Award Articles and Special Reports Philip Levine Award Lecture Chromosome Translocations and Oncogenes in Human Lymphoid Tumors PETER C. NOWELL, M.D. AND CARLO M. CROCE, M.D.

IN THE LAST several decades, chromosome studies have made significant contributions to our understanding of tumor biology. They have helped to establish the clonal nature of most neoplasms and the important role of sequential somatic genetic changes in both the initiation and subsequent progression of many forms of cancer. More recently, the identification of nonrandom chromosomal alterations associated with specific neoplasms has helped to provide important clues to the location of genes involved in the pathogenesis of these tumors and to the mechanisms by which their function has been critically altered. Much of this work has been done in hematopoietic neoplasms, in which the study of certain nonrandom reciprocal translocations has proved particularly valuable. Perhaps most actively investigated has been the t(9;22) chromosome translocation associated with chronic myelogenous leukemia and some acute myeloid and lymphoid leukemias. This rearrangement, which results in

Received September 25, 1989; received revised manuscript and accepted for publication December 4, 1989. Presented in part at the Fall Meeting of the American Society of Clinical Pathologists as the Philip Levine Award Lecture, Washington, D.C., October 1989. Address reprint requests to Dr. Nowell: Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6082.

Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine and Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania

altered structure and function of the c-abl proto-oncogene, has been described elsewhere2354 and will not be considered here. Two other common translocations, also extensively studied, are associated with B-cell neoplasms: the t(8; 14) (q24;q32) rearrangement in Burkitt's lymphoma (BL) and the t( 14; 18) (q32;q21) translocation seen in many low-grade follicular lymphomas. 28 The t(8; 14) translocation in the Burkitt's tumor involves the c-myc oncogene, which appears to be important in the pathogenesis of many types of human cancer; and study of the t( 14; 18) translocation has led to identification of a previously unrecognized oncogene, the bcl-2 gene.12 In this brief review, we will first summarize findings concerning involvement of the c-myc gene in BLs and in other B-cell and T-cell tumors and then describe the evidence for a number of other, previously unknown, growth regulatory genes (of which bcl-2 is the prototype) that can apparently function as oncogenes or "activators" of oncogenes in B- or T-lymphocytes. Chromosome Translocations Involving the C-myc Gene Burkitt 's Lymphomas and Immunoglobulin Genes In approximately 75% of BLs, both endemic and sporadic, the neoplastic cells are characterized by a t(8;14) (q24;q32) translocation. The other 25% of BL cases have a "variant" translocation that includes the same region of chromosome #8, but the other chromosome involved is either #2 or #22. 12 ' 28 Molecular studies of the t(8;14) translocation have shown that it splits the immunoglobulin heavy chain locus and brings the c-myc proto-oncogene, the human homologue of the v-myc oncogene and normally located on chromosome band 8q24, into juxtaposition with immunoglobulin gene sequences on band 14q32.12'31 In the variant translocations, the immunoglobulin kappa light chain gene (from chromosome

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Chromosome studies are helping to identify oncogenes, both known and previously unknown, involved in the pathogenesis of human lymphocytic tumors; and mechanisms by which the function of these genes is critically altered. Most extensively studied have been the chromosome translocations involving the myc gene in Burkitt's lymphomas and the bcl-2 gene in low-grade lymphomas, where "activation" of the oncogene results from association with a transcriptionally active immunoglobulin gene. Other putative oncogenes, similarly involved in translocations with immunoglobulin genes (in B-cell tumors) or T-cell receptor genes (in T-cell tumors), are currently being investigated, as well as alternative mechanisms of myc gene activation in these neoplasms. Limited clinical applications of these studies have already been forthcoming, and they should eventually lead to improvements in diagnosis, prognosis, and even therapy. (Key words: Lymphoma; Leukemia; Chromosome; Translocation; Oncogene, myc; bcl-2) Am J Clin Pathol 1990;94:229-237

