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Abstract

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

This study aimed to clarify the clinicopathological prognostic parameters of de novo diffuse large B-cell lymphoma (DLBCL) in the rituximab era. We examined the correlation of 22 clinicopathological parameters with progression-free survival (PFS), overall survival (OS), and primary refractory disease in 285 DLBCL patients treated with rituximab-containing chemotherapy. Complete response rate was 87%, overall response rate was 91%, 5-year PFS rate was 72%, and 5-year OS rate was 91%. By log–rank test, higher International Prognostic Index (IPI) (P < 0.0001), Bcl-2 positivity (P = 0.0013), Bcl-6 negativity (P = 0.0112), and no irradiation (P = 0.0371) were significantly correlated with shorter PFS; higher IPI (P = 0.0107), starry sky pattern (P = 0.0466), and no irradiation (P = 0.0264) correlated with shorter OS. In multivariate analyses, higher IPI (P = 0.0006), Bcl-2 positivity (P = 0.0015), and Bcl-6 negativity (P = 0.04) were significantly correlated with shorter PFS; higher IPI (P = 0.0045) correlated with shorter OS. Bcl-2 (P = 0.0029), Bcl-6 (P = 0.002), and IPI (P < 0.0001) were significantly correlated with primary refractory disease. In conclusion, Bcl-2 positivity, Bcl-6 negativity, and higher IPI were indicators of shorter PFS and OS plus primary refractory disease in patients with DLBCL in the rituximab era.

Diffuse large B-cell lymphoma (DLBCL) represents the largest and most widely heterogeneous category of aggressive non-Hodgkin lymphomas.[1] Several histopathological prognostic parameters of DLBCL have been reported. In the World Health Organization (WHO) Classification of Tumours of Haematopoietic and Lymphoid Tissues published in 2008, several DLBCL variants, subgroups, and subtypes were proposed.[1] Morphological variants include centroblastic, immunoblastic, and anaplastic variants. Subtypes of DLBCL include T cell/histiocyte-rich large B-cell lymphoma and Epstein–Barr virus (EBV)-positive DLBCL of the elderly. Among these morphological variants and subtypes, immunoblastic variant DLBCL,[2, 3] T cell/histiocyte-rich large B-cell lymphoma,[4] and EBV-positive DLBCL of the elderly[5] have been reported to have poor prognoses. Immunohistochemical expression of Bcl-2[3, 6-9] and CD5[10] has been reported to be associated with an unfavorable prognosis; expression of Bcl-6[11] and CD10[3] are associated with a favorable prognosis. High Ki-67 index[12] has also been reported to be a poor prognostic parameter. However, these results were obtained mainly in the pre-rituximab era. Therefore, these prognostic parameters should be re-evaluated in the rituximab era. As anti-CD30 mAb therapy was found to be effective for classical Hodgkin's lymphoma and anaplastic large cell lymphoma,[13] the prognostic implication of CD30 expression should be examined in DLBCLs. cMYC rearrangement was reported to be a poor prognostic parameter of DLBCL in the rituximab era,[14] and cMYC rearrangement and immunohistochemical cMyc expression were reported to be correlated.[15] Therefore, immunohistochemical cMyc expression should also be evaluated.

Since 2000, DLBCL has been subdivided into germinal center B-cell (GCB) phenotype and non-GCB phenotype (including the activated B-cell phenotype and type 3 phenotype) using the cDNA microarray technique.[16, 17] For use in clinical practice, Hans et al.[18] showed that a panel of immunohistochemical markers comprising CD10, Bcl-6, and MUM1 could be used on paraffin-embedded tissues to classify DLBCL into tumors with a GCB or non-GCB phenotype. The GCB phenotype showed a better outcome in the pre-rituximab era;[5, 16, 17] however, it was reported that the addition of rituximab to the cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) regimen eliminated the prognostic value of the GCB phenotype.[19] Therefore, the use of the GCB phenotype as a prognostic marker is controversial in the rituximab era.

