The serial cultivation of human diploid cell strains.

It is proposed that most neoplasms arise from a single cell of origin, and tumor progression results from acquired genetic variability within the original clone allowing sequential selection of more aggressive sublines. Tumor cell populations are apparently more genetically unstable than normal cells, perhaps from activation of specific gene loci in the neoplasm, continued presence of carcinogen, or even nutritional deficiencies within the tumor. The acquired genetic insta0ility and associated selection process, most readily recognized cytogenetically, results in advanced human malignancies being highly individual karyotypically and biologically. Hence, each patient's cancer may require individual specific therapy, and even this may be thwarted by emergence of a genetically variant subline resistant to the treatment. More research should be directed toward understanding and controlling the evolutionary process in tumors before it reaches the late stage usually seen in clinical cancer.

http://science.sciencemag.org/content/sci/194/4260/23.full.pdf

The clonal evolution of tumor cell populations

Disaggregated mouse embryo cells, grown in monolayers, underwent a progressive decline in growth rate upon successive transfer, the rapidity of the decline depending, among other things, on the inoculation density. Nevertheless, nearly all cultures developed into established lines within 3 months of culture. The first sign of the emergence of an established line was the ability of the cells to maintain a constant or rising potential growth rate. This occurred while the cultures were morphologically unchanged. The growth rate continued to increase until it equaled or exceeded that of the original culture. The early established cells showed an increasing metabolic autonomy, as indicated by decreasing dependence on cell-to-cell feeding. It is suggested that the process of establishment involves an alteration in cell permeability properties. Chromosome studies indicated that the cells responsible for the upturn in growth rate were diploid, but later the population shifted to the tetraploid range, often very rapidly. Still later, marker chromosomes appeared. Different lines acquired different properties, depending on the culture conditions employed; one line developed which is extremely sensitive to contact inhibition.

/pdf/quantitative-studies-of-the-growth-of-mouse-embryo-cells-in-4t852yup42.pdf

Quantitative studies of the growth of mouse embryo cells in culture and their development into established lines

Evolution of the Cancer Stem Cell Model

The success of anticancer therapy is usually limited by the development of drug resistance. Such acquired resistance is driven, in part, by intratumoural heterogeneity - that is, the phenotypic diversity of cancer cells co-inhabiting a single tumour mass. The introduction of the cancer stem cell (CSC) concept, which posits the presence of minor subpopulations of CSCs that are uniquely capable of seeding new tumours, has provided a framework for understanding one dimension of intratumoural heterogeneity. This concept, taken together with the identification of the epithelial-to-mesenchymal transition (EMT) programme as a critical regulator of the CSC phenotype, offers an opportunity to investigate the nature of intratumoural heterogeneity and a possible mechanistic basis for anticancer drug resistance. In fact, accumulating evidence indicates that conventional therapies often fail to eradicate carcinoma cells that have entered the CSC state via activation of the EMT programme, thereby permitting CSC-mediated clinical relapse. In this Review, we summarize our current understanding of the link between the EMT programme and the CSC state, and also discuss how this knowledge can contribute to improvements in clinical practice.

EMT, CSCs, and drug resistance: the mechanistic link and clinical implications

The development of modern cancer cytogenetics is briefly reviewed. The observation around 1950 that ascites tumors were excellent materials for chromosome analysis acted as an incitement both of cancer cytogenetics and of mammalian cytogenetics in general.



The Philadelphia chromosome of chronic myelogenous leukemia (CML). discovered in 1960, was long unique in showing strict correlation between specific chromosomal deviation and malignant disease. Detailed chromosome analysis of large series of experimental tumors was needed to establish the distinction between significant and accidental chromosome deviations and to demonstrate that the former were nonrandom and formed predetermined patterns. These were correlated to the inducing factor rather than to the tissue type, from which the tumor developed.



Systematic analysis of chromosomal aberrations in more than 600 human neoplasms, studied by our group or taken from the literature, showed that in 12 different tumor types most chromosomes were unaffected, the significant deviations being clustered to 10 of the 24 human chromosome types. It was suggested that genes of special importance to malignant development were located in a few specific chromosomes.



The 2 obscure, cancer-associated phenomena of homogeneously staining chromosome regions and double minute chromosomes are briefly described and a tentative interpretation is proposed.

https://www.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1601-5223.1977.tb01208.x

Chromosomes and cancer

Endomitotic reduplication mechanisms in ascites tumors of the mouse.

