Molecular genetics in soft tissue


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Molecular genetics in soft tissue

  1. 1.  Special thanks to my resident Dr Babar Yasin for preparing the presentation  ADAPTED FROM THE ABOVE ARTICLE FROM Seminar in diagnostics Pathology 30 (2013) 375-381  Angelo P. Dei Tos, MD  Department of Pathology, Treviso General Hospital, Piazza Ospedale,1 31100 Treviso, Italy
  2. 2. The marriage of Molecular Genetics and soft tissue Pathology.
  3. 3.  More accurate definition of disease entities and validation of classification schemes.  Improved diagnostic accuracy.  Identification of molecular predictive and prognostic markers.  Discovery and validation of therapeutic molecular targets.
  4. 4.  Nucleic acids (RNA and DNA), either via hybridization on a slide(i.e., fluorescent in situ hybridization—FISH) OR  On isolated DNA or RNA via polymerase chain reaction (PCR) techniques (i.e.,reverse transcriptase PCR and quantitative PCR)It is MANDAOTORY that the results get interpreted in context with morphology.  Genetic assessement is an important ADJUNCT not a replacement to conventional morphological tools.
  5. 5. Soft tissue tumors are heterogeneous group of neoplasm which can be :  Benign  Boderline  Malignant Challenge for Pathologists Diagnostic inaccuracy affects the treatment and development of new drugs as clinical trials depend upon tumor classification.
  6. 6.  Rare group of diseases (< 2% of all cancers)  Extremely heterogenous. Over 100 subtypes are being described.  Usual features of malignancy (?) are not always applicable to these. Histologically worrisome leisons may actually be benign e.g. Nodular fascitis Cytollogically innocent neoplasm may behave aggressively e.g. Low-Grade Fibromyxoid Sarcoma.
  7. 7.  Distinguishing specific subtypes of sarcomas.  Supporting diagnosis in non-canonical clinical presentations.  Distinguishing sarcomas from benign mimickers.
  8. 8.  Molecular genetics has proved diagnostically useful in two relatively large groups of mesenchymal malignancies: round cell sarcomas and pleomorphic sarcomas. Round cell sarcomas include :  Ewing sarcoma  Desmo-plastic small round cell tumor  Alveolar rhabdomyo-sarcoma  Poorly differentiated round cell synovial sarcoma  Mesenchymal chondrosarcoma  Minority of cases of round cell liposarcoma Distinction is crucial because the therapeutic approach differs.
  9. 9.  The demonstration by FISH of EWSR1,SS18, and FOXO1 rearrangements in EWS, PDSS, and ARMS, respectively, or, alternatively, of specific chimerical transcriptsby PCR-based techniques is of great help for achieving acorrect diagnosis. EWSR1 FISH results need to be interpreted in context with morphology and IHC findings.
  10. 10.  Sub classification is very important.  myogenic differentiation in pleomorphic sarcomas is associated with a less favorable outcome.  Another important point is the distinction, among retroper-itoneal sarcomas, of dedifferentiated liposarcoma (DDLPS) from other pleomorphic sarcomas, most often pleomorphic leiomyosarcoma.
  11. 11.  The recognition of DDLPS is based on the identification of a well-differentiated lipogenic component associated with a high- grade, most often non-lipogenic, sarcoma.  Core biopsies usedfor diagnostic purposes may leave the lipogenic component unsampled.  DDLPS exhibit better out- comes when compared to other pleomorphic sarcomas, and its accurate recognition may lead to adopt a more aggressive surgical strategy. (Locally aggressive)
  12. 12.  Detection of MDM2 amplification by FISH or quantitative RT-PCR certainly represents a useful diagnostic adjunct.  The MDM2 gene (as well as CDK4 and HMGA2) maps at the 13q12–15 chromosome region and is amplified in both well-differentiated and dedifferentiated liposarcomas.  MDM2 testing is also potentially useful in distinguishing between myxoid liposar-coma (MDM2 negative) and WD/DDPLS with myxoid change.  The separation of the two conditions again allows adoption of proper treatment in consideration of the high sensitivity of myxoid liposarcoma to the marine- derived alkaloid named trabectedin.
