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Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
Soft tissue toumours in children
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Soft tissue toumours in children

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  • 1. Soft Tissue Tumours in Children Gordan M. Vujanić School Of Medicine Cardiff University Cardiff United Kingdom
  • 2. Pathology of TumoursThe Era of Morphology - 1850-1980sThe Era of Immunohistochemistry – 1980 – 2000The Era of Molecular Diagnosis - 2000 -
  • 3. Tumours in Children Challenging to diagnose - relatively rare - show challenging morphological features Different form tumours in adults (carcinomas vs. embryonal) Rare even in tertiary paediatric referral centres Multicentre trials resulted in huge improvement (central pathology review, biology studies, standardized multidisciplinary therapeutic treatments)
  • 4. Soft Tissue Tumours in Children - WHO Classification -1. Fibrous tissue tumours2. Fibrohistiocytic tumours3. Lipomatous tumours4. Smooth muscle tumours5. Skeletal muscle tumours6. Endothelial tumours of blood vessels7. Perivascular tumours8. Synovial tumours9. Mesothelial tumours10. Neural tumours11. Paraganglion tumours12. Cartilage and bone tumours13. Pluripotential mesenchymal tumours14. Miscellaneous tumours15. Unclassified tumours
  • 5. Paediatric Soft Tissue Tumours Historically, classified according to their degree and type of cellular differentiation  Skeletal myogenesis = rhabdomyosarcoma  Spindle cell morphology = fibrosarcoma/myofibrosarcoma  Small round cell tumours = undifferentiated sarcoma Morphological diagnosis of limited value - Tumours with similar histology – markedly different clinical behaviour and outcome
  • 6. Rationale for using molecular techniques in paediatric tumour pathology Diagnostic difficulties (especially with needle biopsies) Immunophenotype often non-diagnostic  CD56 - PNET, RMS, NB, WT  CD99 - PNET, DSRCT, WT  Bcl-2 - PNET, DSRCT, WT, NB, SS  WT1 - PNET, RMS, DSRCT, NB, WT Important role in diagnostic and prognostic stratification of tumours Prognosis / treatment / trial protocols
  • 7. Which ‘molecular’ methods are diagnostically applicable to histopathology samples? Numerous techniques can be utilised in the molecular genetic analysis of tumours  Classical cytogenetics  Fluorescence in situ hybridisation (FISH)  PCR and RT-PCR
  • 8. Paediatric Soft Tissue Tumours When tumour karyotyping introduced:  STT with recurrent chromosomal translocations - more uniform cell population  STT with complex karyotypes lacking recurrent translocations - more likely to be pleomorphic- RT-PCR and FISH – sorted out many problems ass with classic cytogenetics
  • 9. Why molecular diagnostic techniques? Many paediatric tumours have specific translocations with associated functional fusion gene productsTumour Translocation Fusion gene product(s) PrevalenceA-RMS (2;13)(q35;q14) PAX3-FOXO1 ~70% (2;13)(p36;q14) PAX7-FOXO1 ~10%ES/PNET (11;22)(q24;q12) EWS-FLI1 ~85% (21;22)(q22;q12) EWS-ERG ~10% t(7;22)(q33;q12) EWS-FEV ~1%DSRCT (11;22)(p13;q12) EWS-WT1 ~93%Synovial sarcoma (X;18)(p11.2;q12.2) SYS18SSX1; SYS18SSX2; 63%; 37% SS18-SSX4 rareIFS (12;15)(p13;q25) ETV6-NTRK3 ~100%DFSP (17;22)(q22;q13) COL1A1-PDGFB ~92%AlvSPS (X;17)(p11;q25) ASPL-TFE3 ~100%
  • 10. FISH vs. PCR in routine practice PCR  Precise identification of transcripts BUT  Limited to known targets  Reduced sensitivity in FFPE specimens due to lower RNA quality  Usually the housekeeping gene expressed at higher levels than target so possible false FISH  Less specific since FISH detects for example any EWS translocation  Translocation partner NOT identified but also not necessarily required to interpret test !
  • 11. Role of Molecular Diagnostics in Paediatric Tumours  As a diagnostic tool (Ewing’s sarcoma/PNET, or Synovial Sarcoma)  Tumour Classification (Alveolar RMS vs. Embryonal RMS)  As a prognostic marker (Neuroblastoma, RMS, etc.)
