Diffuse large B-cell lymphoma (DLBCL), a histologically well-defined subset of non-Hodgkin lymphoma, is clinically and genetically heterogenous. By G-banding, most cases showed complex hyperdiploid karyotypes and diverse cytogenetic abnormalities that included recurring and nonrecurring translocations, deletions, duplications, and marker chromosomes. While G-banding provided valuable leads to identification of specific rearrangements that enabled gene discovery and clinical correlations, many aberrations remained uncharacterized because of their complexity. The molecular cytogenetic technique spectral karyotyping (SKY), on the other hand, enables complete characterization of all aberrations in a tumor cell karyotype and, hence, precise quantitation of chromosome instability. We report here, for the first time, SKY analysis of a panel of 46 DLBCL cases previously analyzed by G-banding, ascertained at the Memorial Sloan-Kettering Cancer Center. This analysis provided a cytogenetic profile of DLBCL that was characterized by a higher level of instability, qualitatively as well as quantitatively, compared with G-banding. Thus, 551 breakpoints were detected by SKY, in contrast to the 295 by G-banding. Several new recurring breakpoints, translocations, and regions of gain and loss were identified, which included 13 breakpoints not previously identified by G-banding, 10 breakpoints that were underrepresented by G-banding, and 4 previously unrecognized translocations: der(14)t(3;14)(q21;q32), t(1;13)(p32;q14), t(1;7)(q21;q22), and der(6)t(6;8)(q11;q11). We identified new clinical associations involving recurring breakpoints detected by SKY. These studies emphasize the value of SKY analysis for redefinition of chromosomal instability in DLBCL to enhance gene discovery as well as clinical correlation analysis.

Introduction

Diffuse large B-cell lymphoma (DLBCL) is the most common form of non-Hodgkin lymphoma (NHL), representing 40% of all adult cases, and comprises several entities characterized by genetic, morphologic, and clinical features. This heterogeneity has been suggested to derive from the developmental stage of the B cell at which it is transformed—ie, germinal center or post–germinal center B cell—as well as transformation to DLBCL of a lower histologic grade.1-3 Clinically, less than 45% of patients with DLBCL achieve complete remission and become cured, while the remaining succumb to disease. However, very few biologic or genetic markers that define the heterogeneity or precisely predict clinical behavior are available.

The cytogenetic features of DLBCL have been partly defined by G-banding analysis.4 Approximately 50% of cases exhibit chromosomal translocations involving one of the IG gene sites, which lead to deregulated expression of a variety of genes. The remaining cases display diverse types of chromosomal rearrangements that include translocations, deletions, and undefined aberrations such as additions and marker chromosomes.4 The recent introduction of spectral karyotyping (SKY) has made possible the identification of each of the human chromosomes by a different color, facilitating precise identification of all rearrangements in a tumor karyotype.5 A limited number of hematopoietic tumors such as multiple myeloma6,7 and acute leukemia8-11have been studied by SKY. These studies showed that SKY detected a several-fold increase in random as well as nonrandom chromosomal aberrations compared with G-banding. So far, there have been no attempts to examine clinical correlations based on SKY data in hematopoietic or other neoplasms. There have also been no SKY-based studies of NHL. Here we report SKY analysis of a panel of 46 DLBCLs ascertained at the Memorial Sloan-Kettering Cancer Center. We identified 13 new recurring breakpoints; 4 new recurring translocations; a number of cryptic, balanced, and unbalanced translocations; and regions of gain and loss. We identified new clinical associations involving recurring breakpoints detected by SKY.

Materials and methods

Tumor ascertainment

The ongoing ascertainment of consecutive NHL cases for cytogenetic analysis initiated at the Memorial Sloan-Kettering Cancer Center in 1984 has been described.12,13 A subset of 46 cases with DLBCL for which archived methanol-acetic acid–fixed metaphase preparations were available were selected from this ascertainment for SKY analysis. All patients were histologically reclassified according to the World Health Organization classification14 and staged by the Ann Arbor staging system. None of the cases had histologic evidence or were suspected to have preexisting low-grade lymphoma. Of the 46 cases, 29 were ascertained pretreatment and 17 posttreatment.

Cytogenetic and SKY analysis

Cytogenetic analysis was performed on metaphase spreads obtained from lymph node biopsy (43), bone marrow (2), or pleural effusion (1) as previously described.12 Clonal chromosomal abnormalities were described according to the International System of Human Cytogenetic Nomenclature.15 SKY analysis was performed as previously described.5,6 SKY images were acquired with an SD300 Spectracube (Applied Spectral Imaging, Migdal Ha-Emck, Israel) mounted on a Nikon Eclipse E800 microscope using a custom-designed optical filter (SKY-1) (Chroma Technology, Brattleboro, VT). For each case, 5 to 30 metaphases were analyzed. Breakpoints on G-banded chromosomes were scored according to previously established criteria.4 Breakpoints on the SKY-painted chromosomes were determined by comparison with corresponding DAPI and G-banded karyotype of the same tumor as previously described.6 

Fluorescence in situ hybridization analysis

Fluorescence in situ hybridization (FISH) with whole chromosome painting probes for chromosomes 1, 3, and 14 (Vysis, Downers Grove, IL) was performed according to manufacturer-supplied protocols. To confirm 14q32 rearrangements, the PAC clone 120H10, derived from the region immediately telomeric to IGHV, was used in FISH analysis.16 For each case, 5 to 13 metaphases were analyzed using the Quips Pathvision (Applied Imaging, Santa Clara, CA).

Statistical analysis of data

Differences in percentage of variables were tested for significance using the 2-tailed Fischer exact test. Survival times were estimated by Kaplan-Meier analysis and compared by log-rank test. Cytogenetic variables included 32 recurring break sites. Clinical variables included age, sex, performance status, stage, B symptoms, bulky disease, lactate dehydrogenase, extranodal disease, number of extranodal sites, international prognostic index,17 and response. Time to treatment failure was measured from time of initiation of treatment until disease progression. Overall survival was measured from initiation of treatment (45 cases) or the date of diagnosis, if the patient was observed (1 case), until the date last seen.

Results

Clinical and histologic features of patients

The clinical and histologic features of the 46 cases with reconfirmed DLBCL at diagnosis are summarized in Table1. Morphologic variants included 28 centroblastic, 5 plasmablastic, 4 immunoblastic, 3 pleomorphic, and 5 T-cell–rich/histiocyte-rich lymphomas. The 1 case with primary effusion lymphoma (2612) could not be subtyped. Of the 46 cases, 3 (2293, 2473, 1800) were primary mediastinal large B-cell lymphomas. The male:female ratio was 1:1.19 (21 males, 25 females). The age range was 21 to 85 years (median, 57.5). Eight patients had stage I disease, 11 had stage II disease, 10 had stage III disease, and 16 had stage IV disease. Of the 46 patients, 28 were at low or low-intermediate risk, whereas 14 were at high or high-intermediate risk. Clinical data were not available for 1 patient (980), and 3 patients (980, 2358, 2433) were lost to follow-up. All patients, with the exception of cases 2202 and 2627, received anthracycline-containing combination chemotherapy as part of their initial treatment.

