Impact of constitutional TET2 haploinsufficiency on molecular and clinical phenotype in humans.

Clonal hematopoiesis driven by somatic heterozygous TET2 loss is linked to malignant degeneration via consequent aberrant DNA methylation, and possibly to cardiovascular disease via increased cytokine and chemokine expression as reported in mice. Here, we discover a germline TET2 mutation in a lymphoma family. We observe neither unusual predisposition to atherosclerosis nor abnormal pro-inflammatory cytokine or chemokine expression. The latter finding is confirmed in cells from three additional unrelated TET2 germline mutation carriers. The TET2 defect elevates blood DNA methylation levels, especially at active enhancers and cell-type specific regulatory regions with binding sequences of master transcription factors involved in hematopoiesis. The regions display reduced methylation relative to all open chromatin regions in four DNMT3A germline mutation carriers, potentially due to TET2-mediated oxidation. Our findings provide insight into the interplay between epigenetic modulators and transcription factor activity in hematological neoplasia, but do not confirm the putative role of TET2 in atherosclerosis.

High familial risk in nodular lymphocyte-predominant Hodgkin lymphoma.

PURPOSE: Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) is one of the two established Hodgkin lymphoma (HL) subtypes. The risk factors of NLPHL are largely unknown. In general, genetic factors are known to have a modest effect on the risk of HL; however, familial risk in NLPHL has not been previously examined. We conducted a population-based study by using the Finnish registries and evaluated the familial risk in NLPHL. PATIENTS AND METHODS: We launched a population-based search to identify patients with NLPHL and their relatives by examining the records of the Finnish Cancer Registry, established in 1953, and the official Finnish population registries. We collected a data set of 692 patients with NLPHL, identified their 4,280 first-degree relatives, and calculated the registry-based standardized incidence ratios (SIRs) for different cancers in the first-degree relatives. In addition, the primary tumor biopsies of HL-affected relatives were collected when possible, the HL diagnoses were re-reviewed by a hematopathologist, and the SIR for NLPHL was calculated on the basis of confirmed NLPHL diagnoses. RESULTS: On the basis of confirmed NLPHL diagnoses, the SIR for NLPHL was 19 (95% CI, 8.8 to 36) in the first-degree relatives. The risk was most prominent in female relatives of young patients. The registry-based SIR for classical HL was 5.3 (95% CI, 3.0 to 8.8), and for non-Hodgkin lymphoma, it was 1.9 (95% CI, 1.3 to 2.6). CONCLUSION: Our results implicate an unexpectedly high familial component in the development of NLPHL. Research is warranted to identify the putative genetic and environmental factors underlying this finding and to develop strategies for better management of patients with NLPHL and their relatives.

Exome sequencing reveals germline NPAT mutation as a candidate risk factor for Hodgkin lymphoma.

A strong clustering of Hodgkin lymphoma in certain families has been long acknowledged. However, the genetic factors in the background of familial Hodgkin lymphoma are largely unknown. We have studied a family of 4 cousins with a rare subtype of the disease, nodular lymphocyte predominant Hodgkin lymphoma. We applied exome sequencing together with genome-wide linkage analysis to this family and identified a truncating germline mutation in nuclear protein, ataxia-telangiectasia locus (NPAT) gene, which segregated in the family. We also studied a large number of samples from other patients with Hodgkin lymphoma, and a germline variation leading to the deletion of serine 724 was found in several cases suggesting an elevated risk for the disease (odds ratio = 4.11; P = .018). NPAT is thus far the first gene implicated in nodular lymphocyte predominant Hodgkin lymphoma predisposition.

Common SNP in pre-miR-146a decreases mature miR expression and predisposes to papillary thyroid carcinoma.