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location.53 In the case of endemic BL or acquired immune deficiency syndrome (AIDS) -associated BL, this is typically adjacent to D-regions or J-regions of the heavy chain locus, reflecting an error in normal V-D-J joining.25'42'53 In the sporadic Burkitt's tumors, in which the chromosome translocation appears to occur at a later stage of differentiation, during isotype switching, it commonly involves the switch region of the immunoglobulin locus.42,53 This difference may reflect in part the role of the EpsteinBarr virus, as well as of immune defects, in the African and AIDS-related cases in which the chronic viral infection leads to continued expansion of a pool of partially differentiated B-cells within which the translocation event occurs and the tumor subsequently emerges.25'53 The importance of structural changes in the myc gene itself has been a matter of debate. Truncation or point mutations in the gene may contribute to the deregulation phenomenon.31'42'62 However, in the variant translocations, involving chromosome #2 or #22, the activating immunoglobulin light chain gene is translocated to the 3' end of the myc gene and often at some distance from it. In several of these cases the c-myc gene has been completely sequenced and shown to be structurally unaltered.53 It is still not clear which of the factors that normally regulate immunoglobulin gene transcription are acting on the myc gene in these various circumstances, but it does appear that in at least some instances they can

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FIG. 1. Diagram of the three nonrandom chromosome translocations associated with Burkitt's lymphoma. In the common t(8; 14) translocation (A), the c-myc gene from chromosome No. 8 is brought into juxtaposition with the constant region of the immunoglobulin heavy chain locus that remains on chromosome No. 14 (Ch). In the "variant" translocations, t(8;22) and t(2;8), the constant region of an immunoglobulin light chain gene, either lambda (B) or kappa (Q, is placed 3' (distal) to the c-myc gene, which remains on chromosome No. 8. In all cases, the c-myc gene is deregulated as a result of the translocation.

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#2) or the lambda light chain gene (from chromosome #22) is translocated to the 3' end of the c-myc gene, which remains on chromosome #8.12-31 In each of these circumstances, a rearranged and transcriptionally active immunoglobulin gene is brought into association with the c-myc gene (Fig. 1), resulting in inappropriate expression of the oncogene. Normally, transcriptional activity of the myc gene varies with the proliferative state of the cell, and it is believed that the gene product, a nuclear binding protein, is importantly involved in the regulation of a number of other critical genes.45 Unlike its normal counterpart, the translocated myc gene does not show these variations in expression, and presumably this "deregulated" state plays a major role in the altered growth patterns that lead to the expansion of the neoplastic B-cell clone.12-314553 Considerable information has been developed on the specific molecular mechanisms involved in the chromosome translocation itself and in the resultant change in myc gene function, although some important aspects remain to be clarified. It does appear that the t(8;14) translocation results from errors in the immunoglobulin heavy chain rearrangements that occur during normal B-cell differentiation. The recombinase enzymes that mediate this process apparently use short DNA sequences adjacent to the myc gene that are homologous to those normally involved and thus help to bring the oncogene into its new

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act over considerable distance to deregulate an otherwise structurally normal proto-oncogene.9'53'55 Thus, a number of the molecular and biologic details of the pathogenesis of BLs are still being investigated, but the extensive work on this tumor in recent years has left little doubt that chromosome translocation, with resultant activation of the c-myc proto-oncogene, is a major factor in its development.

deregulation of its function. As with the endemic Burkitt's tumors, this translocation appears to result from an error in V-D-J joining, involving the recombinase enzyme system, and the resultant tumors have usually presented as aggressive T-cell leukemias.17'22 It is interesting that there has as yet been no demonstration of T-cell tumors with translocations involving the myc gene and the other two T-cell receptor genes, beta and gamma, which are both located on chromosome #7.

