Characterization of IGH locus breakpoints in multiple myeloma indicates a subset of translocations appear to occur in pregerminal center B cells.

Translocations in myeloma are thought to occur solely in mature B cells in the germinal center through class switch recombination (CSR). We used a targeted captured technique followed by massively parallel sequencing to determine the exact breakpoints in both the immunoglobulin heavy chain (IGH) locus and the partner chromosome in 61 presentation multiple myeloma samples. The majority of samples (62%) have a breakpoint within the switch regions upstream of the IGH constant genes and are generated through CSR in a mature B cell. However, the proportion of CSR translocations is not consistent between cytogenetic subgroups. We find that 100% of t(4;14) are CSR-mediated; however, 21% of t(11;14) and 25% of t(14;20) are generated through DH-JH recombination activation gene-mediated mechanisms, indicating they occur earlier in B-cell development at the pro-B-cell stage in the bone marrow. These 2 groups also generate translocations through receptor revision, as determined by the breakpoints and mutation status of the segments used in 10% and 50% of t(11;14) and t(14;20) samples, respectively. The study indicates that in a significant number of cases the translocation-based etiological events underlying myeloma may arise at the pro-B-cell hematological progenitor cell level, much earlier in B-cell development than was previously thought.

Two new 3' PML breakpoints in t(15;17)(q22;q21)-positive acute promyelocytic leukemia.

In the present article, two new types of PML/RARA junctions are described. Both were identified in diagnostic samples from two t(15;17)(q22;q21)-positive acute promyelocytic leukemia (APL) patients who failed to achieve complete remission. By using different sets of primers, reverse transcriptase polymerase chain reaction (RT-PCR) of PML/RARA junctions showed atypical larger bands compared with those generated from the three classical PML breakpoints already described. Sequence analysis of the fusion region of the amplified cDNAs allowed us to determine the specificity of these fragments in both patients. This analysis showed two new hybrid transcripts that were 53 and 306 base pairs (bp) longer than that expressed by the NB4 cell line (PML breakpoint within intron 6), and are the result of the direct joining of RARA exon 3 with PML exon 7a (patient 2) or the 5' portion of PML exon 7b (patient 1), respectively. In patient 1, RT-PCR analysis of the reciprocal RARA/PML junction showed a smaller transcript than that expected in bcr1 cases, while in patient 2 no amplified fragment was obtained. Cytogenetic analysis and/or fluorescence in situ hybridization (FISH) showed that both patients had the t(15;17) translocation. The clinical and hematological profiles expressed by the two patients carrying these unexpected types of PML/RARA rearrangement did not differ significantly from that commonly seen in other APLs with the exception of the poor outcome. Genes Chromosomes Cancer 27:35-43, 2000.

A global expression-based analysis of the consequences of the t(4;14) translocation in myeloma

Purpose: Our purpose in this report was to define genes and pathways dysregulated as a consequence of the t(4;14) in myeloma, and to gain insight into the downstream functional effects that may explain the different prognosis of this subgroup.Experimental Design: Fibroblast growth factor receptor 3 (FGFR3) overexpression, the presence of immunoglobulin heavy chain-multiple myeloma SET domain (IgH-MMSET) fusion products and the identification of t(4;14) breakpoints were determined in a series of myeloma cases. Differentially expressed genes were identified between cases with (n = 55) and without (n = 24) a t(4;14) by using global gene expression analysis.Results: Cases with a t(4;14) have a distinct expression pattern compared with other cases of myeloma. A total of 127 genes were identified as being differentially expressed including MMSET and cyclin D2, which have been previously reported as being associated with this translocation. Other important functional classes of genes include cell signaling, apoptosis and related genes, oncogenes, chromatin structure, and DNA repair genes. Interestingly, 25% of myeloma cases lacking evidence of this translocation had up-regulation of the MMSET transcript to the same level as cases with a translocation.Conclusions: t(4;14) cases form a distinct subgroup of myeloma cases with a unique gene signature that may account for their poor prognosis. A number of non-t(4;14) cases also express MMSET consistent with this gene playing a role in myeloma pathogenesis.

Antimicrobial susceptibility of Pseudomonas aeruginosa isolated from cystic fibrosis patients through Northern Europe

Pseudomonas aeruginosa is a major cause of morbidity and mortality in cystic fibrosis patients. This study compares the antimicrobial susceptibility of 153 P. aeruginosa isolates from the United Kingdom (UK) (n=58), Belgium (n=44), and Germany (n=51) collected from 120 patients during routine visits over the 2006-2012 period. MICs were measured by broth microdilution. Genes encoding extended spectrum β-lactamases (ESBL), metallo-β-lactamases and carbapenemases were detected by PCR. Pulsed Field Gel Electrophoresis and Multi-Locus Sequence Typing were performed on isolates resistant to ≥ 3 antibiotic classes among penicillins/cephalosporins, carbapenems, fluoroquinolones, aminoglycosides, polymyxins. Based on EUCAST/CLSI breakpoints, susceptibility was ≤ 30%/≤ 40% (penicillins, ceftazidime, amikacin, ciprofloxacin), 44-48%/48-63% (carbapenems), 72%/72% (tobramycin), and 92%/78% (colistin) independently of patient's age. Sixty percent of strains were multidrug resistant (MDR; European Centre for Disease prevention and Control criteria). Genes encoding ESBL (most prevalent BEL, PER, GES, VEB, CTX-M, TEM, SHV, and OXA), metallo β-lactamases (VIM, IMP, NDM), or carbapenemases (OXA-48, KPC) were not detected. The Liverpool Epidemic Strain (LES) was prevalent in UK isolates only (75% of MDR isolates). Four MDR ST958 isolates were found spread over the three countries. The other MDR clones were evidenced in ≤ 3 isolates and localized in a single country. A new sequence type (ST2254) was discovered in one MDR isolate in Germany. Clonal and non-clonal isolates with different susceptibility profiles were found in 21 patients. Thus, resistance and MDR are highly prevalent in routine isolates from 3 countries, with carbapenem (meropenem), tobramycin and colistin remaining the most active drugs.