Complex chromosomal rearrangements (CCRs) are often observed in clinical samples from patients with cancer and congenital diseases but are difficult to induce experimentally. Here, we report the first success in establishing animal models for CCRs. Mutation in Recql5, a crucial member of the DNA helicase RecQ family involved in DNA replication, transcription, and repair, enabled CRISPR/Cas9-mediated CCRs, establishing a mouse model containing triple fusion genes and megabase-sized inversions. Some of these structural features of individual chromosomal rearrangements use template switching and microhomology-mediated break-induced replication mechanisms and are reminiscent of the newly described phenomenon \"chromoanasynthesis.\" These data show that Recql5 mutant mice could be a powerful tool to analyze the pathogenesis of CCRs (particularly chromoanasynthesis) whose underlying mechanisms are poorly understood. The Recql5 mutants generated in this study are to be deposited at key animal research facilities, thereby making them accessible for future research on CCRs.

Osteosarcoma (OS) is the most prevalent type of bone tumor, but slow progress has been achieved in disentangling the full set of genomic events involved in its initiation and progression. We assessed by NGS the mutational spectrum of 28 primary OSs from Brazilian patients, and identified 445 potentially deleterious SNVs/indels and 1176 copy number alterations (CNAs). TP53 was the most recurrently mutated gene, with an overall rate of ~60%, considering SNVs/indels and CNAs. The most frequent CNAs (~60%) were gains at 1q21.2q21.3, 6p21.1, and 8q13.3q24.22, and losses at 10q26 and 13q14.3q21.1. Seven cases presented CNA patterns reminiscent of complex events (chromothripsis and chromoanasynthesis). Putative RB1 and TP53 germline variants were found in five samples associated with metastasis at diagnosis along with complex genomic patterns of CNAs. PTPRQ, KNL1, ZFHX4, and DMD alterations were prevalent in metastatic or deceased patients, being potentially indicative of poor prognosis. TNFRSF11B, involved in skeletal system development and maintenance, emerged as a candidate for osteosarcomagenesis due to its biological function and a high frequency of copy number gains. A protein-protein network enrichment highlighted biological pathways involved in immunity and bone development. Our findings reinforced the high genomic OS instability and heterogeneity, and led to the identification of novel disrupted genes deserving further evaluation as biomarkers due to their association with poor outcomes.

The use of new technologies in the routine diagnosis of constitutional abnormalities, such as high-resolution chromosomal microarray and next-generation sequencing, has unmasked new mechanisms for generating structural variation of the human genome. For example, complex chromosome rearrangements can originate by a chromosome catastrophe phenomenon in which numerous genomic rearrangements are apparently acquired in a single catastrophic event. This phenomenon is named chromoanagenesis (from the Greek \"chromo\" for chromosome and \"anagenesis\" for rebirth). Herein, we report 2 cases of genomic chaos detected at prenatal diagnosis. The terms \"chromothripsis\" and \"chromoanasynthesis\" and the challenge of genetic counseling are discussed.

Cancer genomes evolve in a punctuated manner during tumor evolution. Abrupt genome restructuring at key steps in this evolution has been called \"genome chaos.\" To answer whether widespread genome change is truly chaotic, this review (i) summarizes the limited number of cell and molecular systems that execute genome restructuring, (ii) describes the characteristic signatures of DNA changes that result from activity of those systems, and (iii) examines two cases where genome restructuring is determined to a significant degree by cell type or viral infection. The conclusion is that many restructured cancer genomes display sufficiently unchaotic signatures to identify the cellular systems responsible for major oncogenic transitions, thereby identifying possible targets for therapies to inhibit tumor progression to greater aggressiveness.

Chromoanagenesis, a phenomenon characterized by complex chromosomal rearrangement and reorganization events localized to a limited number of genomic regions, includes the subcategories chromothripsis, chromoanasynthesis, and chromoplexy. Although definitions of these terms are evolving, constitutional chromoanagenesis events have been reported in a limited number of patients with variable phenotypes. We report on 2 cases with complex genomic events characterized by multiple copy number gains and losses confined to a single chromosome region, which are suggestive of constitutional chromoanagenesis. Case 1 is a 43-year-old male with intellectual disability and recently developed generalized tonic-clonic seizures. Chromosomal microarray analysis identified a complex rearrangement involving chromosome region 14q31.1q32.2, consisting of 16 breakpoints ranging in size from 0.2 to 6.2 Mb, with 5 segments of normal copy number present between these alterations. Interestingly, this case represents the oldest known patient with a complex rearrangement indicative of constitutional chromoanagenesis. Case 2 is a 2-year-old female with developmental delay, speech delay, low muscle tone, and seizures. Chromosomal microarray analysis identified a complex rearrangement consisting of 28 breakpoints localized to 18q21.32q23. The size of the copy number alterations ranged from 0.042 to 5.1 Mb, flanked by 12 small segments of normal copy number. These cases add to a growing body of literature demonstrating complex chromosomal rearrangements as a disease mechanism for congenital anomalies.

