Numerical Chromosomal Instability Research Articles

Chromosomal Damage, Chromosome Instability, and Polymorphisms in GSTP1 and XRCC1 as Biomarkers of Effect and Susceptibility in Farmers Exposed to Pesticides.

In the department of Boyacá, Colombia, agriculture stands as one of the primary economic activities. However, the escalating utilization of pesticides within this sector has sparked concern regarding its potential correlation with elevated risks of genotoxicity, chromosomal alterations, and carcinogenesis. Furthermore, pesticides have been associated with a broad spectrum of genetic polymorphisms that impact pivotal genes involved in pesticide metabolism and DNA repair, among other processes. Nonetheless, our understanding of the genotoxic effects of pesticides on the chromosomes (as biomarkers of effect) in exposed farmers and the impact of genetic polymorphisms (as susceptibility biomarkers) on the increased risk of chromosomal damage is still limited. The aim of our study was to evaluate chromosomal alterations, chromosomal instability, and clonal heterogeneity, as well as the presence of polymorphic variants in the GSTP1 and XRCC1 genes, in peripheral blood samples of farmers occupationally exposed to pesticides in Aquitania, Colombia, and in an unexposed control group. Our results showed statistically significant differences in the frequency of numerical chromosomal alterations, chromosomal instability, and clonal heterogeneity levels between the exposed and unexposed groups. In addition, we also found a higher frequency of chromosomal instability and clonal heterogeneity in exposed individuals carrying the heterozygous GSTP1 AG and XRCC1 (exon 10) GA genotypes. The evaluation of chromosomal alterations and chromosomal instability resulting from pesticide exposure, combined with the identification of polymorphic variants in the GSTP1 and XRCC1 genes, and further research involving a larger group of individuals exposed to pesticides could enable the identification of effect and susceptibility biomarkers. Such markers could prove valuable for monitoring individuals occupationally exposed to pesticides.

A whale of a tale: whale cells evade the driving mechanism for hexavalent chromium-induced chromosome instability.

Chromosome instability, a hallmark of lung cancer, is a driving mechanism for hexavalent chromium [Cr(VI)] carcinogenesis in humans. Cr(VI) induces structural and numerical chromosome instability in human lung cells by inducing DNA double-strand breaks and inhibiting homologous recombination repair and causing spindle assembly checkpoint (SAC) bypass and centrosome amplification. Great whales are long-lived species with long-term exposures to Cr(VI) and accumulate Cr in their tissue, but exhibit a low incidence of cancer. Data show Cr(VI) induces fewer chromosome aberrations in whale cells after acute Cr(VI) exposure suggesting whale cells can evade Cr(VI)-induced chromosome instability. However, it is unknown if whales can evade Cr(VI)-induced chromosome instability. Thus, we tested the hypothesis that whale cells resist Cr(VI)-induced loss of homologous recombination repair activity and increased SAC bypass and centrosome amplification. We found Cr(VI) induces similar amounts of DNA double-strand breaks after acute (24 h) and prolonged (120 h) exposures in whale lung cells, but does not inhibit homologous recombination repair, SAC bypass, or centrosome amplification, and does not induce chromosome instability. These data indicate whale lung cells resist Cr(VI)-induced chromosome instability, the major driver for Cr(VI) carcinogenesis at a cellular level, consistent with observations that whales are resistant to cancer.

Profiling Numerical and Structural Chromosomal Instability in Different Cancer Types.

Many cancers display whole chromosome instability (W-CIN) and structural chromosomal instability (S-CIN), referring to increased rates of acquiring numerically and structurally abnormal chromosome changes. This protocol provides detailed steps to analyze the W-CIN and S-CIN across cancer types, intending to leverage large-scale bulk sequencing and SNP array data complemented with the computational models to gain a better understanding of W-CIN and S-CIN.

Hexavalent Chromium Targets Securin to Drive Numerical Chromosome Instability in Human Lung Cells.

