scholarly journals Rapid Detection of Free Cancer Cells in Intraoperative Peritoneal Lavage Using One-Step Nucleic Acid Amplification (OSNA) in Gastric Cancer Patients

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2168
Author(s):  
Katarzyna Gęca ◽  
Karol Rawicz-Pruszyński ◽  
Jerzy Mielko ◽  
Radosław Mlak ◽  
Katarzyna Sędłak ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Nucleic Acid ◽  
Cancer Cells ◽  
Clinical Decision Making ◽  
Peritoneal Lavage ◽  
Nucleic Acid Amplification ◽  
Cytokeratin 19 ◽  
Ck19 Mrna ◽  
One Step ◽  
Osna Assay

Cytokeratin-19 (CK19) has been proven to be commonly expressed by cancer cells in a variety of solid tumors and may serve as a suitable marker of metastases in gastric cancer (GC). Since objective assessment of peritoneal lavage or fluid for free cancer cells (FCC) is essential for clinical decision making in patients with GC, it is important to develop a quantitative and reproducible method for such evaluation. We assessed the possible application of One-Step Nucleic Acid amplification (OSNA) assay as a rapid method for FCC detection in intraoperative peritoneal lavage or fluid of GC patients. Seventy-eight intraoperative peritoneal lavage or fluid samples were eligible for the analysis by conventional cytology and OSNA examination. The concentration of CK19 mRNA in intraoperative peritoneal lavage and fluid was compared with the conventional cytological assessment. CK19 mRNA concentration was detected by OSNA assay. For peritoneal lavage samples, sensitivity and specificity were 83.3% and 87.8%, respectively. In peritoneal fluid, significantly higher CK19 values were observed in patients with serosal infiltration (medians: 100 copies/µL vs. 415.7 copies/µL; p = 0.0335) and lymph node metastases (medians: 2.48 copies/µL vs. 334.8 copies/µL). OSNA assay turns out to be an objective, fast, and reproducible quantitative method of FCC assessment.

scholarly journals Molecular Cytology by One-Step Nucleic Acid Amplification (OSNA) Assay of Peritoneal Washings during D2 Gastrectomy in Advanced Gastric Cancer Patients: Preliminary Results

2021 ◽  
Vol 10 (22) ◽  
pp. 5230
Author(s):  
Katarzyna Gęca ◽  
Karol Rawicz-Pruszyński ◽  
Radosław Mlak ◽  
Katarzyna Sędłak ◽  
Magdalena Skórzewska ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Nucleic Acid ◽  
Mrna Level ◽  
Nucleic Acid Amplification ◽  
Poor Prognostic Factor ◽  
D2 Gastrectomy ◽  
Ck19 Mrna ◽  
Peritoneal Washings ◽  
One Step ◽  
Osna Assay

The presence of peritoneal free cancer cells (FCC) in gastric cancer (GC) patients is a poor prognostic factor. D2 gastrectomy may induce exfoliated FCC spread from the primary tumour or involved lymph nodes (LN). Conventional cytology for FCC detection has several limitations, whereas prophylactic use of extensive intraoperative peritoneal lavage (IPL) does not improve survival. A prospective single-arm observational study was conducted to verify whether D2 gastrectomy causes an intraoperative increase of FCC in peritoneal fluid. Twenty-seven GC patients underwent D2 gastrectomy, followed by objective quantitative measurements of CK19 mRNA level reflecting FCC with One-Step Nucleic Acid Amplification (OSNA) assay. The IPL with 3000 mL of saline was performed twice: (1) after gastrectomy with D2 lymphadenectomy and (2) after alimentary tract reconstruction. The IPL samples were analysed by initial cytology and four (1–4) consecutive OSNA assays. Initial OSNA measurement (1) revealed positive results (≥24.6 cCP/μL) in 7 (29.6%) patients. Subsequent OSNA measurements showed a significant decrease in the FCC level after D2 gastrectomy (1 vs. 2; p = 0.0012). The first IPL induced a non-significant increase in the FCCs (2 vs. 3, p = 0.3300), but the second IPL reversed it to normal levels (3 vs. 4, p = 0.0.0574). The OSNA assay indicates a temporal intraoperative increase in the peritoneal FCC in advanced GC patients undergoing D2 gastrectomy. Two consecutive IPLs are necessary to reverse the increase of CK19 mRNA level in peritoneal washings.