T-Cell Tumors and T-Cell Receptor Genes

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Other Translocations Involving the C-myc Gene Recently, studies of two other lymphomas have indicated that under some circumstances the c-myc gene may be inappropriately activated through mechanisms that do not involve either immunoglobulin genes or T-cell receptor genes. The first was a neoplastic T-cell line, HUT-78, derived from a patient with Sezary's syndrome. This line has a complex karyotype and was demonstrated, at the molecular level, to have a translocation involving the 3' region of the c-myc gene and a previously unidentified locus on the long arm of chromosome #2 at band q23-24. 118 This rearrangement resulted not only in overexpression of the c-myc gene, but also in the production of a fused transcript encompassing sequences both from the myc gene and from the locus on chromosome #2 (designated tcl-4).iS This example of a "hybrid" gene product resulting from a translocation involving the c-myc gene resembles the t(9;22) chromosome translocation in Ph-positive leukemias, where a hybrid gene is formed between the c-abl

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FlG. 2. Diagram of four abnormal chromosomes that result from nonrandom translocations occurring in T-cell tumors. A. The 8q+ chromosome derived from the t(8; 14) translocation that brings a portion of the T-alpha gene (J-alpha, C-alpha) or the T-delta gene into juxtaposition with c-myc. B, C. and D. The 10q+, 1 lp+, and 14q+ chromosomes derived from the translocations noted under the diagrams and discussed in the text. In each case, a putative oncogene (lcl-1, lcl-2, tcl-\) is associated with the T-alpha/delta locus.

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Much less is known about the role of the myc gene in the pathogenesis of T-cell neoplasms. Several years ago, the mapping of the genes coding for subunits of the Tcell antigen receptor provided the opportunity to ask whether chromosome translocations leading to oncogene activation might be involved in the development of these tumors in a manner similar to that just described for Bcell neoplasms.22,29'53 It was known that in certain T-cell leukemias and lymphomas, karyotypic alterations are relatively common that involve the proximal portion of the long arm of chromosome #14, at band q l l , where the gene for the delta-chain of the T-cell receptor is located within the gene that codes for the alpha-chain. 22 ' 2829 In several cases the translocation was a t(8;14) (q24;ql 1) rearrangement, which was shown to bring a portion of either the alpha or the delta chain gene adjacent to the cmyc gene on chromosome #8 (Fig. 2A).22S3 The association is similar to that of the variant translocations in the Burkitt's tumor, except that here it is the T-cell receptor gene that is placed 3' of the c-myc gene, with resultant

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Translocations Involving Previously Unknown Oncogenes Although 30 or 40 human proto-oncogenes, homologous to retroviral oncogenes, have been identified, only the c-myc gene, as discussed in the preceding section, and the c-ablgene in Ph-positive acute lymphocytic leukemia10 have been shown to be significantly involved in the pathogenesis of human lymphoid tumors through "acti-

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vation" mechanisms mediated by chromosome translocation.6,53 Cytogenetic studies, however, have provided clues to the location of a number of other growth regulatory genes that were previously unknown but may be similarly activated through translocation. Only one of these, the bcl-2 gene, has been isolated and partially characterized, but a number of others are currently being investigated and will be discussed briefly. Putative Oncogenes in B-Cell Tumors The bcl-2 Gene. As indicated earlier, a t(14;18) (q32;q21) translocation occurs very commonly in association with low-grade follicular lymphomas. 28 From the involved region on chromosome #18, a gene, designated bcl-2, was cloned and shown to be deregulated through juxtaposition with the immunoglobulin heavy chain locus in a manner analogous to that described earlier for the cmyc gene in the BL."' 2 ' 5 2 (Fig. 3). The same molecular mechanisms appear to be operative, involving errors in V-D-J joining, with the recombinase enzymes recognizing homologous signal sequences adjacent to the bcl-2 gene.53 Unlike deregulation of the myc gene, however, the inappropriately activated bcl-2 gene appears to confer only a slight growth advantage on the affected B-cell, resulting in a relatively slowly expanding neoplastic clone. Clinical applications have already been found, in that a probe for the translocated bcl-2 gene is being used to look for residual tumor cells with the t( 14; 18) translocation in patients who have apparently been treated successfully for these indolent tumors. 32 It is also now clear that involvement of the bcl-2 gene through the t( 14; 18) translocation can be an early step in a neoplasm that ultimately evolves to a much more aggressive malignancy. It has been shown in several cases that progression from a low-grade to a high-grade lymphoma can occur when a tumor that originally had only the t( 14; 18) translocation acquires additional genetic changes, such as the t(8; 14) translocation involving the myc gene.14'21 In other circumstances, the disease may

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FlG. 3. Diagram of the t(14; 18) translocation found in many B-cell neoplasms. The putative oncogene bcl-2 is moved from its normal location on chromosome No. 18 into association with sequences of the immunoglobulin heavy chain locus on chromosome No. 14, resulting in deregulation of bcl-2.

