Chromosomal translocation t(14;18)(q32;q21) involving the immunoglobulin heavy chain gene (IGH) and the BCL2 gene (t[14;18][q32;q21]/IGH-BCL2) is present in 60% to 90% of nodal follicular lymphomas. To the authors' knowledge, the prevalence and clinical significance of this translocation have not been examined previously in gastrointestinal follicular lymphomas.
Clinicopathologic and molecular features were investigated in 48 patients who had gastrointestinal follicular lymphoma. The site of involvement was the duodenum in 54% of patients, the jejunum in 52%, the ileum in 52%, the stomach in 29%, and the colorectum in 15%. The presence of the t(14;18)/IGH-BCL2 translocation was detected by interphase fluorescence in situ hybridization.
Treatment modalities included surgical resection (n = 16), rituximab plus chemotherapy (n = 13), rituximab alone (n = 6), antibiotics (n = 5), and watchful waiting (n = 8). Complete remission (CR) of lymphoma was achieved in 31 patients (65%). The overall survival and event-free survival rates after 5 years were 93% and 68%, respectively. The t(14;18)/IGH-BCL2 was detected in 39 patients (81%). The involvement of multiple sites (69% vs 0%), manifestation of the lymphomatous polyposis type (72% vs 22%), and histologic grade 1 or 2 tumors (92% vs 56%) were more frequent in the t(14;18)-positive group than in the negative group. In addition, the CR rate was lower in the t(14;18)-positive group than in the negative group (56% vs 100%; P = .0179), and a trend was observed toward poorer event-free survival in the positive group (P = .089).
Although follicular lymphoma (FL) is 1 of the most common subtypes of nodal non-Hodgkin lymphoma,1 primary FL of the gastrointestinal tract is uncommon, accounting for only 3% to 7.8% of cases among all gastrointestinal lymphomas.2-4 Recently, balloon-assisted enteroscopies—that is, double-balloon endoscopy (DBE) and single-balloon endoscopy (SBE)—have enabled the examination of an extensive area of the small bowel and the detection of diminutive small intestinal lesions. The usefulness of DBE for the practical diagnosis of intestinal FL has been reported in several publications.5-9 However, detailed information about the clinical features of gastrointestinal FL remains limited.
The chromosomal translocation t(14;18)(q32;q21) involving the immunoglobulin heavy chain gene (IGH) and the BCL2 gene (t[14;18][q32;q21]/IGH-BCL2) is a genetic hallmark of nodal FL with a prevalence that ranges from 60% to 90%.1, 10 However, the prevalence of this translocation is not high in FL arising in the extranodal regions, such as the skin.11-13 To date, the presence of t(14;18)/IGH-BCL2 in gastrointestinal FL has been examined only in a few investigations in which the clinical significance of the translocation was not specified.14-17 In the current study, we retrospectively analyzed the clinicopathologic and molecular features of 48 patients who had gastrointestinal FL, paying special attention to the prevalence and clinical significance of t(14;18)/IGH-BCL2.
MATERIALS AND METHODS
Between 1966 and 2009, 634 consecutive Japanese patients with primary gastrointestinal lymphoma were treated at our institutions. Among them, 52 patients (8.2%) were diagnosed with FL. Of these, 48 patients who had formalin-fixed, paraffin-embedded tissues available and who had tumor cells that were positive immunohistochemically for both cluster of differentiation 10 (CD10) (membrane metalloendopeptidase, neutral endopeptidase) and BCL2, were selected for the current study.
