Clinical and pathological characteristics associated with the presence of the IS6110 Mycobacterim tuberculosis transposon in neoplastic cells from non-small cell lung cancer patients.

Lung cancer (LC) and pulmonary tuberculosis (TB) are the deadliest neoplastic and bacterial infectious diseases worldwide, respectively. Clinicians and pathologists have long discussed the co-existence of LC and TB, and several epidemiologic studies have presented evidence indicating that TB could be associated with the development of LC, particularly adenocarcinoma. Nonetheless, this data remains controversial, and the mechanism which could underlie the association remains largely unexplored. Some bioinformatic studies have shown that human cancer biopsies have a very high frequency of bacterial DNA integration; since Mycobacterium Tuberculosis (MTb) is an intracellular pathogen, it could play an active role in the cellular transformation. Our group performed an exploratory study in a cohort of 88 LC patients treated at the Instituto Nacional de Cancelorogía (INCan) of Mexico City to evaluate the presence of MTb DNA in LC tissue specimens. For the first time, our results show the presence of the MTb IS6110 transposon in 40.9% (n = 36/88) of patients with lung adenocarcinomas. Additionally, through in-situ PCR we identified the presence of IS6110 in the nuclei of tumor cells. Furthermore, shotgun sequencing from two samples identified traces of MTb genomes present in tumor tissue, suggesting that similar Mtb strains could be infecting both patients.

Targeted RNA-Seq Reveals the M. tuberculosis Transcriptome from an In Vivo Infection Model.

The study of host-pathogen interactions using in vivo models with intracellular pathogens like Mycobacterium tuberculosis (Mtb) entails technical limitations, such as: (i) Selecting an efficient differential lysis system to enrich the pathogen cells; (ii) obtaining sufficient high-quality RNA; and (iii) achieving an efficient rRNA depletion. Thus, some authors had used flow cytometers to separate infected cells or significantly increase the sequencing depth of host-pathogen RNA libraries to observe the pathogens' gene expression. However, these options carry additional expenses in specialized equipment typically not available for all laboratories. Here, we propose an experimental protocol involving differential cell lysis and a probe-based ribosomal depletion to determine the gene expression of Mtb and its host during in vivo infection. This method increased the number of observed pathogen-expressed genes from 13 using the traditional RNA-seq approach to 702. After eliminating rRNA reads, we observed that 61.59% of Mtb sequences represented 702 genes, while 38.41% represented intergenic regions. Some of the most expressed genes codified for IS1081 (Rv2512c) transposase and eight PE-PGRS members, such as PGRS49 and PGRS50. As expected, a critical percent of the expressed genes codified for secreted proteins essential for infection, such as PE68, lppN, and LpqH. Moreover, three Mtb ncRNAs were highly expressed (small RNA MTS2823, transfer-messenger RNA RF00023, and ribozyme RF00010). Many of the host-expressed genes were related to the inflammation process and the expression of surfactant proteins such as the Sftpa and Sftpc, known to bind Mtb to alveolar macrophages and mi638, a microRNA with no previous associations with pulmonary diseases. The main objective of this study is to present the method, and a general catalog of the Mtb expressed genes at one point of the in vivo infection. We believe our method represents a different approach to the existing ones to study host-pathogen interactions in tuberculosis and other similar intracellular infections, without the necessity of specialized equipment.

Impact of detecting plasma EGFR mutations with ultrasensitive liquid biopsy in outcomes of NSCLC patients treated with first- or second-generation EGFR-TKIs.