In the pre-rituximab era, the International Prognostic Index (IPI), which is based on clinical parameters such as age, stage, serum lactate dehydrogenase (LDH) level, performance status (PS), and extent of extranodal involvement (EN), proved to be highly valuable for the prediction of prognosis in patients with DLBCL.[20] However, the IPI seems to have lost some of its high predictive value in the rituximab era.[21]

The aim of this study was to clarify the clinicopathological prognostic parameters of de novo DLBCL in the rituximab era. Thirteen histopathological parameters including DLBCL morphological variant, necrosis, starry sky pattern, CD5, CD10, CD30, Bcl-2, Bcl-6, MUM1, GCB/non-GCB, cMyc, Ki-67, and EBV-encoded RNA (EBER)-1, as well as nine clinical parameters, including IPI, influencing progression-free survival (PFS) and/or overall survival (OS) were evaluated by log–rank tests and multivariate analyses. Correlation of primary refractory disease and these clinicopathological parameters was also examined.

Materials and Methods

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

Patient selection

The study subjects were 285 consecutive patients with de novo DLBCL, treated at the National Cancer Center Hospital (Tokyo, Japan) between 2003 and 2010. Clinical information was extracted from medical records. The Ann Arbor system was used for staging. The staging procedures included bone marrow aspiration or biopsy, endoscopy of the upper gastrointestinal tract, computed tomography, and optionally, PET. After initial diagnoses, all of the patients received rituximab-containing chemotherapy that consisted of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) with or without involved-field radiotherapy (258 patients), R-CHOP-like regimen (three patients), or modified rituximab, cyclophosphamide, vincristine, doxorubicin, and methotrexate (R-CODOX-M)/ifosfamide, etoposide, and cytarabine (IVAC) (24 patients).[22] High-dose chemotherapy with autologous peripheral blood stem cell transplantation was carried out in three patients. Four groups of clinical course were defined as follows: group 1, patients achieving complete response (CR) or partial response (PR) with the initial therapy and no relapse; group 2, patients achieving CR or PR with the initial therapy and relapse after 1 year or later (late relapse); group 3, patients achieving CR or PR with the initial therapy and relapse within 1 year (early relapse); and group 4, patients showing no change or progressive disease after the initial therapy. Group 4 were defined as having primary refractory disease in this study. Median follow-up time was 41 months (range, 1–97 months). Informed consent was obtained from each patient. The study was approved by the institutional review board of the National Cancer Center.

Morphological review

Biopsy materials were fixed in 10% neutral-buffered formalin, embedded in paraffin, cut into sections 4-μm thick, and stained with H&E for histopathological evaluation. All specimens were diagnosed by two pathologists (AMM and HT) according to the WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues, 2008.[1] Diffuse large B-cell lymphoma was subclassified as centroblastic, anaplastic, immunoblastic, or T cell/histiocyte-rich variant. Diffuse large B-cell lymphoma preceded by low-grade B-cell lymphoma or DLBCL with coexisting low-grade B-cell lymphoma was excluded. Necrosis and starry sky pattern were also evaluated.