Table 1.

Clinical and histologic features of the 46 DLBCLs

Subject no. Case Age/Sex Rx Study Specimen Morphology Stage IPI Response TTP, mo OS, mo Status 
980 32/M UT ID Cervical node CB — — — — — LF 
1942 29/M UT ID Colonic mass CB II XEA LR CR 47.0 47.0 NED 
1965 58/F UT ID Jugular node CB II A LR CR 40.0 40.0 NED 
1997 44/M UT ID Axillary node CB III B HR CR 9.6 16.3 DOD 
2045 68/F UT ID Tonsil CB II EA LR CR 39.4 39.4 NED 
2180 33/M UT ID Stomach CB I EA LR CR 15.4 15.4 NED 
2202 82/M UT ID Retroperitoneal mass CB III A LR NR* 4.8 4.8 DOD 
2270 59/M UT PRD Inguinal node CB I A LR NR 2.9 28.8 NED 
2326 50/M UT ID Neck node CB III A LR CR 12.9 12.9 NED 
10 2358 74/F UT ID Neck mass TCR/HR IV A HR — — — LF 
11 2417 21/F UT ID Cervical node PL IV A LR CR 6.8 8.1 AWD 
12 2438 57/F UT ID Inguinal node CB IV B HR CR 11.7 11.7 NED 
13 2447 65/F UT ID Right thigh PB II EX HR CR 9.4 9.4 NED 
14 2451 71/M UT ID Inguinal node CB IV A HR CR 10.3 10.3 NED 
15 2453 64/F UT ID Axillary node TCR/HR IV A HR POD 2.7 5.6 DOD 
16 2511 55/M UT ID Axillary node TCR/HR III A LR CR 6.8 6.8 NED 
17 2526 77/F UT ID Cervical node PB I A LR CR 5.1 5.1 NED 
18 2536 72/F UT ID Axillary node CB IV A HR CR 4.8 4.8 AWD 
19 2545 83/M UT ID Inguinal node TCR/HR III A LR No Rx 5.3 5.3 AWD 
20 2625 61/F UT ID Neck mass CB I A LR CR 5.1 5.1 NED 
21 2627 82/F UT ID Spleen CB I A HR CR* 4.3 4.3 NED 
22 2629 59/F UT ID Hepatic artery node CB II A LR CR 5.6 5.6 AWD 
23 2609 53/M UT ID Bone marrow IM IV B LR POD 4.6 5.9 DOD 
24 2612 52/F UT ID Ascitic fluid — IV A LR CR 18.7 18.7 NED 
25 2433 73/F UT ID Jugular node TCR/HR III A HR — — — LF 
26 2411 60/M UT ID Neck mass IM IV A LR CR 8.5 8.5 NED 
27 2055 71/F UT ID Epiglottis PB I EA LR CR 25.7 25.7 DOD 
28 2308 77/F UT ID Pelvic node CB III BE HR CR 17.8 17.8 NED 
29 2549 85/F UT ID Tonsil CB IV A LR CR 5.8 5.8 NED  
30 2216 35/F 2nd RP Omentum PL II XA LR NR 4.4 10.2 DOD 
31 2293 37/F 2nd RP Level II neck node CB II XA LR CR 20.6 58.8 AWD 
32 2616 43/M PRD Bone marrow CB IV A LR POD 4.6 10.1 DOD 
33 2473 37/M PRD Supraclavicular node CB II XEEB HR POD 6.7 14.7 NED 
34 2631 57/M 2nd RP Antecubital fossa CB IV A HR CR 11 24.2 AWD 
35 2369 39/M PRD Pancreas IM IV XB HR NR 4.7 10.5 DOD  
36 2552 46/M 2nd RP Cervical node CB III A LR CR 11.7 22.8 AWD 
37 2498 57/F 1st RP Axillary node IM I A LR CR 12.3 NED  
38 2005 42/F 1st RP Axillary node PL III ARE LR CR 8.7 37.8 DOD 
39 1778 37/F 2nd RP Jugular node CB IV A HR CR 15.7 30.1 DOD  
40 2020 63/M 1st RP Spleen CB IV A HR CR 16.5 36.8 DOD 
41 1983 66/F 1st RP Scalene node CB I A LR CR 8.6 33.0 DOD 
42 1800 28/M PRD Supraclavicular node CB II A LR PR 5.6 45.7 NED 
43 2006 23/M 1st RP Axillary node PB III A LR CR 7.2 12.1 DOD  
44 2287 48/M 2nd RP Scalene node CB IV A HR PR 9.1 16.7 DOD 
45 1686 61/F 1st RP Cervical node PB II XA LR CR 243.4 319.2 NED 
46 2635 82/F 2nd RP Axillary node CB II A HR CR 155.9 226.2 DOD 
Subject no. Case Age/Sex Rx Study Specimen Morphology Stage IPI Response TTP, mo OS, mo Status 
980 32/M UT ID Cervical node CB — — — — — LF 
1942 29/M UT ID Colonic mass CB II XEA LR CR 47.0 47.0 NED 
1965 58/F UT ID Jugular node CB II A LR CR 40.0 40.0 NED 
1997 44/M UT ID Axillary node CB III B HR CR 9.6 16.3 DOD 
2045 68/F UT ID Tonsil CB II EA LR CR 39.4 39.4 NED 
2180 33/M UT ID Stomach CB I EA LR CR 15.4 15.4 NED 
2202 82/M UT ID Retroperitoneal mass CB III A LR NR* 4.8 4.8 DOD 
2270 59/M UT PRD Inguinal node CB I A LR NR 2.9 28.8 NED 
2326 50/M UT ID Neck node CB III A LR CR 12.9 12.9 NED 
10 2358 74/F UT ID Neck mass TCR/HR IV A HR — — — LF 
11 2417 21/F UT ID Cervical node PL IV A LR CR 6.8 8.1 AWD 
12 2438 57/F UT ID Inguinal node CB IV B HR CR 11.7 11.7 NED 
13 2447 65/F UT ID Right thigh PB II EX HR CR 9.4 9.4 NED 
14 2451 71/M UT ID Inguinal node CB IV A HR CR 10.3 10.3 NED 
15 2453 64/F UT ID Axillary node TCR/HR IV A HR POD 2.7 5.6 DOD 
16 2511 55/M UT ID Axillary node TCR/HR III A LR CR 6.8 6.8 NED 
17 2526 77/F UT ID Cervical node PB I A LR CR 5.1 5.1 NED 
18 2536 72/F UT ID Axillary node CB IV A HR CR 4.8 4.8 AWD 
19 2545 83/M UT ID Inguinal node TCR/HR III A LR No Rx 5.3 5.3 AWD 
20 2625 61/F UT ID Neck mass CB I A LR CR 5.1 5.1 NED 
21 2627 82/F UT ID Spleen CB I A HR CR* 4.3 4.3 NED 
22 2629 59/F UT ID Hepatic artery node CB II A LR CR 5.6 5.6 AWD 
23 2609 53/M UT ID Bone marrow IM IV B LR POD 4.6 5.9 DOD 
24 2612 52/F UT ID Ascitic fluid — IV A LR CR 18.7 18.7 NED 
25 2433 73/F UT ID Jugular node TCR/HR III A HR — — — LF 
26 2411 60/M UT ID Neck mass IM IV A LR CR 8.5 8.5 NED 
27 2055 71/F UT ID Epiglottis PB I EA LR CR 25.7 25.7 DOD 
28 2308 77/F UT ID Pelvic node CB III BE HR CR 17.8 17.8 NED 
29 2549 85/F UT ID Tonsil CB IV A LR CR 5.8 5.8 NED  
30 2216 35/F 2nd RP Omentum PL II XA LR NR 4.4 10.2 DOD 
31 2293 37/F 2nd RP Level II neck node CB II XA LR CR 20.6 58.8 AWD 
32 2616 43/M PRD Bone marrow CB IV A LR POD 4.6 10.1 DOD 
33 2473 37/M PRD Supraclavicular node CB II XEEB HR POD 6.7 14.7 NED 
34 2631 57/M 2nd RP Antecubital fossa CB IV A HR CR 11 24.2 AWD 
35 2369 39/M PRD Pancreas IM IV XB HR NR 4.7 10.5 DOD  
36 2552 46/M 2nd RP Cervical node CB III A LR CR 11.7 22.8 AWD 
37 2498 57/F 1st RP Axillary node IM I A LR CR 12.3 NED  
38 2005 42/F 1st RP Axillary node PL III ARE LR CR 8.7 37.8 DOD 
39 1778 37/F 2nd RP Jugular node CB IV A HR CR 15.7 30.1 DOD  
40 2020 63/M 1st RP Spleen CB IV A HR CR 16.5 36.8 DOD 
41 1983 66/F 1st RP Scalene node CB I A LR CR 8.6 33.0 DOD 
42 1800 28/M PRD Supraclavicular node CB II A LR PR 5.6 45.7 NED 
43 2006 23/M 1st RP Axillary node PB III A LR CR 7.2 12.1 DOD  
44 2287 48/M 2nd RP Scalene node CB IV A HR PR 9.1 16.7 DOD 
45 1686 61/F 1st RP Cervical node PB II XA LR CR 243.4 319.2 NED 
46 2635 82/F 2nd RP Axillary node CB II A HR CR 155.9 226.2 DOD 

