic lymphadenectomy with at least 6 lymph node groups examined, no neoadjuvant therapy, exact data of all aspects of relapse, exact data about the outcome of the treatment.Resultspostoperative lung cancer relapse occurred in 50(56.8%) patients. Locoregional, distant and both types of relapse occurred in 26%, 70% and 4% patients respectively. Postoperative cancer relapse occurred in 27/35(77.1%) pts. in the stage IIIA and in 21/40(52.55) pts in the stage IIB. In none of four pts. in the stage IIA cancer relapse occurred, unlike 22.22% pts. with relapse in the stage IB. The mean disease free interval in the analysed group was 34.38 ± 3.26 months.The mean local relapse free and distant relapse free intervals were 55 ± 3.32 and 41.62 ± 3.47 months respectively Among 30 pts. with the relapse onset inside the first 12 month after the lung resection, in 20(66.6%) pts. either T3 tumours or N2 lesions existed. In patients with N0, N1 and N2 lesions, cancer relapse occurred in 30%, 55.6% and 70.8% patients respectivelyRadiographic aspect T stage, N stage and extent of resection were found as significant in terms of survival. Related to the relapse occurrence, although radiographic aspect and extent of resection followed the same trend as in the survival analysis, only T stage and N stage were found as significant in the same sense as for survival. On multivariate, only T and N stage were found as significant in terms of survival.Specific oncological treatment of relapse was possible in 27/50(54%) patients.Conclusionthe intensified follow up did not increase either the proportion of patients detected with asymptomatic relapse or the number of patients with specific oncological treatment of relapse.

Intestinal Type of Lung Adenocarcinoma in Younger Adults

Intestinal type of lung adenocarcinoma (ILADC) was initially described by Tsao and Fraser in 1991. Morphology and immunophenotype of ILADC are the same as in colorectal adenocarcinoma. Rectocolonoscopy must be performed to exclude colorectal origin of adenocarcinoma. Colorectal adenocarcinoma claimed to be genetically similar to an ILADC. Patients. We describe 24- and 26-year-old patients of both genders who went under surgery because of a lung tumor mass detected on CT scan. ILADC was diagnosed on resected lung specimens. According to positivity of Cytokeratin20, CDX-2, and Villin, respectively, and negativity of Cytokeratin7, TTF-1, Napsin-A, SurfactantB, MUC-1, and MUC-2, respectively, ILADC was diagnosed. KRAS mutation was detected in tumor tissue of the male patient. Conclusion. Rectocolonoscopy is the only relevant method for distinguishing the intestinal type of lung adenocarcinoma from metastatic colorectal carcinoma because immunohistochemistry and detection of mutation status are frequently the same in both types of adenocarcinoma. More investigations are needed for further understanding of ILADC in purpose of personalized lung carcinoma therapy particularly introducing detection of mutation status, especially in younger patients.

Large Cell Lung Carcinoma with Unusual Imaging Feature, Immunophenotype and Genetic Finding

We present a case of large cell lung carcinoma in sixty-one year old male with typical lung cancer symptoms but unusual radiological presentation and immunophenotype. Tumor morphological finding related to its radiological finding was suggestive for large cell lymphoma or carcinoma, but its immunophenotype made confusion for pathological diagnosis. No p53 mutations were detected in genetic investigation. Multidisciplinar diagnostic approach to some tumors is useful for their final diagnosis.

Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis

BackgroundIdiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease of unknown etiology. A major limitation in transcriptomic profiling of lung tissue in IPF has been a dependence on snap-frozen fresh tissues (FF). In this project we sought to determine whether genome scale transcript profiling using RNA Sequencing (RNA-Seq) could be applied to archived Formalin-Fixed Paraffin-Embedded (FFPE) IPF tissues.ResultsWe isolated total RNA from 7 IPF and 5 control FFPE lung tissues and performed 50 base pair paired-end sequencing on Illumina 2000 HiSeq. TopHat2 was used to map sequencing reads to the human genome. On average ~62 million reads (53.4% of ~116 million reads) were mapped per sample. 4,131 genes were differentially expressed between IPF and controls (1,920 increased and 2,211 decreased (FDR < 0.05). We compared our results to differentially expressed genes calculated from a previously published dataset generated from FF tissues analyzed on Agilent microarrays (GSE47460). The overlap of differentially expressed genes was very high (760 increased and 1,413 decreased, FDR < 0.05). Only 92 differentially expressed genes changed in opposite directions. Pathway enrichment analysis performed using MetaCore confirmed numerous IPF relevant genes and pathways including extracellular remodeling, TGF-beta, and WNT. Gene network analysis of MMP7, a highly differentially expressed gene in both datasets, revealed the same canonical pathways and gene network candidates in RNA-Seq and microarray data. For validation by NanoString nCounter® we selected 35 genes that had a fold change of 2 in at least one dataset (10 discordant, 10 significantly differentially expressed in one dataset only and 15 concordant genes). High concordance of fold change and FDR was observed for each type of the samples (FF vs FFPE) with both microarrays (r = 0.92) and RNA-Seq (r = 0.90) and the number of discordant genes was reduced to four.ConclusionsOur results demonstrate that RNA sequencing of RNA obtained from archived FFPE lung tissues is feasible. The results obtained from FFPE tissue are highly comparable to FF tissues. The ability to perform RNA-Seq on archived FFPE IPF tissues should greatly enhance the availability of tissue biopsies for research in IPF.

Concurrent alteration of p16 and PTEN tumor suppressor genes could be considered as potential molecular marker for specific subgroups of NSCLC patients

p16 and PTEN are tumor suppressors that are commonly inactivated in human cancers. Loss of each of these molecules is widely studied in lung cancer, including non-small cell lung carcinoma (NSCLC), its most common clinical form. However, the importance of their mutual alterations for NSCLC pathogenesis has been barely examined so far. In this study we tested hypothesis that aberrant p16 might cooperate with inactive PTEN during pathogenesis of NSCLC, particularly in promoting tumor aggressiveness and invasiveness. Initially, we screened NSCLC tumor samples from patients for the presence of the most common genetic and epigenetic alterations of p16 and further correlated them with previously detected aberrations in PTEN gene. Statistical analyses showed that aberrant p16 directly correlated with altered PTEN. Such significant correlation was also observed in groups of patients with high genomic instability, with squamocellular histological subtype, with disease grade 2 and with lymph node invasion. Finally, survival analyses revealed dramatic decrease in survival rate of patients with mutual alterations of p16 and PTEN, but without prognostic significance. Overall results implicate cooperation between aberrant p16 and PTEN in pathogenesis of NSCLC and suggest that their combination might be considered as potential molecular marker for specific subgroups of NSCLC patients.

Solitary fibrous pleural tumor associated with loss of consciousness due to hypoglycemia

The patient suffered loss of consciousness, dysarthria and right sided hemiparesis. The CT scan and MRI scans were negative. These findings are more in keeping with a diagnosis of Transient Ischemic Attack (TIA) or mild CVA. Hypoglycemia per se does not usually cause hemiparesis. The blood glucose level was low but I am not sure if one can conclude that hypoglycemia caused the above noted neurological signs and symptoms. The authors do not present any data to prove that this patient had “hypoglycemic coma”.

Liver and splenic sarcoidosis: Diagnostic procedures

INTRODUCTION Clinical studies indicate involvement of the liver and spleen in approximately 20-30% of patients affected with sarcoidosis and their detection should be based on a standardized diagnostic procedure. DIAGNOSTIC PROCEDURES Ultrasonography is a reliable and safe method to assess changes related to size and structure of the affected organs that are pathognomonic for sarcoidosis. Further evaluation may include computerized tomography or magnetic resonance, while percutaneous needle aspiration biopsy or laparoscopy may also be applied when indicated. The most important criterion used for final diagnosis is pathohistological evidence of epithelioid noncaseating granuloma in bioptic material along with already established sarcoidosis of the lungs or some other organ. MATERIAL AND METHODS The study on the incidence of liver and spleen sarcoidosis included a group of 130 patients affected with sarcoidosis hospitalized at the Institute of Pulmonary Diseases and Tuberculosis, Clinical Center of Serbia, over the period 2002-2003. RESULTS The analysis evidenced that 31.5% of sarcoidosis patients had pathognomonic echographic findings of abdominal organs: splenomegaly (13%), hepatomegaly (10.8%) and hepatosplenomegaly (7.7%). Three patients underwent surgical treatment of liver and spleen sarcoidosis. CONCLUSION Pathognomonic findings of liver and spleen sarcoidosis were evidenced in approximately one third of sarcoidosis cases and they represented a significant parameter for further therapy, particularly in chronic patients.