Immunohistochemistry and in situ hybridization

Immunohistochemistry on formalin-fixed paraffin-embedded tissues was carried out using a panel of mAbs. The number of available specimens differed for each stain. Sections 4-μm thick were cut from each paraffin block, deparaffinized, and incubated at 121°C in citrate buffer, pH 6.0, for 10 min for antigen retrieval. The antibodies used included those against the following antigens: CD3 (PS1, ×25; Novocastra, Newcastle-upon-Tyne, UK), CD5 (4C7, ×100; Novocastra), CD10 (56C6, ×100; Novocastra), CD20 (L26, ×200; Dako, Glostrup, Denmark), CD30 (Ber-H2, ×100; Dako), Bcl-2 (124, ×100; Dako), Bcl-6 (PG-B6p, ×20; Dako), cMyc (Y69, ×50; Epitomics, Burlingame, CA, USA), cyclin D1 (SP4, ×25; Nichirei, Tokyo, Japan), Ki-67 (MIB-1, ×100; Dako), and MUM1 (MUM1p, ×200; Dako). An autostainer was used with the standard polymer method (Dako Autostainer Plus). All cases were positive for CD20 and negative for CD3. Immunoreactivity for CD5, CD10, CD30, Bcl-2, Bcl-6, and MUM1 was judged positive if more than 20% of the tumor cells were stained. Labeling index was counted for Ki-67 and cMyc; 90% or more was defined as high labeling index for Ki-67, and 80% or more as high labeling index for cMyc. The area with the greatest staining was selected, 500 nuclei were counted manually, and the proportion of positive cells was calculated. To classify each case as having either a GCB phenotype or a non-GCB phenotype, a panel of three antigens (CD10, Bcl-6, and MUM1) was used according to the protocol reported by Hans et al.[18] If the tumor revealed CD5 positivity, cyclin D1 negativity was confirmed. In situ hybridization with an EBER-1 probe (Dako) was carried out to detect possible EBV infection.

Statistical analysis

All survival curves were estimated by the Kaplan–Meier method. Statistical differences between survival curves were compared using the log–rank test for clinicopathological parameters. Multivariate analysis was carried out using Cox's proportional hazard model for the significant parameters detected by the log–rank test. Correlation between primary refractory disease and clinicopathological parameters was analyzed by Spearman's rank correlation coefficient test. Differences were considered significant when the P-value was <0.05.

Results

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

Patient characteristics and histopathological results

Clinical information and histopathological results are summarized in Table 1. Patients comprised 148 men and 137 women, ranging in age from 17 to 88 years with a median age of 55 years. Initial site was nodal in 164 patients and extranodal in 121 patients. The %CR for initial treatment was 87%, and the overall response rate was 91%. The 5-year PFS rate was 72%, and the 5-year OS rate was 91%.

Table 1. Prognostic significance of 22 clinicopathological parameters in 285 patients with diffuse large B-cell lymphoma
ParametersNo. of patients (%)5-year PFS (%)P-valuea5-year OS (%)P-valuea
  1. a

    P-value was calculated by log–rank test. A, anaplastic; C, centroblastic; EN, extranodal involvement; GCB, germinal center B-cell phenotype; H, high; HI, high intermediate; I, immunoblastic; IPI, international prognostic index; ISH, in situ hybridization; L, low; LDH, lactate dehydrogenase (normal range, 119–229 U/L); LI, low intermediate; NS, not significant; OS, overall survival; PFS, progression-free survival; PS, performance status; T/H, T-cell/histiocyte rich.

Total28572 90 
Clinical parameters
Age
≤60 years158 (55)74NS94NS
>60 years127 (45)7085
Gender
Male148 (52)69NS88NS
Female137 (48)7693
IPI
L, LI218 (76)79<0.0001940.0107
HI, H67 (24)5179
Stage
I, II198 (69)790.000393NS
III, IV87 (31)5884
LDH
Normal range144 (51)82<0.0001950.0121
High141 (49)6285
PS
0, 1249 (87)750.031791NS
2–436 (13)5884
EN
0–1236 (83)760.000892NS
≥249 (17)5583
Initial site
Nodal164 (58)74NS89NS
Extranodal121 (42)7092
Irradiation
Not performed151 (53)780.0371940.0264
Performed134 (47)6885
Histopathological parameters
DLBCL variant
C or A259 (91)73NS91NS
I or T/H26 (9)6791
Necrosis
Absent223 (78)72NS90NS
Present62 (22)7793
Starry sky
Absent263 (92)73NS910.0466
Present22 (8)6584
CD5
Negative245 (92)72NS90NS
Positive22 (8)7185
CD10
Negative192 (74)74NS92NS
Positive67 (26)6490
Bcl-6
Negative69 (33)650.011285NS
Positive140 (67)7693
MUM1
Negative96 (46)70NS94NS
Positive112 (54)7588
GCB/non-GCB
GCB100 (43)73NS93NS
Non-GCB132 (57)7189
Bcl-2
Negative102 (46)860.001396NS
Positive122 (54)6792
cMyc index
<80%190 (90)73NS90NS
≥80%20 (10)7294
Ki-67 index
<90%139 (70)76NS94NS
≥90%60 (30)6786
CD30
Negative156 (91)75NS93Not calculated
Positive15 (9)87100
EBER-1 ISH
Negative197 (94)74NS91Not calculated
Positive13 (6)66100