Rx indicates treatment; IPI, international prognostic index; TTP, time to progression; OS, overall survival; UT, untreated; T, treated; ID, initial diagnosis; PRD, primary refractory disease; RP, relapse; CB, centroblastic; TCR/HR, T-cell-rich/histiocyte-rich; PL, pleomorphic; PB, plasmablastic; IM, immunoblastic; HR, high and high-intermediate risk; LR, low and low-intermediate risk; CR, complete remission; NR, no response; POD, progressive disease; PR, partial remission; DOD, died of disease; NED, no evidence of disease; LF, lost to follow-up; AWD, alive with disease.

*

Did not receive anthracycline-containing combination chemotherapy as part of their initial treatment.

Primary effusion lymphoma.

Mediastinal large B-cell lymphoma.

Chromosome involvement and breakpoints identified by SKY analysis versus G-banding

Figure 1 shows the percentage involvement of each chromosome. With the exception of chromosome 1, all chromosomes were affected more frequently in the SKY analysis than in the G-banding analysis; however, the difference in involvement only of chromosomes 2, 5, 22, and X was significant (P ≤ .05).

Fig. 1.

Involvement of chromosomes by SKY versus G-banding.

The difference was significant for chromosomes 2, 5, 22, and X.

Fig. 1.

Involvement of chromosomes by SKY versus G-banding.

The difference was significant for chromosomes 2, 5, 22, and X.

Figure 2 shows the distribution of breakpoints, and a representative SKY karyotype with multiple and cryptic rearrangements is shown in Figure3A. A total of 551 breakpoints were identified by SKY, compared with 295 by G-banding. SKY analysis confirmed 234 (80%) breakpoints identified by G-banding and revised 53 (20%). Most of the revisions comprised breakpoints that were classified by G-banding as terminal, which were identified by SKY to be interstitial. Breakpoints on chromosomes 1, 2, 3, 6, and 8 were misidentified most frequently. In addition, SKY analysis identified 264 new breakpoints, including 60 that could not be precisely assigned to a band. The 551 breakpoints identified by SKY analysis were located at 171 bands and, of these, 125 (73%) were recurring (≥ 2 cases). In the case of G-banding, the 295 breakpoints were located at 144 bands and, of these, 63 (44%) were recurring. While the number of recurring sites noted in more than 10% of the cases was 32 in SKY analysis, it was only 10 in G-banding. With the exception of 1q32, 6q23, 8q24, 14q32, and 19q13, all the recurring sites (27) were noted to be involved more frequently in SKY analysis than in G-banding. Breaks at 2 sites, 16q11-13 and 17q11-21, were significantly more frequent in SKY analysis (P ≥ .05). Sites most frequently identified to be involved in rearrangement in SKY analysis were 14q32 (57%), 6q11-15 (30%), 18q21 (30%), 1q11-21 (28%), 3q27 (28%), and 9p13 (22%); sites most frequently identified in G-banding analysis were 14q32 (57%), 1q11-21 (24%), 3q27 (20%), 6q11-15 (17%), and 18q21 (15%).

Fig. 2.

Distribution of breakpoints.

Idiogram shows distribution of breakpoints identified by G-banding (left) and SKY (right). The number of breakpoints in each chromosome that were identified by SKY, but could not be precisely assigned to a band, are noted on top of the chromosomes. Each dot represents a single breakpoint. ★ represents 10 breakpoints; ✖ represents a single breakpoint misidentified by G-banding; ✜ represents a single breakpoint misidentified by G-banding but within the same chromosome.

Fig. 2.

Distribution of breakpoints.

Idiogram shows distribution of breakpoints identified by G-banding (left) and SKY (right). The number of breakpoints in each chromosome that were identified by SKY, but could not be precisely assigned to a band, are noted on top of the chromosomes. Each dot represents a single breakpoint. ★ represents 10 breakpoints; ✖ represents a single breakpoint misidentified by G-banding; ✜ represents a single breakpoint misidentified by G-banding but within the same chromosome.

Fig. 3.

Representative karyotypes illustrating the various chromosomal rearrangements identified in SKY analysis.

