- Email: [email protected]
The role of molecular testing in soft tissue sarcoma diagnosis
Comment
What is the true fidelity of the expert human eye and brain in affirming an accurate sarcoma diagnosis? The study by Antoine Italiano and colleagues1 aims to answer this question by linking diagnoses made by sarcoma experts to molecular genetic testing done with fluorescence in-situ hybridisation (FISH), PCR, or reverse transcriptase-PCR for six soft tissue sarcoma subtypes. This level IV diagnostic and therapeutic study included 384 patients at 32 centres managed by the French Sarcoma Group. Sarcoma subtypes included dermatofibrosarcoma protuberans, dedifferentiated liposarcoma, Ewing’s sarcoma, synovial sarcoma, alveolar rhabdomyosarcoma, and myxoid or round cell liposarcoma. Diagnoses were first made based solely on histological characteristics and re-assessed after molecular genetic testing had been done. The histological diagnoses were altered in 53 patients (23%), with an effect on primary management or patient prognosis in 45 cases. It should be noted that because molecular genetic testing was excluded entirely from the initial pathology impression (to better assess the contribution towards a final diagnosis), the diagnostic discrepancy of 23% cannot be compared directly with other studies. However, the aim of the study was to report the necessity of molecular testing and not to provide a comparison study. As all sarcomatogists acknowledge, there is a deceiving heterogeneity within soft tissue sarcoma that is not necessarily conveyed solely by histopathology, and molecular diagnostics are necessary to confirm the diagnosis with any stringency.2 With more than 50 different soft tissue sarcoma subtypes, all mesenchymal in origin by definition (with concessions for Ewing’s and nerve tumours), distinguishing between them histologically is difficult. Adding to this predicament, these distinguishing characteristics are often more readily identified in low-grade soft tissue sarcoma but can become less distinct in higher grade and dedifferentiated soft tissue sarcoma,3 with tumour characteristics overlapping like a Venn diagram. The study design used by Italiano and colleagues cleverly and realistically shows the pragmatic responsibility of the pathologist, with diagnoses listed as certain, probable, or possible before molecular testing.
Obtaining certain soft tissue sarcoma diagnoses can be difficult. For example, pathologists were not especially confident in diagnosing synovial sarcoma (split widely with 29 [30%] certain, 42 [43%] probable, and 26 [27%] possible of 97 cases), and molecular testing of these samples was also split with 56 (58%) positive and 41 (42%) negative. Reassuringly, 24 of 26 possible synovial sarcoma diagnoses were negative with molecular testing. Furthermore, pathologists only listed nine (20%) of 46 alveolar rhabdomyosarcoma diagnoses as certain, and 26 (56%) as possible, suggesting this tumour really is a diagnosis of exclusion by histological standards. Molecular testing showed only 19 (41%) positive and 27 (59%) negative in all samples tested for alveolar rhabdomyosarcoma. With so much dependent on an accurate diagnosis, molecular testing can dissuade potential frustration by the pathologists. The advent and further evolution of molecular genetic testing is continuously exposing important soft tissue sarcoma subtypes.4 After the replacement of malignant fibrous histiocytoma in favour of undifferentiated pleomorphic sarcoma,5,6 several subtypes have been delineated from the morphological undifferentiated pleomorphic sarcoma pool using cytogenetics. These include angiomatoid fibrous histiocytoma (EWSR1-CREB1 or EWSR1-ATF1)7 and myxofibrosarcoma (FUS-CREB3L2 and FUS-CREB3L1);8 however, giant cell rich pleomorphic sarcoma and undifferentiated pleomorphic sarcoma with prominent inflammation remain diagnoses of exclusion. These distinctions have proven valuable to surgical management, as an infiltrative myxofibrosarcoma typically requires a much larger resection margin than the physical exam and imaging would suggest, and this would be grossly underestimated were the lesion incorrectly called undifferentiated pleomorphic sarcoma using histology alone. The hope is that future treatment of soft tissue sarcoma, and not just diagnosis, can be tailored to molecular characteristics. Genotyping has shown that most soft tissue sarcomas have cytogenetic abnormalities that are either exclusive or plethoric, with no middle ground. For plethoric sarcomas, it is still unknown whether molecular testing will help to find unique differences in soft tissue sarcomas for diagnoses, or converge on common oncogenic pathways for
