Cryptic aberrations may allow more accurate prognostic classification of patients with myelodysplastic syndromes and clonal evolution.

The karyotype of bone-marrow cells at the time of diagnosis is one of the most important prognostic factors in patients with myelodysplastic syndromes (MDS). In some cases, the acquisition of additional genetic aberrations (clonal evolution [CE]) associated with clinical progression may occur during the disease. We analyzed a cohort of 469 MDS patients using a combination of molecular cytogenomic methods to identify cryptic aberrations and to assess their potential role in CE. We confirmed CE in 36 (8%) patients. The analysis of bone-marrow samples with a combination of cytogenomic methods at diagnosis and after CE identified 214 chromosomal aberrations. The early genetic changes in the diagnostic samples were frequently MDS specific (17 MDS-specific/57 early changes). Most progression-related aberrations identified after CE were not MDS specific (131 non-MDS-specific/155 progression-related changes). Copy number neutral loss of heterozygosity (CN-LOH) was detected in 19% of patients. MDS-specific CN-LOH (4q, 17p) was identified in three patients, and probably pathogenic homozygous mutations were found in TET2 (4q24) and TP53 (17p13.1) genes. We observed a statistically significant difference in overall survival (OS) between the groups of patients divided according to their diagnostic cytogenomic findings, with worse OS in the group with complex karyotypes (P = .021). A combination of cytogenomic methods allowed us to detect many cryptic genomic changes and identify genes and genomic regions that may represent therapeutic targets in patients with progressive MDS.

Mantle cell lymphoma-variant Richter syndrome: Detailed molecular-cytogenetic and backtracking analysis reveals slow evolution of a pre-MCL clone in parallel with CLL over several years.

Richter syndrome represents the transformation of the chronic lymphocytic leukemia (CLL) into an aggressive lymphoma, most frequently the diffuse large B-cell lymphoma (DLBCL). In this report we describe a patient with CLL, who developed a clonally-related pleomorphic highly-aggressive mantle cell lymphoma (MCL) after five cycles of a fludarabine-based second-line therapy for the first relapse of CLL. Molecular cytogenetic methods together with whole-exome sequencing revealed numerous gene alterations restricted to the MCL clone (apart from the canonical t(11;14)(q13;q32) translocation) including gain of one copy of ATM gene or emergence of TP53, CREBBP, NUP214, FUBP1 and SF3B1 gene mutations. Similarly, gene expression analysis revealed vast differences between the MCL and CLL transcriptome, including overexpression of cyclin D1, downregulation of cyclins D2 and D3, or downregulation of IL4R in the MCL clone. Backtracking analysis using quantitative PCR specifically detecting an MCL-restricted focal deletion of TP53 revealed that the pre-MCL clone appeared in the bone marrow and peripheral blood of the patient approximately 4 years before the clinical manifestation of MCL. Both molecular cytogenetic and sequencing data support the hypothesis of a slow development of the pre-MCL clone in parallel to CLL over several years, and thereby exclude the possibility that the transformation event occurred at the stage of the CLL relapse clone by mere t(11;14)(q13;q32) acquisition.

Primary and recurrent diffuse astrocytomas: genomic profile comparison reveals acquisition of biologically relevant aberrations.

BACKGROUND: Diffuse astrocytomas are characterized by their highly variable biological behavior. The possibility that tumors develop novel aberrations, with relevant biological properties, is often neglected. In this study, we present two cases of diffuse astrocytoma in which additional cytogenetic and epigenetic markers with potential influence on cell proliferation or differentiation were detected at relapse. FINDINGS: The biopsies taken from the primary and recurrent tumors of two patients were analyzed with molecular methods to detect copy number variations (CNVs), gene mutations and epigenetic changes. Both cases were characterized by the R132H mutation in the isocitrate dehydrogenase 1 (IDH1) gene. Features typical of astrocytomas, such as copy-neutral loss of heterozygosity at 17p and the deletion of the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene, were also detected in both cases. These markers were present in the primary and recurrent lesions. Other aberrations, predominantly deletions or amplifications of chromosomal segments and the hypermethylation of gene promoters, were detected in the recurrent lesions. CONCLUSIONS: The IDH1 mutation was the primary event, as previously reported. According to our observations, the methylation of promoters constituted later events, which may have further disrupted cell proliferation and/or differentiation, together with additional CNVs.

