Rôle du laboratoire d’anatomie pathologique dans l’approche pré-analytique des examens de biologie moléculaire réalisés en pathologie tumorale

Hofman, Véronique; Gavric-Tanga, Virginie; Lespinet, Virginie; Mari, Mireille; Lassalle, Sandra; Butori, Catherine; Coëlle, Céline; Bordone, Olivier; Selva, Éric; Lamy, Aude; Sabourin, Jean‐Christophe; Hofman, Paul

doi:10.1016/j.annpat.2010.03.003

Cited by 33 publications

(11 citation statements)

References 38 publications

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“…High-quality tumor tissue is the key to optimal molecular and proteomic analysis. Therefore, the collection, processing, storage, and shipment of biosamples for analysis must be performed under well-documented and rigorously quality-controlled procedures as each of one of these critical steps can modify the nucleic acid and protein structures [33]. Moreover, new fixatives may improve such analyses.…”

Section: Discussionmentioning

confidence: 99%

“…Regarding the methodology used for EGFR analysis, the TMA methodology could be a good approach to reduce such variability, as we showed that all the major mechanisms of EGFR activation can be detected and analyzed on TMAs. For this purpose, a stringent quality control of samples is needed, as awareness of the importance of preanalytical factors is becoming apparent [33,57]. High-quality tumor tissue is the key to optimal molecular and proteomic analysis.…”

Section: Discussionmentioning

confidence: 99%

“…To build TMAs, selection of the cases to be included in the study was based on parameters such as collection by rigorous adherence to banking protocols [32], the absence of neoadjuvant treatment, and the availability of sufficient tissue to examine molecular abnormalities and protein expression by IHC. A well-qualified sample is one where there is over 60% cancer cells and no necrosis [33]. The study was done with the approval of the ethics committee (CHU of Nice).…”

Section: Patientsmentioning

confidence: 99%

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Usefulness of tissue microarrays for assessment of protein expression, gene copy number and mutational status of EGFR in lung adenocarcinoma

et al. 2010

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Specific inhibitors targeting the epidermal growth factor receptor (EGFR) can increase survival rates in certain lung adenocarcinoma patients with mutations in the EGFR gene. Although such EGFR-targeted therapies have been approved for use, there is no general consensus among surgical pathologists on how the EGFR status should be tested in lung adenocarcinoma tissues and whether the results of immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and mutational analysis by molecular methods correlate. We evaluated the EGFR status in 61 lung adenocarcinomas by IHC (using total and mutant-specific antibodies against EGFR), by FISH analysis on tissue microarrays (TMAs), and by direct sequencing. The results of each method were compared using χ² and κappa statistics. The sensitivity and negative predictive value estimating the presence of abnormal EGFR for each test was calculated. The results show that, with respect to expression patterns and clinicopathological parameters, the total and mutant-specific EGFR detected by immunohistochemistry and FISH analysis on TMAs are valid and are equivalent to conventional methods performed on whole-tissue sections. Abnormal EGFR was detected in 52.4% of patients by IHC, FISH, and sequencing. The best sensitivity (100%) and negative predictive value (100%) was determined by evaluating the EGFR status with all methods. Testing for molecular changes in EGFR using a single test is likely to underestimate the presence of EGFR abnormalities. Taken together, these results demonstrate the high potential of TMAs to test for the major mechanisms of EGFR activation in patients with lung adenocarcinoma.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Patientsmentioning

confidence: 99%

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Usefulness of tissue microarrays for assessment of protein expression, gene copy number and mutational status of EGFR in lung adenocarcinoma

et al. 2010

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“…In most molecular diagnostic laboratories, this is done by cutting one or two hematoxylin and eosin (H&E) stained slides and 5-10 unstained sections, 6-10 microns in thickness, from FFPE tissue blocks. The H&E slide is reviewed by a pathologist; the area of interest is circled; and the slide is used to guide macro- or microdissection of the tissue from the unstained sections for tumor enrichment [7-10]. When resection specimens with large areas of tumor are available, this technique is facile.…”

Section: Introductionmentioning

confidence: 99%

A simple and cost-effective method of DNA extraction from small formalin-fixed paraffin-embedded tissue for molecular oncologic testing

et al. 2014

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BackgroundExtraction of DNA from formalin-fixed, paraffin-embedded (FFPE) tissue is a critical step in molecular oncologic testing. As molecular oncology testing becomes more important for prognostic and therapeutic decision making and tissue specimens become smaller due to earlier detection of suspicious lesions and the use of fine needle aspiration methods for tissue collection, it becomes more challenging for the typical molecular pathology laboratory to obtain reliable test results. We developed a DNA extraction method to obtain sufficient quantity and high quality genomic DNA from limited FFPE tissue for molecular oncology testing using a combination of H&E stained slides, a matrix capture method and the Qiagen DNA column.MethodsThree DNA extraction methods were compared: our standard procedure of manually scraping tissue from unstained slides followed by DNA extraction using the QIAamp FFPE column (Qiagen, Valencia, CA), a glue capture method (Pinpoint Solution, Zymo Research Corp, Inc) on H&E stained slides followed by DNA extraction using either the QIAamp column or the column included with the Pinpoint kit (Zymo Research). The DNA extraction protocol was optimized. Statistical analysis was performed using the paired two-sample student’s t-test.ResultsThe combination of the matrix capture method with the QIAamp column gave an equivalent amount of DNA as our standard extraction method using the unstained slides and a 4.6-fold higher DNA yield than using the Zymo column included in the Pinpoint Slide Solution kit. Several molecular tests were performed and DNA purified using the new method gave the same results as for the previous methods.ConclusionsUsing H&E stained slides allows visual confirmation of tumor cells during microdissection. The Pinpoint solution made removal of specific tissue from the slides easier and reduced the risk of contamination and tissue loss. This DNA extraction method is simple, cost-effective, and blends with our current workflow requiring no additional equipment.

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“…Son investissement peut également s'étendre jusqu'à la phase d'extraction des acides nucléiques, à leur qualification et à l'analyse moléculaire. L'ensemble de ces étapes est précisément illustré dans l'article de Véronique Hofman [7]. Il y est parfaitement souligné que « le contrôle en amont des étapes présentes entre le prélèvement d'un échantillon tumoral et l'analyse moléculaire conditionne la fiabilité des résultats de biologie moléculaire et donc la stratégie thérapeutique ».…”

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Pathologie moléculaire : au commencement était la phase pré-analytique !

Bibeau

Boissière‐Michot

2010

Annales de Pathologie

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Rôle du laboratoire d’anatomie pathologique dans l’approche pré-analytique des examens de biologie moléculaire réalisés en pathologie tumorale

Cited by 33 publications

References 38 publications

Usefulness of tissue microarrays for assessment of protein expression, gene copy number and mutational status of EGFR in lung adenocarcinoma

Usefulness of tissue microarrays for assessment of protein expression, gene copy number and mutational status of EGFR in lung adenocarcinoma

A simple and cost-effective method of DNA extraction from small formalin-fixed paraffin-embedded tissue for molecular oncologic testing

Pathologie moléculaire : au commencement était la phase pré-analytique !

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