(A) SKY karyotype of case 2612 with primary effusion lymphoma showing several multiple complex and cryptic translocations, notably, t(3;14)(q27;q32) and der(14)t(1;14)(?;q32). The display colors are shown on the left and the classification colors on the right. (B) Confirmation of t(3;14)(q27;q32) by FISH using PAC probe 120H10 (14q32). The lack of green signal on chromosome 1 confirms the nonreciprocal nature of der(14)t(1;14). (C) The same metaphase was washed and rehybridized with whole chromosome painting probe for chromosomes 1 (green) and 14 (red). Presence of green signal on chromosome 14 confirms der(14)t(1;14)(?;q32). (D) Partial karyotypes (from left to right) showing the 3 new and 2 potentially new recurring translocations. Inverted DAPI band image of the involved chromosome is shown on the left and the classification color on the right.

Fig. 3.

Representative karyotypes illustrating the various chromosomal rearrangements identified in SKY analysis.

(A) SKY karyotype of case 2612 with primary effusion lymphoma showing several multiple complex and cryptic translocations, notably, t(3;14)(q27;q32) and der(14)t(1;14)(?;q32). The display colors are shown on the left and the classification colors on the right. (B) Confirmation of t(3;14)(q27;q32) by FISH using PAC probe 120H10 (14q32). The lack of green signal on chromosome 1 confirms the nonreciprocal nature of der(14)t(1;14). (C) The same metaphase was washed and rehybridized with whole chromosome painting probe for chromosomes 1 (green) and 14 (red). Presence of green signal on chromosome 14 confirms der(14)t(1;14)(?;q32). (D) Partial karyotypes (from left to right) showing the 3 new and 2 potentially new recurring translocations. Inverted DAPI band image of the involved chromosome is shown on the left and the classification color on the right.

To identify new recurring breakpoints, all breakpoints identified in SKY and G-banding analyses were compared. Thirteen new recurring breakpoints were identified by SKY, with 16q11-13 (13%), 12p11, and 11p11 (9% each) as the most frequent (Table2). Because the number of cases in the present cohort was small, the 551 breakpoints identified by SKY also were compared with the 1021 breakpoints identified by G-banding in our previously reported cohort of 363 consecutively ascertained DLBCLs.4 All the new recurring sites identified in the present study by SKY were either nonrecurring or were undetected by G-banding in this larger cohort.

Table 2.

New recurring breakpoints and their corresponding incidence by G-banding in this cohort and the previous cohort (Cigudosa et al, 1999)

Subject no. Site SKY G-banding  
This cohort Cigudosa et al 
16q11-13 6  (13.0) — 1  (0.46)  
12p11 4  (8.7) — 1  (0.46)  
11p11 4  (8.7) 1  (2.2) —  
18p11 3  (6.5) 1  (2.2) —  
20q11-13 3  (6.5) 1  (2.2) —  
17q11 3  (6.5) — —  
4p11 2  (4.3) — —  
4p14 2  (4.3) 1  (2.2) —  
5q11 2  (4.3) — 1  (0.46)  
10 16p11 2  (4.3) — —  
11 17q23 2  (4.3) — —  
12 18q11 2  (4.3) — —  
13 Xp11 2  (4.3) — — 
Subject no. Site SKY G-banding  
This cohort Cigudosa et al 
16q11-13 6  (13.0) — 1  (0.46)  
12p11 4  (8.7) — 1  (0.46)  
11p11 4  (8.7) 1  (2.2) —  
18p11 3  (6.5) 1  (2.2) —  
20q11-13 3  (6.5) 1  (2.2) —  
17q11 3  (6.5) — —  
4p11 2  (4.3) — —  
4p14 2  (4.3) 1  (2.2) —  
5q11 2  (4.3) — 1  (0.46)  
10 16p11 2  (4.3) — —  
11 17q23 2  (4.3) — —  
12 18q11 2  (4.3) — —  
13 Xp11 2  (4.3) — — 

To determine the effect of therapy on the incidence of breakpoints, data from treated and untreated cases were compared. The mean number of breaks per case was higher in treated cases (16) than untreated cases (9). Most (24 of 32) of the recurring breakpoints were noted to be affected more frequently in the treated cases; however, the difference was significant only for breaks at 17q11-21 (P ≤ .05).

Chromosome structural aberrations identified by SKY analysis versus G-banding

Multiple complex structural aberrations (translocations, deletions, duplications, isochromosomes, inversions, insertions, additions, and markers) were present in 85% of the cases in this study. Of the 46 cases with clonal chromosome abnormalities, the G-banded karyotype of only 4 cases remained unchanged after SKY analysis. In the remaining 42 cases, SKY provided additional cytogenetic information. G-banding analysis detected a total of 240 aberrations, of which 98 were undefined (73 additions, 24 markers, and 1 ring chromosome). All the undefined aberrations were resolved by SKY. Of the 240 aberrations, SKY confirmed 74 and revised 166. Of the marker and addition chromosomes detected by G-banding, 75% represented translocations. In addition, SKY identified 79 new aberrations. A total of 211 translocations were identified by SKY. Of these, 64 were new, including 30 that were cryptic. Several of the latter were confirmed by FISH using whole chromosome painting or locus-specific probes (Figure 3B,C).

While rearrangements affecting one of the IG gene sites were the most common aberrations detected by both the techniques, they were detected at a slightly higher rate by SKY (66%) than by G-banding (61%) (Table 3). Of the 25 cases with translocations affecting 14q32 detected by SKY, 6 had t(3;14)(q27;q32), 5 had t(14;18)(q32;21), 4 had t(9;14)(p13;q32), and 1 each had t(8;14)(q24;q32) and t(8;14;18)(q24;q32;q21), while 10 had other 14q32-associated translocations (Table4). The incidence of translocations affecting 14q32 detected by G-banding analysis was the same as that detected by SKY (25 cases); however, 3 of the 14q32 breakpoints were misidentified by G-banding. Translocations affecting 22q11 were noted in 7 cases by SKY, compared with 2 by G-banding. Of the former 7 cases, 3 cases had t(3;22)(q27;q11) and 4 had other 22q11 translocations (Table 4). Translocations involving 2p11 were less frequent and were detected in 2 cases by SKY alone (Table 4).

Table 3.