Copy number neutral loss of heterozygosity at 17p and homozygous mutations of TP53 are associated with complex chromosomal aberrations in patients newly diagnosed with myelodysplastic syndromes.

Complex karyotypes are seen in approximately 20% of patients with myelodysplastic syndromes (MDS) and are associated with a high risk of transformation to acute myeloid leukemia and poor outcomes in patients. Copy number neutral loss of heterozygosity (CN-LOH, i.e., both copies of a chromosomal pair or their parts originate from one parent) might contribute to increased genomic instability in the bone-marrow cells of patients with MDS. The pathological potential of CN-LOH, which arises as a clonal aberration in a proportion of somatic cells, consists of tumor suppressor gene and oncogene homozygous mutations. The aim of our study was to evaluate the frequency of CN-LOH at 17p in bone-marrow cells of newly diagnosed MDS patients with complex chromosomal aberrations and to assess its correlation with mutations in the TP53 gene (17p13.1). CN-LOH was detected in 40 chromosomal regions in 21 (29%) of 72 patients analyzed. The changes in 27 of the 40 regions identified were sporadic. The most common finding was CN-LOH of the short arm of chromosome 17, which was detected in 13 (18%) of 72 patients. A mutational analysis confirmed the homozygous mutation of TP53 in all CN-LOH 17p patients, among which two frameshift mutations are not registered in the International Agency for Research on Cancer TP53 Database. CN-LOH 17p correlated with aggressive disease (median overall survival 4 months) and was strongly associated with a complex karyotype in the cohort studied, which might cause rapid disease progression in high-risk MDS. No other CN-LOH region previously recorded in MDS or AML patients (1p, 4q, 7q, 11q, 13q, 19q, 21q) was detected in our cohort of patients with complex karyotype examined at the diagnosis of MDS. The LOH region appeared to be balanced (i.e., with no DNA copy number change) when examined with conventional and molecular cytogenetic methods. Therefore, a microarray that detects single-nucleotide polymorphisms is an ideal method with which to identify and further characterize CN-LOH. Our data should specify the prognosis and should lead to the identification of potential targets for therapeutic interventions.

Genetic and epigenetic characterization of low-grade gliomas reveals frequent methylation of the MLH3 gene.

Diffuse astrocytomas and oligodendrogliomas (WHO grade II) are the most common histological subtypes of low-grade gliomas (LGGs). Several molecular and epigenetic markers have been identified that predict tumor progression. Our aim was in detail to investigate the genetic and epigenetic background of LGGs and to identify new markers that might play a role in tumor behavior. Twenty-three patients with oligodendroglioma or oligoastrocytoma (LGO) and 22 patients with diffuse astrocytoma (LGA) were investigated using several molecular-cytogenetic and molecular methods to assess their copy number variations, mutational status and level of promoter methylation. The most frequent findings were a 1p/19q codeletion in 83% of LGO and copy-neutral loss of heterozygosity (CN-LOH) of 17p in 72% of LGA. Somatic mutations in the isocitrate dehydrogenase 1 or 2 (IDH1/IDH2) genes were detected in 96% of LGO and 91% of LGA. The O-6-methylguanine-DNA-methyltransferase (MGMT) promoter was methylated in 83% of LGO and 59% of LGA. MutL homolog 3 (MLH3) promoter methylation was observed in 61% of LGO and 27% of LGA. Methylation of the MGMT promoter, 1p/19q codeletion, mutated IDH1, and CN-LOH of 17p were the most frequent genetic aberrations in LGGs. The findings were more diverse in LGA than in LGO. To the best of our knowledge, this is the first time description of methylation of the MLH3 gene promoter in LGGs. Further studies are required to determine the role of the methylated MLH3 promoter and the other aberrations detected.

