Hematological malignancies with t(9;11)(p21-22;q23)--a laboratory and clinical study of 125 cases. European 11q23 Workshop participants

This paper reports clinical and cytogenetic data from 125 cases with t(9;11)(p21-22;q32) which were accepted for a European Union Concerted Action Workshop on 11q23. This chromosome abnormality is known to occur predominantly in acute myeloid leukemia (AML) FAB type M5a and less often in AML M4; in this series it was also found to occur, uncommonly, in other AML FAB types, in childhood acute lymphoblastic leukemia (ALL) (nine cases), in relatively young patients with myelodysplastic syndrome (MDS) (five cases), acute biphenotypic leukemia (two cases), and acute undifferentiated leukemia (one case). All age groups were represented but 50% of the patients were aged less than 15 years. The t(9;11) was the sole abnormality in 57 cases with AML; trisomy 8 was the most common additional abnormality (23 cases, including seven with further abnormalities), and 28 cases had other additional abnormalities. Among the t(9;11)+ve patients with AML, the white cell count (WBC) and age group were significant predictors of event-free survival; central nervous system (CNS) involvement or karyotype class (sole, with trisomy 8, or with other), also contributed to prognosis although our data could not show these to be independent factors. The best outcome was for patients aged 1-9 years, with low WBC, and with absence of CNS disease or presence of trisomy 8. For patients aged less than 15 years, the event-free survival for ALL patients was not significantly worse than that of AML patients.     

High incidence of translocations t(11;14)(q13;q32) and t(4;14)(p16;q32) in patients with plasma cell malignancies

Abnormalities involving the 14q32 region are recurrent chromosomal changes in plasma cell malignancies. Recent preliminary molecular analyses found IGH rearrangements in almost 100% of human myeloma cell lines and in 75% of patients. However, no systematic study analyzing the nature of the partner chromosomal regions have been reported thus far. To define the exact incidence of illegitimate IGH rearrangements and the respective incidence of partner genes cloned to date, we analyzed 141 patients with either multiple myeloma (MM, n = 127) or primary plasma cell leukemia (PCL, n = 14) using fluorescence in situ hybridization. The overall incidence of illegitimate recombinations was 57% (80 of 141 patients). Analysis of this incidence according to Durie and Salmon stage, patients' status, i.e., MM versus primary PCL and diagnosis versus relapse, immunoglobulin type and subtype, and beta2-microglobulin value, did not show any correlation. To analyze the nature of the partner chromosomal region, we selected probes specific for the following genes: FGFR3 (4p16), MYC (8q24), CCND1 (11q13), MAF (16q23), and BCL2 (18q21). These probes, combined with differentially labeled 14q32 probes, were used for dual-color fluorescence in situ hybridization on interphase plasma cells. Among the 80 patients with illegitimate IGH rearrangement, we identified 23 IGH-CCND1 fusion cases [i.e., t(11;14)], 17 IGH-FGFR3 fusion cases [i.e., t(4;14)], 3 IGH-MYC fusion cases [i.e., t(8;14)], and only one IGH-MAF fusion case. No IGH-BCL2 fusion case was detected. In 37 of 80 patients, none of these partner genes was involved. Analysis of cases with specific translocations according to their bioclinical features at diagnosis did not show any correlation. This study demonstrated that CCND1 and FGFR3 genes are involved together in about 50% of MM and primary PCL patients with illegitimate IGH rearrangements.     

Prevention of maternal and developmental toxicity in rats via dietary inclusion of common aflatoxin sorbents: potential for hidden risks

The chromosomal translocation t(11;14)(q13;q32) fuses the IGH and CCND1 genes and leads to cyclin D1 overexpression. This genetic abnormality is the hallmark of mantle cell lymphoma (MCL), but is also found in some cases of atypical chronic lymphocytic leukemia (CLL), characterized by a poor outcome. For an unequivocal assessment of this specific chromosomal rearrangement on interphase cells, we developed a set of probes for fluorescence in situ hybridization (FISH). Northern blotting was performed for analysis of the cyclin D1 expression in 18 patients. Thirty-eight patients, with either a typical MCL leukemic phase (17 patients) or atypical CLL with an MCL-type immunophenotype, i.e., CD19-, CD5+, CD23-/low, CD79b/sIgM(D)++, and FMC7+ (21 patients), were analyzed by dual-color interphase FISH. We selected an IGH-specific BAC probe (covering the JH and first constant regions) and a commercially available CCND1 probe. An IGH-CCND1 fusion was detected in 28 of the 38 patients (17 typical MCL and 11 cases with CLL). Cyclin D1 was not overexpressed in two patients with typical MCL and an IGH-CCND1 fusion. In view of the poor prognosis associated with MCL and t(11;14)-positive CLL, we conclude that this set of probes is a valuable and reliable tool for a rapid diagnosis of these entities.     