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proto-oncogene from chromosome #9 and bcr sequences from chromosome #22. 54 How frequently such a phenomenon involves the myc gene and how important such hybrid myc genes may be in the pathogenesis of various lymphocytic neoplasms remain to be determined. Recently, we investigated a case that suggests still another mechanism for the abnormal activation of c-myc in lymphoid tumors. 20 We examined cells from a patient with an aggressive B-cell leukemia that may have evolved from an earlier indolent lymphoma, because the complex karyotype included a t(14;18) translocation involving the bcl-2 gene, as will be discussed later. Molecular studies confirmed the presence, also, of a translocation involving c-myc. In this case a truncated myc gene was associated with promoter elements of a previously unknown locus on the long arm of chromosome #17 at band q22. This association resulted in unusually high levels of myc expression and presumably contributed to the aggressive nature of the patient's leukemia. The newly identified gene on chromosome #17, designated bcl-3, was found to be expressed in a variety of hematopoietic lineages but not in other cell types, suggesting that it may play an important growth regulatory role in hemic cells. As with the findings in the HUT-78 cell line, it remains to be determined how frequently myc gene activation through this translocation involving chromosome #17 is an important factor in the progression of lymphoid tumors, because visible abnormalities of this chromosome are only moderately frequent in these neoplasms.

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Some effort has been made to demonstrate the oncogenic potential of the bcl-2 gene in classical transfection assays as well as in transgenic mice. The results indicate a weak transforming ability in murine fibroblasts, requiring cotransfection with a ras gene,47 and similar weakly oncogenic effects in other assays.34'58 In general, these findings are consistent with the characteristics in vivo of the low-grade tumors with which this gene has been identified, and additional experimental studies are underway to define better the normal role of the bcl-2 gene and exactly how its "activated" form leads to abnormal growth. Thebc\-1 Gene. At(l 1 ;14)(ql3;q32) translocation has been demonstrated in a variety of B-cell tumors, both lymphomas and chronic lymphocytic leukemia.28 This translocation has also been shown to involve the immunoglobulin heavy chain locus, and it has been postulated that another putative oncogene, designated bcl-1, may be activated in a manner similar to that demonstrated for the myc and bcl-2 genes, by an error in V-D-J joining mediated through normal recombinational mecha-

nisms. ' ' Although several breakpoint cluster regions have been identified in the chromosome band, l l q l 3 , that is involved in this translocation,43-53 a transcriptional unit has not been successfully isolated, and so attempts to study this gene have not progressed as rapidly as with bcl-2. It is interesting that this gene appears to be closely linked to two members of the fibroblast growth factor supergene family, int-2 and hst, because all three have been shown to co-amplify in some human breast tumors, 2 but the specific nature of the bcl-\ gene remains to be demonstrated. Putative Oncogenes in T-Cell Tumors Although nonrandom chromosome translocations are generally less frequent in T-cell neoplasms than in B-cell tumors,28 a number of rearrangements have recently been identified that involve T-cell receptor genes, particularly the alpha/delta locus, and donor sites that may harbor a previously unknown oncogene. None of these has yet been well characterized, but considerable preliminary information has been developed concerning three of them, which have been designated tcl-l, tcl-2, and tcl-3. The tcl-1 Gene. Two of the most common cytogenetic rearrangements described in low-grade T-cell leukemias and lymphomas, as well as in certain preneoplastic T-cell clones in patients with ataxia telangiectasia (AT), involve both the proximal (band ql 1) and distal (band q32) regions of chromosome #14. 3,28 These have been observed either as an inverted segment within the chromosome, with the breakpoints in these two bands, or as a "tandem" t( 14; 14) (qll;q32) translocation (Fig. 2D). Molecular studies have shown that in some instances these rearrangements have resulted in a juxtaposition between the T alpha/delta locus and the immunoglobulin heavy chain locus4'22 but that more frequently the breakpoint in band 14q32 is proximal to the immunoglobulin locus.13'37'50 This has led to the suggestion that an oncogene, important in T-cell neoplasia, might be located in this region, and this possibility has been strengthened by our recent demonstration that a t(7;14) (q35;q32) translocation in leukemic T-cells from a patient with AT involved the T-cell receptor beta-chain locus on chromosome #7 and the suspected region of chromosome #14 just proximal to the immunoglobulin heavy chain locus.51 Isolation and characterization of the putative oncogene at this site, designated tcl-\, have been complicated by the fact that these translocations frequently appear to involve small duplications and inversions in the immediate vicinity of the gene. 5051 Two known oncogenes that map to this region are akt-l, a homologue of the viral oncogene v-akt, and the elk-2 gene, a member of the ets oncogene superfamily.44 Whether either of these, or related sequences, constitutes the putative tcl-1 gene remains to be demonstrated.