By log–rank test, higher IPI (P < 0.0001), advanced stage (P = 0.0003), a high level of LDH (≥230 U/L) (P < 0.0001), high PS (2–4) (P = 0.0317), more than two incidences of EN (P = 0.0008), no irradiation (P = 0.0371), Bcl-6 negativity (P = 0.0112; Figs 1a,2), and Bcl-2 positivity (P = 0.0013; Figs 1b,3) were significant parameters of shorter PFS (Table 1). Likewise, higher IPI (P = 0.0107), a high level of LDH (P = 0.0121), no irradiation (P = 0.0264), and starry sky pattern (P = 0.0466; Fig. 1c) were significant parameters of shorter OS (Table 1). Bcl-6 negativity was marginal as a prognostic parameter of OS (P = 0.0691). Patients with CD30 positivity and EBER-1 positivity revealed 100% 5-year OS; however, the number of CD30-positive and EBER-1-positive cases was small.

image

Figure 1. Diffuse large B-cell lymphoma with a Bcl-6-positive phenotype (magnification, ×400) (A), a Bcl-2-positive phenotype (magnification, ×400) (B), and with starry sky pattern (H&E staining; magnification, ×200) (C).

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image

Figure 2. Progression-free survival (PFS) curves for 224 patients with diffuse large B-cell lymphoma stratified by Bcl-2 immunoreactivity. Five-year PFS rates were 67% in the Bcl-2-positive group and 86% in the Bcl-2-negative group. The PFS rate of 122 patients with Bcl-2 positivity was significantly worse than that of 102 patients with Bcl-2 negativity (P = 0.0013).

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image

Figure 3. Progression-free survival (PFS) curves for 209 patients with diffuse large B-cell lymphoma stratified by Bcl-6 immunoreactivity. Five-year PFS rates were 76% in the Bcl-6 positive group and 65% in the Bcl-6 negative group. The PFS rate of 69 patients with Bcl-6 negativity was significantly worse than that of 140 patients with Bcl-6 positivity (P = 0.0112).

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In the multivariate analyses carried out for significant parameters detected by log–rank test, with the exception of parameters such as stage, LDH, PS, and EN already included in the IPI scoring system, higher IPI, Bcl-2 positivity, and Bcl-6 negativity were independently correlated with lower PFS rate (Table 2), and only higher IPI was independently correlated with lower OS rate (Table 3).

Table 2. Results of Cox's multivariate proportional hazards analysis for progression-free survival in 285 patients with diffuse large B-cell lymphoma
ParametersHazard ratio95% confidence intervalP-value
  1. IPI, International Prognostic Index.

IPI1.6101.226–2.1160.0006
Bcl-21.8331.260–2.6680.0015
Bcl-60.5160.275–0.9700.0400
Table 3. Results of Cox's multivariate proportional hazards analysis for overall survival in 285 patients with diffuse large B-cell lymphoma
ParameterHazard ratio95% confidence intervalP-value
  1. IPI, International Prognostic Index.