Hexavalent chromium [Cr(VI)] is a known human lung carcinogen with widespread exposure in environmental and occupational settings. Despite well-known cancer risks, the molecular mechanisms of Cr(VI)-induced carcinogenesis are not well understood, but a major driver of Cr(VI) carcinogenesis is chromosome instability. Previously, we reported Cr(VI) induced numerical chromosome instability, premature centriole disengagement, centrosome amplification, premature centromere division, and spindle assembly checkpoint bypass. A key regulator of these events is securin, which acts by regulating the cleavage ability of separase. Thus, in this study we investigated securin disruption by Cr(VI) exposure. We exposed human lung cells to a particulate Cr(VI) compound, zinc chromate, for acute (24 h) and prolonged (120 h) time points. We found prolonged Cr(VI) exposure caused marked decrease in securin levels and function. After prolonged exposure at the highest concentration, securin protein levels were decreased to 15.3% of control cells, while securin mRNA quantification was 7.9% relative to control cells. Additionally, loss of securin function led to increased separase activity manifested as enhanced cleavage of separase substrates; separase, kendrin, and SCC1. These data show securin is targeted by prolonged Cr(VI) exposure in human lung cells. Thus, a new mechanistic model for Cr(VI)-induced carcinogenesis emerges with centrosome and centromere disruption as key components of numerical chromosome instability, a key driver in Cr(VI) carcinogenesis.

Replication stress causes delayed mitotic entry and chromosome 12 fragility at the ANKS1B large neuronal gene in human induced pluripotent stem cells.

Substantial background level of replication stress is a feature of embryonic and induced pluripotent stem cells (iPSCs), which can predispose to numerical and structural chromosomal instability, including recurrent aberrations of chromosome 12. In differentiated cells, replication stress-sensitive genomic regions, including common fragile sites, are widely mapped through mitotic chromosome break induction by mild aphidicolin treatment, an inhibitor of replicative polymerases. IPSCs exhibit lower apoptotic threshold and higher repair capacity hindering fragile site mapping. Caffeine potentiates genotoxic effects and abrogates G2/M checkpoint delay induced by chemical and physical mutagens. Using 5-ethynyl-2'-deoxyuridine (EdU) for replication labeling, we characterized the mitotic entry dynamics of asynchronous iPSCs exposed to aphidicolin and/or caffeine. Under the adjusted timing of replication stress exposure accounting revealed cell cycle delay, higher metaphase chromosome breakage rate was observed in iPSCs compared to primary lymphocytes. Using differential chromosome staining and subsequent locus-specific fluorescent in situ hybridization, we mapped the FRA12L fragile site spanning the large neuronal ANKS1B gene at 12q23.1, which may contribute to recurrent chromosome 12 missegregation and rearrangements in iPSCs. Publicly available data on the ANKS1B genetic alterations and their possible functional impact are reviewed. Our study provides the first evidence of common fragile site induction in iPSCs and reveals potential somatic instability of a clinically relevant gene during early human development and in vitro cell expansion.

Chromosomal instability (CIN) in HAP1 cell lines revealed by multiplex fluorescence in situ hybridisation (M-FISH)