Although papillary thyroid carcinoma (PTC) displays strong heritability, no predisposing germ-line mutations have been found. We show that a common G/C polymorphism (rs2910164) within the pre-miR-146a sequence reduced the amount of pre- and mature miR-146a from the C allele 1.9- and 1.8-fold, respectively, compared with the G allele. This is matched by a similar decrease in the amount of each pre-miR generated from the corresponding pri-miR-146a in an in vitro processing reaction. The C allele also interfered with the binding of a nuclear factor to pre-miR-146a. The reduction in miR-146a led to less efficient inhibition of target genes involved in the Toll-like receptor and cytokine signaling pathway (TRAF6, IRAK1), and PTC1 (also known as CCDC6 or H4), a gene frequently rearranged with RET proto-oncogene in PTC. In an association study of 608 PTC patients and 901 controls, we found marked differences in genotype distribution of rs2910164 (P = 0.000002), the GC heterozygous state being associated with an increased risk of acquiring PTC (odds ratio = 1.62, P = 0.000007), and both homozygous states protective with odds ratio = 0.42 for the CC genotype (P = 0.003) and odds ratio = 0.69 for the GG genotype (P = 0.0006). Moreover, 4.7% of tumors had undergone somatic mutations of the SNP sequence. Thus, our data suggest that a common polymorphism in pre-miR-146a affects the miR expression, contributes to the genetic predisposition to PTC, and plays a role in the tumorigenesis through somatic mutation. Preliminary evidence suggests that these effects are mediated through target genes whose expression is affected by the SNP status.

Platelet-derived growth factor receptor family mutations in gastrointestinal stromal tumours.

OBJECTIVE: Activating mutations of either KIT or platelet-derived growth factor receptor alpha (PDGFRA) genes are present in the majority of gastrointestinal stromal tumours (GISTs). The type of gene mutation is associated with the aggressiveness of the disease, response to imatinib therapy, and the tumour site in the gastrointestinal tract. However, a subgroup of GISTs does not harbour these mutations. MATERIAL AND METHODS: Thirty-three GISTs were studied for mutations in exons encoding the juxtamembrane and the activation loop domains of KIT, PDGFRA, PDGFRB, CSF1R, and FLT3 genes using denaturing high-performance liquid chromatography and gene sequencing. RESULTS: Twenty-two (67%) GISTs had mutation in KIT and 3 (9%) in PDGFRA. The three PDGFRA mutations were all detected in exon 18 of the gene. Three of the 5 GISTs that had weak to moderate KIT expression had a PDGFRA mutation as compared to none of the 26 cases with strong KIT immunopositivity (p=0.022). No mutations were found in PDGFRB, CSF1R or FLT3 in the 8 cases that did not harbour KIT or PDGFRA mutations. CONCLUSIONS: KIT and PDGFRA are the most commonly mutated type III receptor tyrosine kinase genes in GIST. GISTs with PDGFRA mutations often have reduced expression of the KIT protein in immunohistochemistry, suggesting that immunohistochemistry may be potentially useful in identification of such GISTs.

The role of microRNA genes in papillary thyroid carcinoma.

Apart from alterations in the RET/PTC-RAS-BRAF pathway, comparatively little is known about the genetics of papillary thyroid carcinoma (PTC). We show that numerous microRNAs (miRNAs) are transcriptionally up-regulated in PTC tumors compared with unaffected thyroid tissue. A set of five miRNAs, including the three most up-regulated ones (miR-221, -222, and -146), distinguished unequivocally between PTC and normal thyroid. Additionally, miR-221 was up-regulated in unaffected thyroid tissue in several PTC patients, presumably an early event in carcinogenesis. Tumors in which the up-regulation (11- to 19-fold) of miR-221, -222, and -146 was strongest showed dramatic loss of KIT transcript and Kit protein. In 5 of 10 such cases, this down expression was associated with germline single-nucleotide changes in the two recognition sequences in KIT for these miRNAs. We conclude that up-regulation of several miRs and regulation of KIT are involved in PTC pathogenesis, and that sequence changes in genes targeted by miRNAs can contribute to their regulation.

MET receptor tyrosine kinase sequence alterations in differentiated thyroid carcinoma.

Activating mutations affecting the MET receptor tyrosine kinase are present in several types of human cancer, particularly in papillary renal cell carcinoma. Papillary thyroid carcinomas commonly express high levels of MET mRNA and protein, suggesting that increased MET signaling may be of importance in the molecular pathogenesis of differentiated thyroid carcinoma. To evaluate the role of MET mutations in thyroid carcinoma, we screened MET exons 2 to 21 for mutations in sporadic papillary, follicular, medullary, and anaplastic thyroid carcinomas using denaturing high-performance chromatography. A missense MET sequence alteration T1010I, located in exon 14 encoding for the juxtamembrane domain of MET, was found in 6 (6%) of the 104 thyroid carcinomas examined, whereas all 92 goiter samples had wild-type exon 14 (P = 0.031). Three (6%) of the 53 papillary, 2 (10%) of the 21 follicular, 1 (8%) of the 13 medullary, and none of the 17 anaplastic carcinomas studied had MET(T1010I). Four of the 6 T1010I sequence alterations were present also in the germline. MET protein expression showed no apparent association with the presence of MET(T1010I), and the clinical features of the patients with cancer with MET(T1010I) were similar to those of patients whose cancer did not harbor MET(T1010I). We conclude that MET(T1010I) sequence alteration is relatively frequent in differentiated thyroid carcinoma. The clinical and the molecular pathologic significance of this MET sequence alteration is not known.