  • 12. Soft Tissue Tumours in ChildrenTumour category Children (%) Adults (%) Vascular 29 9 Neurogenic 15 9 Myogenic 14 5 Fibroblastic-myofibroblastic 12 7 Fibrohistiocytic 12 17 Lipocytic 6 16 Other 12 38
  • 13. Rhabdomyosarcoma (RMS) the most common soft tissue sarcoma in childhood peak incidence in the first decade familial form is associated with Li-Faumeni syndrome with increased risk for development of various neoplasms most common in soft tissues, but can occur anywhere (cutaneous RMS in association with epidermal naevus & von Recklinghausen’s disease)
  • 14. International Classification of RMSI Superior prognosis Botryoid RMS Spindle cell RMSII Intermediate prognosis Embryonal RMS (classical)III Poor prognosis Alveolar RMSIV Unknown prognosis RMS with rhabdoid features
  • 15. Diagnosis of RMS Morphology Demonstration of myogenic differentiation Molecular characterisation
  • 16. Embryonal RMS Common in head & neck, genitourinary tract, paraspinal/parameningeal sites Incidence declines after age 3 yrs Prognosis is dependant on a number of factors:  Site  Age  Staging  Histology All are important and form part of an IRS grouping for patient management
  • 17. Histopathology Embryonal RMS has a variable pattern Varying degree of cellularity with alternating dense hypercellular areas and more loose myxoid areas A mixture of round cells, spindle cells and more differentiated cells showing rhabdomyoblastic differentiation Anaplasia can be present in embyonal and alveolar RMS (?worst prognosis)
  • 18. Embryonal RMS
  • 19. Embryonal RMS
  • 20. Alveolar rhabdomyosarcoma A-RMS accounts for at least 30-40% of all RMS Alveolar RMS displays a bimodal age incidence A peak at 3 years and 15 years of age Higher incidence in upper and lower limbs
  • 21. A-RMS - Pathology Tumour composed of ill defined aggregate/nests of poor differentiated round to oval cells Some cells may show rhabdomyoblastic differentiation The nests usually show central loss of cellular cohesion forming irregular alveolar spaces
  • 22. A-RMS - Pathology (continued) Fibrovascular septa separate the nests of cells, with single cells adherent to these septa Mitotic figures are common Wreath like giant cells are a characteristic and diagnostically useful feature of alveolar RMS The alveolar pattern is also seen in metastatic sites Solid variant of alveolar RMS is an important subtype in which fibrous septa may be absent or inconspicuous
  • 23. Alveolar RMS
  • 24. Alveolar RMS
  • 25. Wreath like giant cells in A-RMS
  • 26. Solid variant A-RMS
  • 27. Reticulin
  • 28. A-RMS - Differential diagnosis Any small round cell tumour but in particular pseudo-alveolar pattern in  PNET  alveolar soft part sarcoma  desmoplastic small round cell tumour
  • 29. Pseudoalveolar pattern in PNET
  • 30. Immunohistochemistry in RMSRecommended panel: Desmin (in 99%) Myogenin (in 100%) – most specific MyoD1 (in 100%) Muscle Specific Actin (in 94%) CD99 (weak granular) (in 14%) CAM5.2 EMA SynaptophysinOther useful markers Myoglobin (in 78%) in more differentiated forms Sarcomeric actin Myosin
  • 31. Desmin staining
  • 32. Myo D1 nuclear staining
  • 33. Myogenin CD99
  • 34. Rhabdomyosarcoma - immunohistochemistry vs prognosis Myogenin - >80% ARMS vs >25% ERMS (>50% of tumour cells) Diffuse myogenin + a marker of poor prognosis (independent of fusion status and histology) PAX-5 – 67% ARMS vs 0% ERMS
  • 35. Rhabdomyosarcoma - molecular biology ERMS associated with multiple numeric and structural chromosomal changes (extra copies of chromosomes 2, 8 and 13; LOH at 11p15 – BW region) ARMS associated with two specific gene fusions  PAX3-FOXO1 (correlated with t(2;13)(q35;q14) – 3x more common  PAX7-FOXO1 (correlates with t(1;13)(p36;q14) – better prognosis  20-25% ARMS have neither of these translocations – their genetic profile identical to ERMS - ?prognosis
  • 36. Synovial Sarcoma 5-10% of STS 42% of paediatric non-RMS STS 85-95% arise in the extremities, then head and neck Biphasic pattern – spindle cells and epithelial gland components In children – monophasic SS more common
  • 37. Focal epithelial differentiation
  • 38. Cytokeratin staining
  • 39. SS - Immunoprofile Cytokeratin + (some are -) EMA + Vimentin + S100 + in 30% of cases CD99 + in 60% of cases BCL2 + in 75% - 100% of cases CD34 - Calponin + Desmin - TLE1 - nearly 100%