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present initially as an aggressive lymphoma or leukemia with a complex karyotype that includes a t( 14; 18) translocation, suggesting evolution from an earlier, more indolent phase that was not apparent clinically.20 There is even some recent evidence indicating that this translocation, with activation of the bcl-2 gene, may contribute to the development of as many as one-third of the cases of Hodgkin's disease (Cossman J. Personal communication). It is becoming increasingly apparent that the bcl-2 gene plays a major role in the pathogenesis of a large proportion of all B-cell neoplasms. Some information has also been gained on the role of this gene in the regulation of normal growth within the lymphoid system. The two protein products of the gene have been partially characterized, and unlike the myc gene product that functions within the nucleus, the bcl-2 protein appears to have its growth regulatory role at the cell membrane. 24,57 Recent evidence has suggested that it functions as a guanosine triphosphate (GTP)-binding protein, involved in transmitting growth regulatory signals from external receptors to other sites within the cell.24 Like the myc gene, it is not expressed in resting normal human B-cells or T-cells, but is transcribed after antigenic or mitogenic stimulation, with subsequent reduction in activity as the cell progresses to DNA synthesis.46 The time course and transcriptional regulation are somewhat different than for the c-myc gene, but the pattern of activity strongly supports the view that this gene does play an important growth regulatory role in both normal B-cells and T-cells. It is interesting that involvement of this gene in T-cell tumors has not been described, and it is also not yet clear whether it has any function in cell lineages outside the lymphoid system.

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The tcl-2 Gene. Another relatively common translocation in T-cell neoplasms, particularly T-cell acute lymphocytic leukemia (ALL), is a t(ll;14) (pl3;qll) rearrangement (Fig. 2C). This also has been shown to involve the T delta receptor J region, 826 suggesting its origin from a recombinational error during T-cell differentiation, but the putative oncogene, located at band 11 p 13 and designated tcl-2, has not been characterized.16'22 Several laboratories have now cloned the breakpoint region from chromosome #1l, 8 - 26 and efforts are underway to isolate the transcriptional unit. It is interesting that deletions of this same region of chromosome #11 (band pi 3) are associated with Wilms' tumor, 28 but presumably this involves a tumor suppressor gene rather than the growth stimulatory oncogene postulated for T-cell neoplasms.

Discussion This brief survey indicates how much has been learned recently and how much remains to be learned concerning genes involved in the pathogenesis of human B-cell and T-cell neoplasms, and the importance of chromosomal translocation as a mechanism for altering their growth regulatory functions in a pathologic manner. One question often raised in connection with these translocations is the basis for their nonrandom observance: does this simply reflect positive selection for those random events that confer a growth advantage on the cell in which they occur, or are there specific mechanisms that increase the probability of breakage and of rejoining of particular sites in the genome? The answer appears to be a combination of the two, because within lymphocytic lineages there does appear to be an increased likelihood for certain rearrangements to take place. This is true of the immunoglobulin and T-cell receptor gene loci, where the recombinational and deletional events associated with normal differentiation definitely increase