KIT and platelet-derived growth factor receptor alpha tyrosine kinase gene mutations and KIT amplifications in human solid tumors.

PURPOSE: Mutated KIT and platelet-derived growth factor receptor alpha (PDGFRalpha) tyrosine kinases are the principal targets for imatinib mesylate in the treatment of gastrointestinal stromal tumors (GISTs). The frequency of activating KIT and PDGFRA gene mutations in most other histologic types of human cancer is not known. MATERIALS AND METHODS: KIT exons 9, 11, 13, and 17 and PDGFRA exons 11 and 17 of 334 human cancers were screened for mutations using sensitive denaturing high-performance liquid chromatography (DHPLC). In addition, all KIT exons from 9 to 21 of 115 tumors were screened. Thirty-two histologic tumor types were examined. Samples with abnormal findings in DHLPC were sequenced. Immunostaining for the KIT protein (CD117) was performed in 322 (96.4%) of the 334 cases. RESULTS: Of the 3,039 exons screened, only 17 had mutation. All 17 cases with either mutated KIT (n = 15) or PDGFRA (n = 2) were histologically GIST tumors, whereas none of the other histologic types of cancer (n = 316) harbored KIT or PDGFRA mutation. KIT immunostaining was rarely positive except in GISTs (18 of 18), small-cell lung cancer (10 of 30; 33%), and testicular teratocarcinoma (four of 17; 24%). Wild-type KIT gene amplification or chromosome 4 aneuploidy was common (seven of 12) in non-GIST tumors with strong KIT protein expression when studied with fluorescence in situ hybridization. CONCLUSION: Despite frequent KIT protein expression in some tumor types, KIT and PDGFRA gene mutations are uncommon in most human cancers. Cancer KIT expression is frequently associated with multiple copies of the wild-type KIT gene.

AMES, MACIS and TNM prognostic classifications in papillary thyroid carcinoma.

The need for total thyroidectomy and extended for lymphadenectomy and the need for postoperative radioiodine ablation in the treatment of papillary thyroid carcinoma is continuously debated. Since less aggressive treatment in low-risk patients has been suggested, several scoring systems have been developed to identify low-risk patients. In the current study, we compared the AMES, MACIS and TNM staging systems in predicting carcinoma-specific mortality in papillary thyroid carcinoma. Between 1967 and 1994, 495 patients with papillary thyroid carcinoma were treated at the Department of Surgery, Helsinki University Central Hospital. Carcinoma-specific mortality in the AMES low-risk group, comprising 89.7% of these patients, was 2.4%. Corresponding figures for the MACIS were 89.9%, and 2.4%, and for the TNM 85.9% and 1.2%. The mortality ratio, at 10 years, between low-risk and high-risk patients was 22.2 for the AMES, 25.0 for the MACIS and 41.8 for the TNM system. The proportion of explained variance in the Cox model was 16.3 for the AMES, 30.0 for the MACIS taken as a conitinuous variable and 28.9 for the TNM stage. The TNM stage was on average superior to the MACIS or AMES score in predicting cancer-specific mortality of patients with papillary thyroid carcinoma. This may be explained by the fact that the TNM system includes the prognostic effect of nodal metastases, which is included in neither the MACIS nor AMES systems.

Lymphomas diagnosed by percutaneous kidney biopsy.