The histologic diagnosis, including the grade of FL (grade 1or 2, 3A, or 3B), of each tumor was reviewed according to the criteria described by the World Health Organization classification.1 The clinical stage of lymphoma was determined according to the Lugano International Conference classification.18 The staging workup included computed tomography scans of the neck, chest, and abdomen; esophagogastroduodenoscopy (EGD); colonoscopy; small-bowel barium radiography; bone marrow aspiration or biopsy; and fluorine-18 fluorodeoxyglucose-positron emission tomography or gallium scintigraphy studies. Endoscopic examinations of the small bowel were performed in 29 patients using DBE or SBE. Among them, the entire gastrointestinal tract was evaluated endoscopically in 26 patients. The macroscopic type of lymphoma was classified as superficial, polypoid, ulcerative, lymphomatous polyposis, diffuse, or mixed type.4Helicobacter pylori status was assessed in 32 patients by histology, culture, the rapid urease test, the 13C urea breath test, and/or serology. Each patient was examined at least with the 13C urea breath test or serology. H. pylori infection was judged positive if 1 of any of the pretreatment tests for H. pylori produced a positive result, and it was judged negative only when all of the test results were negative.
The initial treatment modalities included surgical resection; chemotherapy with a cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP)-based regimen; or rituximab with CHOP (R-CHOP) or without CHOP; antibiotic treatment with the same regimen that was used for H. pylori eradication; or watchful waiting. Complete remission (CR) was defined as the complete disappearance of clinical evidence of lymphoma. Partial remission (PR) was defined as a tumor reduction ≥50%. Patients who did not attain a CR or a PR, including those who had progressive disease (PD), were considered to be unresponsive. Overall survival (OS) was measured from the date of diagnosis to the date of death from any cause, and event-free survival (EFS) was measured from the date of diagnosis to the date of PD, recurrence, or death from any cause. The study protocol was approved by the Ethics Committee at Kyushu University Hospital (no. 20-105), and the study was conducted in accordance with the Helsinki Declaration.
Interphase Fluorescence In Situ Hybridization
The presence of t(14;18)(q32;q21)/IGH-BCL2 was investigated by using interphase fluorescence in situ hybridization (FISH) with a IGH/BCL2 dual-color, dual-fusion translocation probe (Vysis-Abbott, Des Plaines, Ill) on formalin-fixed, paraffin-embedded tissue sections.19 In tumors that were negative for the IGH-BCL2 translocation, additional FISH analyses were performed. FISH with the BCL6 dual-color, break-apart probe (Vysis-Abbott) was used for the detection of translocations involving BCL6 and numerical gains at chromosome 3q27. For the detection of trisomy 3 and 18, FISH also was performed with the centromere-specific probes chromosome enumeration probe 3 (CEP3) (Spectrum Orange) and CEP18 (Spectrum Green; Vysis-Abbott). In each tumor, the hybridization signals were evaluated in at least 100 nuclei. The threshold for positivity using each probe was the mean percentage of cells with false-positive signals plus 3 standard deviations as assessed in tissue specimens that were obtained from 10 reactive tonsils.19
Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissues using a Histofine Simple Stain MAX-PO kit (Nichirei, Tokyo, Japan) according to the manufacturer's instructions.20 Monoclonal antibodies for CD10 (clone 56C6; Novocastra, Newcastle upon Tyne, United Kingdom) and BCL2 (clone 124; Dako, Carpinteria, Calif) were stained in all 48 tumors. In tumors that were negative for t(14;18)/IGH-BCL2 by FISH, immunostaining with a monoclonal antibody for BCL6 (clone PG-B6p; Dako) was added.
The probabilities of OS and EFS were calculated by using the Kaplan-Meier method, and the values were compared using the log-rank test. Other differences were evaluated with the Fisher exact test, the chi-square test, or the Mann-Whitney U test, as appropriate. Probabilities <.05 were regarded as statistically significant. For multiple comparisons, however, P values were interpreted after the Bonferroni correction.
Clinical Features of Gastrointestinal FL
The clinicopathologic features of the patients are summarized in Table 1. The entire patient cohort included 27 men and 21 women who ranged in age from 32 years to 82 years (mean age, 60.2 years) at the time of diagnosis. The most frequently involved site was the duodenum (54%), followed by the jejunum (52%), the ileum (52%), the stomach (29%), and the colorectum (15%). In 11 patients (23%), FL was localized in the stomach, whereas 37 patients (77%) had intestinal involvement. Of the 37 patients who had intestinal involvement, 10 had FL localized to a single site: 5 patients had a lesion in the duodenum, 1 patient had a lesion in the jejunum, and 4 patients had a lesion in the ileum. In 27 patients (56%), multiple sites within the gastrointestinal tract were involved (Table 2).
