Validation of the Complexity INdex in SARComas prognostic signature on formalin-fixed, paraffin-embedded, soft-tissue sarcomas

Canine oral malignant melanomas (OMMs) exhibit a variety of morphologic phenotypes, including a spindloid variant. The microscopic diagnosis of spindloid OMMs is based on junctional activity and/or the presence of melanin pigment. In the absence of these features, spindloid OMMs are difficult to differentiate from soft tissue sarcomas (STS). An antibody cocktail (MDX) that includes Melan-A, PNL2, and tyrosinase-related proteins 1 and 2 (TRP-1 and TRP-2) is the current gold standard for identifying amelanotic OMMs by immunohistochemistry (IHC). However, MDX is less sensitive for diagnosing spindloid amelanotic OMMs. This raises concern for biopsy specimens that lack overlying epithelium, making it potentially difficult to differentiate OMM from STS by IHC. The goal of this study was to identify additional markers to help differentiate between STS and OMMs that lack pigment and junctional activity. SOX-10 has recently been proposed as a sensitive marker for melanocytes in humans but has not been validated in dogs. Similarly, RNA expression for various genes has been analyzed in humans, but not in the context of diagnosing canine melanocytic neoplasms. For this retrospective study, formalin-fixed, paraffin-embedded tissues from 20 OMMs, 20 STS, and 20 oral spindle cell tumors (OSCTs) that lacked junctional activity and pigmentation were selected. IHC for MDX, SOX-10, and laminin, in parallel with RT-qPCR of TYR, SOX10, CALD1, CD34, DES, and LAMA1, was performed in all cases. TYR, CD34, and CALD1 were the most discriminatory genes in differentiating between OMM and STS, all having 100% specificity and 65, 95, and 60% sensitivity, respectively. While all 20 OMMs were immunohistochemically labeled for SOX-10, two STS were also labeled (100% sensitivity and 90% specificity). MDX IHC labeled all 20 OMMs and no STS. Surprisingly, none of the 20 OSCTs expressed TYR RNA above the cutoff, and 14/20 OSCTs expressed CALD1 or CD34 RNA above the cutoff, thereby confirming them as STS. Four OSCT were suspect STS, and no OSCTs were confirmed as OMMs based on IHC and RNA expression patterns. In conclusion, the RNA levels of TYR, CD34, and CALD1 should be evaluated in suspected amelanotic OMMs that are negative for MDX to accurately differentiate between OMM and STS.

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Detection of Disease-specific Fusion Genes of Soft Tissue Tumors Using Formalin-fixed Paraffin-embedded Tissues; Its Diagnostic Usefulness and Factors Affecting the Detection Rates

Yonago Acta Medica ◽

10.33160/yam.2019.03.016 ◽

2019 ◽

Vol 62 (1) ◽

pp. 115-123

Author(s):

Takahiro Matsushige ◽

Satoshi Kuwamoto ◽

Michiko Matsushita ◽

Lusi Oka Wardhani ◽

Yasushi Horie ◽

...

Keyword(s):

Soft Tissue ◽

Soft Tissue Tumors ◽

Fusion Genes ◽

Detection Rates ◽

Factors Affecting ◽

Formalin Fixed Paraffin ◽

Formalin Fixed Paraffin Embedded ◽

Diagnostic Usefulness ◽

Formalin Fixed ◽

Disease Specific

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Evaluation of the optimal provision of formalin-fixed, paraffin-embedded material for reverse transcription-PCR in soft-tissue tumour diagnosis

Journal of Clinical Pathology ◽

10.1136/jclinpath-2016-203691 ◽

2016 ◽

Vol 70 (1) ◽

pp. 20-24 ◽

Cited By ~ 8

Author(s):

Khin Thway ◽

Dorte Wren ◽

Jasmin Lee ◽

Lisa Thompson ◽

Cyril Fisher ◽

...

Keyword(s):

Soft Tissue ◽

Reverse Transcription ◽

Rna Extraction ◽

Histopathological Diagnosis ◽

Technical Failure ◽

Rt Pcr ◽

Reverse Transcription Pcr ◽

Formalin Fixed Paraffin ◽

Formalin Fixed Paraffin Embedded ◽

Formalin Fixed

AimsMolecular genetic analysis is now a routine ancillary diagnostic modality to the histopathological diagnosis of soft-tissue neoplasms, many of which harbour characteristic gene fusions detectable by reverse transcription-PCR (RT-PCR). As the final diagnosis often depends on the molecular result, it is important to obtain the optimal yield of patient RNA.MethodsWe assessed the most reliable method of providing formalin-fixed, paraffin-embedded material for optimal RNA yield by comparing three consecutive periods in which different preparations (5×10 μm scrolls, 5×5 μm sections and 1×10 μm sections) were used for RNA extraction for RT-PCR, with its technical success rate.ResultsFor ‘2011’, ‘2012’ and ‘2013’, RT-PCR technical failure rates were 13.4%, 4.4% and 7.9%, respectively. The percentage of failed referral cases was 71.4%, 85.7% and 31.3%, and the proportion of core biopsy to excision specimens was 3:15, 2:5 and 13:3.ConclusionsThis study shows that the effectiveness of RNA extraction and purification is dependent on both specimen type and the tissue sectioning strategy. The failure rate has improved over recent years, particularly for large specimens as large numbers of thick 10 μm scrolls can saturate RNA extraction columns. In contrast, recent technical fails are more frequent in core biopsies, where 1×10 μm sections are insufficient for adequate RNA extraction. While previous technical fails occurred mostly in referred cases, this appears no longer the case due to the better fixation and processing of specimens in external surgical pathology departments because of the widespread recognition of the importance of molecular diagnostics as an important part of the patient pathway.

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