Frequency of IG gene site and 3q27 involvement detected by SKY and by G-banding

Translocation Number of cases (%) 
SKY G-banding Misidentified by G-banding 
14q32    
 t(3;14)(q27;q32) 6  (13.04) 5  (10.87)  
 t(14;18)(q32;q21) 5  (10.87) 5  1  (2.17)  
 t(8;14;18)(q24;q32;q21) 1  (2.17) —  
 t(8;14)(q24;q32) 1  (2.17) 1  (2.17)  
 t(9;14)(p13;q32) 4  (8.70) 6  (13.04) 2  (4.35)  
 Other 14q323-150 10  (21.74) 8  (17.39) 1  (2.17)  
 Overall 14q32 26  (56.52)3-151 26  (56.52)3-151 4  (8.70) 
22q11    
 t(3;22)(q27;q11) 3  (6.52) 2  (4.35)  
 Other 22q11 4  (8.70) —  
 Overall 22q11 7  (15.23) 2  (4.35)  
2p11 2  (4.35) —  
Overall IG gene site 31  (67.40)3-152 28  (60.87) 3  (6.52) 
3q27    
 Other 3q27 5  (10.87) 2  (4.35)  
 Overall 3q27 13  (28.26)3-153 9  (19.55)  
Translocation Number of cases (%) 
SKY G-banding Misidentified by G-banding 
14q32    
 t(3;14)(q27;q32) 6  (13.04) 5  (10.87)  
 t(14;18)(q32;q21) 5  (10.87) 5  1  (2.17)  
 t(8;14;18)(q24;q32;q21) 1  (2.17) —  
 t(8;14)(q24;q32) 1  (2.17) 1  (2.17)  
 t(9;14)(p13;q32) 4  (8.70) 6  (13.04) 2  (4.35)  
 Other 14q323-150 10  (21.74) 8  (17.39) 1  (2.17)  
 Overall 14q32 26  (56.52)3-151 26  (56.52)3-151 4  (8.70) 
22q11    
 t(3;22)(q27;q11) 3  (6.52) 2  (4.35)  
 Other 22q11 4  (8.70) —  
 Overall 22q11 7  (15.23) 2  (4.35)  
2p11 2  (4.35) —  
Overall IG gene site 31  (67.40)3-152 28  (60.87) 3  (6.52) 
3q27    
 Other 3q27 5  (10.87) 2  (4.35)  
 Overall 3q27 13  (28.26)3-153 9  (19.55)  
F3-150

One case had dup(14)(q11q32).

F3-151

Both the homologs of chromosome 14 were involved in 1 case.

F3-152

Two IG gene sites were affected in 4 cases (14q32 and 22q11 in 3 cases and 14q32 and 2p11 in 1 case).

F3-153

One case had both t(3;14)(q27;q32) and t(3;22)(q27;q11).

Table 4.

Translocations affecting IG gene sites other than those involving 3q27, 8q24, and 18q21

IG gene sites Case Translocation/derivative 
14q32 2545 der(14)t(3;14)(q21;q32) 
 2433 der(1)ins(1;12)(p34-36;q13q24)t(1;3;14)
(q32;q21;q32) 
 2552 der(3)t(3;21)(p13;q11)t(3;14)(q21;q11)
t(14;17)(q32;?)t(4;17)(?;?) 
 2612 der(14)t(1;14)(p?;q32)  
 2629 t(1;14)(q42;q32) 
 2438 der(11;14)(p11;q32) 
 2453 der(14)t(19;14)(q12;q32) 
 1800 der(14)t(14;15)(q32;q?) 
 980 der(14)t(2;14)(p11;q32) 
22q11 2473 t(16;22)(p11;q11) 
 2552 t(19;22)(q13.3;q11) 
 2020 der(12)t(12;22)(p11;q11) 
 1778 der(22)t(6;22)(p11;q11) 
2p11 980 der(14)t(2;14)(p11;q32) 
 2616 der(17)t(2;17)(p11;q25) 
IG gene sites Case Translocation/derivative 
14q32 2545 der(14)t(3;14)(q21;q32) 
 2433 der(1)ins(1;12)(p34-36;q13q24)t(1;3;14)
(q32;q21;q32) 
 2552 der(3)t(3;21)(p13;q11)t(3;14)(q21;q11)
t(14;17)(q32;?)t(4;17)(?;?) 
 2612 der(14)t(1;14)(p?;q32)  
 2629 t(1;14)(q42;q32) 
 2438 der(11;14)(p11;q32) 
 2453 der(14)t(19;14)(q12;q32) 
 1800 der(14)t(14;15)(q32;q?) 
 980 der(14)t(2;14)(p11;q32) 
22q11 2473 t(16;22)(p11;q11) 
 2552 t(19;22)(q13.3;q11) 
 2020 der(12)t(12;22)(p11;q11) 
 1778 der(22)t(6;22)(p11;q11) 
2p11 980 der(14)t(2;14)(p11;q32) 
 2616 der(17)t(2;17)(p11;q25) 

Translocations involving 3q27 were the next most frequent after those affecting the IG gene sites. Eight cases exhibited t(3;14)(q27;q32) or t(3;22)(q27;q11), and 5 involved other sites (Table5). Only 9 of these translocations were detected by G-banding. Translocations affecting 1q11-21 (15%), 1p32-36 (11%), and 1q42-44 (9%) were also common and noted equally in SKY and G-banding analyses. Of note were 10 other non-IGH sites identified by SKY that were promiscuously involved in balanced and unbalanced translocations: 2q31, 12q11-13 (15%), 1p11-13, 2p13, 7q11, 16q11-13, 17q11-21 (13% each), 3p21, 7q22, and 15q13-15 (11% each). These sites were either undetected or underrepresented in G-banding analysis.

Table 5.

Translocations affecting band 3q27 other than those involving 14q32 and 22q11

Case Translocation/derivative 
1778 t(3;5)(q27;p13) 
2451 t(3;12)(q27;q22)  
2552 der(3)t(3;6)(q27;p21) 
2545 der(3)t(3;7)(p21;p13)t(X:3)(q27;q27) 
2308 der(3)t(2;3)(?;q27)del(3)(p21) 
Case Translocation/derivative 
1778 t(3;5)(q27;p13) 
2451 t(3;12)(q27;q22)  
2552 der(3)t(3;6)(q27;p21) 
2545 der(3)t(3;7)(p21;p13)t(X:3)(q27;q27) 
2308 der(3)t(2;3)(?;q27)del(3)(p21) 

SKY is known for its ability to unambiguously characterize complex chromosomal aberrations and thus facilitate identification of new or hidden recurring translocations. Indeed, 4 new recurring translocations were identified in 2 cases each (Table 6) (Figure 3D). The translocations t(3;14)(q21;q32) and t(1;7)(q21;q22) were also identified by G-banding; the former was nonrecurring. Interestingly, 3 translocations, t(1;13)(p32;q14), t(1;7)(q21;q22), and der(6)t(6;8)(q11;q11), were observed only in cases without t(3q27) or t(14q32). In addition, 2 potentially new recurring translocations were identified in 2 cases each: t(5;16)(?;q11-12) and t(19;22)(q13;q11-13). A search in our database revealed another t(14;18)(q32;q21)-negative follicular large cell lymphoma with a t(19;22)(q13;q13). The ability to precisely delineate chromosomal abnormalities also facilitated the correct identification of known specific recurring translocations. Thus, in 2 cases, SKY revealed the presence of t(3;14)(q27;q32) and t(3;22)(q27;q11), respectively, that were overlooked by G-banding due to poor morphology. In 3 other cases, t(9;14)(p13;q32) (2 cases) and t(14;18)(q32;q21) (1 case), identified by G-banding, were revised by SKY and shown to be otherwise.

Table 6.