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Several other T-cell tumors have been described in which there is a translocation involving the T alpha/delta locus and a more distal site on the short arm of chromosome #11, band pl5. 7 These are currently being investigated at the molecular level, and it will be of interest to see whether two different genes on 1 lp are in fact involved in these T-cell neoplasms and whether they have any structural or functional relationship. The tcl-5 Gene. The third putative oncogene in T-cell tumors that has been most actively studied thus far is the so-called tcl-3 gene, identified through a t( 10; 14) (q24;q 11) translocation (Fig. 2B). This rearrangement is associated with a subgroup of aggressive T-cell leukemias in children and young adults, often associated with a thymic lymphoma. 15 The chromosomal breakpoint has been cloned from a number of cases and shown to involve the T delta locus and a relatively circumscribed breakpoint cluster region at band 10q24.30 The cloned breakpoint region is already being used clinically as a probe for residual disease in treated patients whose leukemic cells carry this t( 10; 14) translocation. However, the coding region for the putative tcl-3 gene appears to be some distance from the breakpoint, as in some of the BL translocations, and has not yet been isolated and characterized. This region of chromosome #10 also includes the terminal transferase (TDT) gene and perhaps a locus involved in the pathogenesis of malignant melanoma, but neither seems to be closely associated with the putative tcl-3 gene. 3041 Other Sites Involved in T-Cell Neoplasms. Another translocation that has been observed occasionally in tumors with a T-cell phenotype is a t(l;14) (p32;ql 1) rearrangement. 33 Analysis of this translocation in a poorly differentiated leukemic cell line (DU.528) has demonstrated involvement of the T delta locus and a transcriptional unit at chromosome band lp32 that has been designated tcl-5 (and also SCL).5A9 Differential expression of this putative oncogene was observed in the DU.528 cell

line as compared with other T-cell and B-cell lines that did not contain this translocation,19 suggesting an alteration in function as the result of the juxtaposition with the T-cell receptor gene and presumably a role in the proliferation of this stem cell leukemia. The tcl-5 gene has been shown to be rearranged in a melanoma cell line carrying a deletion at lp32, 19 suggesting that it may have a growth regulatory role in nonhematopoietic lineages as well. As indicated previously, translocations involving the T beta locus, at 7q34-35, and the T gamma locus, at 7pl315, appear to be much less common than those involving the T alpha/delta region. A few cases have been reported, however, and several may involve additional new oncogenes that remain to be fully characterized. For example, a t(7;9) (q34;q34) translocation has been shown to involve the T beta locus and a previously unrecognized gene just proximal to the c-abl locus on chromosome #9.48-59 In addition, Mellentin and associates35 have recently isolated a putative oncogene, lyl-l, from a t(7;19) translocation in T-ALL, demonstrating that it codes for a DNA binding protein and that the translocation brings it into association with the constant region of the T beta gene. Related genes on chromosome #19 may be involved in the t(l;19) and t( 11; 19) translocations seen in leukemias of different phenotype.36 Thus, there are a number of clues to other growth regulatory genes that may be activated within the T-cell lineage by association with T-cell receptor genes. Further study will be required to determine how frequently they are involved in various T-cell tumors and also their normal role in lymphocyte function.