BackgroundHAP1, a near-haploid human leukemic cancer cell line is often used in combination with CRISPR-Cas9 gene editing technology for genetic screens. HAP1 carries the Philadelphia chromosome (Ph) and an additional ~ 30 Mb fragment of chromosome 15 inserted into chromosome 19. The potential use of an in vitro cell line as a model system in biomedical research studies depends on its ability to maintain genome stability. Being a cancer cell line with a near-haploid genome, HAP1 is prone to genetic instability, which is further compounded by its tendency to diploidise in culture spontaneously. Moreover, CRISPR-Cas9 gene editing coupled with prolonged in-vitro cell culturing has the potential to induce unintended ‘off-target’ cytogenetic mutations.To gain an insight into chromosomal instability (CIN) and karyotype heterogeneity, 19 HAP1 cell lines were cytogenetically characterised, 17 of which were near-haploids and two double-haploids, using multiplex fluorescence in situ hybridisation (M-FISH), at single cell resolution. We focused on novel numerical (N) and structural (S) CIN and discussed the potential causal factors for the observed instability. For each cell line we examined its ploidy, gene editing status and its length of in-vitro cell culturing.ResultsSixteen of the 19 cell lines had been gene edited with passage numbers ranging from 10 to 35. Diploidisation in 17 near-haploid cell lines ranged from 4 to 35% and percentage of N- and S-CIN in [1n] and [2n] metaphases ranged from 7 to 50% with two cell lines showing no CIN. Percentage of cells with CIN in the two double-haploid cell lines were 96% and 100% respectively. The most common S-CIN observed was deletion followed by translocation of both types, non-reciprocal and Robertsonian. Interestingly, we observed a prevalence of S-CIN associated with chromosome 13 in both near-and double-haploid cell lines, with a high incidence of Robertsonian translocation involving chromosome 13. Furthermore, locus-specific BAC (bacterial artificial chromosome) FISH enabled us to show for the first time that the additional chromosome 15 fragment is inserted into the p-arm rather than the q-arm of chromosome 19 of the HAP1 genome.ConclusionOur study revealed a high incidence of CIN leading to karyotype heterogeneity in majority of the HAP1 cell lines with the number of chromosomal aberrations varying between cell lines. A noteworthy observation was the high frequency of structural chromosomal aberrations associated with chromosome 13. We showed that CRISPR-Cas9 gene editing technology in combination with spontaneous diploidisation and prolonged in-vitro cell culturing is potentially instrumental in inducing further chromosomal rearrangements in the HAP1 cell lines with existing CIN. We highlight the importance of maintaining cell lines at low passage and the need for regular monitoring to prevent implications in downstream applications. Our study also established that the additional fragment of chromosome 15 in the HAP1 genome is inserted into chromosome 19p rather than 19q.

Suppression of Chromosome Instability Limits Acquired Drug Resistance.

Numerical chromosome instability, or nCIN, defined as the high frequency of whole chromosome gains and losses, is prevalent in many solid tumors. nCIN has been shown to promote intratumor heterogeneity and corresponds with tumor aggressiveness, drug resistance, and tumor relapse. Although increased nCIN has been shown to promote the acquisition of genomic changes responsible for drug resistance, the potential to modulate nCIN in a therapeutic manner has not been well explored. Here we assess the role of nCIN in the acquisition of drug resistance in non-small cell lung cancer. We show that the generation of whole chromosome segregation errors in non-small cell lung cancer cells is sensitive to manipulation of microtubule dynamics and that enhancement of chromosome cohesion strongly suppresses nCIN and reduces intratumor heterogeneity. We demonstrate that suppression of nCIN has no impact on non-small cell lung cancer cell proliferation in vitro nor in tumor initiation in mouse xenograft models. However, suppression of nCIN alters the timing and molecular mechanisms that drive acquired drug resistance. These findings suggest mechanisms to suppress nCIN may serve as effective cotherapies to limit tumor evolution and sustain drug response.

Distinct and Common Features of Numerical and Structural Chromosomal Instability across Different Cancer Types.

Simple SummaryMany cancer cells are chromosomally unstable, a phenotype describing a tendency for accumulating chromosomal aberrations. Entire chromosomes tend to be gained or lost, which is called whole chromosome instability (W-CIN). Structural chromosomal instability (S-CIN) describes an increased rate of gaining, losing or translocating smaller parts of chromosomes. Here, we analyse data from 33 cancer types to find differences and commonalities between W-CIN and S-CIN. We find that W-CIN is strongly linked to whole genome doubling (WGD), whereas S-CIN is associated with a specific DNA damage repair pathway. Both W-CIN and S-CIN are difficult to target using currently available compounds and have distinct prognostic values. The activity of the drug resistance gene CKS1B is associated with S-CIN, which merits further investigation. In addition, we identify a potential copy number-based mechanism promoting signalling of the important PI3K cancer pathway in high-S-CIN tumours.A large proportion of tumours is characterised by numerical or structural chromosomal instability (CIN), defined as an increased rate of gaining or losing whole chromosomes (W-CIN) or of accumulating structural aberrations (S-CIN). Both W-CIN and S-CIN are associated with tumourigenesis, cancer progression, treatment resistance and clinical outcome. Although W-CIN and S-CIN can co-occur, they are initiated by different molecular events. By analysing tumour genomic data from 33 cancer types, we show that the majority of tumours with high levels of W-CIN underwent whole genome doubling, whereas S-CIN levels are strongly associated with homologous recombination deficiency. Both CIN phenotypes are prognostic in several cancer types. Most drugs are less efficient in high-CIN cell lines, but we also report compounds and drugs which should be investigated as targets for W-CIN or S-CIN. By analysing associations between CIN and bio-molecular entities with pathway and gene expression levels, we complement gene signatures of CIN and report that the drug resistance gene CKS1B is strongly associated with S-CIN. Finally, we propose a potential copy number-dependent mechanism to activate the PI3K pathway in high-S-CIN tumours.