BACKGROUND: Few trials have evaluated the utility of liquid biopsies to detect epidermal growth factor receptor mutations (EGFRm) at the time of response evaluation and its association with the clinical characteristics and outcomes of non-small-cell lung cancer (NSCLC) patients. OBJECTIVE: This study aimed to evaluate, in a real-world clinical setting, the prevalence of plasma EGFRm and its association with the clinical characteristics, response and survival outcomes of NSCLC patients under treatment with EGFR-tyrosine kinase inhibitors (EGFR-TKIs). METHODS: This observational study enrolled advanced or metastatic NSCLC patients, with confirmed tumor EGFRm, receiving treatment with first- or second-generation EGFR-TKIs. Blood samples for the detection of plasma EGFRm were collected at the time of response evaluation and processed using the Target Selector™ assay. The main outcomes were the detection rate of plasma EGFRm, median Progression-Free Survival (PFS) and Overall Survival (OS) according to plasma EGFR mutational status. RESULTS: Of 84 patients, 50 (59.5%) had an EGFRm detected in plasma. After a median follow-up of 21.1 months, 63 patients (75%) had disease progression. The detection rate of plasma EGFRm was significantly higher in patients with disease progression than in patients with partial response or stable disease (68.3% versus 33.3%; P< 0.01). PFS and OS were significantly longer in patients without plasma EGFRm than among patients with plasma EGFRm (14.3 months [95% CI, 9.25-19.39] vs 11.0 months [95% CI, 8.61-13.46]; P= 0.034) and (67.8 months [95% CI, 39.80-95.94] vs 32.0 months [95% CI, 17.12-46.93]; P= 0.006), respectively. A positive finding in LB was associated with the presence of ⩾ 3 more metastatic sites (P= 0.028), elevated serum carcinoembryonic (CEA) at disease progression (P= 0.015), and an increase in CEA with respect to baseline levels (P= 0.038). CONCLUSIONS: In NSCLC patients receiving EGFR-TKIs, the detection of plasma EGFRm at the time of tumor response evaluation is associated with poor clinical outcomes.

Evidence for the Effect of Vaccination on Host-Pathogen Interactions in a Murine Model of Pulmonary Tuberculosis by Mycobacterium tuberculosis.

The global control of Tuberculosis remains elusive, and Bacillus Calmette-Guérin (BCG) -the most widely used vaccine in history-has proven insufficient for reversing this epidemic. Several authors have suggested that the mass presence of vaccinated hosts might have affected the Mycobacterium tuberculosis (MTB) population structure, and this could in turn be reflected in a prevalence of strains with higher ability to circumvent BCG-induced immunity, such as the recent Beijing genotype. The effect of vaccination on vaccine-escape variants has been well-documented in several bacterial pathogens; however the effect of the interaction between MTB strains and vaccinated hosts has never been previously described. In this study we show for the first time the interaction between MTB Beijing-genotype strains and BCG-vaccinated hosts. Using a well-controlled murine model of progressive pulmonary tuberculosis, we vaccinated BALB/c mice with two different sub-strains of BCG (BCG-Phipps and BCG-Vietnam). Following vaccination, the mice were infected with either one of three selected MTB strains. Strains were selected based on lineage, and included two Beijing-family clinical isolates (strains 46 and 48) and a well-characterized laboratory strain (H37Rv). Two months after infection, mice were euthanized and the bacteria extracted from their lungs. We characterized the genomic composite of the bacteria before and after exposure to vaccinated hosts, and also characterized the local response to the bacteria by sequencing the lung transcriptome in animals during the infection. Results from this study show that the interaction within the lungs of the vaccinated hosts results in the selection of higher-virulence bacteria, specifically for the Beijing genotype strains 46 and 48. After exposure to the BCG-induced immune response, strains 46 and 48 acquire genomic mutations associated with several virulence factors. As a result, the bacteria collected from these vaccinated hosts have an increased ability for immune evasion, as shown in both the host transcriptome and the histopathology studies, and replicates far more efficiently compared to bacteria collected from unvaccinated hosts or to the original-stock strain. Further research is warranted to ascertain the pathways associated with the genomic alterations. However, our results highlight novel host-pathogen interactions induced by exposure of MTB to BCG vaccinated hosts.

Tuberculosis and lung cancer.

