Treatment of aggressive glioblastoma multiforme (GBM) must be based on very precise histological and molecular diagnostic of GBM type. According to the WHO guidelines, only tissue biopsy is a relevant source of cellular material evaluated in the diagnostic process to specify the tumor features. Nevertheless, obtaining a GBM biopsy is complicated and relies mostly on resection surgery. Evaluating circulating free DNA and/or circulating tumor cells (CTCs) in the clinic, using a liquid biopsy could represent a non-invasive cancer care optimization. In the present study, the peripheral blood of patients undergoing GBM resection (n = 18) was collected and examined for CTCs. The feasibility of GBM molecular diagnostics from a simple non-invasive peripheral blood withdrawal was evaluated. The size-based enriched CTCs were analyzed using cytomorphology and their origin confirmed based on mutational analysis. In addition, shared DNA mutations in CTCs and in primary tumor tissue were searched. For the identification of CTCs, next generation sequencing (NGS) was used. The GeneReader™ sequencing platform enables targeted sequencing of a 12-gene panel and direct evaluation of detected gene variations using QIAGEN Clinical Insight Analyze (QCI-A) software with a special algorithm for liquid biopsy sequencing analysis. Herein, we present a standard operating procedure for CTC enrichment in GBM patients, CTC in vitro culture, CTC cytomorphological evaluation, and NGS analysis of CTCs using the QIAGEN Actionable Insights Tumor (ATP) Panel. CTCs were present in all tested patients (18/18). The NGS data generated for formalin-fixed paraffin-embedded (FFPE) primary tumor tissues and CTCs reached significantly high-quality parameters. The comparisons between different sample types (CTCs vs. primary tumors) and sampling area (different primary tumor regions) showed a significant level of concordance, indicating CTC testing could be used for patient monitoring and recurrence awareness. Notably, more mutations were detected when analyzing CTC samples compared with the paired primary tumors (n = 3). The results confirm the feasibility of using CTCs as a source of tumor DNA in a diagnostic process, especially when evaluating the molecular characteristics of GBMs. A major advantage of the presented NGS approach for detecting CTCs is the simultaneous identification of several markers relevant for GBM diagnostics, allowing molecular diagnostics on cytological specimens and potential administration of innovative targeted therapies.
- Publikační typ
- časopisecké články MeSH
BACKGROUND: The occurrence of catamenial pneumothorax (CP) is rare, and the awareness of this diagnosis among physicians is insufficient. CP is highly correlated with pelvic endometriosis and remains the most common form of thoracic endometriosis syndrome. Circulating endometrial cells (CECs) have been previously detected in patients with pelvic endometriosis. Could CECs bring new insights into pneumothorax management? METHODS: This study aims to describe the occurrence and molecular characteristics of CECs in women with spontaneous pneumothorax (SP) (N = 20) with high suspicion of its catamenial character. CECs were enriched from peripheral blood by size-based separation (MetaCell). In addition to cytomorphology, gene expression profiling of captured cells was performed for 24 endometriosis-associated genes. RESULTS: CECs were present in all 20 patients with SP. Enriched CECs exhibited four character features: epithelial, stem cell-like, stroma-like, and glandular. However, not all of them were present in every sampling. Gene expression profiling revealed two distinct phenotypes of CECs in SP and/or CP: one of them refers to the diaphragm openings syndrome and the other to endometrial tissue pleural implantations. Comparisons of the gene expression profiles of CECs in pneumothorax (CECs-SP group) with CECs in pelvic endometriosis (CECs-non-SP group) have revealed significantly higher expression of HER2 in the CECs-SP group compared with the CECs-non-SP group. CONCLUSIONS: This proof-of-concept study demonstrates successful isolation and characterization of CECs in patients with SP. Identification of CECs in SP could alert endometriosis involvement and help early referral to gynecologic consultation for further examination and treatment.
- MeSH
- antigen CA-125 genetika MeSH
- dospělí MeSH
- endometrióza krev genetika MeSH
- endometrium cytologie MeSH
- keratin-18 genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- membránové proteiny genetika MeSH
- mladý dospělý MeSH
- mucin 1 genetika MeSH
- nemoci pleury krev genetika MeSH
- pneumotorax krev diagnóza genetika MeSH
- receptor erbB-2 genetika MeSH
- studie případů a kontrol MeSH
- tekutá biopsie MeSH
- transkriptom MeSH
- vimentin genetika MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The presence of circulating tumor cells (CTCs) in patients with solid tumors is associated with poor prognosis. However, there are limited data concerning the detection of CTCs in renal cell cancer (RCC). The aim of this study is to evaluate the presence of CTCs in peripheral blood of patients with RCC undergoing surgery (n = 186). CTCs were tested before and after surgery as well as during the follow-up period afterwards. In total 495 CTC testing in duplicates were provided. To enrich CTCs, a size-based separation protocol and tube MetaCell® was used. CTCs presence was evaluated by single cell cytomorphology based on vital fluorescence microscopy. Additionally, to standardly applied fluorescence stains, CTCs viability was controlled by mitochondrial activity. CTCs were detected independently on the sampling order in up to 86.7% of the tested blood samples in patients undergoing RCC surgery. There is higher probability of CTC detection with growing tumor size, especially in clear cell renal cell cancer (ccRCC) cases. Similarly, the tumor size corresponds with metastasis presence and lymph node positivity and CTC detection. This paper describes for the first-time successful analysis of viable CTCs and their mitochondria as a part of the functional characterization of CTCs in RCC.
- Publikační typ
- časopisecké články MeSH
New non-invasive approaches have developed for diagnosis and treatment of malignant diseases. Cells shed from the primary tumor circulating in the bloodstream with metastasis potential are called Circulating Tumor Cells (CTCs). These cells are easily acquired from the peripheral blood of patients, while several enrichment and isolation methods are available nowadays with different benefits and positive detection rates. A brief characterization of three major categories of detection is described (nucleic acid-based, physical properties-based, antibody-based). In this review we concentrate on gynecological malignancies and how CTCs could be used in the diagnosis of cancer, treatment management and its effective prognosis and early recurrence detection. Presence of CTCs in endometrial cancer patients show worse overall survival, while gene analysis could identify patients in need of systemic therapy after surgical treatment to prevent metastasis and recurrence. Based on the influence of human papillomavirus (HPV) in the etiology of cervical cancer, viral oncogene transcripts could be used as an ideal marker for cervical cancer cells detection. In ovarian cancer, CTCs could help in the differentiation from benign adnexal masses and show a high independence from other biomarkers such as CA125 and HE4. While detection of CTC after complete cytoreductive surgery could indicate invisible lesions, combination of tumor associated genes rises the specificity of CTC detection.
- MeSH
- lidé MeSH
- nádorové cirkulující buňky metabolismus MeSH
- nádory ženských pohlavních orgánů metabolismus MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