Evidence of chromosomal damage in scleroderma

Evidence of chromosomal damage in scleroderma

Abstract 595: Patterns and dynamics of genome instability drive metastatic activity in non-small cell lung cancer (NSCLC) circulating tumor cell (CTC)-derived xenograft (CDX) models

Abstract Background: CDX models have emerged as tractable systems to explore mechanisms involved in metastatic progression and tumor-initiating properties of CTCs. However, their development is challenging due to low prevalence of CTCs in patient blood. We report the genomic and functional characterization of 4 NSCLC CDX models and their derived cell lines. We focus on genome and chromosomal instability (CIN) mechanisms operating in the CDX-derived cell lines and decipher relevant therapeutic targets. Methods: CTCs were enriched from 30mL blood samples of 58 advanced NSCLC patients and implanted subcutaneously into Nod/Scid-IL2Rγ-/- (NSG) mice. Tumors were palpable within a median of 108 days. Among the 4 CDX models established, 3 CDX-derived cell lines (GR-CDXL1, GR-CDXL3 and GR-CDXL4) were obtained. Cell lines were characterized by immunofluorescence (IF), immunohistochemistry and whole-exome sequencing (WES). CIN and DNA damage response (DDR) activity were evaluated using IF and western blot. Tumorigenic potential of CDX-derived cell lines was assessed in the chick embryo chorioallantoic membrane (CAM) and NSG mice engrafted intravenously. IC50 was assessed using CellTiter-Glo®. Results: All CDX models had an epithelial phenotype. CDX and cell lines recapitulated the corresponding tumor histological features (available for 3 patients). WES revealed multiple copy number alterations (CNAs) in driver genes implicated in diverse genome instability mechanisms. GR-CDXL1 presented BRCA2 mutation and FANCA promoter deletion, while AKT gain was detected in GR-CDXL3 in addition to whole genome doubling. This led us to investigate the DDR in the cell lines. Homologous recombination deficiency and unrepaired DNA damage post-mitosis were observed in GR-CDXL1. GR-CDXL3 presented numerical CIN and centrosome clustering. GR-CDXL4 showed high levels of DNA damage and CIN. CNA and functional analysis provided a biological rationale for the selection of drug candidates in pharmacological testings on the CDX-derived cell lines. The assays mirrored patients' response to chemotherapy and revealed olaparib efficiency in GR-CDXL1 and GR-CDXL4 cells, which was validated in the CAM. GR-CDXL3 cells were sensitive to phosphoinositide 3-kinase-α inhibitor alpelisib, suggesting tumor dependence on PI3K/AKT pathway. Cell lines were tumorigenic in the CAM and mice. Notably, GR-CDXL3 seeded multiple metastases, which is concordant with its distinguished CIN characteristics and WGD. In ovo and in vivo validation of candidate therapeutic strategies is ongoing and will be presented. Conclusion: This study reveals distinct genome and CIN patterns operating in our CDX-derived cell lines that may play a critical role in CTC seeding capacity. Our CDX models offer new tools for designing therapeutic strategies targeting metastatic progression in NSCLC. Citation Format: Tala Tayoun, Vincent Faugeroux, Marianne Oulhen, Emma Pailler, Olivier Deas, Laura Mezquita, Laura Brulle-Soumare, Stephano Cairo, Jean-Yves Scoazec, Virginie Marty, Agathe Aberlenc, Maud NgoCamus, Claudio Nicotra, David Planchard, Patricia Kannouche, Benjamin Besse, Jean-Gabriel Judde, Patrycja Pawlikowska, Françoise Farace. Patterns and dynamics of genome instability drive metastatic activity in non-small cell lung cancer (NSCLC) circulating tumor cell (CTC)-derived xenograft (CDX) models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 595.

