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

Abstract

Background: Accurate histopathological evaluation of lymph nodes (LNs) is essential for reliable staging in prostate cancer. In routine practice, conventional techniques only examine parts of the LN. Molecular nodal staging methods are limited by their high costs and extensive time requirement. One-step nucleic acid amplification (OSNA) determines the metastatic status of the complete LN and allows for rapid intraoperative detection of LN metastases. OSNA has been proposed for diagnosis of LN metastases from breast cancer by quantifying the CK19 mRNA copy number. To provide basic data for OSNA development for prostate cancer, we conducted an investigation of CK19 and OSNA in prostate cancer specimens.Methods: OSNA is based on a short homogenization step and subsequent automated amplification of CK19 mRNA directly from the sample lysate, with results available in 30-40 min. A total of 20 prostate cancer specimens from consecutive patients with intermediate or high-risk prostate cancer (Gleason-Score ≥7) were investigated by both OSNA and conventional histopathology (H&E staining, CK19 immunohistochemistry). OSNA was performed on frozen samples using a ready-to-use amplification kit in an automated real-time detection system. Samples were defined as 'negative' or 'positive' according to mRNA copy number: >5000 copies/µl (++), 250-5000 copies/µl (+), and <250 copies/µl (-).Results: Histopathological analysis confirmed prostate cancer in all samples: Gleason score 7 (n=11), Gleason score 8 (n=2), and Gleason score 9 (n=6). Gleason score could not be given for one patient who previously underwent hormonal treatment. OSNA analysis detected CK19 expression in 100% of the specimens and high numbers of CK19 mRNA copies in all cases (9 samples ++; 11 samples +). Immunohistochemistry confirmed CK19 expression in 19 of 20 cases. In the immunohistochemistry CK19-negative patient, a Gleason score 9 prostate cancer was diagnosed.Conclusions: This is the first study using OSNA to detect CK19 expression in prostate cancer. Initial data indicate that this rapid method for molecular LN staging reliably identifies CK19 mRNA in prostate cancer. These results suggest that the OSNA assay may be suitable to improve (intraoperative) LN staging in prostate cancer. For further verification, OSNA analysis of LN specimens from prostate cancer patients is required.

Evaluation of Fast Molecular Detection of Lymph Node Metastases in Prostate Cancer Patients Using One-Step Nucleic Acid Amplification (OSNA)

“…Therefore, it seems that in some cases, advanced and poorly differentiated tumors might show a lack of CK19 expression, so that metastases are not detectable by OSNA. However, in our pilot study analyzing primary PCa specimens, CK19 mRNA could be detected by OSNA in all cases, regardless of the degree of tissue differentiation [ 21 ]. Further investigations are needed to clarify this discrepancy.…”

“…Immunohistochemical studies revealed that CK19 expression frequently occurs in basal as well as in luminal cells of normal, dysplastic, and benign hyperplastic prostatic tissues and in PCa [ 20 ]. Using the OSNA assay, our group recently detected a high number of CK19 mRNA copies in all PCa specimens examined [ 21 ]. This suggests that OSNA may be suitable to identify LNM and to improve LN staging in PCa patients.…”

Ultrastaging Using Ex Vivo Sentinel Lymph Node Mapping and One-Step Nucleic Acid Amplification (OSNA) in Gastric Cancer: Experiences of a European Center

“…It was initially developed for intra-operative SLN evaluation in breast cancer [ 13 ], and its suitability was demonstrated in a meta-analysis of 19 studies with over 6000 LNs [ 14 ]. It can evaluate LNs within 30 min, and its feasibility in several other cancer entities, including lung, thyroid, uterus, head and neck, and prostate cancers, has been demonstrated previously [ 15 , 16 , 17 , 18 , 19 ]. Another focus was on tumors of the gastrointestinal tract, mainly on colorectal cancers [ 20 ].…”

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

“…Regarding urological tumours, based on previous studies that demonstrated the expression of CK19 in PCa tissue, Winter et al showed that the OSNA method can detect CK19 mRNA in 100% of primary PCa tumours regardless of Gleason score and even more effectively than CK19 IHC expression, suggesting the valid application of this technique in LN evaluation [23]. In a very recent study, Engels et al [24] demonstrated that OSNA can identify nodal metastases at an equivalent or, in cases of micrometastases, better rate than enhanced histological examination in PCa patients, con rming its promising use in intraoperative decision-making in personalized LN surgery.…”