Specificity of TLS-CHOP rearrangement for classic myxoid/round cell liposarcoma: absence in predominantly myxoid well-differentiated liposarcomas

Abstract

Myxoid liposarcoma (LS), the most common subtype of LS, is known to be characterized by the specific t(12;16) resulting in a TLS-CHOP fusion in almost all cases. We wished to address the following questions: (i) Is this genetic hallmark also present in other types of LS with predominant myxoid change? (ii) What is the proportion of cases with the variant EWS-CHOP fusion? (iii) What is the optimal approach for Southern blot detection of TLS breakpoints? We identified 59 LS characterized histologically by >90% myxoid component, in which frozen tissue tumor was available for DNA extraction. These 59 LS with myxoid features were divided into 2 groups: 42 LS with classic myxoid/round cell appearance (myxoid LS) and 17 well-differentiated LS (WDLS) with a predominant (>90%) myxoid component. Within the myxoid LS group, 29 tumors were low grade and 13 high grade (>20% round cell component). Among the 17 predominantly myxoid WDLS, there were 15 low grade and 2 focally high grade tumors. In addition, we selected as control group, 20 LS of other histological types with minimal or no myxoid change (17 WDLS and 3 pleomorphic LS) and 13 myxofibrosarcomas. Southern blot analysis was performed in all cases using a CHOP cDNA probe, and in all CHOP rearranged cases using a TLS cDNA probe. Probe/enzyme combinations for Southern blot analysis were CHOP exon 3-4 cDNA probe with BamHI or SacI, TLS exon 3-6 cDNA probe with BclI. All 42 cases of myxoid LS showed a CHOP rearrangement and 38 of them also had a TLS rearrangement. Among the 4 myxoid LS without Southern blot evidence of TLS rearrangement, 1 showed an EWS-CHOP fusion by Southern blotting and reverse transcriptase-polymerase chain reaction and in another case, reverse transcriptase-polymerase chain reaction detected a TLS-CHOP fusion transcript. None of the predominantly myxoid WDLS and none of the tumors included in the control group showed rearranegements with CHOP probe. In addition, 12 predominantly myxoid WDLS, 10 other LS, and 5 myxofibrosarcoma from the control group were also tested for TLS rearrangement; all were negative. The TLS-CHOP fusion is highly sensitive and specific for the entity of classic myxoid/round cell LS. Other types of LS, even with a predominant myxoid component, lack the TLS-CHOP rearrangement, confirming that they represent a genetically distinct group of LS. The prevalence of the EWS-CHOP variant fusion was approximately 2% in this series. The optimal enzyme for TLS genomic breakpoint detection is BclI.

Figure 1.

A: Classic myxoid LS, low grade, with small, uniform tumor cells in a background of myxoid stroma and delicate “chicken-wire” type vascular network. Of note is the absence of the pleomorphic giant tumor cells characteristic of WDLS with myxoid changes. B: WDLS, predominantly myxoid. Although the myxoid background and the branching vasculature might suggest the diagnosis of classic myxoid LS, the tumor cells are larger and less uniform. C: WDLS, predominantly myxoid, with focal areas of lipoma-like LS and scattered tumor giant cells. D: Myxofibrosarcoma, low grade. Most of the tumor cells are uniform, but predominantly spindly, in contrast with the round to oval appearance of myxoid LS. Rare pleomorphic tumor cells are scattered within the myxoid stroma, which helps in distinguishing this tumor from classic myxoid LS.

Figure 2.

TLS genomic map with selected restriction enzyme sites (B, BamHI; E, EcoRI; Bc, BclI; P, PstI; S, SacI). The map is based on the complete genomic sequence of TLS (GenBank no. AF071213 17 ). Exons encoding translated sequences are shown as filled boxes, and the 3′ untranslated portion of exon 15 is shown as an empty box. The TLS breakpoint cluster regions are based on references 18 and 19. The TLS probe is described in the text (see Methods). Partial restriction maps of the CHOP gene have been previously published.

Figure 3.

CHOP rearrangement analysis by Southern blotting. Rearranged bands are seen in both SacI- and BamHI-digested genomic DNA in four cases of classic myxoid LS (LS4, LS13, LS24, LS37) using a CHOP exon 3 and 4 cDNA probe (see Methods).

Figure 4.

TLS rearrangement analysis by Southern blotting. Rearranged bands are seen in BclI-digested genomic DNA in two cases of classic myxoid LS (LS2 and LS15), but not in a case of predominantly myxoid WDLS (LS56) using a TLS exon 3–6 cDNA probe (see Methods and Figure 2 ).