An unusual case of high hyperdiploid childhood ALL with cryptic BCR/ABL1 rearrangement.

BACKGROUND: Both high hyperdiploidy (HeH) and the translocation t(9;22)(q34;q11) are recurrent abnormalities in childhood B-cell acute lymphoblastic leukemia (ALL) and both are used in current classification to define different genetic and prognostic subtypes of the disease. The coexistence of these two primary genetic aberrations within the same clone is very rare in children with ALL. Here we report a new case of a 17-year-old girl with newly diagnosed ALL and uncommon cytogenetic and clinical finding combining high hyperdiploidy and a cryptic BCR/ABL1 fusion and an inherited Charcot-Marie-Tooth neuropathy detected during the induction treatment. RESULTS: High hyperdiploid karyotype 51,XX,+X,+4,+14,+17,+21 without apparent structural aberrations was detected by conventional cytogenetic analysis and multicolor FISH. A cryptic BCR/ABL1 fusion, which was caused by the insertion of part of the ABL1 gene into the 22q11 region, was proved in HeH clone by FISH, RT-PCR and CGH-SNP array. In addition, an abnormal FISH pattern previously described as the deletion of the 3'BCR region in some BCR/ABL1 positive cases was not proved in our patient. CONCLUSION: A novel case of extremely rare childhood ALL, characterized by HeH and a cryptic BCR/ABL1 fusion, is presented and to the best of our knowledge described for the first time. The insertion of ABL1 into the BCR region in malignant cells is supposed. Clearly, further studies are needed to determine the genetic consequences and prognostic implications of these unusual cases.

Change in ploidy status from hyperdiploid to near-tetraploid in multiple myeloma associated with bortezomib/lenalidomide resistance.

Ploidy is an important prognostic factor in the risk stratification of multiple myeloma (MM) patients. Patients with MM can be divided into two groups according to the modal number of chromosomes: nonhyperdiploid (NH-MM) and hyperdiploid (H-MM), which has a more favorable outcome. The two ploidy groups represent two different oncogenetic pathways determined at the premalignant stage. The ploidy subtype also persists during the course of the disease, even during progression after the therapy, with only very rare cases of ploidy conversion. The clinical significance of ploidy conversion and its relation to drug resistance have been previously discussed. Here, we describe a female MM patient with a rare change in her ploidy status from H-MM to NH-MM, detected by cytogenetic and molecular cytogenetic examinations of consecutive bone marrow aspirates. We hypothesize that ploidy conversion (from H-MM to NH-MM) is associated with disease progression and acquired resistance to bortezomib/lenalidomide therapy.

Jumping-like translocation-a rare chromosomal rearrangement in a patient with Burkitt lymphoma/leukemia.

Chromosomal translocations are acquired genetic rearrangements in human cancers. Jumping translocations are rare nonreciprocal rearrangements involving the same donor chromosome segment translocated to two or more recipient chromosomes. In this report, we describe a patient with Burkitt lymphoma/leukemia (BL) and a complex karyotype including a t(2;8)(p12;q24), copy-neutral loss of heterozygosity at 17p13.1-p13.3 and 19q13.1-q13.2, trisomy 20, and two uncommon chromosomal aberrations. The first uncommon aberration was a complex rearrangement of chromosome 15 (probably the consequence of chromothripsis) masked by an apparently balanced reciprocal translocation, t(11;15)(p11.2;q21). The second one was a special type of unbalanced "vice versa" jumping translocation, which involved the same acceptor chromosome arm (13q) and various donor chromosome segments. It is unclear whether both atypical rearrangements are the consequence of the TP53 alteration or whether assumed chromothripsis influenced the development of the jumping-like translocation. However, the presence of the t(11;15)(p11.2;q21) in all pathological cells suggests that it occurred in the early stage of the disease, whereas the jumping-like translocation, as an additional change, subsequently accelerated the progression of the disease.