IPI1.7921.199–2.6800.0045

We carried out subgroup analyses to examine the predictive value of Bcl-2 and Bcl-6 with PFS and OS. Bcl-2 was a significant prognostic factor of PFS in the low (L)/low-intermediate (LI) IPI group (P = 0.009), but not in the high-intermediate (HI)/high (H) IPI group (Fig. 4a). Bcl-6 was a significant prognostic factor of PFS in the HI/H IPI group (P = 0.0451), but not in the L/LI IPI group (Fig. 4b). The PFS was compared among the following four groups: Bcl2+/Bcl6; Bcl2/Bcl6+; Bcl2+/Bcl6+; and Bcl2/Bcl6. The Bcl2/Bcl6+ group had a significantly better PFS than the other three groups (P = 0.0486), but significant differences were not found among the other three groups (Fig. 5).

image

Figure 4. (A) Progression-free survival (PFS) curves for 224 patients with diffuse large B-cell lymphoma stratified by the International Prognostic Index (IPI) and Bcl-2 immunoreactivity. Five-year PFS rates were 92% in the low (L)/low-intermediate (LI) IPI and Bcl-2 group, 72% in the L/LI and Bcl-2+ group, 58% in the high-intermediate (HI)/high (H) and Bcl-2+ group, and 56% in the HI/H and Bcl-2 group. The PFS of the L/LI and Bcl-2 group was significantly better than that of the other three groups (P = 0.009). (B) The PFS curves for 209 patients stratified by IPI and Bcl-6 immunoreactivity. Five-year PFS rates were 75% in the L/LI and Bcl-6 group, 80% in the L/LI and Bcl-6+ group, 72% in the HI/H and Bcl-6 group, and 33% in the HI/H and Bcl-6+ group. The PFS of the HI/H and Bcl-6+ group was significantly worse than that of the other three groups (P = 0.0451).

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image

Figure 5. Progression-free survival (PFS) curves for 195 patients stratified by Bcl-2 and Bcl-6 immunoreactivity. Five-year PFS rates were 92% in the Bcl-2/Bcl-6+ group, 79% in the Bcl-2/Bcl-6 group, 68% in the Bcl-2+/Bcl-6+ group, and 63% in the Bcl-2+/Bcl-6 group. The PFS of the Bcl-2/Bcl-6+ group was significantly better than that of the other three groups (P = 0.0486).

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Correlation of primary refractory disease with clinicopathological parameters

Group 1 comprised 220 (77%) patients, group 2 comprised 13 (5%) patients, group 3 comprised 25 (9%) patients, and group 4 comprised 27 (9%) patients (Table 4). The PFS curves for all 285 patients stratified by these four groups are shown in Figure 6. Five-year PFS rates were 100% in group 1, 8% in group 2, and 0% in groups 3 and 4. The OS curves for all 285 patients stratified by the four groups are shown in Figure 7. Five-year OS rates were 97% in group 1, 92% in group 2, 62% in group 3, and 52% in group 4. The OS rates significantly differed between groups 1 and 2 versus group 3 (P < 0.0001), and between group 3 versus group 4 (P = 0.0499). Correlation of primary refractory disease with clinicopathological parameters is shown in Table 4. Primary refractory disease was significantly correlated with higher IPI, advanced stage, high LDH, PS (2–4), EN (≥2), Bcl-2 positivity, and Bcl-6 negativity.

image

Figure 6. Progression-free survival (PFS) curves for 285 patients with diffuse large B-cell lymphoma stratified into four groups. Five-year PFS rates were 100%, 8%, 0%, and 0% in groups 1 (220 patients), 2 (13 patients), 3 (25 patients), and 4 (27 patients), respectively.

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image

Figure 7. Overall survival (OS) curves for 285 patients with diffuse large B-cell lymphoma stratified into four groups. The OS rates differed significantly (P < 0.0001) between groups 1 (220 patients) and 2 (13 patients) versus group 3 (25 patients); OS rates differed significantly (P = 0.0499) between group 3 versus group 4 (27 patients). Five-year OS rates were 97%, 92%, 62%, and 52% in groups 1, 2, 3, and 4, respectively.

