Unstable genomes elevate transcriptome dynamics.

The challenge of identifying common expression signatures in cancer is well known, however the reason behind this is largely unclear. Traditionally variation in expression signatures has been attributed to technological problems, however recent evidence suggests that chromosome instability (CIN) and resultant karyotypic heterogeneity may be a large contributing factor. Using a well-defined model of immortalization, we systematically compared the pattern of genome alteration and expression dynamics during somatic evolution. Co-measurement of global gene expression and karyotypic alteration throughout the immortalization process reveals that karyotype changes influence gene expression as major structural and numerical karyotypic alterations result in large gene expression deviation. Replicate samples from stages with stable genomes are more similar to each other than are replicate samples with karyotypic heterogeneity. Karyotypic and gene expression change during immortalization is dynamic as each stage of progression has a unique expression pattern. This was further verified by comparing global expression in two replicates grown in one flask with known karyotypes. Replicates with higher karyotypic instability were found to be less similar than replicates with stable karyotypes. This data illustrates the karyotype, transcriptome, and transcriptome determined pathways are in constant flux during somatic cellular evolution (particularly during the macroevolutionary phase) and this flux is an inextricable feature of CIN and essential for cancer formation. The findings presented here underscore the importance of understanding the evolutionary process of cancer in order to design improved treatment modalities.

Genome chaos: survival strategy during crisis.

Genome chaos, a process of complex, rapid genome re-organization, results in the formation of chaotic genomes, which is followed by the potential to establish stable genomes. It was initially detected through cytogenetic analyses, and recently confirmed by whole-genome sequencing efforts which identified multiple subtypes including "chromothripsis", "chromoplexy", "chromoanasynthesis", and "chromoanagenesis". Although genome chaos occurs commonly in tumors, both the mechanism and detailed aspects of the process are unknown due to the inability of observing its evolution over time in clinical samples. Here, an experimental system to monitor the evolutionary process of genome chaos was developed to elucidate its mechanisms. Genome chaos occurs following exposure to chemotherapeutics with different mechanisms, which act collectively as stressors. Characterization of the karyotype and its dynamic changes prior to, during, and after induction of genome chaos demonstrates that chromosome fragmentation (C-Frag) occurs just prior to chaotic genome formation. Chaotic genomes seem to form by random rejoining of chromosomal fragments, in part through non-homologous end joining (NHEJ). Stress induced genome chaos results in increased karyotypic heterogeneity. Such increased evolutionary potential is demonstrated by the identification of increased transcriptome dynamics associated with high levels of karyotypic variance. In contrast to impacting on a limited number of cancer genes, re-organized genomes lead to new system dynamics essential for cancer evolution. Genome chaos acts as a mechanism of rapid, adaptive, genome-based evolution that plays an essential role in promoting rapid macroevolution of new genome-defined systems during crisis, which may explain some unwanted consequences of cancer treatment.

Chromosomal instability (CIN): what it is and why it is crucial to cancer evolution.

Results of various cancer genome sequencing projects have "unexpectedly" challenged the framework of the current somatic gene mutation theory of cancer. The prevalence of diverse genetic heterogeneity observed in cancer questions the strategy of focusing on contributions of individual gene mutations. Much of the genetic heterogeneity in tumors is due to chromosomal instability (CIN), a predominant hallmark of cancer. Multiple molecular mechanisms have been attributed to CIN but unifying these often conflicting mechanisms into one general mechanism has been challenging. In this review, we discuss multiple aspects of CIN including its definitions, methods of measuring, and some common misconceptions. We then apply the genome-based evolutionary theory to propose a general mechanism for CIN to unify the diverse molecular causes. In this new evolutionary framework, CIN represents a system behavior of a stress response with adaptive advantages but also serves as a new potential cause of further destabilization of the genome. Following a brief review about the newly realized functions of chromosomes that defines system inheritance and creates new genomes, we discuss the ultimate importance of CIN in cancer evolution. Finally, a number of confusing issues regarding CIN are explained in light of the evolutionary function of CIN.

The evolutionary mechanism of cancer.