  13. 13. Tumor Gene Mutation Gene Involved Alveolar rhabdomyosarcoma t(2;13)(q35;q14) t(1;13)(p36;q14) PAX3–FOXO1A PAX7–FOXO1A Alveolar soft part sarcoma (X;17)(p11.2;q25) ASPL–TFE3 Angiomatoid fibrous histiocytoma t(12;16)(q13;p11) t(12;22)(q13;q12) t(2;22)(q34;q12) FUS–ATF1 ATF1–EWSR1 CREB1–EWSR1 Aneurysmal bone cyst t16;17)(q22;p13) CDH11–USP6 Atypical lipomatous tumour/dedifferentiated liposarcoma Amplification MDM2, CDK4, and HMGA2 Central/periosteal osteosarcoma Point mutation IDH1/IDH2 Clear cell sarcoma t(12;22)(q13;q12) t(2;22)(q34;q12) ATF1–EWSR1 CREB1–EWSR1 Dermatofibrosarcoma protuberans t(17;22)(q22;q13) COL1A1–PDGFB Desmoid-type fibromatosis Activating mutation BCTN1
  14. 14. Desmoplastic round cell tumor t(11;22)(p13;q12) WT1–EWSR1 Endometrial stromal sarcoma t(7;17)(p15;q21) t(6;7)(p21;p15) t(6;10)(p21;p11) t(10;17)(q22;p13) JAZF1–JJAZ1 PHF1–JAZF1 PHF1–EPC1 YWHAE–FAM22 Ewing sarcoma/PNET t(11;22)(q24;q12) t(21;22)(q22;q12) t(7;22)(p22;q12) t(17;22)(q12;q12) t(16;21) (q13;q22) t(2;22)(q33;q12) EWSR1–FLI1 EWSR1–ERG ETV1–EWS EIAF–EWS FUS–ERG FEV–EWS Extraskeletal Myxoid Chondrosarcoma t(9;22)(q22;q12) t(9;15)(q22;q21) EWSR1–NR4A3 TCF12–NR4A3 TFG–NR4A3 GIST Activating mutation KIT and PDGFRA Intramuscular myxoma Activating mutation GNAS1
  15. 15. Pericytoma with t(7;12) t(7;12)(p22;q13) ACTB–GLI Pigmented villonodular synovitis t(1;2)(p13;q37) COL6A–CSF1 Soft tissue myoepithelioma t(1;22)(q23;q12) t(19;22)(q13;q12) EWSR1–PBX1 EWSR1–ZNF444 Synovial sarcoma t(X;18)(p11;q11) SS18–SSX1 SS18–SSX2 SS18–SSX4
  16. 16.  The combination of morphological criteria and genetics validates the recognition of rare diseases even when arising at non-canonical anatomic locations.  This is particularly true for referral centers wherein challenging cases tend inevitably to concentrate.  Molecular genetics has greatly  Contributed to the identification of primary Ewing sarcoma of the skin, kidney, and dura mater, as well as of viscerally located synovial sarcomas.
  17. 17.  Morphological appearance of mesenchymal lesions does not always reflect the clinical behavior.  The distinction of sarcomas from benign mimics most often relies on morphology.  In a minority of cases molecular genetics may also prove diagnostically helpful.
  18. 18. Fibroid and myxoid areas. Swirling whorled growth pattern. Low to moderate cellularity. Bland cells. Minimal nuclear pleomorphism.