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progression of certain lymphomas, and the cells of these aggressive tumors may also carry point mutations in ras genes, as well as other somatic genetic abnormalities.38,49-61 In some circumstances, a critical submicroscopic alteration, allowing clonal expansion, may precede visible cytogenetic changes; in other instances a chromosomal translocation may be the initial event that allows a single cell to become the progenitor of an expanding neoplastic clone.38-39-4961 In either case, specific conditions within the host may help to determine the probability that a tumor will develop. Chronic infection with a virus such as Epstein-Barr virus (EBV) or human T-cell leukemia virus-1 (HTLV1) may provide an expanded pool of proliferating B-cells or T-cells and so increase the chance of a critical genetic alteration occurring within one cell and being expressed.40-53 Similarly, defects in the immune system, either inherited or acquired, may prevent the elimination of incipient neoplastic clones and enhance the likelihood of lymphoid tumors.25-40 This is also true of inherited defects in chromosomal stability that lead to chromosome breakage, as already noted in AT and Bloom's syndrome, and increase the chances that a tumorigenic translocation will occur.3-40 All of these factors must be considered when attempting to fully understand the pathogenesis of specific lymphoid tumors in particular patients and populations. The purpose of this review, however, is to emphasize the progress that has been made in recent years, through molecular analysis of nonrandom chromosome translocations, in beginning to identify both known and previously unknown genes significantly involved in the development of many of these neoplasms; and the very real hope that from the continuation of such studies will come additional applications in clinical diagnosis and prognosis, and ultimately specific therapy. References 1. Aghib D, Ottolenghi S, Rocchi M, et al. A novel type of e-myc translocation in a T lymphoma cell line. Ann NY Acad Sci 1987;511:338-342. 2. AH IU, Merlo G, Callahan R. et al. The amplification unit on chromosome 1 lq 13 in aggressive primary human breast tumors entails the bcl-1. int-2 and list loci. Oncogene 1989;4:89-92. 3. Aurias A, Dutrillaux B, Buriot D, et al. High frequencies of inversions and translocations of chromosomes 7 and 14 in ataxia telangiectasia. Mutat Res 1980;69:369-673. 4. Baer R. Chen K-C, Smith SD, et al. Fusion of an immunoglobulin variable gene and a T-cell receptor constant gene in the chromosome 14 inversion associated with T-cell tumors. Cell 1985;43: 705-709. 5. Begley CG, Apian PD. Davey MP. et al. Chromosomal translocation in a human leukemic stem-cell line disrupts the T-cell antigen receptor delta-chain diversity region and results in a previously unreported fusion transcript. Proc Natl Acad Sci USA 1989:86: 2031-2035. 6. Bishop JM. The molecular genetics of cancer. Science 1987:235: 305-311.

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the fragility of these sites. In any routine cytogenetic preparation from the peripheral blood of normal people, in which the metaphases are derived from mitogen-stimulated T-cells, the most frequently observed chromosome breakages involve the T-cell receptor gene loci: and translocations involving two of these genes, or between a Tcell receptor locus and an immunoglobulin gene locus, are the most common structural rearrangements seen.27 These same phenomena are enhanced in patients with an inherited defect in DNA repair, such as AT or Bloom's syndrome.3 In addition to this "physiologic" fragility, at least one other mechanism has been demonstrated that may further increase the probability that immunoglobulin genes and T-cell receptor genes will become associated with the cmyc gene or with putative oncogenes such as bcl-2, once chromosome breakage has occurred. This involves the short heptamer/nonamer signal sequences that have been identified adjacent to not only immunoglobulin and Tcell receptor genes, but also the c-myc gene and several of the newly defined oncogenes.53 As already noted, these short sequences provide the basis for the function of recombinase enzymes during normal lymphocyte differentiation, and the chromosome translocation events presumably reflect erroneous recognition by this enzyme system of homologous sequences in proximity to oncogenes. Thus, the fragility of immunoglobulin and T-cell receptor genes and the recognition sequences adjacent to oncogenes probably both contribute significantly to the nonrandom nature of these tumorigenic translocations. There are other "fragile sites" in the genome that have been identified, some present in most people and others that appear limited to certain families,60 but none of these has yet been specifically associated with the genetic loci that we have been discussing in connection with lymphoid tumors. 56 It will be of interest to see whether additional study can uncover any other factors relevant to the nonrandom nature of these chromosomal translocations. As a final consideration, it is also important to emphasize that these translocations, although critical in the pathogenesis of many human lymphoid tumors, frequently represent only one step in a sequence of somatic genetic alterations necessary to produce an aggressive malignancy and that various factors in the host may also be important in the ultimate development of a clinical neoplasm. It is now well documented, in both hematopoietic and nonhemic tumors, that the function of growth regulatory genes (oncogenes, tumor suppressor genes) can be critically altered not only by karyotypic rearrangements, but also by submicroscopic genetic changes such as point mutations and even potentially reversible variation in gene methylation.6-39 We have mentioned examples of sequential karyotypic changes in the clinical

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