www.thelancet.com/oncology Published online March 9, 2016 http://dx.doi.org/10.1016/S1470-2045(15)00625-7
Steve Gschmeissner/Science Photo Library
The role of molecular testing in soft tissue sarcoma diagnosis
Lancet Oncol 2016 Published Online March 9, 2016 http://dx.doi.org/10.1016/ S1470-2045(15)00625-7 See Online/Articles http://dx.doi.org/10.1016/ S1470-2045(15)00583-5
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Comment
possible treatments. For example, MDM2 amplification is used to confirm liposarcoma but is also expressed in many other tumour types including myxofibrosarcoma, leiomyosarcoma, and MPNST.9 Therefore, conventional histology remains relevant. It will be interesting when the investigators publish a future study containing their results of molecular testing done on fresh frozen versus formalin-fixed paraffin-embedded tissue. The development of next-generation sequencing techniques capable of using formalin-fixed and paraffin-embedded tissues might enable a more streamlined multi-institutional approach for centres with insufficient resources to do testing themselves.10 This study also provided an ancillary point involving biopsy technique, with an altered diagnosis not associated with either core needle or open biopsy. The brain might eventually supersede the microscope and the expert human eye—but not yet. Before we can treat soft tissue sarcoma appropriately, we have to at least identify it correctly, and molecular genetic testing will play an integral role alongside histopathology moving forward to maximise our fidelity. *Lor Randall, Matthew G Cable
LR receives honorarium for a speaking engagement with Biomet, and institutional support for research from the Musculoskeletal Transplant Foundation. MGC declares no competing interests. 1
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Italiano A, Di Mauro I, Rapp J, et al. Clinical effect of molecular methods in sarcoma diagnosis (GENSARC): a prospective, multicentre, observational study. Lancet Oncol 2016; published online March 9. http://dx.doi. org/10.1016/S1470-2045(15)00583-5. Randall RL, Bruckner JD, Papenhausen MD, et al. Errors in diagnosis of soft-tissue sarcomas initially treated at non-tertiary centers. Orthopedics 2004; 27: 209–12. Coindre JM, Trojani M, Contesso G, et al. Reproducibility of a histopathologic grading system for adult soft tissue sarcoma. Cancer 1986; 58: 306–09. Henderson-Jackson EB, Bui MM. Molecular pathology of soft-tissue neoplasms and its role in clinical practice. Cancer Control 2015; 22: 186–92. Matushansky I, Charytonowicz E, Mills J, et al. MFH classification: differentiating undifferentiated pleomorphic sarcoma in the 21st century. Expert Rev Anticancer Ther 2009; 9: 1135–44. Randall RL, Albritton KH, Ferney BJ, et al. Malignant fibrous histiocytoma of soft tissue: an abandoned diagnosis. Am J Orthop 2004; 33: 602–08. Thway K, Gonzalez D, Wren D, et al. Angiomatoid fibrous histiocytoma: comparison of fluorescence in situ hybridization and reverse transcription polymerase chain reaction as adjunct diagnostic modalities. Ann Diagn Pathol 2015; 19: 137–42. Rose B, Tamvakopoulos GS, Dulay K, et al. The clinical significance of the FUS-CREB3L2 translocation in low-grade fibromyxoid sarcoma. J Orthop Surg Res 2011; 6: 15. Bihn MB, Sastre-Garau X, Guillou L, et al. MDM2 and CDK4 immunostainings are useful adjuncts in diagnosing well-differentiated and dedifferentiated liposarcoma subtypes: a comparative analysis of 559 soft tissue neoplasms with genetic data. Am J Surg Pathol 2005; 29: 1340–47. van de Rijn M, Guo X, Sweeney RT, et al. Molecular pathological analysis of sarcomas using paraffin-embedded tissue: current limitations and future possibilities. Histopathology 2014; 64: 163–170.