OBJECTIVE: To describe the epidemiological studies about the relationship between lung cancer (LC) and pulmonary tuberculosis (Tb) and its possible molecular mechanisms. MATERIALS AND METHODS: We reviewed research databases in search of publications that included keywords LC and Tb. RESULTS: It has been proposed that chronic inflammation in the lungs due to Tb could cause clastogenic activity in the DNA of bronchial epithelium. Another possibility is lateral gene transfer; since Mycobacterium tuberculosis (MTb) is an intracellular organism, bacterial DNA could integrate to bronchial epithelial cells inducing neoplastic transformation. CONCLUSIONS: There are epidemiological reports, particularly from Asian countries, which confirm a relationship between LC and Tb. MTb could play an active role in cellular transformation and it is important to elucidate the mecha- nism involved.

OBJETIVO: Describir los estudios que documentan la relación entre el cáncer de pulmón (CP) y la tuberculosis pulmonar (Tb) y sus posibles mecanismos moleculares. MATERIAL Y MÉTODOS: Se revisaron bases de datos de publicaciones, usando como palabras clave CP y Tb. RESULTADOS: Se ha propuesto que la inflamación crónica en el pulmón provocada por la Tb podría producir actividad clastogénica. Otra posibilidad es la transferencia lateral de genes; Mycobacterium tuberculosis (MTb) es un organismo intracelular facultativo cuyo DNA podría integrarse al material genético del epitelio bronquial induciendo transformación neoplásica. CONCLUSIONES: Existen evidencias epidemiológicas, particularmente en países asiáticos, que documentan la relación entre CP y Tb. MTb podría desempeñar un papel activo en la transformación neoplásica cuyo mecanismo debe de ser elucidado.

Tuberculosis and lung cancer / Tuberculosis y cáncer de pulmón

Abstract: Objective: To describe the epidemiological studies about the relationship between lung cancer (LC) and pulmonary tuberculosis (Tb) and its possible molecular mechanisms. Materials and methods: We reviewed research databases in search of publications that included keywords LC and Tb. Results: It has been proposed that chronic inflammation in the lungs due to Tb could cause clastogenic activity in the DNA of bronchial epithelium. Another possibility is lateral gene transfer; since Mycobacterium tuberculosis (MTb) is an intracellular organism, bacterial DNA could integrate to bronchial epithelial cells inducing neoplastic transformation. Conclusions: There are epidemiological reports, particularly from Asian countries, which confirm a relationship between LC and Tb. MTb could play an active role in cellular transformation and it is important to elucidate the mechanism involved.

Resumen: Objetivo: Describir los estudios que documentan la relación entre el cáncer de pulmón (CP) y la tuberculosis pulmonar (Tb) y sus posibles mecanismos moleculares. Material y métodos: Se revisaron bases de datos de publicaciones, usando como palabras clave CP y Tb. Resultados: Se ha propuesto que la inflamación crónica en el pulmón provocada por la Tb podría producir actividad clastogénica. Otra posibilidad es la transferencia lateral de genes; Mycobacterium tuberculosis (MTb) es un organismo intracelular facultativo cuyo DNA podría integrarse al material genético del epitelio bronquial induciendo transformación neoplásica. Conclusión: Existen evidencias epidemiológicas, particularmente en países asiáticos, que documentan la relación entre CP y Tb. MTb podría desempeñar un papel activo en la transformación neoplásica cuyo mecanismo debe de ser elucidado.

Differential gene expression profiles according to the Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society histopathological classification in lung adenocarcinoma subtypes.

The current lung cancer classification from the Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society has considerably changed the pathologic diagnosis of lung invasive adenocarcinoma, identifying disease subtypes with substantial implications for medical practice, such as clinical, radiological, molecular, and prognostic differences. We analyzed the differences in the genetic expression of adenocarcinoma subtypes according to the new classification. Microarray gene expression analysis was performed on a cohort of 29 adenocarcinoma patients treated at the Instituto Nacional de Cancerología of Mexico from 2008 to 2011. All patients had an available biopsy sample and were classified into 4 different subtypes of adenocarcinoma (2015 World Health Organization classification). Lepidic-predominant adenocarcinoma was the only pattern that exhibited a marked gene expression difference compared with other predominant histologic patterns, revealing genes with significant expression (P < .01). Moreover, we identified 13 genes with specific differential expression in the lepidic-predominant adenocarcinoma that could be used as a gene signature. The lepidic-predominant histologic pattern has a differential gene expression profile compared with all predominant histologic patterns. Additionally, we identified a gene expression signature of 13 genes that have a unique behavior in the lepidic histologic pattern; these 13 genes are candidates for follow-up studies for their potential use as biomarkers or therapeutic targets. Results from this study highlight the importance of the new Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society classification and exemplify the potential clinical implications of correlating histopathology with exclusive molecular beacons.