Internal tandem duplication of the FLT3 gene is preferentially seen in acute myeloid leukemia and myelodysplastic syndrome among various hematological malignancies. A study on a large series of patients and cell lines

In this study, we examined a large number of patients to clarify the distribution and frequency of a recently described FLT3 tandem duplication among hematopoietic malignancies, including 112 acute myelocytic leukemia (AML), 55 acute lymphoblastic leukemia (ALL), 37 myelodysplastic syndrome (MDS), 20 chronic myelogenous leukemia (CML), 30 non-Hodgkin's lymphoma (NHL), 14 adult T cell leukemia, 15 chronic lymphocytic leukemia (CLL) and 38 multiple myeloma (MM). We also evaluated 71 cell lines derived from 11 AML, 31 ALL, two hairy cell leukemia, three acute unclassified leukemia, 10 CML, 12 NHL including six Burkitt's lymphoma, and two MM. Using genomic PCR of exon 11 coding for the juxtamembrane (JM) domain and first amino acids of the 5'-tyrosine kinase (TK) domain, this length mutation was found only in AML (22/112, 20%) and MDS (1/37). According to the FAB subclassification, they were 5/18 (28%) of M1, 4/29 (14%) of M2, 3/17 (18%) of M3, 6/24 (25%) of M4, 4/20 (20%) of M5 and 1/9 of refractory anemia with excess of blast in transformation. In the various cell lines examined, this abnormality was determined in only one derived from AML and never found in other hematological malignancies. The sequence analysis of the abnormal PCR products revealed that 23 of 24 showed internal tandem duplication with or without insertion of nucleotides. In one AML, insertion and deletion without duplication was determined. All 24 lengthened sequences were in-frame. Duplication takes place in the sequence coding for the JM domain and leaves the TK domain intact. In conclusion, we emphasize that the length mutation of FLT3 at JM/TK-I domains were restricted to AML and MDS. Since all these mutations resulted in in-frame, this abnormality might function for the proliferation of leukemic cells.     

t(8;21)急性髓系白血病患者MICM分型及预后的回顾性分析

目的 分析t(8;21)急性髓系白血病(AML)患者的细胞形态学、免疫表型、遗传学、分子生物学(MICM)分型及临床治疗疗效.方法 运用瑞特染色法、FAB细胞形态分类标准、流式细胞术(FCM)直接免疫荧光标记技术、遗传学染色体吉姆萨显带技术及RT-PCR技术对70例确认有t(8;21)与AML1-ETO融合基因双阳性的AML患者及70例正常染色体核型的AML患者进行分析和比较.结果 70例t(8;21)AML患者中M11例,M2 64例,M4 3例,无法分型的急性白血病(AL)2例;免疫表型分析发现CD13、CD33、CD34、CD117高表达,40%表达CD19,11%表达CD15,10%表达CD11b,7%表达CD7;遗传学显示50%的t(8;21)AML患者有附加染色体异常,主要为性染色体丢失、9q-及超二倍体;RT-PCR检测AML1-ETO融合基因100%阳性.CD+19t(8;21)AML患者完全缓解(CR)率72%,CD+19伴CD+7t(8;21)AML患者CR率为0,正常核型CR率31%.结论 t(8;21)AML患者主要在M2中集中出现,附加染色体异常较多见.CD19表达较高,而CD7表达极低,CD34、CD117高表达,这些抗原的表达可能与核型密切相关.CD+19是预后良好的指标,但同时出现CD+7,则预后不良.     

Phenotypic conversion from t(8;21) acute myeloid leukemia to MLL gene rearrangement-positive acute lymphoblastic leukemia

Phenotypic conversion from acute myeloid leukemia (AML) to acute lymphoblastic leukemia (ALL) is rare. A 38-year-old man was initially diagnosed as having AML (FAB-M2) associated with the t(8;21)(q22;q22) chromosomal abnormality. The blasts showed myeloperoxidase (MPO) activity and CD13 antigen expression. He showed complete remission after standard chemotherapy for AML. However, the patient relapsed with blasts showing ALL morphology (FAB-L1), MPO negativity, and CD19 antigen expression 33 months after cessation of AML therapy. Cytogenetic analysis at relapse was unsuccessful. Molecular analysis of ALL blasts revealed immunoglobulin heavy-chain gene and MLL gene rearrangements but no AML1 gene. MLL gene rearrangement or the 11q23 chromosomal abnormality has been associated with therapy-related leukemia. The subsequent ALL in our patient may have been induced by the chemotherapy including daunorubicin, known as a topoisomerase II inhibitor.     