New recurring translocations in DLBCL

Case SKY G-banding t(14q32) t(3q27) 
New     
2545 der(14)t(3;14)(q21;q32) der(14)t(4;14)(q21;q32) 
2433 der(1)ins(1;12)(p34-36;
q13q24)t(1;3;14)(q32;
q21;q32) 
der(1)add(1)(p36)t(1;3;
14)(q32;q21;q32) 
− 
2631 der(1)t(1;13)(p32;q14)t
(X;1)(?;q32) 
der(1)add(1)(p32)add(1)
(q32) 
− − 
2369 t(1;13)(p32;q14) add(1)(p34) − − 
1983 der(1)t(1;7)(q21;q22) der(1)t(1;7)(q21;q22) − − 
2616 t(1;7)(q21;q22) t(1;7)(q21;q22) − − 
2005 der(6)t(6;8)(q11;q11) add(6)(q23) − − 
2498 der(6)t(6;8)(q11;q11) del(6)(q21q23) − − 
Potentially new     
2438 der(16)t(5;16)(?;q11) mar − 
1983 der(16)t(5;16)(?;q11-12) inv(16)(q22q24) − − 
2552 t(19;22)(q13;q11-13) add(19)(q13) − 
2616 der(22)t(19;22)(q?;q13) 22 − − 
 9816-150 − t(19;22)(q13;q13) − − 
Case SKY G-banding t(14q32) t(3q27) 
New     
2545 der(14)t(3;14)(q21;q32) der(14)t(4;14)(q21;q32) 
2433 der(1)ins(1;12)(p34-36;
q13q24)t(1;3;14)(q32;
q21;q32) 
der(1)add(1)(p36)t(1;3;
14)(q32;q21;q32) 
− 
2631 der(1)t(1;13)(p32;q14)t
(X;1)(?;q32) 
der(1)add(1)(p32)add(1)
(q32) 
− − 
2369 t(1;13)(p32;q14) add(1)(p34) − − 
1983 der(1)t(1;7)(q21;q22) der(1)t(1;7)(q21;q22) − − 
2616 t(1;7)(q21;q22) t(1;7)(q21;q22) − − 
2005 der(6)t(6;8)(q11;q11) add(6)(q23) − − 
2498 der(6)t(6;8)(q11;q11) del(6)(q21q23) − − 
Potentially new     
2438 der(16)t(5;16)(?;q11) mar − 
1983 der(16)t(5;16)(?;q11-12) inv(16)(q22q24) − − 
2552 t(19;22)(q13;q11-13) add(19)(q13) − 
2616 der(22)t(19;22)(q?;q13) 22 − − 
 9816-150 − t(19;22)(q13;q13) − − 
F6-150

Identified from our database. The case was histologically diagnosed as follicular large cell lymphoma and was t(14;18)-negative.

Gains and losses of chromosomes

Resolution of all derived, addition, and marker chromosomes by SKY resulted not only in the detection of a 2-fold increase in the gains and losses but also in their correct identification. Fifty percent of deletions and 11% of translocations identified by G-banding were found to represent translocations and duplications, respectively, by SKY. Figure 4 shows the regions of partial- or whole-arm gains and losses identified by SKY. Gains (89%) were noted more frequently than losses (80%). While gains were noted equally in both treated and untreated groups (90% vs 88%), losses were more frequent in the treated group than the untreated (94% vs 72%). All the chromosomes were affected by gains. Chromosomes frequently involved in gains were 7 (39%), 1 (37%), 3 (35%), 12 (33%), 2 (30%), and 18 (28%). Region of common cytogenetic gain in these chromosomes comprised 7q11, 7q22, 1q11-23, 3q21-29, 12q13-15, 12q22-24, 2q21, 2p13-21, and 18q21-23. Most of the chromosomes also were affected by deletions, with del(6q) (35%) being the most frequent. Five regions of common cytogenetic deletions, in decreasing order of incidence, were observed: 6q23, 6q21, 6q25-27, 6q15, and 6q11-13. Other chromosomes involved in deletions were 2 (22%), 13q (17%), 1 (15%), and 17p (13%). The region commonly deleted on chromosome 13 was 13q22-32. Deletions affecting chromosomes 1 and 2 were highly heterogenous and involved both arms of each chromosome.

Fig. 4.

Idiogram showing distribution of gain and loss of genetic material.

The bars on the right indicate gains, and the bars on the left indicate loss.

Fig. 4.

Idiogram showing distribution of gain and loss of genetic material.

The bars on the right indicate gains, and the bars on the left indicate loss.

Correlation of the recurring breakpoints with clinical features

A correlation analysis of recurring breakpoints identified by SKY with clinical features was performed. Of 32 such sites, 5 showed a significant association with one of the clinical features (P ≤ .05): 7q11 with female, 3p21 with male, 3q27 with stage III-IV disease, 3q27 and 2q31 with more than 2 high or high-intermediate risk, and 2q31 and 7q22 with poor response to treatment (Table 7). Correlation with time to treatment failure and overall survival were not performed because the median follow-up was short.

Table 7.

Summary of significant correlations between the clinical features and recurring breakpoints

Clinical feature Recurring site Present (%) Absent (%) P 
Female 7q11 06/06  (100) 19/40  (47) .025 
Male 3p21 05/05  (100) 14/41  (34) .008  
Stage III-IV 3q27 11/13  (85) 15/32  (47) .024 
High or high-intermediate risk 2q31 06/06  (100) 23/36  (63) .016  
 3q27 13/13  (100) 17/30  (57) .004 
Poor response to treatment 2q31 04/07  (57) 06/36  (18) .040 
 7q22 03/05  (60) 07/38  (18) .033 
Clinical feature Recurring site Present (%) Absent (%) P 
Female 7q11 06/06  (100) 19/40  (47) .025 
Male 3p21 05/05  (100) 14/41  (34) .008  
Stage III-IV 3q27 11/13  (85) 15/32  (47) .024 
High or high-intermediate risk 2q31 06/06  (100) 23/36  (63) .016  
 3q27 13/13  (100) 17/30  (57) .004 
Poor response to treatment 2q31 04/07  (57) 06/36  (18) .040 
 7q22 03/05  (60) 07/38  (18) .033 

Discussion

DLBCL is a histologically well-defined entity that is clinically and genetically heterogeneous. Previous G-banding studies by us and others have shown that most cases are characterized by complex hyperdiploid karyotypes. The types of chromosomal rearrangements are diverse and include translocations, deletions, duplications, and several undefined aberrations such as additions and marker chromosomes.4,12,18-21 This study is the first reported to use SKY to fully characterize the chromosomal rearrangements in a panel of histologically reconfirmed DLBCL. In more than 90% of the cases, SKY provided additional cytogenetic information. Resolution of all the derived, addition, and marker chromosomes by SKY revealed several new recurring breakpoints, translocations, and regions of gains and losses.