A survey of cancer genome signatures identifies genes connected to distinct chromosomal instability phenotypes.

Certain breast and ovarian cancers are characterised by high levels of chromosomal instability. We established a suite of eleven SNP array-based signatures of various forms of chromosomal instability. To understand what biological mechanisms might underpin these signatures, we generated and assembled genetic-feature data including allele-specific expression, fusion genes, gene expression, methylation, somatic coding mutations and protein expression. For each signature, we extracted a compendium of significantly associated genetic features using machine learning. We established an association between subchromosomal allelic imbalance-based measures and DNA repair genes. Numerical chromosomal instability and chromothripsis were found to have distinct genetic associations but shared a relationship to mitotic processes, while whole-genome doubling was characterised by TP53 mutation, and high AURKA and GINS1 expression. Furthermore, we identified two genetically distinct forms of tandem duplicator phenotypes. These findings identify potentially novel genomic targets for validation and drug development for specific subsets of breast and ovarian cancer.

CHROMOSOMAL ASSOCIATIONS AND GENOMIC INSTABILITY IN IDIOPATHIC RECURRENT PREGNANCY LOSS(RPL), A PILOT SCALE STUDY

The emotional issues adjoining pregnancy loss become magnified exponentially when miscarriage take place on a repetitive basis. But there are very limited studies correlated with stress in recurrent pregnancy loss (RPL). Telomeres maintain the integrity and stability of our genetic material by protecting our chromosomes. We hypothesized that stress induced background genomic instability and telomere loss plays a pivotal role in unexplained RPL. We aimed to investigate any possible link between background genomic instability, telomere loss and early unexplained recurrent pregnancy loss in a cohort of subjects along with age and sex matched controls. Participation of a cohort of 50 couples (100 individuals) experiencing with 2 or more earlier pregnancy losses, and struggling for natural conception in a age and sex matched study for elucidating any possible role of chromosomal associations and genomic instability in recurrent pregnancy loss. Blood samples were collected from each RPL couple in heparin and EDTA vials along with a consent form for participation in this study. Karyotyping was performed from heparinized blood to screen common/unique chromosomal anomalies associated with unexplained RPL. The lymphocytes were isolated from EDTA blood and subjected for isolation of DNA, RNA and preparation of whole cell protein lysate. The background genomic instability status was examined by using cytokinesis blocked micronucleus (CBMN) assay. We also checked the telomere length of the individuals by quantitative real time PCR (qRT-PCR) and Quantitative-Fluorescent In-Situ Hybridization (Q-FISH). The functional telomeric integrity of telomeric shelterin complex were evaluated through western blot analysis. Karyotyping analysis demonstrated few novel translocations responsible for RPL, which were not reported previously. In our cohort, we observed that 83 individuals out of 100 were reported karyotypically normal. This prompted us to further examine these karyotpically normal individuals for their background genomic instability. We found that 38.5% (32 out of 83) individuals have high background of genomic instability as compared with the control samples. Next, we investigated the telomere length of the patients with high genomic instability by qRT-PCR and observed that 28.125% (9 out of 32) exhibited loss of telomeres. Q-FISH (PNA-FISH) analysis of these subjects also showed loss in Telomeric signals. Furthermore, western blot analysis of some of the key players of the shelterin components showed reduced level of expression of these proteins in the RPL subjects. Apart from numerical or structural chromosomal anomalies background genomic instability leading to loss of telomeres may play a critical role in promoting RPL. This work will further lead to identification of novel biomarkers associated with unexplained recurrent pregnancy loss and it will further help in real time monitoring based on molecular analysis of the couple experiencing recurrent pregnancy failure and aid them with better possibility of conception.