Identification of the general molecular mechanism of cancer is the Holy Grail of cancer research. Since cancer is believed to be caused by a sequential accumulation of cancer gene mutations, the identification, characterization, and targeting of common genetic alterations and their defined pathways have dominated the field for decades. Despite the impressive data accumulated from studies of gene mutations, epigenetic dysregulation, and pathway alterations, an overwhelming amount of diverse molecular information has offered limited understanding of the general mechanisms of cancer. To solve this paradox, the newly established genome theory is introduced here describing how somatic cells evolve within individual patients. The evolutionary mechanism of cancer is characterized using only three key components of somatic cell evolution that include increased system dynamics induced by stress, elevated genetic and epigenetic heterogeneity, and genome alteration mediated natural selection. Cancer progression represents a macro-evolutionary process where karyotype change or genome replacement plays the key dominant role. Furthermore, the recently identified relationship between the evolutionary mechanism and a large number of diverse individual molecular mechanisms is discussed. The total sum of all the individual molecular mechanisms is equal to the evolutionary mechanism of cancer. Individual molecular mechanisms including all the molecular mechanisms described to date are stochastically selected and unpredictable and are therefore clinically impractical. Recognizing the fundamental importance of the underlying basis of the evolutionary mechanism of cancer mandates the development of new strategies in cancer research.

Genetic and epigenetic heterogeneity in cancer: a genome-centric perspective.

Genetic and epigenetic heterogeneity (the main form of non-genetic heterogeneity) are key elements in cancer progression and drug resistance, as they provide needed population diversity, complexity, and robustness. Despite drastically increased evidence of multiple levels of heterogeneity in cancer, the general approach has been to eliminate the "noise" of heterogeneity to establish genetic and epigenetic patterns. In particular, the appreciation of new types of epigenetic regulation like non-coding RNA, have led to the hope of solving the mystery of cancer that the current genetic theories seem to be unable to achieve. In this mini-review, we have briefly analyzed a number of mis-conceptions regarding cancer heterogeneity, followed by the re-evaluation of cancer heterogeneity within a framework of the genome-centric concept of evolution. The analysis of the relationship between gene, epigenetic and genome level heterogeneity, and the challenges of measuring heterogeneity among multiple levels have been discussed. Further, we propose that measuring genome level heterogeneity represents an effective strategy in the study of cancer and other types of complex diseases, as emphasis on the pattern of system evolution rather than specific pathways provides a global and synthetic approach. Compared to the degree of heterogeneity, individual molecular pathways will have limited predictability during stochastic cancer evolution where genome dynamics (reflected by karyotypic heterogeneity) will dominate.

Genome based cell population heterogeneity promotes tumorigenicity: the evolutionary mechanism of cancer.

Cancer progression represents an evolutionary process where overall genome level changes reflect system instability and serve as a driving force for evolving new systems. To illustrate this principle it must be demonstrated that karyotypic heterogeneity (population diversity) directly contributes to tumorigenicity. Five well characterized in vitro tumor progression models representing various types of cancers were selected for such an analysis. The tumorigenicity of each model has been linked to different molecular pathways, and there is no common molecular mechanism shared among them. According to our hypothesis that genome level heterogeneity is a key to cancer evolution, we expect to reveal that the common link of tumorigenicity between these diverse models is elevated genome diversity. Spectral karyotyping (SKY) was used to compare the degree of karyotypic heterogeneity displayed in various sublines of these five models. The cell population diversity was determined by scoring type and frequencies of clonal and non-clonal chromosome aberrations (CCAs and NCCAs). The tumorigenicity of these models has been separately analyzed. As expected, the highest level of NCCAs was detected coupled with the strongest tumorigenicity among all models analyzed. The karyotypic heterogeneity of both benign hyperplastic lesions and premalignant dysplastic tissues were further analyzed to support this conclusion. This common link between elevated NCCAs and increased tumorigenicity suggests an evolutionary causative relationship between system instability, population diversity, and cancer evolution. This study reconciles the difference between evolutionary and molecular mechanisms of cancer and suggests that NCCAs can serve as a biomarker to monitor the probability of cancer progression.

Mitotic cell death by chromosome fragmentation.

Cell death plays a key role for both cancer progression and treatment. In this report, we characterize chromosome fragmentation, a new type of cell death that takes place during metaphase where condensed chromosomes are progressively degraded. It occurs spontaneously without any treatment in instances such as inherited status of genomic instability, or it can be induced by treatment with chemotherapeutics. It is observed within cell lines, tumors, and lymphocytes of cancer patients. The process of chromosome fragmentation results in loss of viability, but is apparently nonapoptotic and further differs from cellular death defined by mitotic catastrophe. Chromosome fragmentation represents an efficient means of induced cell death and is a clinically relevant biomarker of mitotic cell death. Chromosome fragmentation serves as a method to eliminate genomically unstable cells. Paradoxically, this process could result in genome aberrations common in cancer. The characterization of chromosome fragmentation may also shine light on the mechanism of chromosomal pulverization.

