Molecular characterization and heterogeneity of circulating tumor cells in breast cancer.

INTRODUCTION: This study analyzes peripheral blood samples from breast cancer (BC) patients. CTCs from peripheral blood were enriched by size-based separation and were then cultivated in vitro. The primary aim of this study was to demonstrate the antigen independent CTC separation method with high CTC recovery. Subsequently, CTCs enriched several times during the treatment were characterized molecularly. METHODS: Patients with different stages of BC (N = 167) were included into the study. All patients were candidates for surgery, surgical diagnostics, or were undergoing chemotherapy. In parallel, 20 patients were monitored regularly and in addition to CTC presence, also CTC character was examined by qPCR, with special focus on HER2 and ESR status. RESULTS: CTC positivity in the cohort was 76%. There was no significant difference between the tested groups, but the highest CTC occurrence was identified in the group undergoing surgery and similarly in the group before the start of neoadjuvant treatment. On the other hand, the lowest CTC frequencies were observed in the menopausal patient group (56%), ESR+ patient group (60%), and DCIS group (44.4%). It is worth noting that after completion of neoadjuvant therapy (NACT) CTCs were present in 77.7% of cases. On the other hand, patients under hormonal treatment were CTC positive only in 52% of cases. DISCUSSIONS: Interestingly, HER2 and ESR status of CTCs differs from the status of primary tumor. In 50% of patients HER2 status on CTCs changed not only from HER2+ to HER2-, but also from HER2- to HER2+ (33%). ESR status in CTCs changed only in one direction from ESR+ to ESR-. CONCLUSIONS: Data obtained from the present study suggest that BC is a heterogeneous disease but CTCs may be detected independently of the disease characteristics in 76% of patients at any time point during the course of the disease. This relatively high CTC occurrence in BC should be considered when planning the long-term patient monitoring.

Circulating tumor cells in different stages of colorectal cancer.

INTRODUCTION: Liquid biopsies are noninvasive tests using blood or body fluids to detect circulating tumor cells (CTCs) or the products of tumor cells, such as fragments of nucleic acids or proteins that are shed into biological fluids from primary tumor or its metastates. The analysis of published clinical studies provides coherent evidence that the presence of CTCs detected in peripheral blood is a strong prognostic factor in patients with colorectal carcinoma (CRC). The aim of the study was to implement size-based separation protocol of CTCs in CRC patients. MATERIAL AND METHODS: Patients diagnosed with different stages of CRC (n = 98) were included in the study. All patients have been diagnosed for colorectal adenocarcinoma by pathology examination, 45 patients with colon carcinoma and 53 with rectosigmoid cancer. A size-based separation method (MetaCell®) for viable CTC enrichment from peripheral blood was used to assess the presence of CTCs by cytomorphological evaluation using vital fluorescence microscopy. RESULTS: Cytomorphological analysis revealed that 81 (83%) tested samples were CTC-positive and 17 (17%) were CTC-negative. We report a successful isolation of CTCs with proliferation potential in patients with CRC. The CTCs were cultured in vitro for further downstream applications. Some of the isolated CTCs were able to grow in vitro for 6 months as a standard cell culture. CONCLUSIONS: We established a reliable, inexpensive and relatively fast protocol for CTCs enrichment in CRC patients by means of vital fluorescence staining which enables their further analysis in vitro.

Circulating tumor cells in pancreatic cancer patients: enrichment and cultivation.

AIM: To investigate the feasibility of separation and cultivation of circulating tumor cells (CTCs) in pancreatic cancer (PaC) using a filtration device. METHODS: In total, 24 PaC patients who were candidates for surgical treatment were enrolled into the study. Peripheral blood samples were collected before an indicated surgery. For each patient, approximately 8 mL of venous blood was drawn from the antecubital veins. A new size-based separation MetaCell technology was used for enrichment and cultivation of CTCs in vitro. (Separated CTCs were cultured on a membrane in FBS enriched RPMI media and observed by inverted microscope. The cultured cells were analyzed by means of histochemistry and immunohistochemistry using the specific antibodies to identify the cell origin. RESULTS: CTCs were detected in 16 patients (66.7%) of the 24 evaluable patients. The CTC positivity did not reflect the disease stage, tumor size, or lymph node involvement. The same percentage of CTC positivity was observed in the metastatic and non-metastatic patients (66.7% vs 66.7%). We report a successful isolation of CTCs in PaC patients capturing proliferating cells. The cells were captured by a capillary action driven size-based filtration approach that enabled cells cultures from the viable CTCs to be unaffected by any antibodies or lysing solutions. The captured cancer cells displayed plasticity which enabled some cells to invade the separating membrane. Further, the cancer cells in the "bottom fraction", may represent a more invasive CTC-fraction. The CTCs were cultured in vitro for further downstream applications. CONCLUSION: The presented size-based filtration method enables culture of CTCs in vitro for possible downstream applications.

Implant site development in the distal region of the mandible: bone formation and its stability over time.

INTRODUCTION: The purpose of this study was to determine the dimensional changes that occur in the alveolar ridge of patients upon distalization of the mandibular first premolars into the place of congenitally missing mandibular second premolars. The amount of bone formation that accompanied orthodontic treatment and the long-term stability of the new bone were assessed. METHODS: Fifty-five patients were included in the study, representing 71 congenitally missing mandibular second premolars. The dimensional changes were evaluated by comparing the dental stone casts and panoramic radiographs taken at treatment initiation (T1) and end (T2) and at follow-ups of 2 years (T3A) and 5 years (T3B). RESULTS: During the treatment period (T1-T2), the alveolar ridge width increased by an average of 28.5%, and the height increased by an average of 1.1 mm. During the retention periods (T2-T3A, T2-T3B), the alveolar ridge decreased by an average of 4.2%, but the height decreased only slightly (by an average of 0.07 mm). CONCLUSIONS: Orthodontic tooth movement created a significant amount of new bone that was stable in both the horizontal and vertical directions.

Clinical studies monitoring circulating and disseminated tumor cells in gastrointestinal cancers.

Circulating tumor cells (CTCs) and disseminated tumor cells (DTCs) are responsible for the development of metastatic disease, and may also hold the key to determining tailored therapies of advanced cancer disease. Our review summarizes the prognostic significance of the detection of CTCs and DTCs in various gastrointestinal cancers with an overview of their possible use as prognostic biomarkers. This could be used inthe future as a starting point for new clinical trials focusing on the predictive potential of circulating and disseminated tumor cells.