DNA Fiber Assay for the Analysis of DNA Replication Progression in Human Pluripotent Stem Cells.

Human pluripotent stem cells (PSC) acquire recurrent chromosomal instabilities during prolonged in vitro culture that threaten to preclude their use in cell-based regenerative medicine. The rapid proliferation of pluripotent cells leads to constitutive replication stress, hindering the progression of DNA replication forks and in some cases leading to replication-fork collapse. Failure to overcome replication stress can result in incomplete genome duplication, which, if left to persist into the subsequent mitosis, can result in structural and numerical chromosomal instability. We have recently applied the DNA fiber assay to the study of replication stress in human PSC and found that, in comparison to somatic cells states, these cells display features of DNA replication stress that include slower replication fork speeds, evidence of stalled forks, and replication initiation from dormant replication origins. These findings have expanded on previous work demonstrating that extensive DNA damage in human PSC is replication associated. In this capacity, the DNA fiber assay has enabled the development of an advanced nucleoside-enriched culture medium that increases replication fork progression and decreases DNA damage and mitotic errors in human PSC cultures. The DNA fiber assay allows for the study of replication fork dynamics at single-molecule resolution. The assay relies on cells incorporating nucleotide analogs into nascent DNA during replication, which are then measured to monitor several replication parameters. Here we provide an optimized protocol for the fiber assay intended for use with human PSC, and describe the methods employed to analyze replication fork parameters. © 2020 Wiley Periodicals LLC. Basic Protocol 1: DNA fiber labeling Basic Protocol 2: DNA fiber spreading Basic Protocol 3: Immunostaining Support Protocol 1: Microscopy/data acquisition Support Protocol 2: Data analysis.

DNA Replication Stress and Chromosomal Instability: Dangerous Liaisons.

Chromosomal instability (CIN) is associated with many human diseases, including neurodevelopmental or neurodegenerative conditions, age-related disorders and cancer, and is a key driver for disease initiation and progression. A major source of structural chromosome instability (s-CIN) leading to structural chromosome aberrations is “replication stress”, a condition in which stalled or slowly progressing replication forks interfere with timely and error-free completion of the S phase. On the other hand, mitotic errors that result in chromosome mis-segregation are the cause of numerical chromosome instability (n-CIN) and aneuploidy. In this review, we will discuss recent evidence showing that these two forms of chromosomal instability can be mechanistically interlinked. We first summarize how replication stress causes structural and numerical CIN, focusing on mechanisms such as mitotic rescue of replication stress (MRRS) and centriole disengagement, which prevent or contribute to specific types of structural chromosome aberrations and segregation errors. We describe the main outcomes of segregation errors and how micronucleation and aneuploidy can be the key stimuli promoting inflammation, senescence, or chromothripsis. At the end, we discuss how CIN can reduce cellular fitness and may behave as an anticancer barrier in noncancerous cells or precancerous lesions, whereas it fuels genomic instability in the context of cancer, and how our current knowledge may be exploited for developing cancer therapies.

Mild replication stress causes chromosome mis-segregation via premature centriole disengagement

Replication stress, a hallmark of cancerous and pre-cancerous lesions, is linked to structural chromosomal aberrations. Recent studies demonstrated that it could also lead to numerical chromosomal instability (CIN). The mechanism, however, remains elusive. Here, we show that inducing replication stress in non-cancerous cells stabilizes spindle microtubules and favours premature centriole disengagement, causing transient multipolar spindles that lead to lagging chromosomes and micronuclei. Premature centriole disengagement depends on the G2 activity of the Cdk, Plk1 and ATR kinases, implying a DNA-damage induced deregulation of the centrosome cycle. Premature centriole disengagement also occurs spontaneously in some CIN+ cancer cell lines and can be suppressed by attenuating replication stress. Finally, we show that replication stress potentiates the effect of the chemotherapeutic agent taxol, by increasing the incidence of multipolar cell divisions. We postulate that replication stress in cancer cells induces numerical CIN via transient multipolar spindles caused by premature centriole disengagement.

Micronucleus formation in human cancer cells is biased by chromosome size.