Secretome Prediction of Two M. tuberculosis Clinical Isolates Reveals Their High Antigenic Density and Potential Drug Targets.

The Excreted/Secreted (ES) proteins play important roles during Mycobacterium tuberculosis invasion, virulence, and survival inside the host and they are a major source of immunogenic proteins. However, the molecular complexity of the bacillus cell wall has made difficult the experimental isolation of the total bacterial ES proteins. Here, we reported the genomes of two Beijing genotype M. tuberculosis clinical isolates obtained from patients from Vietnam (isolate 46) and South Africa (isolate 48). We developed a bioinformatics pipeline to predict their secretomes and observed that ~12% of the genome-encoded proteins are ES, being PE, PE-PGRS, and PPE the most abundant protein domains. Additionally, the Gene Ontology, KEGG pathways and Enzyme Classes annotations supported the expected functions for the secretomes. The ~70% of an experimental secretome compiled from literature was contained in our predicted secretomes, while only the 34-41% of the experimental secretome was contained in the two previously reported secretomes for H37Rv. These results suggest that our bioinformatics pipeline is better to predict a more complete set of ES proteins in M. tuberculosis genomes. The predicted ES proteins showed a significant higher antigenic density measured by Abundance of Antigenic Regions (AAR) value than the non-ES proteins and also compared to random constructed secretomes. Additionally, we predicted the secretomes for H37Rv, H37Ra, and two M. bovis BCG genomes. The antigenic density for BGG and for isolates 46 and 48 was higher than the observed for H37Rv and H37Ra secretomes. In addition, two sets of immunogenic proteins previously reported in patients with tuberculosis also showed a high antigenic density. Interestingly, mice infected with isolate 46 showed a significant lower survival rate than the ones infected with isolate 48 and both survival rates were lower than the one previously reported for the H37Rv in the same murine model. Finally, after a druggability analysis of the secretomes, we found potential drug targets such as cytochrome P450, thiol peroxidase, the Ag85C, and Ribonucleoside Reductase in the secreted proteins that could be used as drug targets for novel treatments against Tuberculosis.

Gene-expression profiles in lung adenocarcinomas related to chronic wood smoke or tobacco exposure.

BACKGROUND: Tobacco-smoke is the major etiological factor related to lung cancer. However, other important factor is chronic wood smoke exposure (WSE). Approximately 30 % of lung cancer patients in Mexico have a history of WSE, and present different clinical, pathological and molecular characteristics compared to tobacco related lung cancer, including differences in mutational profiles. There are several molecular alterations identified in WSE associated lung cancer, however most studies have focused on the analysis of changes in several pathogenesis related proteins. METHODS: Our group evaluated gene expression profiles of primary lung adenocarcinoma, from patients with history of WSE or tobacco exposure. Differential expression between these two groups were studied through gene expression microarrays. RESULTS: Results of the gene expression profiling revealed 57 statistically significant genes (p < 0.01). The associated biological functional pathways included: lipid metabolism, biochemistry of small molecules, molecular transport, cell morphology, function and maintenance. A highlight of our analysis is that three of the main functional networks represent 37 differentially expressed genes out of the 57 found. These hubs are related with ubiquitin C, GABA(A) receptor-associated like protein; and the PI3K/AKT and MEK/ERK signaling pathways. CONCLUSION: Our results reflect the intrinsic biology that sustains the development of adenocarcinoma related to WSE and show that there is a different gene expression profile of WSE associated lung adenocarcinoma compared to tobacco exposure, suggesting that they arise through different carcinogenic mechanisms, which may explain the clinical and mutation profile divergences between both lung adenocarcinomas.