By SKY, 13 new recurring translocation breakpoints (Xp11, 4p11, 4p14, 5q11, 11p11, 12p11, 16p11, 16q11-13, 17q11, 17q23, 18p11, 18q11, 20q11-13) were detected that were noted as unique occurrences by G-banding (this study and Cigudosa et al4). The frequency of 10 other recurring translocation breakpoints (1p11-13, 2p13, 2q31, 3p21, 7q11, 7q22, 12q11-13, 15q13-15, 17q11-21, 22q11) was higher in SKY analysis compared with G-banding (11%-17% vs 1%-5%).4,12,18-20 Based on G-banding analysis, we have previously shown that translocations involving 14q32 and 3q27 were the most frequent and together affected 50% of cases. Among these, the partner chromosomes involved remained unidentified in 5% to 15% of the cases due to poor morphology or karyotype complexity. SKY unambiguously identified all the partner chromosomes involved in these translocations, and this in turn enabled the detection of a new recurring translocation, t(3;14)(q21;q32). We have previously observed t(3;12)(q27;q22) (case 862) as nonrecurring in our larger G-banded series.4 The identification of a t(3;12)(q27;q22), designated by G-banding as t(3;12;14)(q29;q22;q32) in the present cohort (2541), makes this a recurring translocation.

The incidence of the known recurring translocations, t(3;14)(q32;q27), t(14;18)(q32;21), t(3;22)(q27;q11), and t(8;14)(q24;q32), was in keeping with the literature. The incidence of t(9;14)(p13;q32) was comparatively higher in the present cohort and may be due to case selection. Apart from detecting some of these recurring translocations with a higher frequency, SKY also enabled their correct identification. Rearrangements affecting bands 3q27 and/or 14q32 were absent in 39% of the cases. We have previously shown, by G-banding analysis, that this subset is characterized by deletions and numerous unidentified additions.4 SKY facilitated the complete cytogenetic characterization of this subset and identified 3 new recurring translocations, t(1;13)(p32;q14), t(1;7)(q21;q22), and der(6)t(6;8)(q11;q11). We also identified 2 potentially new recurring translocations in this group: t(5;16)(?;q11-12) and t(19;22)(q13;q11-13). A search of our database revealed another t(14;18)(q32; q21)-negative follicular large cell lymphoma with a t(19;22)(q13;q13).

Recurring chromosomal changes that lead to gain and/or loss of genetic material constitute important events in both disease transformation and progression.1,22 In this study, SKY identified gains and losses in more than 80% of the cases, a higher frequency of detection than by G-banding. Delineation of all the unbalanced translocations, additions, and marker chromosomes by SKY allowed us to narrow the regions of gain and loss in several chromosomes, namely, gain of 2p13-22, 2q21, 3q27-29, 9p11-24, 13q22-34, 18q21, and 19q11-13 and loss of 6q11-13 and 18p11-13. Although these regions have previously not been identified by G-banding, they have been delineated more precisely by comparative genomic hybridization.22-25 Interestingly, all the cases without translocations of 14q32 or 3q27 showed gains. Gain of 3 (53%), 7q (65%), and 18q21 (41%) and loss of 6q11-13 (59%) were significantly more frequent, suggesting that gain and/or loss of genetic material may play a more proximal role in the development of this subset.

Several studies have attempted to correlate cytogenetic findings with clinical features and/or patient outcome, and the results have been generally contradictory. Most studies failed to show an association between the recurring translocations t(3q27)/BCL6, t(14;18)/BCL2, and t(8;14)/MYC and clinical outcome.26-31 In some studies, changes such as trisomies of chromosomes 2/2p, 3/3p, 5, 6, or 18, monosomy 7, del(6q), abnormalities of 17 [−17, del(17p), i(17q)], and breaks or duplications of 1q have been shown to correlate with advanced stage disease and poor prognosis.13,19,32-36 In the present study, SKY analysis identified 3 sites (2q31, 3q27, 7q22) that significantly associated with advanced-stage disease or poor treatment response. Two of these, 2q31 and 7q22, have previously not been described by G-banding studies. However, we recognize that as interesting as these results are, they need to be confirmed on larger series of consecutively ascertained and uniformly treated DLBCLs. As shown by SKY in this study, most patients, even prior to treatment, harbored multiple genetic changes, making consistent correlations difficult.

The SKY studies reported here and the previously reported comparative genomic hybridization studies by us and others22,23,25present a picture of impressive genetic instability of DLBCL manifested at the cytogenetic level. Much of it remained undetected by conventional G-banding studies that most likely contributed to the conflicting reports in the literature, especially in the clinical correlation analyses. Therefore, the studies reported here emphasize the need for redefinition of chromosomal instability in DLBCL and other NHL subsets applying modern molecular cytogenetic techniques, together with conventional G-banding, for gene discovery as well as meaningful clinical correlations.

We thank Jane Houldsworth for careful reading of the manuscript and constructive criticism.

Supported by the National Cancer Institute grants CA34775, CA66999, and CA80814 (R.S.K.C.).

G.N. and P.H.R. contributed equally to this work.

The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked “advertisement” in accordance with 18 U.S.C. section 1734.