  • 40. Synovial Sarcoma Characterised by a translocation: t(x;18)(p11.2;q11.2) (SS18-SSX1); (SS18-SSX2); (SS18-SSX4) The identification of a specific translocation has resulted in the ability to diagnose these tumours in unusual sites (viscera) It is also possible to identify poorly or undifferentiated SS Is an important tool in differentiating SS from other spindle cell tumours (congenital fibrosarcoma)
  • 41. Congenital Infantile Fibrosarcoma Diagnosed in the 1st year of life (~50% congenital) Usually occur in distal extremities & head and neck Histologically simulates adult-type fibrosarcoma 5-year survival of >90% Recurrence rate 30% Metastases rare Chromosomal translocation t(12;15)(p13;q26) (ETV6/NTRK3 genes)
  • 42. Congenital Fibrosarcoma
  • 43. Vimentin staining
  • 44. EWS/PNET Ewing Family of Tumours EWS/PNET - a group of tumours with overlapping clinical & pathological features peak incidence in childhood & young adolescence EWS – classically a primary bone neoplasm (rarely extra-osseous) PNET – usually arises in soft tissues (exceptionally in bone)
  • 45. PNET - Pathology i) lobular, ii) diffuse & iii) incohesive growth pattern foci of haemorrhage but NO calcification tumour lobules divided by fibro-connective septae Homer-Wright & perivascular pseudo-rosettes rare small cells, round-oval nuclei, coarse chromatin, small nucleoli, narrow rim of eosinophilic cytoplasm intercellular fibrillary material or ganglion cells exceptionally rare
  • 46. IHC of EWS/PNET – essentialPOSITIVE for: CD99 ~95%, perimembranous VIM ~95%, focal or diffuse, dot-like or perinuclear NSE, S100, synaptophysin, NFP ~40% Chromogranin ~20% GFAP <10% EMA, Cytokeratin, Desmin <10%
  • 47. CD99
  • 48. Ewing’s Family of Tumours – molecular biology t(11;22) (involving EWS gene) – 85-90% of ESF - t(11;22)(q24;q12) (EWSR1-FLI1) - Type 1 (exon 7 – exon 6) - ?better prognosis - Type 2 (exon 7 – exon 5)- t(21;22)(q22;q12) (EWSR1-ERG) – 5-10% of ESF- 4-9% of ESF – other fusions
  • 49. Desmoplastic Small Round Cell Tumour - DSRCT rare, highly aggressive tumour of uncertain origin M>F presents with abdominal distension, pain & mass at laparotomy – variable sized mass with numerous smaller nodular peritoneal implants throughout the peritoneal cavity
  • 50. DSRCT - ImmunohistochemistryNot specific but distinct co-expression of i) epithelial (AE1/AE3, CAM5.2) ii) neural (NSE, PGP9.5; CD99 in 20-35%) iii) muscle markers (desmin; negative for MyoD1, myoglobin)
  • 51. DSRCT – Molecular biology This tumour is defined by a reciprocal translocation t(11;22)(p13;q12) (EWS-WT1 genes) This translocation creates a fusion gene that transcribes a chimeric protein containing portions of WT1 protein & EWS protein
  • 52. DSRCT - Prognosis a very aggressive neoplasm chemotherapy may induce temporary remission the ultimate outcome remains dismal
  • 53. Molecular findings are the Gold Standard for diagnosis ? ‘Non-specificity’ of immunomorphology  CD56 - PNET, RMS, NB, WT  CD99 - PNET, DSRCT, WT  Bcl-2 - PNET, DSRCT, WT, NB, SS  WT1 - PNET, RMS, DSRCT, NB, WT vs. specific diagnosis with molecular techniques  ETV6/NTRK3 in Infantile fibrosarcoma  SS18-SSX1 in Synovial sarcoma  EWS-FLI1 / other translocations in atypical PNET
  • 54. Histomorphology is the Gold Standard for diagnosis? ‘Non-specificity’ of molecular markers  12;15 ETV6-NTRK3 in CMN/IFS and secretory breast carcinoma ‘A FISH study of ETV-NTRK3 fusion gene in secretry breats carcinoma’ Marketsov et al. Genes Chrom Cancer 2004; 40: 152-7  X;18 in SS and MPNST ‘Lack of SYT-SSX fusion transcripts in malignant peripheral nerve sheath tumors on RT- PCR analysis of 34 archival cases’ Tamborini et al. Lab Invest 2002; 82: 609-18  Xp1 RCC and AlvSPS ‘Primary renal neoplasms with the ASPL-TFE3 gene fusion of alveolar soft part sarcoma: a distinctive entity previously included among renal cell carcinoma of children and adolescents’ Argani et al. Am J Surg Pathol 2001; 159: 179-92
  • 55. SUMMARY Molecular studies invaluable for diagnostic and clinical management of a range of tumours In the future, clinical decisions will increasingly be based on a combination of histological criteria and the molecular identification of genetic abnormalities Future advances likely to lead to even greater diagnostic and prognostic information, and identification of new therapeutic targets in treatment Gold standard is interpretation of molecular findings in the context of the histopathological features

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