Table 1. Clinicopathologic Features of 48 Patients With Gastrointestinal Follicular Lymphoma
No. of Patients (%)
GI indicates gastrointestinal; R-CHOP, rituximab plus combined cyclophosphamide, doxorubicin, vincristine, and prednisolone; CR, complete remission.
Site of involvement
No. of involved site within GI tract
1 or 2
Surgery plus chemotherapy
Response to treatment
Table 2. Anatomic Distribution of Gastrointestinal Follicular Lymphomas
Involved Site (n = 48)
No. of Patients
Single intestinal involvement
Multiple intestinal involvement
Duodenum, jejunum, and ileum
Duodenum and jejunum
Stomach, duodenum, jejunum, ileum, and colorectum
Duodenum, jejunum, ileum, and colorectum
Jejunum and ileum
Ileum and colorectum
Jejunum, ileum, and colorectum
Stomach and ileum
Macroscopically, 28 tumors (58%) were classified as lymphomatous polyposis, 8 tumors (17%) were classified as polypoid, 6 tumors (13%) were classified as ulcerative, 3 tumors (6%) were classified as diffuse, 2 tumors (4%) were classified as mixed type, and 1 tumor (2%) was classified as superficial. The 2 tumors that were classified as mixed type had a component of lymphomatous polyposis. The histologic grade of FL was grade 1 or 2 in 41 patients (85%) (Fig. 1A-C), grade 3A in 5 patients (10%), and grade 3B in the other 2 patients (4%). The clinical stage was stage I in 21 patients, stage II1 in 8 patients, stage II2 in 6 patients, and stage IV in 13 patients. Tests for H. pylori were positive in 17 of the 32 patients who were examined (53%).
In total, 16 patients underwent surgical resection with chemotherapy (n = 9) or without chemotherapy (n = 7), whereas 24 patients received treatment with nonsurgical strategies, including 13 patients who received R-CHOP, 6 patients who received rituximab monotherapy, and 5 patients who received antibiotic therapy. Four of the 5 patients who received antibiotic therapy were positive for H. pylori infection, whereas the other patient was negative for the infection. The remaining 8 patients were subjected to a watchful waiting strategy without any antitumor therapy. Overall, CR was achieved in 31 patients (65%), including 15 of 16 patients who underwent surgery, 10 of 13 patients who received R-CHOP, 4 of 6 patients who received rituximab alone, 1 of 5 patients who received antibiotics alone, and 1 of 8 patients who were subjected to watchful waiting.
Follow-up after the initial diagnosis of FL ranged from 6 months to 148 months (median, 33 months). During follow-up, the disease progressed or recurred in 9 patients, including 2 patients who had transformation to diffuse large B-cell lymphoma (DLBCL). The initial treatments for those 9 patients were watchful waiting (4 patients), antibiotic treatment (1 patient), and R-CHOP chemotherapy (4 patients). Five patients died, including 2 who died of primary disease. However, the causes of death were unknown for the other 3 patients. The OS and EFS rates after 5 years were 93% and 68%, respectively (Fig. 2).
T(14;18)/IgH-BCL2 in Gastrointestinal FL
FISH with the IGH/BCL2 dual-color, dual-fusion translocation probe was successful in all patients. Thirty-nine patients (81%) were positive for the t(14;18)/IGH-BCL2 translocation (Fig. 1D). Table 3 compares the clinicopathologic features between the t(14;18)-positive group and the t(14;18)-negative group. No differences were observed between the 2 groups with regard to age, sex, duodenal involvement, clinical stage, H. pylori infection, or initial management with watchful waiting, surgical resection, or chemotherapy. However, tumors that involved multiple sites within the gastrointestinal tract were more frequent in the positive group than in the negative group (69% vs 0%; P = .0002). The macroscopic type of lymphomatous polyposis and the mixed type with areas of lymphomatous polyposis were more frequent in the positive group than in the negative group (72% vs 22%; P = .0091), although the difference was not significant after a Bonferroni correction. In addition, histologic grade 1/2 tumors tended to be more frequent in the t(14;18)-positive positive group than in the negative group (92% vs 56%; P = .0170). In contrast, the CR rate was lower in the positive group (56%) compared with the negative group (100%; P = .0179). However, differences in the histologic grade and the CR rate did not reach statistical significance. Two patients who achieved CR after antibiotic treatment or after watchful waiting (spontaneous regression) were negative for t(14;18)/IgH-BCL2 (Patients 1 and 3) (Table 4). Although the difference was not statistically significant, PD and recurrent disease were more frequent in the t(14;18)-positive group (9 of 39 patients; 23%) than in the negative group (zero of 9 patients; 0%; P = .176).