Stochastic cancer progression driven by non-clonal chromosome aberrations.

Cancer research has previously focused on the identification of specific genes and pathways responsible for cancer initiation and progression based on the prevailing viewpoint that cancer is caused by a stepwise accumulation of genetic aberrations. This viewpoint, however, is not consistent with the clinical finding that tumors display high levels of genetic heterogeneity and distinctive karyotypes. We show that chromosomal instability primarily generates stochastic karyotypic changes leading to the random progression of cancer. This was accomplished by tracing karyotypic patterns of individual cells that contained either defective genes responsible for genome integrity or were challenged by onco-proteins or carcinogens that destabilized the genome. Analysis included the tracing of patterns of karyotypic evolution during different stages of cellular immortalization. This study revealed that non-clonal chromosomal aberrations (NCCAs) (both aneuploidy and structural aberrations) and not recurrent clonal chromosomal aberrations (CCAs) are directly linked to genomic instability and karyotypic evolution. Discovery of "transitional CCAs" during in vitro immortalization clearly demonstrates that karyotypic evolution in solid tumors is not a continuous process. NCCAs and their dynamic interplay with CCAs create infinite genomic combinations leading to clonal diversity necessary for cancer cell evolution. The karyotypic chaos observed within the cell crisis stage prior to establishment of the immortalization further supports the ultimate importance of genetic aberrations at the karyotypic or genome level. Therefore, genomic instability generated NCCAs are a key driving force in cancer progression. The dynamic relationship between NCCAs and CCAs provides a mechanism underlying chromosomal based cancer evolution and could have broad clinical applications.

Cancer progression by non-clonal chromosome aberrations.

The establishment of the correct conceptual framework is vital to any scientific discipline including cancer research. Influenced by hematologic cancer studies, the current cancer concept focuses on the stepwise patterns of progression as defined by specific recurrent genetic aberrations. This concept has faced a tough challenge as the majority of cancer cases follow non-linear patterns and display stochastic progression. In light of the recent discovery that genomic instability is directly linked to stochastic non-clonal chromosome aberrations (NCCAs), and that cancer progression can be characterized as a dynamic relationship between NCCAs and recurrent clonal chromosome aberrations (CCAs), we propose that the dynamics of NCCAs is a key element for karyotypic evolution in solid tumors. To support this viewpoint, we briefly discuss various basic elements responsible for cancer initiation and progression within an evolutionary context. We argue that even though stochastic changes can be detected at various levels of genetic organization, such as at the gene level and epigenetic level, it is primarily detected at the chromosomal or genome level. Thus, NCCA-mediated genomic variation plays a dominant role in cancer progression. To further illustrate the involvement of NCCA/CCA cycles in the pattern of cancer evolution, four cancer evolutionary models have been proposed based on the comparative analysis of karyotype patterns of various types of cancer.

Clonal and non-clonal chromosome aberrations and genome variation and aberration.

The theoretical view that genome aberrations rather than gene mutations cause a majority of cancers has gained increasing support from recent experimental data. Genetic aberration at the chromosome level is a key aspect of genome aberration and the systematic definition of chromosomal aberrations with their impact on genome variation and cancer genome evolution is of great importance. However, traditionally, efforts have focused on recurrent clonal chromosome aberrations (CCAs). The significance of stochastic non-clonal chromosome aberrations (NCCAs) is discussed in this paper with emphasis on the simple types of NCCAs that have until recently been considered "non-significant background". Comparison of various subtypes of transitional and late-stage CCAs with simple and complex types of NCCAs has uncovered a dynamic relationship among NCCAs, CCAs, overall genomic instability, and karyotypic evolution, as well as the stochastic nature of cancer evolution. Here, we review concepts and methodologies to measure NCCAs and discuss the possible causative mechanism and consequences of NCCAs. This study raises challenging questions regarding the concept of cancer evolution driven by stochastic chromosomal aberration mediated genome irregularities that could have repercussions reaching far beyond cancer and organismal genomes.