University of Utah, Huntsman Cancer Institute, UT 84103, USA [email protected]
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www.thelancet.com/oncology Published online March 9, 2016 http://dx.doi.org/10.1016/S1470-2045(15)00625-7
What is the true fidelity of the expert human eye and brain in affirming an accurate sarcoma diagnosis? The study by Antoine Italiano and colleagues1 aims to answer this question by linking diagnoses made by sarcoma experts to molecular genetic testing done with fluorescence in-situ hybridisation (FISH), PCR, or reverse transcriptase-PCR for six soft tissue sarcoma subtypes. This level IV diagnostic and therapeutic study included 384 patients at 32 centres managed by the French Sarcoma Group. Sarcoma subtypes included dermatofibrosarcoma protuberans, dedifferentiated liposarcoma, Ewing’s sarcoma, synovial sarcoma, alveolar rhabdomyosarcoma, and myxoid or round cell liposarcoma. Diagnoses were first made based solely on histological characteristics and re-assessed after molecular genetic testing had been done. The histological diagnoses were altered in 53 patients (23%), with an effect on primary management or patient prognosis in 45 cases. It should be noted that because molecular genetic testing was excluded entirely from the initial pathology impression (to better assess the contribution towards a final diagnosis), the diagnostic discrepancy of 23% cannot be compared directly with other studies. However, the aim of the study was to report the necessity of molecular testing and not to provide a comparison study. As all sarcomatogists acknowledge, there is a deceiving heterogeneity within soft tissue sarcoma that is not necessarily conveyed solely by histopathology, and molecular diagnostics are necessary to confirm the diagnosis with any stringency.2 With more than 50 different soft tissue sarcoma subtypes, all mesenchymal in origin by definition (with concessions for Ewing’s and nerve tumours), distinguishing between them histologically is difficult. Adding to this predicament, these distinguishing characteristics are often more readily identified in low-grade soft tissue sarcoma but can become less distinct in higher grade and dedifferentiated soft tissue sarcoma,3 with tumour characteristics overlapping like a Venn diagram. The study design used by Italiano and colleagues cleverly and realistically shows the pragmatic responsibility of the pathologist, with diagnoses listed as certain, probable, or possible before molecular testing.
Obtaining certain soft tissue sarcoma diagnoses can be difficult. For example, pathologists were not especially confident in diagnosing synovial sarcoma (split widely with 29 [30%] certain, 42 [43%] probable, and 26 [27%] possible of 97 cases), and molecular testing of these samples was also split with 56 (58%) positive and 41 (42%) negative. Reassuringly, 24 of 26 possible synovial sarcoma diagnoses were negative with molecular testing. Furthermore, pathologists only listed nine (20%) of 46 alveolar rhabdomyosarcoma diagnoses as certain, and 26 (56%) as possible, suggesting this tumour really is a diagnosis of exclusion by histological standards. Molecular testing showed only 19 (41%) positive and 27 (59%) negative in all samples tested for alveolar rhabdomyosarcoma. With so much dependent on an accurate diagnosis, molecular testing can dissuade potential frustration by the pathologists. The advent and further evolution of molecular genetic testing is continuously exposing important soft tissue sarcoma subtypes.4 After the replacement of malignant fibrous histiocytoma in favour of undifferentiated pleomorphic sarcoma,5,6 several subtypes have been delineated from the morphological undifferentiated pleomorphic sarcoma pool using cytogenetics. These include angiomatoid fibrous histiocytoma (EWSR1-CREB1 or EWSR1-ATF1)7 and myxofibrosarcoma (FUS-CREB3L2 and FUS-CREB3L1);8 however, giant cell rich pleomorphic sarcoma and undifferentiated pleomorphic sarcoma with prominent inflammation remain diagnoses of exclusion. These distinctions have proven valuable to surgical management, as an infiltrative myxofibrosarcoma typically requires a much larger resection margin than the physical exam and imaging would suggest, and this would be grossly underestimated were the lesion incorrectly called undifferentiated pleomorphic sarcoma using histology alone. The hope is that future treatment of soft tissue sarcoma, and not just diagnosis, can be tailored to molecular characteristics. Genotyping has shown that most soft tissue sarcomas have cytogenetic abnormalities that are either exclusive or plethoric, with no middle ground. For plethoric sarcomas, it is still unknown whether molecular testing will help to find unique differences in soft tissue sarcomas for diagnoses, or converge on common oncogenic pathways for