We present the genetic profile of kidney giant leiomyosarcoma characterized by sequencing of 409 cancer related genes and chromosomal microarray analysis. Renal leiomyosarcomas are extremely rare neoplasms with aggressive behavior and poor survival prognosis. Most frequent somatic events in leiomyosarcomas are mutations in the TP53, RB1, ATRX, and PTEN genes, chromosomal instability (CIN) and chromoanagenesis. 67-year-old woman presented with a right kidney completely replaced by tumor. Immunohistochemical reaction on surgical material was positive to desmin and smooth muscle actin. Molecular genetic analysis revealed that tumor harbored monosomy of chromosomes 3 and 11, gain of Xp (ATRX) arm and three chromoanasynthesis regions (6q21-q27, 7p22.3-p12.1, and 12q13.11-q21.2), with MDM2 and CDK4 oncogenes copy number gains, whereas no copy number variations (CNVs) or tumor specific single nucleotide variants (SNVs) in TP53, RB1, and PTEN genes were present. We hypothesize that chromoanasynthesis in 12q13.11-q21.2 could be a trigger of observed CIN in this tumor.

Micronuclei, small spatially-separated, nucleus-like structures, are a common feature of human cancer cells. There are considerable heterogeneities in the sources, structures and genetic activities of micronuclei. Accumulating evidence suggests that micronuclei and main nuclei represent separate entities with respect to DNA replication, DNA damage sensing and repairing capacity because micronuclei are not monitored by the same checkpoints nor covered by the same nuclear envelope as the main nuclei. Thus, micronuclei are spatially restricted \"mutation factories.\" Several large-scale DNA sequencing and bioinformatics studies over the last few years have revealed that most micronuclei display a mutational signature of chromothripsis immediately after their generation and the underlying molecular mechanisms have been dissected extensively. Clonal expansion of the micronucleated cells is context-dependent and is associated with chromothripsis and several other mutational signatures including extrachromosomal circular DNA, kataegis and chromoanasynthesis. These results suggest what was once thought to be merely a passive indicator of chromosomal instability is now being recognized as a strong mutator phenotype that may drive intratumoral genetic heterogeneity. Herein, we revisit the actionable determinants that contribute to the bursts of mutagenesis in micronuclei and present the growing number of evidence which suggests that micronuclei have distinct short- and long-term mutational and functional effects to cancer genomes. We also pose challenges for studying the long-term effects of micronucleation in the upcoming years.

We report a case of a de novo ring 21 complex chromosomal rearrangement in a fetus presenting with hydrops. Noninvasive prenatal testing (NIPT) failed to detect the imbalance. This case highlights the need to understand the various limitations and strengths of NIPT technology when counseling patients.

UNASSIGNED: Chromoanagenesis events encompassing chromoanasynthesis, chromoplexy, and chromothripsis are described in cancers and can result in highly complex chromosomal rearrangements derived from \'all-at-once\' catastrophic cellular events. The complexity of these rearrangements and the original descriptions in cancer cells initially led to the assumption that it was an acquired anomaly. While rare, these phenomena involving chromosome 1 have been reported a few individuals in a constitutional setting.
UNASSIGNED: Here, we describe a newborn baby who was initially referred for cytogenetic testing for multiple congenital anomalies including cystic encephalomalacia, patent ductus arteriosus, inguinal hernia, and bilateral undescended testicles. Chromosome analysis was performed and revealed a derivative chromosome 1 with an 1q24-q31 segment inserted into 1q42.13 resulting in gain of 1q24-q31. Whole genome SNP microarray analysis showed a complex pattern of copy number variants with four gains and one loss involving 1q24-q31. Mate pair next-generation sequencing analysis revealed 18 chromosome breakpoints, six gains along an 1q24-q31 segment, one deletion of 1q31.3 segment and one deletion of 1q42.13 segment, which is strongly evocative of a chromoanasynthesis event for developing this complex rearrangement. Parental chromosome analyses were performed and showed the same derivative chromosome 1 in the mother.
UNASSIGNED: To our knowledge, our case is the first case with familial constitutional chromoanagenesis involving chromosome 1q24-q42. This report emphasizes the value of performing microarray and mate pair next-generation sequencing analysis for individuals with germline abnormal or complex chromosome rearrangements.

UNASSIGNED: During the last decade, genome sequencing projects in cancer genomes as well as in patients with congenital diseases and healthy individuals have led to the identification of new types of massive chromosomal rearrangements arising during single chaotic cellular events. These unanticipated catastrophic phenomenon are termed chromothripsis, chromoanasynthesis and chromoplexis., and are grouped under the name of \"chromoanagenesis\".
UNASSIGNED: For each process, several specific features have been described, allowing each phenomenon to be distinguished from each other and to understand its mechanism of formation and to better understand its aetiology. Thus, chromothripsis derives from chromosome shattering followed by the random restitching of chromosomal fragments with low copy-number change whereas chromoanasynthesis results from erroneous DNA replication of a chromosome through serial fork stalling and template switching with variable copy-number gains, and chromoplexy refers to the occurrence of multiple inter-and intra-chromosomal translocations and deletions with little or no copy-number alterations in prostate cancer. Cumulating data and experimental models have shown that chromothripsis and chromoanasynthesis may essentially result from lagging chromosome encapsulated in micronuclei or telomere attrition and end-to-end telomere fusion.
UNASSIGNED: The concept of chromanagenesis has provided new insight into the aetiology of complex structural rearrangements, the connection between defective cell cycle progression and genomic instability, and the complexity of cancer evolution. Increasing reported chromoanagenesis events suggest that these chaotic mechanisms are probably much more frequent than anticipated.