Chromosomal instability is one of the hallmarks of cancer and caused by chromosome missegregation during mitosis, a process frequently associated with micronucleus formation. Micronuclei are formed when chromosomes fail to join a daughter nucleus during cell division and are surrounded by their own nuclear membrane. Although it has been commonly assumed that the gain or loss of specific chromosomes is random during compromised cell division, recent data suggest that the size of chromosomes can impact on chromosome segregation fidelity. To test whether chromosome missegregation rates scale with chromosome size in primary human cancer cells, we assessed chromosome sequestration into micronuclei in patient-derived primary NCH149 glioblastoma cells, which display high-level numerical chromosome instability (CIN), pronounced spontaneous micronucleus formation but virtually no structural CIN. The cells were analyzed by interphase fluorescence in situ hybridization using chromosome-specific painting probes for all chromosomes. Overall, 33% of early passage NCH149 cells harbored micronuclei. Entrapment within a micronucleus clearly correlated with chromosome size with larger chromosomes being significantly more frequently missegregated into micronuclei than smaller chromosomes in primary glioblastoma cells. These findings extend the concept that chromosome size determines segregation fidelity by implying that size-specific micronucleus entrapment occurs in primary human cancer cells as well.

A Markov chain for numerical chromosomal instability in clonally expanding populations.

Cancer cells frequently undergo chromosome missegregation events during mitosis, whereby the copies of a given chromosome are not distributed evenly among the two daughter cells, thus creating cells with heterogeneous karyotypes. A stochastic model tracing cellular karyotypes derived from clonal populations over hundreds of generations was recently developed and experimentally validated, and it was capable of predicting favorable karyotypes frequently observed in cancer. Here, we construct and study a Markov chain that precisely describes karyotypic evolution during clonally expanding cancer cell populations. The Markov chain allows us to directly predict the distribution of karyotypes and the expected size of the tumor after many cell divisions without resorting to computationally expensive simulations. We determine the limiting karyotype distribution of an evolving tumor population, and quantify its dependency on several key parameters including the initial karyotype of the founder cell, the rate of whole chromosome missegregation, and chromosome-specific cell viability. Using this model, we confirm the existence of an optimal rate of chromosome missegregation probabilities that maximizes karyotypic heterogeneity, while minimizing the occurrence of nullisomy. Interestingly, karyotypic heterogeneity is significantly more dependent on chromosome missegregation probabilities rather than the number of cell divisions, so that maximal heterogeneity can be reached rapidly (within a few hundred generations of cell division) at chromosome missegregation rates commonly observed in cancer cell lines. Conversely, at low missegregation rates, heterogeneity is constrained even after thousands of cell division events. This leads us to conclude that chromosome copy number heterogeneity is primarily constrained by chromosome missegregation rates and the risk for nullisomy and less so by the age of the tumor. This model enables direct integration of karyotype information into existing models of tumor evolution based on somatic mutations.

Hexavalent Chromium-Induced Chromosome Instability Drives Permanent and Heritable Numerical and Structural Changes and a DNA Repair-Deficient Phenotype.

A key hypothesis for how hexavalent chromium [Cr(VI)] causes cancer is that it drives chromosome instability (CIN), which leads to neoplastic transformation. Studies show chronic Cr(VI) can affect DNA repair and induce centrosome amplification, which can lead to structural and numerical CIN. However, no studies have considered whether these outcomes are transient or permanent. In this study, we exposed human lung cells to particulate Cr(VI) for three sequential 24-hour periods, each separated by about a month. After each treatment, cells were seeded at colony-forming density, cloned, expanded, and retreated, creating three generations of clonal cell lines. Each generation of clones was tested for chromium sensitivity, chromosome complement, DNA repair capacity, centrosome amplification, and the ability to grow in soft agar. After the first treatment, Cr(VI)-treated clones exhibited a normal chromosome complement, but some clones showed a repair-deficient phenotype and amplified centrosomes. After the second exposure, more than half of the treated clones acquired an abnormal karyotype including numerical and structural alterations, with many exhibiting deficient DNA double-strand break repair and amplified centrosomes. The third treatment produced new abnormal clones, with previously abnormal clones acquiring additional abnormalities and most clones exhibiting repair deficiency. CIN, repair deficiency, and amplified centrosomes were all permanent and heritable phenotypes of repeated Cr(VI) exposure. These outcomes support the hypothesis that CIN is a key mechanism of Cr(VI)-induced carcinogenesis.Significance: Chromium, a major public health concern and human lung carcinogen, causes fundamental changes in chromosomes and DNA repair in human lung cells. Cancer Res; 78(15); 4203-14. ©2018 AACR.