BACKGROUND: Occult renal lymphoma clinically mimicking renal medical disease constitutes a diagnostic challenge to nephrologists, radiologists, and renal pathologists. The clinical and radiological findings, mostly nonspecific or inconclusive, seldom obviate the need for a kidney biopsy. METHODS AND RESULTS: We report 5 new cases of diffuse bilateral renal lymphoma diagnosed by percutaneous kidney biopsy, all presenting with acute renal failure (ARF) of unknown cause. Three cases showed an interstitial and 2 an intraglomerular/intravascular type of lymphomatous infiltration. All tumors were of B-cell lineage. Our cases add to 50 similar cases reported since 1980. Considering all 55 cases together, 39 (87%) of the 44 cases with interstitial and 5 of 11 (45%) of those with intraglomerular lymphoma presented with ARF. In contrast, 5 of 10 cases with intraglomerular but none with interstitial infiltration presented with nephrotic range proteinuria. All but 2 cases (95%) with ARF and interstitial lymphoma but none with ARF and intraglomerular lymphoma showed bilaterally enlarged kidneys. Signs of extrarenal lymphomatous involvement were detected in 24 cases (44%) at the time of kidney biopsy or shortly thereafter. However, in only 10 cases (18%), all with interstitial lymphoma, was a tumor suspected prior to biopsy, mainly based on radiographical evidence of enlarged kidneys. CONCLUSION: Both types of diffuse bilateral renal lymphoma may clinically mimic renal medical disease. ARF in interstitial and in intraglomerular lymphoma may be due to increased intrarenal pressure and intraglomerular obstruction, respectively. Percutaneous kidney biopsy provides the most expedient means of establishing the diagnosis. Differential diagnosis includes interstitial nephritis and proliferative glomerulonephritis.

Cyclin A and Ki-67 expression as predictors for locoregional recurrence and outcome in laryngeal cancer patients treated with surgery and postoperative radiotherapy.

PURPOSE: To investigate the association between the cancer cell proliferation fraction and the risk of recurrence in laryngeal cancer patients treated without systemic therapy. METHODS AND MATERIALS: Paraffin-embedded tumor samples from 90 laryngeal cancer patients were stained for cyclin A and the Ki-67 antigen by immunohistochemistry. The patients were treated with partial or total laryngectomy followed by postoperative radiotherapy to a total dose of 50 Gy or greater. The median follow-up time was 91 months (minimum 48 months). RESULTS: High cyclin A expression (>19% positive cancer cell nuclei, the highest tertile) was associated with a high rate of locoregional tumor recurrence and unfavorable disease-free and overall survival as compared with cases with a lower expression (p = 0.025, 0.032, and 0.042, respectively). In a multivariate analysis, high cyclin A expression was an independent predictor of poor disease-free survival (RR 2.4, 95% CI 1.2-4.9, p = 0.013) and overall survival (RR 2.1, 1.2-3.6, p = 0.012), together with a poor Karnofsky's performance status and the presence of positive margins at surgery. Ki-67 expression was not an independent predictor of survival, but cancers with high Ki-67 expression (>34% nuclei positive, the highest tertile) recurred more frequently locoregionally when treated with split-course radiotherapy than when treated with a continuous course of therapy (p = 0.035), whereas the presence of a planned split did not influence the frequency of locoregional recurrences when Ki-67 expression was lower (p = 0.93). CONCLUSION: Cancer cell cyclin A expression is a novel predictive factor for outcome in laryngeal cancer treated with surgery and postoperative radiotherapy. Planned gaps in the radiotherapy course are deleterious in patients with a high proliferative fraction, and immunostaining for the Ki-67 antigen may be useful in identification of such patients.

Primary treatment with autologous stem cell transplantation in mantle cell lymphoma: outcome related to remission pretransplant.

OBJECTIVES: The aim of the first Nordic mantle cell lymphoma (MCL) protocol was to study the clinical significance of an augmented CHOP induction chemotherapy followed by high-dose chemotherapy and autologous stem cell transplantation (ASCT) and to examine the prognostic significance of stem cell contamination rates in newly diagnosed patients with MCL. PATIENTS AND METHODS: Forty-one newly diagnosed patients below 66 yr were enrolled and given three series of an augmented CHOP regimen. Responders underwent stem cell mobilization with a fourth course of CHOP, stem cell harvest and ASCT. Stem cell purging was optional in the protocol and followed the routine of each participating centre. The number of tumour cells in the reinfused autografts was estimated by flow cytometry or quantitative PCR. RESULTS: Induction therapy led to complete remission (CR) in 11 of 41 patients (27%), partial remission (PR) in 20 of 41 patients (49%) and no response in nine patients (22%), whereas one patient was not evaluable. Twenty-seven of the 31 responders underwent ASCT and 24 achieved or maintained a CR. The overall and failure-free 4-yr survival on intention-to-treat basis were 51% and 15%, respectively. Among the transplanted patients, a significantly increased failure-free (P<0.03) and overall survival (P=0.03) was noted among patients transplanted in CR compared with PR, respectively. By contrast, reinfusion of highly variable numbers of tumour cells with the autografts (range 0.71-80 x 10(6) tumour cells), did not affect outcome. CONCLUSION: In MCL, an important strategy to improve the outcome will be to intensify the induction chemotherapy.