Induction of death (CD95/FAS), activation and adhesion (CD54) molecules on blast cells of acute myelogenous leukemias by TNF-alpha and IFN-gamma

Leukemic growth is determined by the balance of cell proliferation, differentiation and cell death. In vitro, the blasts of acute myelogenous leukemia (AML) proliferate under the influence of certain positive and negative regulators (cytokines). We conducted this study to determine whether cytokines could induce markers of cell death (FAS/Apo-1/CD95), of cell activation (HLA-DR) and cell adhesion (ICAM-1, CD54) in AML cell lines and primary AML samples. As inducers, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma were chosen. At baseline, CD95 and CD54 were weakly and HLA-DR was strongly expressed. CD95 was induced by TNF in 6/12 myeloid leukemia cell lines, and by IFN in 9/12 cell lines. Taken together, CD95 was upregulated by at least one cytokine in 11/12 cell lines. HLA-DR was inducible in 10/12 cell lines, with IFN being more potent than TNF. CD54 showed the strongest induction: TNF resulted in a more than 20-fold induction in positive cell lines, and IFN resulted in a more than 20-fold induction. In primary AML samples, CD95 was induced in 14/14 samples examined, with TNF being more potent than IFN. HLA-DR expression was increased by IFN in 12/15 samples and by TNF in 11/13 samples. The inducibility of HLA-DR by IFN was inversely correlated with baseline expression. As in the cell lines, CD54 was induced in most cases of AML. In addition to the induction of surface markers by cytokines, the culture of leukemia cells with fetal calf serum increased the expression of these markers, especially CD95 and CD54. Our results demonstrate that CD95 is not downregulated when TNF binds to its receptors, but is induced in cell lines and patient samples. Despite the induction of expression of CD95 (all cases of AML and most cell lines), 7/8 myelogenous leukemia lines and 6/7 patient samples remained resistant to CD95 triggering by antibody or by CD95 ligand, which suggests a lesion in normal cell signaling. As a positive control, a T-cell line (Jurkat) with 60% to > 90% apoptotic cells after a 22 h incubation was used. The number of CD95-binding sites was not correlated with the induction of apoptosis. The resistance of most cases of AML to CD95 triggering despite inducible expression may also be related to leukemia-specific antagonists of CD95 signal transduction, and requires further investigation. Altogether, our results indicate that surface markers related to apoptosis, activation and adhesion can be induced on AML blasts, and could be relevant to treatment strategies that exploit ligand binding to these surface epitopes.     

Analysis of myeloid and lymphoid markers on the surface and in the cytoplasm of mononuclear bone marrow cells in patients with myelodysplastic syndrome

Mononuclear cells of the bone marrow (BM) of patients in various subgroups of the myelodysplastic syndrome (MDS) were studied by flow cytometry for the expression of myeloid and lymphoid markers both on the surface and in the cytoplasm. A significantly higher percentage of the BM cells of MDS patients reacted with monoclonal antibodies (mAbs) to myeloid antigens (CD13, CD15 and CD33) by cytoplasmic staining as compared with cell surface staining. The percentage of BM cells expressing CD34 was markedly elevated in patients with RAEB-T. A distinct finding in MDS patients was the expression of myeloid antigens on mononuclear BM cells. The proportion of individuals whose mononuclear BM cells were positive for surface reactivity with anti-CD13 and anti-CD33 mAbs was highest among RAEB-T patients while none of the patients with RA expressed these surface antigens. Cytoplasmic staining significantly increased the percentage of CD13+ and CD33+ BM cells among RAEB and RAEB-T patients. The proportion of individuals whose BM cells possessed myeloid antigens was increased by cytoplasmic staining in all subgroups of MDS. The BM of a considerable proportion of RAEB-T and RAEB patients showed cells which coexpressed the CD7 and CD3 lymphoid markers along with the CD13 and CD33 myeloid antigens. The present study indicates the importance of comparative surface and cytoplasmic immunophenotyping with CD13 and CD33 mAbs for the diagnosis of subgroups of MDS. The coexpression of CD3 and CD7 with markers of the myeloid lineage may reflect derangement of the differentiation of pluripotent stem cells characteristic for MDS.     