Basing our views on observations of chromosomal behavior in cancers and following the classical views of Boveri, Winge, and others, we tend to regard the development of malignancy as a process of selection on the cellular level that favors cells with the acquired capacity of escaping the control of the host organism. The regular appearance of new karyotypes in cancer stemlines supports this view. In our opinion the significant feature in some kinds of cancerogenesis is the induction of genotypic variability. Any random genotype variation within a tissue may in time produce cells capable of neoplastic growth. Many facts indicate that the genotypic changes leading to this end are highly complex. Therefore, considerable experimentation with various genotypes will be necessary, and the transition from normal cells over precancerous to malignant cells will be floating. Natural selection within the cell population should automatically lead to cells with ever-greater independence of the host control. Because gross chromosomal disturbances and somatic segregation for chromatid and subchromatid changes will take place a t mitosis, i t is evident that any chromosomal instability will have an especially strong effect in tissues with high mitotic activity. According to this view any mutagenic agent could be carcinogenic, provided the mutagenic action is extended over many mitotic cycles. Even one single exposure to X radiation would thus have a chance to bring about this effect if a specific type of mutation is induced in either the nucleus or the cytoplasm, that is, a mutation giving a permanent increase in mutability in all cells carrying this gene. This view is compatible with the chemical induction of cancer and perhaps will turn out to be so with the viral induction and transfer of the disease. The change in environment connected with the explantation of mammalian cells from their normal position in the body into culture i l z vitro is indeed a drastic one. The fact that a high percentage of cells may survive such a change and may even start reproduction under the new conditions is good evidence of the considerable adaptability of mammalian cells. In many Cornell University Medical College, New I.ork, N . Y .

Role of chromosomes in cancerogenesis, as studied in serial tissue culture of mammalian cells.

AN investigation has been carried on for some time in this Laboratory into the immediate cytological effect of salt solutions on root meristems of Allium Cepa. Salts, mostly nitrates, of some forty metals were tested. A dilution series of 10–16 concentrations were studied for each substance, covering the whole range from total lethality down to such weak concentrations that no cytological effect could be detected. Fixations of root tips were made after 4 and 24 hours respectively. All slides were stained with crystal-violet.

Cytological reactions induced by inorganic salt solutions.

The cancer literature contains a great number of papers providing information about chromosomes in cancer tissue. Very few of these investigations, however, have chromosomes as their main issue, or are written by cytologists. In the majority of papers, the information about chromosomes is given more or less in passing, as side-products of pathological, chemotherapeutic, or other investigations. Data on cancer chromosomes have been gradually accumulating, however, and it was realized early that chromosomes in cancer tissue behaved somewhat differently from those of normal tissue: the chromosomal mechanism functioned less accurately, mitotic deviations of all kinds being frequent. Ever since the end of the last century, etiological connections have been suggested between chromosomal aberrations and the origin of malignancy. Names such as von Hansemann and Boveri are associated with this idea. is the first modern cytogeneticist to stress the tumor tissue as a genetically heterogeneous cell population and to visualize the origin and growth of tumors as a result of selective processes among the constituents of this population. He based his ideas on careful and extensive chromosome studies in plant and animal tumors. The followiiig quotation out of Winge’s 1930 paper2 (p. 727) illustrates his view of the origin and growth of tumors: “Ich betrachte den malignen Tumor als bestehend aus Abkommlingszellen von einer oder mehreren Zellen, die durch eine lokale Reizung einen abnormen Chromosomenbestand bekommen haben, in dem besonders haufig ein Verlust verschiedener Chromosomen und eventuell Verdoppelungen ariderer Chromosomen eingetreten ist, was eine vergrosserte Wachstumsintensitat und im iibrigen cinen gestohrten Metabolismus mit sich fiihren kann.” This quotation shows that Winge was well aware of the stemliiie concept. Through genetic variation in the tumor, manifested as asymmetric or multipolar mitoses, the basis for an increase ic virulence is available: “Es geschieht wahrend des Vordringens des malignen Gewebes einc Selektion unter den aberranteii Zellen” (p. 728). Winge considers chromosome doublings as especially significant for the development of malignancy. In his analysis of tar carcinomas of micc, he observed that the most malignant neoplasms usually had hypotetraploid chromosome numbers, while precancerous stages still maintained near-diploid numbers. Winge’s view of the cancer tissue as genetically a mosaic, in which the most viable cell lineages are seIected to propagate the tumor, is still largely valid. On the other hand, it does not seem to follow necessarily that the original, primary induction of malignancy is caused by the same factors as those responsible for changes in the established tumors, as evidently assumed by Winge. T t is one of the most urgent tasks for chromosome research in cancer to focus on the early stages of cancer growth, or even on precancerous stages. The discovery during recent years that ascites tumors permit a much more Winge’p

Albert Levan

Papers

Chromosomes and cancer

Endomitotic reduplication mechanisms in ascites tumors of the mouse.

Role of chromosomes in cancerogenesis, as studied in serial tissue culture of mammalian cells.

Cytological reactions induced by inorganic salt solutions.

Chromosomes in cancer tissue.