ABSTRACT
Circulating tumor cells (CTC) have been studied in various solid tumors but clinical utility of CTC in small cell lung cancer (SCLC) remains unclear. The aim of the CTC-CPC study was to develop an EpCAM-independent CTC isolation method allowing isolation of a broader range of living CTC from SCLC and decipher their genomic and biological characteristics. CTC-CPC is a monocentric prospective non-interventional study including treatment-naïve newly diagnosed SCLC. CD56+ CTC were isolated from whole blood samples, at diagnosis and relapse after first-line treatment and submitted to whole-exome-sequencing (WES). Phenotypic study confirms tumor lineage and tumorigenic properties of isolated cells for the 4 patients analyzed with WES. WES of CD56+ CTC and matched tumor biopsy reveal genomic alteration frequently impaired in SCLC. At diagnosis CD56+ CTC were characterized by a high mutation load, a distinct mutational profile and a unique genomic signature, compared to match tumors biopsies. In addition to classical pathways altered in SCLC, we found new biological processes specifically affected in CD56+ CTC at diagnosis. High numeration of CD56+ CTC (> 7/ml) at diagnosis was associated with ES-SCLC. Comparing CD56+ CTC isolated at diagnosis and relapse, we identify differentially altered oncogenic pathways (e.g. DLL3 or MAPK pathway). We report a versatile method of CD56+ CTC detection in SCLC. Numeration of CD56+ CTC at diagnosis is correlated with disease extension. Isolated CD56+ CTC are tumorigenic and show a distinct mutational profile. We report a minimal gene set as a unique signature of CD56+ CTC and identify new affected biological pathways enriched in EpCAM-independent isolated CTC in SCLC.
Subject(s)
Lung Neoplasms , Neoplastic Cells, Circulating , Small Cell Lung Carcinoma , Humans , Epithelial Cell Adhesion Molecule , Clinical Relevance , Prospective Studies , Genomics , Carcinogenesis , Membrane Proteins , Intracellular Signaling Peptides and ProteinsABSTRACT
Aspergillus pathogens usually infect immunocompromised patients with lethal outcome. We report a multiplex PCR assay for the discrimination of the most frequent Aspergillus pathogens, A. fumigatus, A. flavus, A. niger and A. terreus, through distinct amplicons of 250 bp, 200 bp, 150 bp and 450 bp respectively, derived from the rDNA gene of A. terreus and the aspergillopepsin genes of the remaining species.
Subject(s)
Aspergillosis/diagnosis , Aspergillosis/microbiology , Aspergillus flavus/isolation & purification , Aspergillus fumigatus/isolation & purification , Aspergillus niger/isolation & purification , DNA, Fungal/genetics , DNA, Fungal/isolation & purification , Mycological Typing Techniques , Polymerase Chain Reaction/methods , Aspergillus flavus/genetics , Aspergillus fumigatus/genetics , Aspergillus niger/genetics , DNA Primers , Humans , Sensitivity and SpecificityABSTRACT
Strain delineation of the emerging opportunistic pathogen Clavispora lusitaniae was studied using 12 strains, including two strains of known opposite mating type, CBS 6936 (h+) and CBS 5094 (h-), and 10 strains isolated between 1998 and 2001 from immunocompromized patients. This retrospective study assessed the occurrence of C. lusitaniae subtypes within and among hospitals, and in outpatients who were regularly screened for fungal infections in the course of radio-chemotherapy. Strain typing was accomplished for the first time using single strand conformation polymorphism (SSCP) analysis of amplicons of the ribosomal DNA internal transcribed spacer (ITS) 1 and 2 regions. The results were compared with those produced by three pulsed-field gel electrophoresis (PFGE) methods and PCR fingerprinting with the minisatellite-specific primer M13. Karyotyping separated 7-9 chromosomes, not 6-8 as previously reported. Pulsotyping of SfiI and NotI digested chromosomes grouped isolates in five and four distinct clusters, respectively. All methods revealed strain heterogeneity, though not as extensive as previously recorded. SSCP analysis of the ITS1 region generated five subtypes, based on a sequencing-confirmed nucleotide polymorphism. The discriminatory power of this method was high. All strains displayed a homogeneous SSCP pattern for the ITS2 region. ITS1 PCR-SSCP appears to allow rapid and reliable delineation of C. lusitaniae strains. Pending examination of a larger sample size and interlaboratory study, this protocol can be recommended for rapid prospective identification of hospital outbreaks.
Subject(s)
DNA, Ribosomal Spacer/analysis , Mycology/methods , Saccharomycetales/classification , Amino Acid Sequence , DNA, Ribosomal Spacer/chemistry , Female , Humans , Male , Molecular Sequence Data , Phylogeny , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Retrospective Studies , Saccharomycetales/geneticsABSTRACT
Developments in medical intervention and the increasing population of patients with immunodeficiencies and transient or long-term immunosuppression have increased the list of yeast species that can cause disease. Candida dubliniensis is a novel species with close genetic relatedness to C. albicans. The two species share many common physiological and biochemical properties thus making their distinction cumbrous. A rapid and inexpensive way to presumptively differentiate between the two species, having previously performed a germ tube test, is the ability of C. dubliniensis to reduce the tetrazolium salt and it is reported for the first time. Microbiological information about new and emerging yeast pathogens, including rapid means for their identification, equips medical microbiologists with the means to identify and physicians to treat effectively infections attributed to unusual yeasts.
