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Figure 3. Classification analysis to follicular lymphoma (FL) subtypes FL3A, FL3B and DLBCL/FL3B into either the category of FL1/2 or germinal center B-cell dif- fuse large B-cell lymphoma (GCB-DLBCL). While the Gene Expression Profiles (GEPs) were clearly separated for FL1/2 and GCB-DLBCL, no clear distinction was seen for FL3A and FL3B. DLBCL/FL3B tended to cluster in the group of GCB-DLBCL (A). Schematic overview of FL subtype categorization based on GEP. While FL1/2 and GCB-DLBCL indicate a homogeneous expression profiling clearly separated from each other, the expression pattern of FL3A and FL3B is in between FL and DLBCL. DLBCL/FL3B are more closely related to DLBCL than to FL (B).
Gene expression profiling of FL subtypes
A
B
approximately 75% of all iterations. Eleven of 16 (69%) FL3A clustered within the group of FL1/2 (Figure 3A). Considering genetic features of the resulting 5 ‘misclassi- fied’ FL3A, the frequency of BCL6 breaks in this group (3 of 5, 60%) was more similar to the DLBCL group than to FL1/2 (GCB-DLBCL: 44% vs. FL1/2: 9%) (Table 1). Of interest in this context, the GEP of 3 FL3B (MPI-661, MPI- 721 and MPI-817; 3 of 6, 50%) was more closely related to DLBCL than to FL1/2, while the remaining 3 FL3B (50%) were clearly classified as FL1/2 (Figure 3A). However, there was no significant difference in genetic features of the 2 FL3B clusters, especially not with respect to the occurrence of the t(14;18). With special regard to immuno- histochemical markers, no differences were detected that could possibly separate the ‘core’ group from the outliers.
The majority of the DLBCL/FL3B samples (8 of 9, 89%) were classified as DLBCL, apart from MPI-650 which was classified as FL1/2 in more than 70% of all iterations (Figure 3A). The only striking finding in this sample was the lack of a BCL6-translocation, while 44% of either DLBCL/FL3B or DLBCL, respectively, harbored the rearrangement.
To summarize, gene expression profiling and classifica- tion analysis showed FL1/2 to be clearly separated from DLBCL, indicating a distinct FL-specific and DLBCL-spe- cific gene expression pattern, as would have been expect- ed. FL3B turned out to be closely related to FL3A, not cat- egorizing within a separate gene expression cluster, and
both FL3A and FL3B showed overlapping GEPs in between FL1/2 and DLBCL. Finally, based upon their expression pattern, DLBCL/FL3B did indeed seem to rep- resent a composite form of FL3B and DLBCL, with the majority of samples more closely resembling DLBCL (Figure 3B).
Gene expression profiles of FL with or without the t(14;18) do not differ significantly in the various histological subtypes
Presence or absence of the t(14;18) was one of the fac- tors most distinguishing FL1-3A from FL3B, DLBCL/FL3B and GCB-DLBCL (see above). We, therefore, asked whether GEP might be different in lymphomas with or without the t(14;18). Interestingly, such a difference was not seen either in the entire FL cohort, nor within different histological subtypes. Only 2 genes (FAM30A and IL17RB) were differentially expressed between t(14;18)-positive and t(14;18)-negative FL1/2, FL3A and FL3B (n=33 and n=11, respectively). Upon classification analysis, trying to separate t(14;18)-positive from negative FL1-3B samples, GEP of t(14;18)-negative FL1-3B was quite homogenous, while the classification profiles of some t(14;18)-positive cases fluctuated quite heavily, with 6 FL with t(14;18) (MPI-600, MPI-604, MPI-640, MPI-659, MPI-667 and MPI- 668) (6 of 33, 18%) showing a GEP more similar to the t(14;18)-negative cohort (Figure 4A). When testing for the differential expression of single genes that varied between
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