Clinicopathological features in HLA-DR-negative acute myeloid leukemia

From a series of 176 patients with acute myeloid leukemia (AML), we have identified 11 patients with HLA-DR-negative AML excluding acute promyelocytic leukemia. All patients showed not (15; 17) or consistent chromosomal abnormalities. According to the French-British-American criteria, seven, three, and one patients were classified into M1, M2, and M4, respectively. Blasts from these patients were CD33+, HLA-DR-, and lymphoid-antigen-. All patients entered complete remission after induction therapy. Of these, blasts from four patients had strikingly invaginated nuclear membrane and finely granular peroxidase activity. The phenotype of the blasts was CD33+, CD34-, CD2-, and HLA-DR-. All four patients showed hyperleukocytosis on initial presentation. One patient received all-trans retinoic acid at relapse, however, the drug did not induce the differentiation of the blasts. These four patients were suspected to have acute myeloid/natural killer (NK) leukemia because of the prominent morphological feature of the blasts and the initial hyperleukocytosis, although NK cell activity of the blasts was not examined. Since HLA-DR-negative AML is heterogeneous, it is necessary for identifying acute myeloid/ NK leukemia within the disorder.     

Multiple sclerosis: use of MRI in evaluating new therapies

11q23 translocations (t(11q23)) are recurring cytogenetic abnormalities in both acute myeloid leukemia (AML) and acute lymphoblastic leukemia, involving the same gene, ALL1 (or MLL). Mixed lineage antigen expression has been reported in these leukemias, but its frequency and clinical significance are unknown. We immunophenotyped leukemia cells from 19 adult de novo AML patients with t(11q23) by multiparameter flow cytometry. Translocations included t(6;11)(q27;q23), t(9;11)(p22;q23), t(9;11;19)(p22;q23;q13.3), t(2;11)(11;17)(q37;q11q23;q11), t(11;17)(q23;q25), t(11;19)(q23;p13.1), t(11;19)(q23;p13.3) and t(11;22)(q23;q11). FAB types were M4 and M5. The committed stem cell and myeloid antigens HLADr, CD4dim, CD11b, CD13, CD15, CD32, CD33, CD38 and CD64 were each expressed in 80-100% of cases, and the early stem cell and lymphoid antigens CD34, CD56, CD3, CD2 and CD7 in 42, 39, 16, 5 and 5%, respectively. Antigen expression frequencies did not differ from those in 443 adequately karyotyped M4 and M5 cases without t(11q23). Fifteen patients (79%) attained complete remission (CR); median CR duration and survival were 10.0 and 15.1 months. CR duration and survival did not correlate with antigen expression. In particular, patients with t(9;11) survived longer than those with other t(11q23) (median not reached vs 7.6 months; P = 0.048), but antigen expression did not differ in the two groups. Thus frequencies of lymphoid antigen expression are similar in AML with t(11q23) and in other FAB M4 and M5 cases, treatment outcome does not differ in t(11q23) cases with and without lymphoid antigen expression, and better outcome of patients with t(9;11) compared to other t(11q23) does not correlate with differences in antigen expression. Mixed lineage antigen expression is not a distinctive feature of AML with t(11q23).     

Tissue inhibitor of metalloproteinase (TIMP)-1 induces differentiation and an antiapoptotic phenotype in germinal center B cells

Tissue inhibitors of metalloproteinases (TIMPs) have been shown to be multifunctional factors. Contrasting with their enzyme-inhibitory activity, TIMPs also promote cell growth. Previously, we have reported an enhanced expression of TIMP-1 by normal reactive B cells and high-grade lymphomas. In the present study, a series of Burkitt's lymphoma (BL) cell lines were analyzed for their expression of TIMP-1. TIMP-1 expression correlates with upregulation of activation and survival markers. TIMP-1-negative cells express the phenotype associated with group I BL lines and Epstein-Barr virus (EBV)-negative, nonendemic BLs (CD10+, CD38+, sIg+, and CD77+). However, TIMP-1+ BL lines showed group II/III BL phenotype, downregulation of the above markers, and upregulation and secretion of the activation marker CD23. Also, TIMP-1+ cells have high levels of CD40 expression. To determine whether TIMP-1 is directly involved in the BL phenotype, an EBV-negative BL line JD38 was infected with timp-1-expressing retrovirus and analyzed. In the absence of EBV, upregulation of TIMP-1 is sufficient to induce the same phenotype seen in TIMP-1+, EBV+ BL lines (CD10-, CD38-, sIg-, CD77-, CD23+, CD40 bright). This study not only suggests a role for TIMP-1 in BLs, but also supports its value as a prognostic factor. This is a US government work. There are no restrictions on its use.