Table 3. Clinical Features of Translocation (14;18)-Positive and Translocation (14;18)-Negative Patients
The detailed clinicopathologic findings from 9 patients who were negative for t(14;18)/IGH-BCL2 are listed in Table 4. In all patients, FL was localized to a single site, including 4 tumors in the stomach, 3 tumors in the ileum, and 2 tumors in the duodenum. Immunohistochemically, all 9 patients were positive for the CD10, BCL2, and BCL6 proteins. Genetic aberrations other than t(14;18) were identified in 3 patients. BCL6-involved translocation was detected in 1 patient with gastric FL (Patient 6). Two patients with intestinal FL (Patients 1 and 8) had extra copies of BCL2 (18q21) identified by FISH with the IGH/BCL2 dual-fusion probe, but not with the CEP18 probe, which indicated the presence of partial trisomy 18. In Patient 8, extra copies also were observed by FISH with both the CEP3 and BCL6 probes, suggesting complete trisomy 3. In the remaining 6 patients, neither BCL6-involved translocation nor trisomy 3 or 18 were detected.
Table 5 provides the results from Kaplan-Meier analyses for possible prognostic factors in our patients. The age, sex, duodenal involvement, gastric localization, lymphomatous polyposis type, histologic grade, clinical stage, and treatment modalities were not associated with OS or EFS. However, there was a trend toward poorer EFS for patients who had multiple involved sites compared with those who had the involvement of a single site (P = .095). Furthermore, patients with t(14;18)/IgH-BCL2 tended to have a poorer EFS than patients without t(14;18)/IgH-BCL2 (P = .089) (Fig. 3).
Table 5. Results From Kaplan-Meier Analysis of Prognosis
OS indicates overall survival; EFS, event-free survival; LP, lymphomatous polyposis; NA, not applicable; t(14;18)/IgH-BCL2, translocation t(14;18)(q32;q21) involving the immunoglobulin heavy chain gene (IGH) and the BCL2 gene.
Assessed by the log-rank test.
≤59, n = 25
≥60, n = 23
Men, n = 27
Women, n = 21
Yes, n = 26
No, n = 22
No. of involved sites
Single, n = 21
Multiple, n = 27
Gastric localization, n = 11
Intestinal involvement, n = 37
Yes, n = 30
No, n = 18
Grade 1 or 2, n = 41
Grade 3A or 3B, n = 7
I, n = 21
II1 or II2 or IV, n = 27
Watch/antibiotics, n = 13
Antitumor treatment, n = 35
Yes, n = 16
No, n = 32
Yes, n = 22
No, n = 26
Yes, n = 39
No, n = 9
The clinicopathologic features of gastrointestinal FL have been described in several publications with a relatively small number of patients.3, 14-17, 21 In our study, the most frequently involved site was the duodenum (54%). Such a predominance of duodenal involvement of FL is in agreement with most previous studies,3, 6, 7, 9, 14 However, LeBrun et al21 and Damaj et al15 reported that the ileum was the most frequent site. In recent studies that evaluated an extensive area of the small intestine by DBE, involvement of the jejunum and/or the ileum was identified in 80% to 100% of patients.5-9 In our current study, jejunoileal lesions were found identified in 90% of the patients who underwent DBE or SBE. In addition, gastric involvement was relatively frequent in our study (29%), in contrast to previous studies, in which the frequency ranged from only 0% to 26%.3, 14-16, 21 Among the various macroscopic types, the lymphomatous polyposis type was the most frequent in our study (58%). Lymphomatous polyposis, which implies that there are multiple, whitish, small, sessile polyps or nodules, is considered 1 of the most characteristic macroscopic findings of intestinal FL.3-7, 14-16