Lymphotoxin beta expression is high in chronic lymphocytic leukemia but low in small lymphocytic lymphoma: a quantitative real-time reverse transcriptase polymerase chain reaction analysis.

BACKGROUND AND OBJECTIVES: The lymphotoxin beta (LTB) gene, localized to the major histocompatibility complex region on chromosome 6p21.3, has an important role in the formation of germinal center reactions and regulation of immune response and apoptosis. Our aim was to determine LTB gene expression in different hematologic neoplasias. DESIGN AND METHODS: We determined the expression of LTB using quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) on a series of RNA samples from CD3(+) T cells and CD19(+) B cells isolated from peripheral blood (n=7); CD19(+) B cells isolated from lymph nodes (n=11) and from patients with acute lymphoblastic leukemia (ALL; n=16), acute myeloid leukemia (AML; n=43), chronic myeloid leukemia (CML; n=12), mantle cell lymphoma (MCL; n=19), chronic lymphocytic leukemia (CLL; n=32) and small lymphocytic lymphoma (SLL; n=22). RESULTS: The expression level of LTB in CD3(+) T cells and CD19(+) B cells isolated from blood was ten to forty times lower than that in normal CD19(+) B cells isolated from lymph nodes. In malignant myeloid cells the expression levels were very low, whereas in malignant lymphoid cells the expression was higher than in myeloid cells, being highest in MCL and CLL (20.2+/-14.0 ng/microL and 81.0+/-116.3 ng/microL) and low in SLL (4.5+/-3.6 ng/microL; p=0.001). We did not find correlations between LTB expression and hematoclinical parameters (risk groups, immunophenotypes and overall survival). INTERPRETATION AND CONCLUSIONS: A distinct difference in expression of LTB in CLL and SLL indicates that these morphologically similar B-cell malignancies are molecularly different. Further studies are needed to investigate the prognostic value of LTB and any role that LTB may have in determining whether the malignant B cells manifest a leukemia or lymphoma.

Abnormal expression of apoptosis-related genes in haematological malignancies: overexpression of MYC is poor prognostic sign in mantle cell lymphoma.

The expression of apoptosis-related genes BCL2, BAX, BCL2L1, BCL2A1, MCL1, DAPK1 and MYC was studied by quantitative real-time polymerase chain reaction on total RNA samples from patients with acute lymphoblastic leukaemia (ALL, n = 16), acute myeloid leukaemia (AML, n = 27), chronic myeloid leukaemia (CML, n = 12), mantle cell lymphoma (MCL, n = 19) and chronic lymphoid leukaemia (CLL, n = 32). BCL2, BAX, BCL2A1, MCL1, DAPK1 and MYC were overexpressed in all patient groups. BCL2L1 was underexpressed in CLL and CML, but not in AML, ALL and MCL. MCL1 levels were significantly higher in CD13 and CD33-positive ALL, and in CD56-positive AML samples. BCL2, BCL2L1, BCL2A1 and MCL1 were overexpressed and DAPK1 was underexpressed in CLL samples with a 11q23 deletion. MYC overexpression was significantly associated with shorter overall survival in MCL (P < 0.01). AML patients with a normal karyotype showed a higher frequency of BCL2A1 overexpression (P < 0.001) than those with an abnormal karyotype.

Hepatocyte growth factor receptor, matrix metalloproteinase-11, tissue inhibitor of metalloproteinase-1, and fibronectin are up-regulated in papillary thyroid carcinoma: a cDNA and tissue microarray study.

