Complex molecular-pathological diagnosis patient NSCLC – case report

Konference: 2012 8. Sympozium a workshop molekulární patologie a histo-cyto-chemie

Kategorie: Kasuistiky; Zhoubné nádory plic a průdušek

Téma: Posters

Číslo abstraktu: 017p

Autoři: RNDr. Ing. Bc. Libor Staněk, PCTM; Prof. MUDr. Pavel Dundr, Ph.D.; MUDr. Martin Syrůček; MUDr. Soňa Lísová; Daniel Tvrdík; doc. MUDr. Marie Ludvíková, Ph.D.; Prof. MUDr. Ctibor Povýšil, DrSc.

Summary: Lung cancer is one of the most common cancer around the world, in men with the highest mortality of all malignant diseases (1), it is often closely linked with troublesome biopsy diagnosis, but high quality and accurate diagnosis is for the treatment and prediction necessary (2). Predictive testing involves not only molecular pathological examination (EGFR mutation, KRAS, BRAF), but also cytogenetic investigation (break ALK/EML-4, HER2 amplification). The biological treatment is usually indicated based on the results of these tests, and may stabilize progression the disease. Our case report presents a patient with lung cancer with an unusual morphology and describes the complete bioptic and predictive diagnostic process using methods available today, with the emphasis on logistic of the process.

The clinical findings: Patient be born 1945, T26 location of disease, the diagnosis C341.

Materials and methods: Investigated material was biopsy specimen from the left lower bronchus taken during bronchoscopy. The specimen was processed by standard procedures, fixed in formalin and embedded in paraffin, was used a standard hematoxilin-eosin staining and immunohistochemical examination with antibodies against TTF1, CK7 and cdx2, followed pathological and molecular cytogenetic analysis (3):

  • Isolation of DNA after previous deparafination with xylene-ethanol series kit (DNA Mini Kit, Qiagen Inc., USA) and measuring concentration (qRT-PCR).
  • EGFR mutations were detected by Real-Time PCR using TheraScreen kit (Roche) and tested for mutations G719X in exon 18, 19 deletion in exon 19, S768I in exon 20, 3 insertions in exon 20 and L861Q, L858R in exon 21 .
  • KRAS mutations were detected by Real-Time PCR using TheraScreen kit (Roche) and were tested for mutations Gly12Ala, Gly12Asp, Gly12Arg, Gly12Cys, Gly12Ser, and Gly12Val Gly13Asp at codon 12 and 13 K-ras gene.
  • BRAF gene mutation were tested by hybridization using Strip, kit KRAS BRAF StripAssay (ViennaLab), searching for mutations in exon 15, caused by transverse T1799A, leading to amino acid substitution of V600E.
  • HER2/neu gene amplification by FISH using probes directly labeled LSI Her2/neu Spectrum Orange and centromeric probes for chromosome 17 directly labeled with Spectrum Green was established Her-2/ neu gene copy number of chromosome 17 (Abbott MolecularTM)
  • Break ALK gene (2p23) by FISH with the LSI ALK probe DC BA Rearrangement Probe (Abbott MolecularTM)


Results: In the microscopic examination of bronchial mucosa was found solidly struktured adenocarcinoma witch shows intracellular mucin production and passing to areas with signet-ring cell features. The tumor grew infiltrative way with evident tumorous lymphangiopathy. In immunohistochemical examination the tumor cells expressed cytokeratin 7. Detection TTF1 and cdx2 was negative. Molecular pathological examination was negative for EGFR, activating mutations were not detected. When KRAS gene was tested, there was found inhibiting mutation of Gly12Ala (GGT ≥ GCT) at codon 12. During mutational analysis of the BRAF gene there was no evident mutation V600E. During cytogenetic examination by FISH method was not detected Her2/neu gene amplification. Examination of ALK gene (2p23) however proved a break in the area of the fusing signal ALK/ EML-4.

Conclusion: From the describing case is obvious the need for comprehensive and complete diagnosis, which is crucial to achieve the adequate response to the currently available biological treatment. In the describing case study is demonstrated that the results of analysis of the signaling pathway EGFR ras / raf / MAPK was interpreted to detriment of the patient. Due to the concurrent use of cytogenetic methods which were confirmed a break and the emergence of ALK gene fusion proteins ALK/EML-4 there can be used treatment by Crizotinib inhibitors (Pfizer) (4). The presentation of the ALK gene break in lung cancer is rare, but may be suspicious already from morphology of the tumor. In the vast majority of these cases uncommon type of adenocarcinoma presents. For example, there is a component of tumor cells with signetring cell features or mucinous cribriform patern organized carcinoma, which are described in almost 80% of cancinomas with ALK gene conversion and occur only in less than 1% of cancers without this alteration of ALK gene (5).

  1. Zdeněk A, Krejčí M, Vorlíček J, et al. Speciální onkologie (1 th ed), Galen, Praha, 2010: 29.
  2. Thomas R, Wolf J. Personalized therapy of lung cancer. Onkologie. 2012, 35 Suppl 1:14-9.
  3. Staněk L, Tvrdík D, Vítková I, Povýšil C. Možnosti molekulární diagnostiky mutací EGFR receptoru pro biologickou léčbu pomocí inhibitorů tyrozinkinázy u pacientů s nemalobuněčným karcinomem plic. Stud. Pneumol. Phthiseol. 2010, 70:110-112.
  4. Katayama R, Shaw AT, Khan TM, et al. Mechanisms of Acquired Crizotinib Resistance in ALK-Rearranged Lung Cancers. Sci Transl Med. 2012, 4(120):120ra17.
  5. Yoshida A, Tsuta K, Harumi N, et al. Comprehensive histologic analysis of ALK-rearranged lung carcinomas. Am J Surg Pathol. 2011, 35: 1226–1234.

Datum přednesení příspěvku: 27. 4. 2012