Acquired clonal chromosomal abnormalities are found in the malignant cells of most patients with leukemia, lymphoma, and solid tumors. Many genes involved in consistent chromosome rearrangements, notably translocations, have already been identified. The identity of most genes affected by these aberrations will likely be determined within the next decade. Completion of the Human Genome Project has increased the pace at which new cancer-related genes are identified. Landmark special issues of Science (Feb 16, 2001) and Nature (Feb. 15, 2001) contained analysis of the initial working draft of the human genome sequence. Moreover, for a number of rearrangements, the changes in gene structure and function have been defined. Thus, general principles that are applicable to all chromosome rearrangements in human malignant disease are beginning to emerge. This review presents the most current data on primary chromosome rearrangements in both hematologic malignancies and solid tumors.

Much of the detailed information regarding the relevant chromosome rearrangements is contained in a number of recent reviews, and only a general summary is presented here.^1–3 Mitelman has published five editions of his Catalog of Chromosome Aberrations in Cancer, which describes the cytogenetic aberrations in more than 46,000 neoplasms.⁴ This catalogue is now available online <http://cgap.nci.nih.gov/Chromosomes/Mitelman>. In addition, a map of recurrent chromosomal rearrangements was published recently, and the National Cancer Institute maintains a Website with an updated map (Cancer Chromosome Aberration Project: <www.ncbi.nlm.nih.gov/CCAP>).² Another online catalog is the Atlas of Genetics and Cytogenetics in Oncology and Haematology <www.infobiogen.fr/services/chromcancer/Tumors>.

Although solid tumors such as lung, breast, and prostate cancer account for the greatest proportion of malignant disease, they represent only a small fraction of the karyotypic data. From the beginning of cytogenetic analysis, it has been clear that virtually all solid tumors, including the non-Hodgkin lymphomas, have an abnormal karyotype and that some of these abnormalities are limited to a given tumor type.^5,6 In the 1960s and 1970s, only 50% of leukemias had identifiable karyotypic abnormalities. With the improvement in cytogenetic culturing techniques and processing methods, an abnormal karyotype is now detected in over 80% of leukemias. The use of fluorescence in situ hybridization (FISH), comparative genomic hybridization (CGH), spectral karyotyping (SKY) (Figure 8-1A and Figure 8-1B) and multicolor FISH (M-FISH) is adding new precision to chromosome identification.^7,8

Figure A

Standard G banded karyotype from a 44-year-old male with acute myelomonocytic leukemia (M4). The original karyotype was 41,add(X)(q22),-Y,-5,del(7)(q11q36),inv(11)(p15q23),dic(12;?)(p12;?),-13,del(15)(q15q22),-16,add(17)(p13),-19,-20,-21,I(21)(q10),+mar1,+mar2. (more...)

Figure B

Analysis of a single cell using spectral karyotyping (SKY). The upper left figure is the metaphase cell stained with DAPI, the upper middle figure is the spectral image, the upper right is the classified image, and the lower panel is the spectral karyotype. (more...)

FISH is a technique in which DNA probes are labeled with various fluorochromes (eg, rhodamine) that are detected by fluorescence microscopy. A large number of chromosome-specific centromere probes are now available that unequivocally mark a pair of chromosomes. Using these probes, gains or losses of chromosomes can be detected not only during metaphase, but also during interphase. Large-size DNA probes that contain specific genes or anonymous DNA sequences (eg, yeast artificial chromosomes, P1, BACs) can be used to screen for recurring translocations and to identify those probes that are split by the translocation breakpoints. This technique has been widely used by many groups in their search for genes involved in different translocations.⁹ CGH is an in situ hybridization-based procedure used to detect and map relative gene copy aberrations (both gains and losses) in the tumor genome onto normal metaphase chromosomes.⁸ This technique is powerful because it does not require advanced knowledge of the existence or genetic location of altered copy-number regions or cell culture, thereby eliminating the possibility of subpopulation selection during the culture. Changes in relative gene copy number detected using CGH may be associated with oncogene amplification or loss of tumor-suppressor gene function. SKY or M-FISH is a relatively new technique in which DNA from each human chromosome pair is labeled with single or multiple fluors and is applied to the sample containing malignant cells. Using various systems to capture and analyze the image, one can detect all of the chromosome abnormalities in a single experiment (see Figure 8-1).¹⁰ SKY analysis identifies the chromosomes involved but not the specific region of the chromosome.

