ALBERT

Description: Abstract 2900 Poster Board II-876 Ph-negative myeloproliferative neoplasms: polycythemia vera (PV), essential thrombocythemia (ET) and primitive myelofibrosis (PMF) carry an acquired somatic mutation JAK2V617F in 95% (PV), and in 50 to 60% (ET or PMF) of the patients. Mutations of the TET2 gene have been observed with roughly similar frequencies in the three MPN, irrespective of the presence of JAK2V617F. Evolution to myelofibrosis or acute leukemia may occur with time in MPN patients. Although its molecular bases are poorly understood, the evolution is likely due to the acquisition of additional mutations. To investigate whether cytogenetic abnormalities are distributed differently according to type of transformation and to the JAK2 and TET2 statuses, the Groupe Francophone de Cytogénétique Hématologique has collected 82 patients with transformation of MPN. There were 66 (80%) acute myeloid leukemia or myelodysplastic syndromes (AML/MDS) and 16 (20%) myelofibroses (MF). Of note pipobroman (Pi) treatment seems to be associated with MF, and hydroxyuera (Hu) with AML/MDS evolution in our series. Statistical analyses of clinical, cytogenetic and molecular data are shown Table 1. On the cytogenetical point of view, several points are noteworthy. Some abnormalities were unevenly distributed: there were significantly more -7/del7q and -5/del5q in AML/MDS and tri1q and tri9 in MF. MF and PMF cytogenetic profile looked similar, suggesting a potential link between cytogenetic markers and the phenotype. Although the derivative chromosome der(1;7), observed in 9 patients, is responsible for a loss of 7q, it seemed different from patients with -7/del7q [excluding der(1;7)]. In the -7/del7q group, AML/MDS patients were more numerous than MF patients and the overall survival was shorter compared with the der(1;7) group (22/22 (100%) vs 6/9 (67%) AML/MDS, p=0.02; median: 4 vs 41 months, p=0.0007 respectively). Some specific associations could be observed, such as 17p deletions with 5q deletion (12/30, 40% vs 4/48, 8%, p=0.0007) and 20q deletion with der(1;7) (4/9 (44%) vs11/69 (16%), p=0.03). We detected 24/40 (60%) JAK2V617F and 8/25 (32%) TET2 mutations in transformed MPN, with all possible combinations between the wildtype and mutated forms of both genes. For one post-ET AML patient, JAK2V617F had been observed in a fraction of the granulocytes at the chronic phase. Analysis of blood cDNA obtained at chronic phase showed the same TET2 mutation as observed at acute phase. Because the blast cells were JAK2wt-TET2mut and carried a t(10;16)(q22;q23) affecting the CBFB gene, it is likely that the resulting non-MYH11 CBFB fusion gene transformed a JAK2wt-TET2 mutated progenitor that predominated in the chronic phase. In conclusion, no specific chromosomal abnormality was associated with TET2 or JAK2 mutations. Chromosomal abnormalities were associated with a type of transformation (AML/MDS or MF), suggesting a specific role in the process. In addition, association between some chromosomal abnormalities suggest a specific oncogenic cooperation.Table 1.n=82AML/MDS n=66 MF n=16 p univariate p multivariate Sex F39 (59%)5 (31%)nsnsPV/ET/PMF30/26/1013/3/0nsnsAge at diagnosis of MPN54 [20-82]55.5 [31-69]nsnsChronic Phase (duration, years)12 [2-34]14.5 [3-28]nsnsPrior treatments (n=73*)57*16..No treatment (n=6)60nsnsOne treatment (n=40)33 (58%)7 (44%)nsnsTreatments with Hu (n=57)48 (73%)9 (56%)0.03Treatments with Pi (n=41)26 (46%)15 (93%)0.00060.05Age at transformation66.5[37-92]68 [45-80]nsnsAbnormal karyotype62 (94%)16 (100%)nsnsComplex karyotype45 (68%)7 (44%)nsns-7/del7q28 (42%)3 (18%)0.07ns-7/del7q[without der(1;7)]22 (33%)00.0040.04-5/del5q28 (42%)2 (12%)0.03ns-13/del13q5 (8%)3 (19%)nsns-20/del20q11 (17%)4 (25%)nsns-17/del17p15 (23%)1(6%)nsns+1q14 (22%)9 (56%)0.01ns+95 (8%)4 (25%)0.04ns+811 (17%)3 (19%)nsnsdic17 (26%)3 (19%)nsnsder(1;7)6 (9%)3 (19%)nsnsAmplification MLL0 (0%)nsnsJAK2mut17/31 (55%)7/9 (78%)nsnsTET2mut6/19 (32%)2/6 (33%)nsnsMedian overall survival (months)448

Description: Abstract 318 Aberrant junctions involving the switch regions of the IGH locus are frequent in the non-GCB subtype of diffuse large B cell lymphomas (DLBCL), suggesting that some deregulation of the class switch recombination process (CSR) participates in the acquisition of genomic abnormalities in this pathology. To address the impact of this defect on the B cell receptor (BCR) itself we characterized the variable and constant regions of the immunoglobulin heavy chain by RT-PCR in 51 cases. Our results show that, in contrast with most GCB cases, almost all non-GCB DLBCL express an IgM (25/28 vs. 9/23; in non-GCB and GCB cases, respectively, P=0.0002; Fisher's exact test). We addressed the molecular bases of this IgM restriction by studying the configuration of the IGH loci. We found frequent legitimate and illegitimate switch junctions in IgM+ DLBCLs by genomic PCR (in 25/34 cases), showing that CSR is activated during lymphomagenesis but does not modify the isotype of the BCR. Interestingly, fluorescence in situ experiments using a probe directed against the Cμ gene showed that class switch recombination (CSR) occurs symmetrically on the two IGH alleles in IgG+ cases (5/5 tested cases) and only asymmetrically in a majority of IgM+ cases (8/11 tested cases). This finding indicates that the expression of an IgM does not result from a global failure of the CSR machinery but instead from a specific lack of recombination on the productive IGH allele. In line with this hypothesis we observed that the large internal deletions in the switch μ region (Sμ) which are known to be frequent in this pathology are specific to IgM+ cases (15/34 IgM+ vs. 0/15 IgG+ cases, P=0.0018; Fisher's exact test) and cluster on the productive IGH alleles (7/7 cases characterized by allele specific PCR), suggesting that the loss of the Sμ repeats plays a direct role in IgM stabilization by reducing the probability that the Sμ region could be involved in CSR. Finally, we observed that the non-GCB signature is associated with a higher level of somatic mutations in the 5'Sμ region (median mutation rates 4.51% vs. 2.07% in non-GCB and GCB cases, P=0.034, Mann-Whitney Test) with a strong bias toward C〉T and G〉A transitions suggesting repeated AID activations. We found the same bias it the BCL6 and PAX5 genes but not in VDJ exons, suggesting that the aberrant mutations in multiple oncogenes in this pathology result specifically from repeated CSR attempts rather than from VDJ hypermutation. Together, our data show that, in non-GCB DLBCL, lymphomagenesis occurs in IgM+ B cells which are repeatedly induced to switch but which fail to modify the isotype of their BCR. We hypothesize that the stabilization of this specific isotype is an important contributor to pathogenesis which allows repeated CSR activations in germinal centers and the accumulation of genomic abnormalities. Disclosures: No relevant conflicts of interest to declare.