www.thelancet.com/oncology Published online March 9, 2016 http://dx.doi.org/10.1016/S1470-2045(15)00625-7
Steve Gschmeissner/Science Photo Library
The role of molecular testing in soft tissue sarcoma diagnosis
Lancet Oncol 2016 Published Online March 9, 2016 http://dx.doi.org/10.1016/ S1470-2045(15)00625-7 See Online/Articles http://dx.doi.org/10.1016/ S1470-2045(15)00583-5
1
Comment
possible treatments. For example, MDM2 amplification is used to confirm liposarcoma but is also expressed in many other tumour types including myxofibrosarcoma, leiomyosarcoma, and MPNST.9 Therefore, conventional histology remains relevant. It will be interesting when the investigators publish a future study containing their results of molecular testing done on fresh frozen versus formalin-fixed paraffin-embedded tissue. The development of next-generation sequencing techniques capable of using formalin-fixed and paraffin-embedded tissues might enable a more streamlined multi-institutional approach for centres with insufficient resources to do testing themselves.10 This study also provided an ancillary point involving biopsy technique, with an altered diagnosis not associated with either core needle or open biopsy. The brain might eventually supersede the microscope and the expert human eye—but not yet. Before we can treat soft tissue sarcoma appropriately, we have to at least identify it correctly, and molecular genetic testing will play an integral role alongside histopathology moving forward to maximise our fidelity. *Lor Randall, Matthew G Cable
LR receives honorarium for a speaking engagement with Biomet, and institutional support for research from the Musculoskeletal Transplant Foundation. MGC declares no competing interests. 1
2
3
4
5
6 7
8
9
10
Italiano A, Di Mauro I, Rapp J, et al. Clinical effect of molecular methods in sarcoma diagnosis (GENSARC): a prospective, multicentre, observational study. Lancet Oncol 2016; published online March 9. http://dx.doi. org/10.1016/S1470-2045(15)00583-5. Randall RL, Bruckner JD, Papenhausen MD, et al. Errors in diagnosis of soft-tissue sarcomas initially treated at non-tertiary centers. Orthopedics 2004; 27: 209–12. Coindre JM, Trojani M, Contesso G, et al. Reproducibility of a histopathologic grading system for adult soft tissue sarcoma. Cancer 1986; 58: 306–09. Henderson-Jackson EB, Bui MM. Molecular pathology of soft-tissue neoplasms and its role in clinical practice. Cancer Control 2015; 22: 186–92. Matushansky I, Charytonowicz E, Mills J, et al. MFH classification: differentiating undifferentiated pleomorphic sarcoma in the 21st century. Expert Rev Anticancer Ther 2009; 9: 1135–44. Randall RL, Albritton KH, Ferney BJ, et al. Malignant fibrous histiocytoma of soft tissue: an abandoned diagnosis. Am J Orthop 2004; 33: 602–08. Thway K, Gonzalez D, Wren D, et al. Angiomatoid fibrous histiocytoma: comparison of fluorescence in situ hybridization and reverse transcription polymerase chain reaction as adjunct diagnostic modalities. Ann Diagn Pathol 2015; 19: 137–42. Rose B, Tamvakopoulos GS, Dulay K, et al. The clinical significance of the FUS-CREB3L2 translocation in low-grade fibromyxoid sarcoma. J Orthop Surg Res 2011; 6: 15. Bihn MB, Sastre-Garau X, Guillou L, et al. MDM2 and CDK4 immunostainings are useful adjuncts in diagnosing well-differentiated and dedifferentiated liposarcoma subtypes: a comparative analysis of 559 soft tissue neoplasms with genetic data. Am J Surg Pathol 2005; 29: 1340–47. van de Rijn M, Guo X, Sweeney RT, et al. Molecular pathological analysis of sarcomas using paraffin-embedded tissue: current limitations and future possibilities. Histopathology 2014; 64: 163–170.
University of Utah, Huntsman Cancer Institute, UT 84103, USA [email protected]
2
www.thelancet.com/oncology Published online March 9, 2016 http://dx.doi.org/10.1016/S1470-2045(15)00625-7