Circulating Tumor Cell Counts in Patients With Localized Prostate Cancer Including Those Under Active Surveillance.

AIM: To evaluate the clinical efficacy of a circulating tumor cell (CTC) test by comparison between healthy volunteers and patients with localized prostate cancer including those under active surveillance. MATERIALS AND METHODS: CTC counts in peripheral blood were compared between patients with prostate cancer (n=45) and healthy volunteers (n=17). CTCs were identified based on the expression of epithelial cell adhesion molecule (EpCAM) and counted using a SMART BIOPSY™ SYSTEM. RESULTS: The number of EpCAM+ cells was significantly higher in patients with cancer than in healthy volunteers. Among the low-risk patients (n=9), two had up-staging and six had up-grading. Among those up-staged, there was one case which was EpCAM+ Among those cases up-graded, three were EpCAM+ In those with stage T2 tumors, the presence of Gleason pattern 5 was positively correlated with EpCAM positivity (rho=0.59, p<0.001). CONCLUSION: CTC counts in localized prostate cancer were associated with Gleason pattern 5. Active treatment should be considered for patients with low-risk disease during active surveillance who are found to have EpCAM+ CTCs because of a risk of up-staging and up-grading.

Identification of circulating tumor cells with EML4-ALK translocation using fluorescence in situ hybridization in advanced ALK-positive patients with lung cancer.

[This retracts the article DOI: 10.3892/ol.2018.8480.].

Identification of circulating tumor cells with EML4-ALK translocation using fluorescence in situ hybridization in advanced ALK-positive patients with lung cancer.

Analysis of anaplastic lymphoma kinase (ALK) rearrangement in non-small cell lung cancer (NSCLC) is considered to be a useful tool when considering predictive biomarker detection for evaluating eligibility for targeted therapy. It is not always possible to perform a tumor biopsy in patients. Isolation and culturing of circulating tumor cells (CTCs) may be an alternative to tumor biopsies for the diagnosis of ALK rearrangement. Blood was collected from 22 patients with NSCLC harboring ALK rearrangement and was divided into two groups: One for immunofluorescence staining and the other for culture. Samples were filtered by size and cultured CTCs were analyzed for echinoderm microtubule-associated protein-like 4-ALK translocation using fluorescence in situ hybridization. CTCs positive for epithelial cell adhesion molecule and CTCs exhibiting ALK rearrangement were detected. Therefore, CTCs may be used as a potential alternative method to tissue biopsy for diagnosing ALK rearrangement. Additionally, this method may have clinical applications including serial blood sampling for the development of personalized cancer therapy based on individual genomic information.

An intrinsically disordered domain in Polaribacter irgensii KOPRI 22228 CspB confers extraordinary freeze-tolerance.

Organisms living in extremely cold environments possess mechanisms to survive low temperatures. Among the known cold-induced genes, cold-shock proteins (Csps) are the most prominent. A csp-homologous gene, cspBPi, has been cloned from the Arctic bacterium Polaribacter irgensii KOPRI 22228, and overexpression of this gene greatly increased the freezing tolerance of its host. This protein consists of a unique N-terminal domain and a well conserved C-terminal cold shock domain. To elucidate the detailed mechanisms involved in the extraordinary freeze-tolerance conferred by CspBPi, we identified the responsible domain by mutational analysis. Changes of residues in the cold shock domain that are crucial for binding RNA or single-stranded DNA did not impair the ability of the host to survive freezing stress. All domain-shuffled CspBPi variants containing the N-terminal domain retained the ability to confer superior freeze-tolerance. Slow electrophoretic mobility and far-UV circular dichroism spectra of the N-terminal domain suggested an intrinsically disordered structure for this region. The N-terminal domain also bound to lipid vesicles in vitro. This lipid vesicle binding characteristic is shared with other intrinsically disordered proteins, such as α-synuclein and plant dehydrins, known to confer cold-tolerance when overexpressed, suggesting a mechanism for cold-survival through membrane binding.