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Table 4. Correlation between groups 1–4 and clinicopathological parameters in 285 patients with diffuse large B-cell lymphoma
ParametersGroup1234P-value
  1. P-value was calculated by Spearman's co-efficiency test. EN, extranodal involvement; Group 1, patients with complete response (CR) or partial response (PR) after the first therapy and no relapse; Group 2, patients with CR or PR after the first therapy and relapse after 1 year or later (late relapse); Group 3, patients with CR or PR after the first therapy and relapse within 1 year (early relapse); Group 4, patients with no change or progressive disease after the first therapy; H, high; HI, high intermediate; IPI, International Prognostic Index; L, low; LDH, lactate dehydrogenase (normal range, 119–229 U/L); LI, low intermediate; NS, not significant; PS, performance status.

Clinical parametersIPI (L, LI/HI, H)183/379/415/1011/16<0.0001
Stage (I, II/III, IV)164/568/512/1314/13<0.0001
LDH (normal/high)126/938/54/215/22<0.0001
PS (0–1/2–4)195/2412/121/420/7<0.0001
EN (0–1/≥2)191/299/418/718/90.0011
Histopathological parametersBcl-6 (negative/positive)4/1163/69/912/90.0029
Bcl-2 (negative/positive)91/882/73/126/150.0020

Histopathology of rebiopsied material at the time of relapse indicated DLBCL in five patients (site: bone marrow, 1; liver, 1; skin, 2; and stomach, 1), follicular lymphoma grade 1 in one patient (site: jejunum), and mucosa-associated lymphoid tissue lymphoma in four patients (site: orbit, 1; parotid gland, 1; and thyroid, 2) in group 2. In group 3, histopathology indicated DLBCL in 16 patients (site: central nervous system, 1; bone marrow and skin, 1; chest wall, 1; lung, 1; lymph node, 5; mediastinum, 2; skin, 2; testis and central nervous system, 2; and urinary bladder, 1), and DLBCL and follicular lymphoma grade 3B in one patient (site: lymph node).

Discussion

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

In this study, clinicopathological prognostic parameters of patients with de novo DLBCL in the rituximab era were examined. Histopathological parameters studied included DLBCL morphological variant, necrosis, starry sky pattern, CD5, CD10, CD30, Bcl-2, Bcl-6, MUM1, GCB/non-GCB, cMyc, Ki-67, and EBER-1. The results revealed that higher IPI, Bcl-2 positivity, and Bcl-6 negativity were significantly correlated with shorter PFS and primary refractory disease in the rituximab era.

Concerning the outcome of patients with DLBCL, Récher et al.[23] reported that the 3-year PFS rate was 73% and the 3-year OS rate was 84%. Sehn et al.[21] reported that the 4-year PFS rate was 70% and the 4-year OS rate was 70%. Patients in the present study might have had better outcomes than the patients in these reports: 5-year PFS rate was 72% and 5-year OS rate was 91%, which might be partly associated with the low median age (55 years) and high incidence of low/low intermediate IPI and stage I/II disease in the current study.

Bcl-2 protein, an antiapoptotic molecule, is expressed on resting B and T cells, but not on normal germinal center cells.[24] Bcl-2 is expressed in 22–80% of DLBCLs,[25] and Bcl-2 positivity has been reported to be associated with an unfavorable prognosis.[3, 6-9] Bcl-6 protein is expressed in B and CD4+ T cells within the germinal center,[26] and is expressed in 47–84% of DLBCLs.[1] The Bcl-6 rearrangement observed in 30–40% of DLBCLs leads to deregulation of Bcl-6 gene expression.[27, 28] Bcl-6 rearrangement was reported to correlate with a favorable outcome.[29] However, the level of Bcl-6 protein expression is not correlated with the presence or absence of Bcl-6 gene rearrangement and mutation.[30, 31] Bcl-6 protein expression was reported to be associated with favorable prognosis.[11] The addition of rituximab to treatment regimens has considerably improved the survival of patients with DLBCL[32] and was reported to have eliminated the negative impact of Bcl-2 expression and the positive impact of Bcl-6 expression on clinical outcome.[33-35] However, in the present study, Bcl-2 positivity and Bcl-6 negativity were found to be parameters predicting a significantly shorter PFS (particularly Bcl-2 in the L/LI IPI group, and Bcl-6 in the HI/H IPI group) and primary refractory disease even in the rituximab era.