  19. 19.  Deceptively bland-looking spindle cell mesenchymal malignancY with an aggressive clinical behavior.  The differential diagnosis of LGFMS includes benign lesions such as perineurioma, neurofibroma, cellular myxoma, and nodular fasciitis, as well locally aggressive neoplasms such as desmoid fibromatosis. MUC4 expression is regarded as key diagnostic feature. Identification of FUS rearrangement via FISH or identification of FUS-CREB3L2 transcript via PCR is very useful. B- Catenin
  20. 20. Several attempts have been made to determine the prognostic value of molecular genetic findings. Focused on Ewing sarcoma, alveolar rhabdomyosarcoma, and synovial sarcoma.  No meaningful molecular prognostic stratification can be foreseen for now.
  21. 21. A notable exception is represented by a molecular signature named CINSARC, which allows better separation of grade 2 sarcomas. This attempt is based on the use of a complex technique (CGH-array) and requires availability of fresh material, which hampers a large scale clinical application of CINSARC.
  22. 22.  Type of mutations involving both the KIT and PDGFRA genes are associated with distinctive outcomes.  Deletions occurring at the exon 11 of the KIT gene are associated with more aggressive disease, whereas mutations of exon 18 of the PDGFRA gene generally identify a more indolent clinical course.
  23. 23.  Distinct mutation types in GIST reflect different objective response rates (greater for KIT exon 11 mutation and much lower for so-called wild-type GIST).  presence of specific mutations in the exon 18 of the PDGFRA gene (D842V) predict primary resistance to tyrosine kinase inhibitors. Molecular assessement in GIST assumes a central role in clinical decision making.
  24. 24. The identification of the two specific fusion products of CHOP/DDIT3 gene with FUS and more rarely with EWSR1 is extremely helpful in distinguishing challenging examples of myxoid liposarcoma from other myxoid sarcomas and therefore to apply the adequate therapeutic regimen.
  25. 25.  Another examples is use of crizotinib in inflammatory myofibroblastic tumors wherein assessment of the ALK gene may represent an important diagnostic confirmatory finding as well as a key biomarker of prediction. Molecular Genetics represents the most valuable tool to identify and validate new therapeutic targets.
  26. 26. Good examples are represented by MDM2, amplified in dedifferentiated liposarcoma and potentially targetable by Nutlin-A3, the mTOR pathway in malignant PEComa and lymphangioleiomyomatosis, PDG FB in DFSP, and KDR in angiosarcoma.
  27. 27. Promiscuity
  28. 28.  Molecular pathology/genetics does not represent an alternative but a complement to surgical diagnostic pathology.  Considering the degree of molecular promiscuity of EWSR1 gene aberrations, the results of FISH analysis need to be mandatorily evaluated in context with morphology as EWSR1 aberrations are described in a variety of unrelated entities.
  29. 29. ETV6–NTRK3/t(12;15) Infantile fibrosarcoma Acute myeloid leukemia Secretory breast carcinoma ALK gene fusions Inflammatory myofibroblastic tumor Anaplastic large cell lymphoma Subsets of lung adenocarcinoma FUS–ERG/t(16;21) Ewing sarcoma Acute myeloid leukemia ASPL–TFE3/t(X;17) Alveolar soft part sarcoma Subset of pediatric renal EWS–ATF1/t(12;22) and EWS- CREB1/t(2;22) Clear cell sarcoma Angiomatoid fibrous histiocytoma
  30. 30.  Genetics has certainly played a key role in allowing a better understanding of many lesions.  The unification of myxoid and round cell liposarcoma within a single tumor entity represents one of the best examples.  Genetics has greatly helped in recognizing the close relationships between several tumors such as giant cell fibroblastoma and DFSP, LGFMS and epithelioid sclerosing fibrosarcoma, and hemosiderotic fibrolipomatous tumor and myxoinflammatory fibroblastic sarcoma.
  31. 31.  The definition of new entities has been strongly supported by genetics.  The identification of t(7;19)(q22;q13) translocation in pseudomyogenic (epithelioid sarcoma-like) hemangioendothelioma.  The future classifications will include more genetic observation.  Genetic aberrations will also contribute to the definition of the specific tumor entities.
  32. 32. The marriage between Pathology and Genetics not only proved to be fruitful but also to be stable.