Detection of CK19 mRNA Using One-step Nucleic Acid Amplification (OSNA) in Prostate Cancer: Preliminary Results

Background: In clinical routine, only fractions of lymph nodes (LNs) are examined histopathologically, often resulting in missed (micro-)metastases and incorrect staging of prostate cancer (PCa). One-step nucleic acid amplification (OSNA) analyzes the entire LN by detecting cytokeratin 19 (CK19) mRNA as a surrogate for LN metastases requiring less effort than conventional biomolecular techniques. We aimed to evaluate performance of OSNA in detecting sentinel LN (SLN) metastases in PCa. Methods: SLNs (n = 534) of 64 intermediate- or high-risk PCa patients undergoing radical prostatectomy with extended and sentinel-guided lymphadenectomy were cut into slices and alternatingly assigned to OSNA and histopathology (hematoxylin-eosin staining, CK19, and CK AE1/AE3 immunohistochemistry). Sensitivity and specificity of OSNA and concordance and measure of agreement (Cohen’s kappa (κ)) between OSNA and histopathology were assessed. Results: Histopathology revealed metastases in 76 SLNs. Sensitivity and specificity of OSNA were 84.2% and 96.1%, respectively. Discordant results were recorded for 30 of 534 SLNs, revealing high concordance (94.4%). Twenty-four discordant cases were classified as micrometastases, indicating a possible allocation bias. In 18 cases, positive results were conferred only by OSNA resulting in seven LN-positive patients who were missed by histopathology. Overall, the level of agreement was high (κ = 0.78). Conclusions: OSNA provided a diagnosis that was as least as accurate as detailed histological examination and might improve LN staging in PCa.

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MP60-20 DETECTION OF LYMPH NODE METASTASES USING POOLING METHOD BY ONE-STEP NUCLEIC ACID AMPLIFICATION (OSNA) ASSAY IN PROSTATE CANCER PATIENTS: PRELIMINARY RESULTS FROM A PROSPECTIVE-MULTICENTRE STUDY

The Journal of Urology ◽

10.1097/ju.0000000000002095.20 ◽

2021 ◽

Vol 206 (Supplement 3) ◽

Author(s):

Mercè Cuadras ◽

Lucas Regis ◽

Jacques Planas ◽

Ana Celma ◽

Ma Eugenia Semidey ◽

...

Keyword(s):

Prostate Cancer ◽

Lymph Node ◽

Nucleic Acid ◽

Multicentre Study ◽

Cancer Patients ◽

Nucleic Acid Amplification ◽

Prospective Multicentre Study ◽

One Step ◽

Node Metastases ◽

Osna Assay

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

Cells ◽

10.3390/cells9102168 ◽

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.

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

Journal of Clinical Medicine ◽

10.3390/jcm10225230 ◽

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.

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PD17-03 FIRST EVALUATION OF ONE-STEP NUCLEIC ACID AMPLIFICATION (OSNA) FOR RAPID MOLECULAR DETECTION OF LYMPH NODE METASTASES IN PROSTATE CANCER PATIENTS

The Journal of Urology ◽

10.1097/01.ju.0000555563.39848.36 ◽

2019 ◽

Vol 201 (Supplement 4) ◽

Author(s):

Alexander Winter* ◽

Svenja Engels ◽

Friederike Hasselmann ◽

Lutz Brautmeier ◽

Rolf-Peter Henke ◽

...

Keyword(s):

Prostate Cancer ◽

Lymph Node ◽

Nucleic Acid ◽

Cancer Patients ◽

Lymph Node Metastases ◽

Molecular Detection ◽

Nucleic Acid Amplification ◽

One Step ◽

Node Metastases

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A new application to one‑step nucleic acid amplification‑discarded sample in breast cancer: Preliminary results

Molecular and Clinical Oncology ◽

10.3892/mco.2021.2380 ◽

2021 ◽

Vol 15 (4) ◽

Author(s):

Carolina Ribeiro ◽

Inês Gante ◽

Margarida Dias ◽

Ana Gomes ◽

Henriqueta Silva

Keyword(s):

Breast Cancer ◽

Nucleic Acid ◽

Nucleic Acid Amplification ◽

Preliminary Results ◽

One Step

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

Journal of Clinical Oncology ◽

10.1200/jco.2009.27.15_suppl.6028 ◽

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.

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Novel One-Step Nucleic Acid Amplification (OSNA) Application in Prostate Cancer. What Can We Learn from its Usage in Other Cancer Entities? A Systematic Review

10.21203/rs.3.rs-840288/v1 ◽

2021 ◽

Author(s):

Mercè Cuadras ◽

Jacques Planas ◽

Ana Celma ◽

Lucas Regis ◽

Inés M. de Torres ◽

...

Keyword(s):

Prostate Cancer ◽

Systematic Review ◽

Nucleic Acid ◽

Rate Sensitivity ◽

Nucleic Acid Amplification ◽

Cochrane Library ◽

Cytokeratin 19 ◽

General Interest ◽

Predictive Values ◽

One Step

Abstract Background: Lymph node (LN) status is a key prognostic factor in the decision-making process of prostate cancer (PCa) management. Sectioning and haematoxylin and eosin (H&E) staining technique remain the gold standard for the evaluation of LN metastases despite some limitations, especially low sensitivity in detecting an accurate tumour burden within the LN, as well as a subjective and time-consuming result. One-step nucleic acid amplification (OSNA) quantifies mRNA copies of cytokeratin 19 (CK19) in a fast, objective, automated, and reproducible way, raising a general interest to explore its utility for lymphatic metastasis identification in different malignancies.Methods: To present the latest evidence related to the detection of LN metastases in several tumours by using OSNA compared with the conventional H&E method, a systematic review of articles published since March 2021 was conducted using PubMed, Cochrane Library, and Web of Science databases. References from primary papers and review articles were checked to obtain further potential studies. Our procedure for evaluating records identified during the literature search followed the Preferred Reporting Items for Systematic Reviews and Meta-analyses criteria.Results: Twenty five studies were included. LN from six different groups of tumours: breast, gastrointestinal, gynecological, lung, head and neck and prostate cancers has been assessed. OSNA was compared with post-operative formalin-fixed paraffin-embedded tissue sections with H&E staining as the reference standard. Contingency tables were created, and concordance rate, sensitivity, specificity and predictive values were reported. Seventeen studies analysed the discordant cases using different techniques.Conclusion: OSNA method has a high diagnostic accuracy for the detection of LN metastases in several CK19 expressing tumours. Available evidence encourages its usage in PCa patients to improve LN staging and prognosis.

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