CD10 shows restricted expression in the germinal center B cells of reactive lymphoid tissue and is expressed in 30–60% of DLBCLs.[1] MUM1 is a lymphoid-specific member of the interferon regulatory factor family of transcription factors.[36] Normally expressed in plasma cells and a minor subset of germinal center B cells, MUM1 has been reported to be expressed in 35–65% of DLBCLs.[1] CD10 has been reported to be a favorable prognostic parameter.[3] CD10, Bcl-6, and MUM1 are included in the panel of markers used to assess GCB or non-GCB phenotype.[18] Some previous studies examining the difference in prognosis between patients with GCB phenotype and those with non-GCB phenotype DLBCL revealed that the former group had a more favorable prognosis.[16, 17] However, Colomo et al.[7] found no prognostic difference between these groups; thus, this has recently become a controversial issue. In the present study, GCB versus non-GCB was not a significant prognostic factor of DLBCL and neither were CD10 or MUM1.

Ki-67 index, cMyc index, starry sky pattern, and necrosis are considered to be correlated immunohistochemical and histopathological findings, and all of them are associated with proliferation activity of tumors. High Ki-67 index and high cMyc index reflecting cMYC rearrangement were reported to be poor prognostic parameters.[12, 14] The predictive value of Ki-67 index was reported in the pre-rituximab era.[12] In the present study, only starry sky pattern was a marginally significant predictor of OS by log–rank test; however, this result was not maintained in multivariate analysis. Our results suggested that the predictive values of these factors are limited in the rituximab era.

Our results suggested that CD5 was not a significant poor prognostic factor in the rituximab era. Yamaguchi et al.[10] reported that CD5 was a significant poor prognostic factor of OS in the pre-rituximab era, but not in the rituximab era.[37] In addition, expression of CD30, and EBER-1 and morphological DLBCL variant were not significant prognostic parameters in the rituximab era.

In the pre-rituximab era, IPI proved to be highly valuable in predicting the prognosis of DLBCLs;[20] however, IPI seems to have lost some of its predictive value in the rituximab era.[21] In the present study, conventional IPI was a significant prognostic parameter for predicting PFS, OS, and primary refractory disease. Several parameters comprising IPI, such as stage, LDH, PS, and EN, were also significant prognostic parameters predicting PFS, OS, or primary refractory disease.

In group 2, histopathology of rebiopsied material at the time of relapse revealed DLBCL in five patients and low-grade B-cell lymphoma in five patients. The latter could have represented transformed low-grade B-cell lymphoma from initial presentation. Therefore, it was speculated that approximately 50% of late relapsed DLBCLs had transformed from low-grade B-cell lymphomas.

In conclusion, our study shows that Bcl-2 positivity, Bcl-6 negativity, and higher IPI are significant indicators of shorter PFS, that IPI is a significant indicator of shorter OS, and that Bcl-2 positivity, Bcl-6 negativity, and higher IPI are indicators of primary refractory disease. Our results clarify the significant clinicopathological prognostic parameters of DLBCL in the rituximab era.

Acknowledgments

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

The authors would like to thank Ms C. Kina and Ms S. Miura for technical assistance with immunohistochemistry. The study was supported in part by the National Cancer Center Research and Development Fund, and a Grant-in-Aid for the Third-Term Comprehensive 10-Year Strategy for Cancer Control from the Ministry of Health, Labor and Welfare, Japan.

Disclosure Statement

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References

Kensei Tobinai received research grants from Zenyaku Kogyo and Chugai Pharmaceutical.

References

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure Statement
  8. References
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