The newest, most exciting tool in cytogenetic analysis is array-CGH. In this technique, metaphase chromosomes are replaced with an array of thousands of BAC clones, each of which contains a segment of the human genome. Each segment is about 150 kb in length, and an array of 3,000 BACs can be constructed. Such an array samples the genome, on average, once every megabase pair. Therefore, array-CGH is similar to conducting thousands of FISH experiments at once. Array-CGH is not only more efficient than FISH, it is superior to conventional CGH in that it allows for better quantification of copy number and more precise information on the breakpoints of segments that are lost or gained. While conventional CGH is insensitive to changes that are present at low frequency, array-CGH enables investigators to evaluate large numbers of tumors for recurrent changes.¹¹

Different chromosome changes have been observed in neoplastic cells. The simplest change is either a gain or a loss of a whole chromosome. Common structural alterations are translocations, which involve the exchange of material between two or more chromosomes, and deletions, which involve the loss of DNA from a chromosome and, thus, from the affected cell (Figure 8-2). Chromosome inversions have also been observed. In this rearrangement, a single chromosome is broken in two places and the central portion inverted and rejoined to the ends of the chromosome. Each chromosome band is numbered. The chromosome number of the clone (modal number) is followed by the sex chromosomes, and gains and losses of whole chromosomes are identified by a “+” or a “-” before the chromosome number, respectively; “p” and “q” represent the short and the long arms, respectively. Deletions are indicated by the abbreviation “del.” Translocations are indicated by “t,” with the chromosomes involved noted in the first set of brackets and the breakpoints in the second set (Table 8-1). A number of international meetings over the last 25 years led to the establishment of a universally accepted system for chromosome nomenclature.¹²

Figure 8-2

Schematic diagram illustrating a normal chromosome and three chromosomal abnormalities observed in human neoplasms. A, Diagram of the banding pattern of a normal chromosome 9. The chromosome arms (p, short arm; q, long arm), regions, and band numbers (more...)

Table 8-1A

Glossary of Cytogenetic Terminology.

Table 8-1B

Karyotype Symbols.

For structural abnormalities, clonal abnormalities are defined as those that are present in at least two cells. Loss of a chromosome must occur in three cells to be considered a clonal abnormality. Banding of chromosomes is essential to cytogenetic investigations because it allows the identification of individual chromosomes. A band is defined as a chromosome area that is distinguished from adjacent segments by appearing darker or lighter through one or more banding techniques. Various banding methods are currently used, including quinacrine-mustard (Q bands) and trypsin-Giemsa banding (G bands).

Chromosomes for analysis must be obtained from tumor cells. For leukemia, bone marrow cells or peripheral blood cells processed directly or after 24- to 72-h culture are used; lymph nodes or solid tumors are minced to yield a single cell suspension that can be harvested immediately or cultured for a short period of time. The cells are exposed to a hypotonic solution, fixed, and stained according to a variety of protocols.¹³ Brief exposure to mitotic inhibitors, DNA-binding agents to elongate chromosomes, or amethopterin or fluorodeoxyuridine to synchronize cells has resulted in longer, more distinct chromosomes. The addition of PHA-stimulated conditioned medium or recombinant colony-stimulating factors to the culture medium has also contributed to the increased rate of successful cytogenetic analysis of different tumors. Cytogenetic analysis requires specimens that contain viable dividing cells; therefore, specimens should be transported without delay to the cytogenetics laboratory in a suitable culture medium at room temperature.

Figure 8-3

Partial karyotypes from trypsin-Giemsa-banded metaphase cells depicting nonrandom chromosomal rearrangements observed in myeloid malignant diseases. (a) t(9;22)(q34;q11), CML; (b) t(8;21)(q22;q22), AML-M2; (c) inv(16)(p13q22), AMMoL-M4Eo; (d) t(15;17)(q22;q11–12), (more...)

Figure 8-4

The locations of translocation breakpoints that involve MLL are indicated by colored bands. The involvement of MLL was determined by FISH, by Southern blot analysis, or by RT-PCR. Symbols to the right of the band indicate that the translocation has been (more...)

Contents

• Myeloproliferative Disorders
• Acute Myeloid Leukemia and Myelodysplastic Syndrome Associated with Prior Cytotoxic Therapy
• How And Why Recurring Chromosomal Abnormalities Occur
• Acknowledgment
• References