An optimal therapeutic strategy for gastrointestinal FL has not been established. Various treatments, including watchful waiting, surgery, chemotherapy, radiotherapy, immunotherapy (rituximab), or a combination of these, have been applied to patients with FL. In 2 reports by Kodama et al6 and Akamatsu et al,9 all of the patients who received R-CHOP therapy achieved CR, and the investigators emphasized the efficacy of immunochemotherapy for gastrointestinal FL. In contrast, Damaj et al15 and Mori et al17 reported that the prognosis (which they assessed as the median time to PD15 or progression-free survival17) did not differ between patients who received a certain antitumor treatment and who received no treatment (watchful waiting). In our current study, we also observed no differences in the OS or EFS between patients who initially were subjected to watchful waiting and those who received antitumor treatment. Although Feuerlein et al22 divided the first-line treatment for FL into “no” treatment (watchful waiting), “soft” treatment (rituximab or single-agent chemotherapy), and “intensive” treatment (eg, R-CHOP therapy), we consider watchful waiting 1 of the appropriate choices for the treatment of asymptomatic patients who have stage I, grade 1/2 gastrointestinal FL.
By using FISH, we identified t(14;18)(q32;q21)/IGH-BCL2 in 81% of our patients with gastrointestinal FL. The prevalence of t(14;18)/IGH-BCL2 in FL differs greatly according to the site of origin. Whereas 60% to 90% of patients with nodal FL harbor t(14;18), the majority of patients with cutaneous FL were negative for the translocation.12 In contrast, 50% of FLs of the thyroid gland had this translocation.13 Shia et al14 detected t(14;18)/IGH-BCL2 in all 4 of their patients with gastrointestinal FL using either the G-banding karyotype technique or polymerase chain reaction (PCR) for the rearrangement of the BCL2 major breakpoint region (MBR). In a study by Damaj et al,15IGH-BCL2 rearrangement at MBR was identified in 11 of 14 patients (79%) by PCR. Kodama et al16 detected t(14;18)/IGH-BCL2 in 7 of 10 patients (70%) using FISH. More recently, Mori et al17 reported that they detected the translocation in 20 of their 24 patients (83%) with duodenal FL by using FISH. The prevalence of the translocation in our patients (81%) was similar to the prevalence reported by Damaj et al15 and Mori et al17
The clinical significance of t(14;18)/IGH-BCL2 in FL is controversial. The translocation reportedly was associated with a favorable prognosis with regard to OS and EFS,23-25 was unrelated to prognosis,26-28 and even was associated with a poor prognosis.12, 29, 30 With regard to prognosis, we observed that t(14;18)/IGH-BCL2-positive patients had a lower CR rate and tended to have a poorer EFS than t(14;18)-negative patients. Matsumoto et al30 reported that the presence of t(14;18) was associated with a poor response to therapy and poor disease-free survival. Similarly, Goodlad et al12 reported that, in patients with nodal or extranodal FL, those with t(14;18)-positive disease had a poorer OS and disease-specific survival than those with t(14;18)-negative disease. In addition, t(14;18)/IGH-BCL2 was identified in 5% to 20% of patients with DLBCL,31-34 and the translocation was associated with poor survival in patients with DLBCL in various clinical observations.29, 31, 32 Furthermore, the transformation from FL to DLBCL reportedly occurs more frequently in t(14;18)-positive patients than in t(14;18)-negative patients,10, 27 which we observed in our current cohort. On the basis of these observations, it appears to be possible that t(14;18)/IGH-BCL2 may be a predictor of an adverse clinical course in patients with gastrointestinal FL. In our study, however, the finding that 7 of 8 patients who initially were subjected to watchful waiting belonged to the t(14;18)-positive group may have influenced the lower CR rate for those patients (Table 3).