One-step nucleic acid amplification for detecting lymph node metastasis of head and neck cancer

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 6028-6028
Author(s):  
H. Goda ◽  
K. Nakashiro ◽  
T. Yoshimura ◽  
T. Sumida ◽  
H. Wakisaka ◽  
...  
Keyword(s):  
Lymph Node ◽  
Nucleic Acid ◽  
Lymph Node Metastasis ◽  
Nucleic Acid Amplification ◽  
Cytokeratin 19 ◽  
Node Metastasis ◽  
Ck19 Mrna ◽  
One Step ◽  
Genetic Detection ◽  
Osna Assay

6028 Background: Lymph node stage is an important prognostic factor in squamous cell carcinoma of the head and neck (SCCHN). We previously reported the clinical usefulness of sentinel lymph node (SLN) biopsy diagnosed by concurrently performing histological examination using semiserial sections and genetic analysis by quantitative RT-PCR. However, these methods took about 3 hours. In this study, we have attempted to develop a more efficient method for intraoperative genetic detection of lymph node metastasis in SCCHN. Methods: A total of 291 lymph nodes (59 patients) resected on SLN biopsy for cN0 SCCHN or neck dissection for cN1/2 SCCHN were diagnosed by one-step nucleic acid amplification (OSNA) method using GD-100. The primary site was tongue, gingiva, oral floor, buccal mucosa, and pharynx in 44% (26), 37% (22), 10% (6), 5% (3), and 3% (2), respectively. OSNA consists of a short homogenization step followed by amplification of cytokeratin 19 (CK19) mRNA directly from the lysate. It is characterized by the use of 4 different primers specifically designed to recognize 6 distinct regions, so the CK19 primers do not amplify the known CK19 pseudogenes. The reaction process proceeds at a constant temperature (65°C) during strand displacement reaction. Amplification and detection of CK19 mRNA can be completed in a single step. Each lymph node was divided into two halves to diagnose metastasis. An alternative half was used for the OSNA assay with cytokeratin 19 (CK19) mRNA, and the remaining block was subjected to semiserial sectioning, sliced at 200-μm intervals and then examined by H&E and cytokeratin AE1/AE3 immunohistochemical staining. Results: Fifty-four of 291 lymph nodes were pathologically metastasis-positive. The optimal cut-off for the copy number of CK19 mRNA in assessing lymph node metastasis was 300 copies/μl, which had the highest diagnostic accuracy. The sensitivity and specificity of OSNA assay with CK19 mRNA was 92.6% (50/54) and 97% (230/237), respectively. An overall concordance rate between the OSNA assay and histopathology was 96.2%. The OSNA assay could be completed within 30 minutes. Conclusions: The OSNA assay showing high sensitivity and specificity can be used as a novel genetic detection tool of lymph node metastasis in SCCHN patients. No significant financial relationships to disclose.

Utility of the OSNA assay in sentinel node mapping for early gastric cancer.

2014 ◽  
Vol 32 (3_suppl) ◽  
pp. 13-13
Author(s):  
Hiroya Takeuchi ◽  
Tomohiko Nishi ◽  
Rieko Nakamura ◽  
Tsunehiro Takahashi ◽  
Norihito Wada ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Nucleic Acid ◽  
Early Gastric Cancer ◽  
Sentinel Node ◽  
Histological Examination ◽  
Nucleic Acid Amplification ◽  
Histopathological Diagnosis ◽  
Frozen Sections ◽  
One Step ◽  
Osna Assay

13 Background: Recent studies on sentinel node (SN) mapping for early gastric cancer have shown acceptable results in terms of detection rate and accuracy to determine lymph node (LN) status. Accurate detection of nodal metastasis including micrometastasis during surgery is indispensable for clinical application of minimized gastrectomy based on SN mapping and biopsy. However, histopathological diagnosis of micrometastasis using frozen sections has to be considered as limitations for intraoperative pathology. We hypothesized that one-step nucleic acid amplification (OSNA) assay may allow the PCR-based intraoperative diagnosis to be applicable to SN mapping for early gastric cancer. Methods: A prospective study of SN mapping using the OSNA (One-step nucleic acid amplification) assay has been conducted for patients with cT1N0M0 gastric cancer at our institution. All SNs and randomly selected non-SNs were examined by routine histopathological diagnosis and the OSNA assay. Results: Histological examination was performed in in 1,732 LNs (286 SNs and 1,446 non-SNs) from 43 patients with cT1N0M0 gastric cancer. We also evaluated 439 LNs (286 SNs and 153 non-SNs) with histological examination and the OSNA assay from the 43 patients. Histological LN metastasis was revealed in 6 (14.0%) of 43 patients. All (100%) of the 6 patients with LN metastasis showed positive SNs by histological examination. The diagnostic accuracy to predict the LN status based on SN concept by histological examination was 100%. The concordance rate between the OSNA assay and the histological examination of paraffin specimens was 0.970 (95 % CI, 0.950–0.984). Discordant results were observed in 13 LNs (3.0 %). Sensitivity and specificity of the OSNA assay compared to the histological examination were 0.636 (95 % CI, 0.407–0.828) and 0.988 (95 % CI, 0.972–0.996), respectively. The OSNA assay to evaluate the SNs status was comparable with intraoperative pathological diagnosis of frozen sections of SNs in terms of concordance rate (0.981), sensitivity (0.800) and specificity (0.995). Conclusions: Our results suggest that the OSNA assay may be a useful tool for the intraoperative detection of metastasis in SNs for patients with early gastric cancer.