FGF-2 is a driving force for chromosomal instability and a stromal factor associated with adverse clinico-pathological features in prostate cancer

BackgroundThere is mounting evidence to suggest that stromal cells play an integral role in the progression of prostate cancer (PCa). One of the most frequently altered growth factors in PCa is fibroblast growth factor-2 (FGF-2). It has previously been proposed that early stages of PCa are characterized by a primarily exogenous, that is, stromal cell-derived FGF-2 production, whereas advanced tumors rely more on an autocrine FGF-2 production. Prostate cancer progression is characterized by an increase of genomic instability including aneuploidy and structural chromosomal alterations. Herein, we address 2 problems that have not been comprehensively answered. First, we ask whether exogenous FGF-2 can directly drive genomic instability to promote PCa progression. Second, we investigate whether and to what extent stromal FGF-2 expression is maintained in advanced PCa and whether this influences tumor progression and patient prognosis. MethodsIn vitro experiments to investigate the role of FGF-2 in numerical and structural chromosomal instability were performed using immunofluorescence microscopy, fluorescence in situ hybridization and single cell electrophoresis. A human patient-derived xenograft mouse model recapitulating osteoblastic PCa bone metastasis was used for in vivo validation experiments. The prognostic role of stromal FGF-2 expression was analyzed using immunohistochemical staining of a tissue microarray with primary tumor specimens from 162 predominantly high-risk patients with PCa. ResultsOur results show that FGF-2 not only rapidly induces mitotic defects and numerical chromosomal imbalances but also an enhanced DNA breakage to promote chromosomal instability. Using the patient-derived xenograft model, we show that a deregulation of the FGF axis results in an increase of mitotic aberrations as well as DNA damage checkpoint activation in vivo. The FGFR inhibitor dovitinib was found to reduce numerical chromosomal instability as well as DNA breakage, thus underscoring the relevance of the FGF axis in promoting genomic instability. An overexpression of tumor cell-associated FGF-2 was detected in 52 of 162 patients (32.1%), whereas a stromal overexpression was found in 27 of 165 patients (16%). Remarkably, a strong stromal FGF-2 expression was associated with a significantly higher clinical stage and higher biochemical recurrence rate. Patients with strong stromal FGF-2 expression also had a significantly worse biochemical recurrence-free survival. ConclusionsOur results underscore that exogenous FGF-2 can shape PCa cell genomes and that stromal FGF-2 expression is detectable in a sizeable proportion of advanced PCa where it is associated with adverse clinico-pathological features. Our results highlight the impact of the tumor stroma on malignant progression and provide a rationale for a further exploration of components of the tumor stroma as therapeutic targets in PCa.

PTEN in the maintenance of genome integrity: From DNA replication to chromosome segregation.

Faithful DNA replication and accurate chromosome segregation are the key machineries of genetic transmission. Disruption of these processes represents a hallmark of cancer and often results from loss of tumor suppressors. PTEN is an important tumor suppressor that is frequently mutated or deleted in human cancer. Loss of PTEN has been associated with aneuploidy and poor prognosis in cancer patients. In mice, Pten deletion or mutation drives genomic instability and tumor development. PTEN deficiency induces DNA replication stress, confers stress tolerance, and disrupts mitotic spindle architecture, leading to accumulation of structural and numerical chromosome instability. Therefore, PTEN guards the genome by controlling multiple processes of chromosome inheritance. Here, we summarize current understanding of the PTEN function in promoting high-fidelity transmission of genetic information. We also discuss the PTEN pathways of genome maintenance and highlight potential targets for cancer treatment.