Several previous studies suggested that gastrointestinal FL has a better prognosis than nodal FL.35 However, our current results indicated a similar prevalence of t(14;18)/IGH-BCL2 in gastrointestinal FL and nodal FL. Thus, the translocation itself may not be associated directly with a poor prognosis. Most patients with gastrointestinal FL manifest localized disease at the time of diagnosis, whereas most patients with nodal FL have multiple involved sites, resulting in an advanced disease at the time of staging procedures. Such a difference in disease stage may account for the different prognoses for nodal FL and gastrointestinal FL.
The t(14;18)/IGH-BCL2 translocation is considered an initial event in the development of FL, resulting in the overexpression of BCL2 protein, which confers a growth advantage to the affected B-cell through inhibition of apoptosis.1 However, this translocation alone is insufficient for the development of lymphoma. The additional accumulation of genetic alterations is detected in 90% of patients and commonly includes loss of chromosomes 1p, 6q, 10q, and 17p (involving p53) and gains of chromosomes 1q, 6p, 7, 8q, 12q, and 18q (involving BCL2).1, 36 In nodal FL, Cheung et al36 reported that 1p36 and 6q23 were independent prognostic indicators for both OS and transformation to DLBCL, as determined by array-based comparative genomic hybridization. In patients with t(14;18)-negative FL, translocations involving BCL6,10, 37, 38 trisomy 3,38, 39 and trisomy 1810, 37 have been observed. It is noteworthy that these genetic aberrations have been associated with histologic grade 3A or 3B disease.10, 13, 37-39 In our current study, patients who had grade 3A or 3B tumors also frequently were t(14;18)-negative rather than t(14;18)-positive (44% vs 8%). However, we were able to identify the alternative genetic alterations (BCL6 translocation, trisomy 3 or 18) in only approximately 33% of the patients without t(14;18). It has been reported that 26% to 68% of FLs without t(14;18) lack expression of the BCL2 protein,10, 37, 38 although we did not enroll such BCL2-negative patients in this study. In our 2 patients who had extra copies of BCL2 (partial trisomy 18), amplification of the BCL2 gene may have contributed to overexpression of the BCL2 protein.
It is noteworthy that our t(14;18)-negative patients tended to have a higher CR rate, although they had higher histologic grade tumors (grade 3A or 3B). Such a favorable clinical course for the translocation-negative group may be explained in part by the finding that the tumors in those patients were localized to a single gastrointestinal site and that approximately 66% of those patients underwent successful surgical resection. It also has been demonstrated that patients with pediatric FL are sensitive to treatment and have a better prognosis,1 although they are characterized by high-grade histology and lack of the t(14;18)/IGH-BCL2. In contrast, patients with typical nodal FL who are positive for t(14;18) usually are resistant to treatment, although their clinical course is indolent. Thus, patients with FL comprise a heterogeneous group with various clinical courses, and their prognosis cannot be predicted only by their t(14;18)/IGH-BCL2 status.
The retrospective nature of our investigation did not allow endoscopic assessment of the entire gastrointestinal tract in half of the patients. Thus, it seems possible that we may have underestimated the prevalence of small bowel involvement. It is also possible that we underestimated the impact of t(14;18)/IGH-BCL2 on the efficacy of treatment, because data collection for a prolonged period revealed heterogeneity in the choice of therapy. Therefore, a prospective study with standardized treatment arms should be undertaken to confirm the clinical significance of the translocation.
In summary, we observed that, in addition to nodal FL, t(14;18)(q32;q21)/IGH-BCL2 frequently occurs in gastrointestinal FL, and this translocation may be a predictor of an adverse clinical disease course. Further clinical studies of a large number of patients with longer follow-up are warranted to clarify whether this genetic alteration is associated with the prognosis of patients with gastrointestinal FL.
We are grateful to Akiko Ohgami and Hideko Noguchi for technical assistance.
CONFLICT OF INTEREST DISCLOSURES
This study was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Sciences, and Technology of Japan (no. 20590744).