scholarly journals Lymph Node Positivity in One-Step Nucleic Acid Amplification is a Prognostic Factor for Postoperative Cancer Recurrence in Patients with Stage II Colorectal Cancer: A Prospective, Multicenter Study

2019 ◽  
Vol 27 (4) ◽  
pp. 1077-1083 ◽  
Author(s):  
Michio Itabashi ◽  
Hirofumi Yamamoto ◽  
Naohiro Tomita ◽  
Masafumi Inomata ◽  
Kohei Murata ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Lymph Node ◽  
Nucleic Acid ◽  
Disease Free Survival ◽  
Nucleic Acid Amplification ◽  
Stage Ii ◽  
Pathological Examination ◽  
Cytokeratin 19 ◽  
One Step ◽  
Osna Assay

Abstract Background For colorectal cancer (CRC) patients, the standard histological lymph node (LN) evaluation has low sensitivity. Our previously developed one-step nucleic acid amplification (OSNA™) assay measures cytokeratin 19 gene expression in whole LNs. We recently showed that 17.6% of pN0 stage II CRC patients were OSNA positive, suggesting a correlation between OSNA results and disease recurrence. This multicenter, prospective study investigateed the prognostic value of the OSNA assay for pStage II CRC patients. Methods We examined 204 CRC patients who were preoperatively diagnosed as cN0 and cN1 and surgically treated at 11 medical institutions across Japan. Nine patients were excluded, and 195 patients (Stage I: n = 50, Stage II: n = 70, Stage III: n = 75) were examined. All LNs, harvested from patients, were examined histopathologically using one-slice hematoxylin–eosin staining. Furthermore, half of the LNs was examined by the OSNA assay. Patients were classified according to the UICC staging criteria and OSNA results, and the 3-year, disease-free survival (DFS) of each cohort was analyzed. Results Average 21.2 LNs/patient were subject to pathological examination. Approximately half of all harvested LNs (average, 9.4 LNs/patient) were suitable for the OSNA assay. Significantly lower 3-year DFS rates were observed in pStage (pathological Stage) II OSNA-positive patients than in OSNA-negative patients (p = 0.005). Among all assessed clinical and pathological parameters, only the OSNA result significantly affected 3-year DFS rates in pStage II CRC patients (p = 0.027). Conclusions This study shows that OSNA positivity is a risk factor for recurrence of the patients with pStage II CRC.

scholarly journals One-step nucleic acid amplification (OSNA) of Sentinel Lymph Node in Early Stage Endometrial Cancer: Spanish Multicenter Study (ENDO-OSNA).

2021 ◽  
Author(s):  
M. D. Diestro ◽  
A. Berjón ◽  
I. Zapardiel ◽  
L. Yébenes ◽  
I. Ruiz ◽  
...  
Keyword(s):  
Endometrial Cancer ◽  
Lymph Node ◽  
Nucleic Acid ◽  
Sentinel Lymph Node ◽  
Early Stage ◽  
Nucleic Acid Amplification ◽  
One Step ◽  
Hematoxylin Eosin ◽  
Early Stage Endometrial Cancer ◽  
Osna Assay

The objective of this study was to evaluate the efficacy of one-step nucleic acid amplification (OSNA) for the detection of sentinel lymph node (SLN) metastasis compared to standard pathological ultrastaging in patients with early-stage endometrial cancer (EC). A total of 526 SLNs from 191 patients with EC were included in the study. 379 SLNs (147 patients) were evaluated by both methods, OSNA and standard pathological ultrastaging. The central 1-mm portion of each lymph node was subjected to semi-serial sectioning at 200-μm intervals and examined by hematoxylin-eosin and immunohistochemistry with CK19; the remaining tissue was analysed by OSNA for CK19 mRNA. The OSNA assay detected metastases in 19.7% of patients (14.9% micrometastasis and 4.8% macrometastasis), whereas pathological ultrastaging detected metastasis in 8.8% of patients (3.4% micrometastasis and 5.4% macrometastasis). Using the established cut-off value for detecting SLN metastasis by OSNA in EC (250 copies/μl), the sensitivity of the OSNA assay was 92%; specificity was 82%; diagnostic accuracy was 83%, and the negative predictive value was 99%. Discordant results between both methods were recorded in 20 patients (13.6%). OSNA resulted in an upstaging in 12 patients (8.2%). OSNA could aid in the identification of patients requiring adjuvant treatment at the time of diagnosis.

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