References

References
1
Chaganti
RS
Nanjangud
G
Schmidt
H
Teruya-Feldstein
J
Recurring chromosomal abnormalities in non-Hodgkin's lymphoma: biologic and clinical significance.
Semin Hematol.
37
2000
396
411
2
Alizadeh
AA
Eisen
MB
Davis
RE
et al. 
Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling.
Nature.
403
2000
503
511
3
Lossos
IS
Alizadeh
AA
Eisen
MB
et al. 
Ongoing immunoglobulin somatic mutation in germinal center B cell-like but not in activated B cell-like diffuse large cell lymphomas.
Proc Natl Acad Sci U S A.
97
2000
10209
10213
4
Cigudosa
JC
Parsa
NZ
Louie
DC
et al. 
Cytogenetic analysis of 363 consecutively ascertained diffuse large B-cell lymphomas.
Genes Chromosomes Cancer.
25
1999
123
133
5
Schrock
E
du Manoir
S
Veldman
T
et al. 
Multicolor spectral karyotyping of human chromosomes.
Science.
273
1996
494
497
6
Rao
PH
Cigudosa
JC
Ning
Y
et al. 
Multicolor spectral karyotyping identifies new recurring breakpoints and translocations in multiple myeloma.
Blood.
92
1998
1743
1748
7
Sawyer
JR
Lukacs
JL
Munshi
N
et al. 
Identification of new nonrandom translocations in multiple myeloma with multicolor spectral karyotyping.
Blood.
92
1998
4269
4278
8
Rowley
JD
Reshmi
S
Carlson
K
Roulston
D
Spectral karyotype analysis of T-cell acute leukemia.
Blood.
93
1999
2038
2042
9
Mohr
B
Bornhauser
M
Thiede
C
et al. 
Comparison of spectral karyotyping and conventional cytogenetics in 39 patients with acute myeloid leukemia and myelodysplastic syndrome.
Leukemia.
14
2000
1031
1038
10
Kerndrup
GB
Kjeldsen
E
Acute leukemia cytogenetics: an evaluation of combining G-band karyotyping with multi-color spectral karyotyping.
Cancer Genet Cytogenet.
124
2001
7
11
11
Elghezal
H
Le Guyader
G
Radford-Weiss
I
et al. 
Reassessment of childhood B-lineage lymphoblastic leukemia karyotypes using spectral analysis.
Genes Chromosomes Cancer.
30
2001
383
392
12
Offit
K
Jhanwar
SC
Ladanyi
M
Filippa
DA
Chaganti
RS
Cytogenetic analysis of 434 consecutively ascertained specimens of non-Hodgkin's lymphoma: correlations between recurrent aberrations, histology, and exposure to cytotoxic treatment.
Genes Chromosomes Cancer.
3
1991
189
201
13
Offit
K
Wong
G
Filippa
DA
Tao
Y
Chaganti
RS
Cytogenetic analysis of 434 consecutively ascertained specimens of non-Hodgkin's lymphoma: clinical correlations.
Blood.
77
1991
1508
1515
14
Harris
NL
Jaffe
ES
Diebold
J
et al. 
The World Health Organization classification of neoplastic diseases of the hematopoietic and lymphoid tissues. Report of the Clinical Advisory Committee meeting, Airlie House, Virginia, November, 1997.
Ann Oncol.
10
1999
1419
1432
15
Mitleman
F
ISCN: An International System for Human Cytogenetic Nomenclature.
1995
Karger
Basel, Switzerland
16
Chen
W
Palanisamy
N
Schmidt
H
et al. 
Deregualtion of FCGR2B expression by 1q21 rearrangements in follicular lymphomas.
Oncogene.
20
2001
7686
7693
17
The International Non-Hodgkin's Lymphoma Prognostic Factors Project
A predictive model for aggressive non-Hodgkin's lymphoma.
N Engl J Med.
329
1993
987
994
18
Levine
EG
Arthur
DC
Frizzera
G
Peterson
BA
Hurd
DD
Bloomfield
CD
There are differences in cytogenetic abnormalities among histologic subtypes of the non-Hodgkin's lymphomas.
Blood.
66
1985
1414
1422
19
Juneja
S
Lukeis
R
Tan
L
et al. 
Cytogenetic analysis of 147 cases of non-Hodgkin's lymphoma: non-random chromosomal abnormalities and histological correlations.
Br J Haematol.
76
1990
231
237
20
Schouten
HC
Sanger
WG
Weisenburger
DD
Anderson
J
Armitage
JO
Chromosomal abnormalities in untreated patients with non-Hodgkin's lymphoma: associations with histology, clinical characteristics, and treatment outcome. The Nebraska Lymphoma Study Group.
Blood.
75
1990
1841
1847
21
Johansson
B
Mertens
F
Mitelman
F
Cytogenetic evolution patterns in non-Hodgkin's lymphoma.
Blood.
86
1995
3905
3914
22
Rao
PH
Houldsworth
J
Dyomina
K
et al. 
Chromosomal and gene amplification in diffuse large B-cell lymphoma.
Blood.
92
1998
234
240
23
Monni
O
Joensuu
H
Franssila
K
Knuutila
S
DNA copy number changes in diffuse large B-cell lymphoma—comparative genomic hybridization study.
Blood.
87
1996
5269
5278
24
Monni
O
Joensuu
H
Franssila
K
Klefstrom
J
Alitalo
K
Knuutila
S
BCL2 overexpression associated with chromosomal amplification in diffuse large B-cell lymphoma.
Blood.
90
1997
1168
1174
25
Palanisamy
N
Abou-Elella
AA
Chaganti
SR
et al. 
Similar patterns of genomic alterations characterize primary mediastinal large B-cell lymphoma and diffuse large B-cell lymphoma.
Genes Chromosomes Cancer.
33
2002
114
122
26
Romaguera
JE
Pugh
W
Luthra
R
Goodacre
A
Cabanillas
F
The clinical relevance of t(14;18)/BCL-2 rearrangement and DEL 6q in diffuse large cell lymphoma and immunoblastic lymphoma.
Ann Oncol.
4
1993
51
54
27
Gascoyne
RD
Pathologic prognostic factors in diffuse aggressive non-Hodgkin's lymphoma.
Hematol Oncol Clin North Am.
11
1997
847
862
28
Bastard
C
Deweindt
C
Kerckaert
JP
et al. 
LAZ3 rearrangements in non-Hodgkin's lymphoma: correlation with histology, immunophenotype, karyotype, and clinical outcome in 217 patients.
Blood.
83
1994
2423
2427
29
Tang
SC
Visser
L
Hepperle
B
Hanson
J
Poppema
S
Clinical significance of bcl-2-MBR gene rearrangement and protein expression in diffuse large-cell non-Hodgkin's lymphoma: an analysis of 83 cases.
J Clin Oncol.
12
1994
149
154
30
Kramer
MH
Hermans
J
Wijburg
E
et al. 
Clinical relevance of BCL2, BCL6, and MYC rearrangements in diffuse large B-cell lymphoma.
Blood.
92
1998
3152
3162
31
Vitolo
U
Gaidano
G
Botto
B
et al. 
Rearrangements of bcl-6, bcl-2, c-myc and 6q deletion in B-diffuse large-cell lymphoma: clinical relevance in 71 patients.
Ann Oncol.
9
1998
55
61
32
Levine
EG
Arthur
DC
Frizzera
G
et al. 
Cytogenetic abnormalities predict clinical outcome in non-Hodgkin lymphoma.
Ann Intern Med.
108
1988
14
20
33
Yunis
JJ
Mayer
MG
Arnesen
MA
Aeppli
DP
Oken
MM
Frizzera
G
bcl-2 and other genomic alterations in the prognosis of large-cell lymphoma.
N Engl J Med.
320
1989
1047
1054
34
Cabanillas
F
Pathak
S
Grant
G
et al. 
Refractoriness to chemotherapy and poor survival related to abnormalities of chromosomes 17 and 7 in lymphoma.
Am J Med.
87
1989
167
172
35
Schlegelberger
B
Zwingers
T
Harder
L
et al. 
Clinicopathogenetic significance of chromosomal abnormalities in patients with blastic peripheral B-cell lymphoma. Kiel-Wien-Lymphoma Study Group.
Blood.
94
1999
3114
3120
36
Whang-Peng
J
Knutsen
T
Jaffe
ES
et al. 
Sequential analysis of 43 patients with non-Hodgkin's lymphoma: clinical correlations with cytogenetic, histologic, immunophenotyping, and molecular studies.
Blood.
85
1995
203
216

Author notes

R. S. K. Chaganti, Memorial Sloan-Kettering Cancer Center, 1275 York Ave, New York, NY 10021; e-mail: chagantr@mskcc.org.