PURPOSE: To screen and validate the global gene expression in papillary thyroid carcinoma (PTC) using cDNA expression arrays and immunohistochemistry on tumor tissue microarrays in an attempt to find genes that may be of importance in the molecular pathogenesis and malignant progression of PTC. EXPERIMENTAL DESIGN: Eighteen PTC tissue specimens were compared with three morphologically normal thyroid specimens by applying Atlas Human Cancer 1.2 Array membranes printed with cDNAs of 1176 human genes involved in cancer. Results for selected genes were confirmed by reverse transcription-PCR. Protein expression of selected genes was further studied using a tissue microarray consisting of 107 PTCs and compared with histologically normal thyroid tissue samples. RESULTS: By cDNA arrays, two genes [c-MET and matrix metalloproteinase (MMP)-11] were expressed only in tumor tissue, where they were present in >50% of cases. Ten genes [macrophage inhibitory cytokine-1, CGD, fibronectin (FN), hypoxia-inducible factor 1, Fc-epsilon-receptor gamma-chain, lactate dehydrogenase A, HLA-DBP1, AH receptor, tissue inhibitor of metalloproteinase (TIMP-1), and glycyl-tRNA-synthetase] were found to be up-regulated >2-fold in 40-100% of cancers, whereas 9 genes (GADD153, polykystic kidney disease-1, CYR61, DPC4, HBA1, gravin, DLG3, protein tyrosine phosphatase sigma, and heterochromatin protein 1 homologue-alpha) were down-regulated to <50% of their normal levels in 40-94% of cases. Conventional reverse transcription-PCR gave consistent results with the cDNA array findings for all four genes selected to be studied (c-MET, FN, TIMP-1, and GADD153). Immunohistochemistry for three selected proteins, FN, MMP-11, and TIMP-1, showed positive staining in 81, 87, and 68% of the tumor samples, respectively. CONCLUSIONS: Several novel and previously undetected tumor promoting/inhibiting genes may be of importance in the molecular pathogenesis and malignant progression of PTC. Transcription of these genes may result in overexpression of proteins, such as c-MET, MMP-11, TIMP-1, and FN, which may contribute to the pathogenesis of PTC.

Investigatory and analytical approaches to differential gene expression profiling in mantle cell lymphoma.

Mantle cell lymphoma (MCL) is a non-Hodgkin's lymphoma of B-cell lineage. The blastoid variant of MCL, characterized by high mitotic rate, is clinically more aggressive than common MCL. We used the cDNA array technology to examine the gene expression profiles of both blastoid variant and common MCL. The data was analysed by regression analysis, principal component analysis and the naive Bayes' classifier. Eight genes were identified as differentially deregulated between the two groups. Oncogenes CMYC, BCL2 and PIM1 were upregulated more frequently in the blastoid variant than in common MCL. This implied that the gp130-mediated signal transducer and activator of transcription 3 (STAT3) signalling pathway was involved in the blastoid variant transformation of MCL. Other differentially deregulated genes were TOP1, CD23, CD45, CD70 and NFATC. By using the eight differentially deregulated genes, we created a classifier to distinguish the blastoid variant from common MCL with high accuracy. We also identified 18 genes that were deregulated in both groups. Among them, BCL1, CALLA/CD10 and GRN were suggested to be oncogenes. The products of RGS1, RGS2, ANX2 and CD44H were suggested to promote tumour metastasis. CD66D was suggested to be a tumour suppressor gene.

Ki-67 expression level, histological subtype, and the International Prognostic Index as outcome predictors in mantle cell lymphoma.

OBJECTIVES: Mantle cell lymphoma (MCL) is characterised by translocation t(11;14) (q13;q32) leading to rearrangement of bcl1/CCND1 and overexpression of the cell-cycle regulatory protein cyclin D1. We assessed the significance of the cell proliferation rate as an outcome predictor with the five components of the International Prognostic Index (IPI) and the histological subtypes of MCL. PATIENTS AND METHODS: The hospital case records and histopathological material of 127 patients diagnosed with MCL in a single centre were reviewed. The cell proliferation rate was assessed by Ki-67 immunostaining and mitosis counting. Cox's multivariate regression model was used in multivariate survival analyses. The median follow-up time was 87 months. RESULTS: The mantle zone/nodular subtype of the common variant (19%) was associated with median survival of 70 months, the diffuse subtype of the common variant (64%) of 35 months, and the blastoid subtype (17%) of 11 months (P < 0.001). Patients with Ki-67 expression in > or = 26% (the upper tertile) of the lymphoma cells had median survival of only 13 months as compared with 45 months in the rest of the patients (P < 0.001). In a multivariate analysis high Ki-67 expression, Ann Arbor stage III-IV, and age over 60 yr had independent influence on survival, whereas serum lactate dehydrogenase level, the number of extranodal disease sites, and performance status did not. CONCLUSIONS: The IPI may not be an optimal tool for outcome prediction in MCL, and a better prognostic index may be obtained by including Ki-67 expression and possibly the histological subtype in the index.