Cancer gene panel analysis of cultured circulating tumor cells and primary tumor tissue from patients with breast cancer.

Although numerous effective therapies have improved the survival rate of patients with breast cancer, a number of patients present with tumor recurrence and metastasis. A liquid biopsy of circulating tumor cells (CTC) is a non-invasive method to obtain tumor cells and may be used as substitute for a tumor tissue biopsy. The present study focuses on determining whether CTC culture is an optimal method of obtaining sufficient amounts of CTCs for molecular analysis. The current study demonstrates a method of isolating and culturing CTCs from patients with breast cancer and the construction of a molecular profile of cultured cells using the Ion AmpliSeq Cancer Gene Panel V2. Gene mutations that were observed in cultured CTCs were compared with those observed in primary tumor tissues. CTCs were isolated and cultured from the blood of six patients with breast cancer. Mutations from the Catalogue Of Somatic Mutation In Cancer (COSMIC) were detected in Platelet-Derived Growth Factor Receptor Alpha, MET (also known as Hepatocyte Growth Factor Receptor), Phosphatase and Tensin Homolog, Harvey Rat Sarcoma Viral Oncogene Homolog, SWI/SNF Related, Matrix Associated, Actin Dependent Regulator of Chromatin Subfamily B Member 1, Cyclin Dependent Kinase Inhibitor 2A and MutL Homolog 1 genes in 5/6 samples. A comparison between mutations detected in cultured CTCs and mutations detected in primary tumor tissues demonstrated that a large number of mutations that were identified in CTCs were also detected in primary tumor tissues. The results from the current study describe a novel cell culture approach that may be used to obtain an optimal number of CTCs for molecular analysis. This novel approach is able to be used as a tool for liquid biopsy during breast cancer treatment.

Overexpression of cold shock protein A of Psychromonas arctica KOPRI 22215 confers cold-resistance.

A polar bacterium was isolated from Arctic sea sediments and identified as Psychromonas artica, based on 16S rDNA sequence. Psychromonas artica KOPRI 22215 has an optimal growth temperature of 10 degrees C and a maximum growth temperature of 25 degrees C, suggesting this bacterium is a psychrophile. Cold shock proteins (Csps) are induced upon temperature downshift by more than 10 degrees C. Functional studies have researched mostly Csps of a mesophilic bacterium Escherichia coli, but not on those of psychrophilic bacteria. In an effort to understand the molecular mechanisms of psychrophilic bacteria that allow it withstand freezing environments, we cloned a gene encoding a cold shock protein from P. artica KOPRI 22215 (CspA(Pa)) using the conserved sequences in csp genes. The 204 bp-long ORF encoded a protein of 68 amino acids, sharing 56% homology to previously reported E. coli CspA protein. When CspA(Pa) was overexpressed in E. coli, it caused cell growth-retardation and morphological elongation. Interestingly, overexpression of CspA(Pa) drastically increased the host's cold-resistance by more than ten times, suggesting the protein aids survival in polar environments.

Rescue of a cold-sensitive mutant at low temperatures by cold shock proteins from Polaribacter irgensii KOPRI 22228.

Exposure to low temperatures induces the biosynthesis of specific sets of proteins, including cold shock proteins (Csps). Since many of the specific functions of pychrophilic Csps are unknown, the roles of Csps from an Arctic bacterium, Polaribacter irgensii KOPRI 22228, were examined. The genes encoding CspA and CspC of P. irgensii were cloned in this study. Sequence analysis showed that these proteins have cold shock domains containing two RNA-binding motifs, RNP1 and RNP2. Both proteins bound oligo(dT)-cellulose resins, suggesting single-stranded nucleic acid-binding activity. When the P. irgensii Csps were overexpressed in Escherichia coli, the cold-resistance of the host was increased by more than five-fold. The P. irgensii Csps also rescued a cold-sensitive E. coli csp-quadruple deletion strain, BX04, at low temperatures. These results suggest that Csps from P. irgensii play a role in survival in polar environments.