Calcium and cholecalciferol supplementation provides no added benefit to nutritional counseling to improve bone mineral density in survivors of childhood acute lymphoblastic leukemia (ALL).

BACKGROUND: We sought to improve lumbar spine bone mineral density (LS-BMD) in long-term survivors of childhood acute lymphoblastic leukemia (ALL) using calcium and cholecalciferol supplementation. PROCEDURE: This double-blind, placebo-controlled trial randomized 275 participants (median age, 17 [9-36.1] years) with age- and gender-specific LS-BMD Z-scores <0 to receive nutritional counseling with supplementation of 1,000 mg/day calcium and 800 International Unit cholecalciferol or placebo for 2 years. The primary outcome was change in LS-BMD assessed by quantitative computerized tomography (QCT) at 24 months. Linear regression models were employed to identify the baseline risk factors for low LS-BMD and to compare LS-BMD outcomes. RESULTS: Pre-randomization LS-BMD below the mean was associated with male gender (P = 0.0024), White race (P = 0.0003), lower body mass index (P < 0.0001), and cumulative glucocorticoid doses of ≥ 5,000 mg (P = 0.0012). One hundred eighty-eight (68%) participants completed the study; 77% adhered to the intervention. Mean LS-BMD change did not differ between survivors randomized to supplements (0.33 ± 0.57) or placebo (0.28 ± 0.56). Participants aged 9-13 years and those 22-35 years had the greatest mean increases in LS-BMD (0.50 ± 0.66 and 0.37 ± 0.23, respectively). Vitamin D insufficiency (serum 25[OH]D <30 ng/ml) found in 296 (75%), was not associated with LS-BMD outcomes (P = 0.78). CONCLUSION: Cholecalciferol and calcium supplementation provides no added benefit to nutritional counseling for improving LS-BMD among adolescent and young adult survivors of ALL (93% of whom had LS-BMD Z-scores above the mean at study entry).

Frequent mutations in SH2D1A (XLP) in males presenting with high-grade mature B-cell neoplasms.

X-linked lymphoproliferative syndrome (XLP) is caused by mutations in SH2D1A, and is associated with overwhelming infectious mononucleosis, aplastic anemia, hypogammaglobulinemia, and B-cell lymphomas. However, the frequency of SH2D1A mutations in males who present with B NHL is unknown. Five cases of XLP were diagnosed among 158 males presenting with B NHL (approximately 3.2%). Four of the patients had two episodes of B NHL and one had a single episode of B NHL followed by aggressive infectious mononucleosis. Prospective screening for XLP in males with B-cell lymphoma at the time of initial diagnosis should be considered.

IDH1 and IDH2 mutations in pediatric acute leukemia.

To investigate the frequency of isocitrate dehydrogenase 1 (IDH1) and 2 (IDH2) mutations in pediatric acute myeloid leukemia (AML) and acute lymphoid leukemia (ALL), we sequenced these genes in diagnostic samples from 515 patients (227 AMLs and 288 ALLs). Somatic IDH1/IDH2 mutations were rare in ALL (N=1), but were more common in AML, occurring in 3.5% (IDH1 N=3 and IDH2 N=5), with the frequency higher in AMLs with a normal karyotype (9.8%). The identified IDH1 mutations occurred in codon 132 resulting in replacement of arginine with either cysteine (N=3) or histidine (N=1). By contrast, mutations in IDH2 did not affect the homologous residue but instead altered codon 140, resulting in replacement of arginine with either glutamine (N=4) or tryptophan (N=1). Structural modeling of IDH2 suggested that codon 140 mutations disrupt the enzyme's ability to bind its substrate isocitrate. Accordingly, recombinant IDH2 R140Q/W were unable to carry out the decarboxylation of isocitrate to α-ketoglutarate (α-KG), but instead gained the neomorphic activity to reduce α-KG to R(-)-2-hydroxyglutarete (2-HG). Analysis of primary leukemic blasts confirmed high levels of 2-HG in AMLs with IDH1/IDH2 mutations. Interestingly, 3/5 AMLs with IDH2 mutations had FLT3-activating mutations, raising the possibility that these mutations cooperate in leukemogenesis.

A minimally hypomorphic mutation in Btk resulting in reduced B cell numbers but no clinical disease.

Reduced B cell numbers and a mutation in Btk are considered sufficient to make the diagnosis of X-linked agammaglobulinaemia. In the process of conducting family studies, we identified a 58-year-old healthy man with an amino acid substitution, Y418H, in the adenosine-5'-triphosphate binding site of Btk. Immunofluorescence studies showed that this man had 0.85% CD19+ B cells (normal 4-18%) in the peripheral circulation and his monocytes were positive for Btk. He had borderline low serum immunoglobulins but normal titres to tetanus toxoid and multiple pneumococcal serotypes. To determine the functional consequences of the amino acid substitution, a Btk- chicken B cell line, DT40, was transfected with expression vectors producing wild-type Btk or Y418H Btk. The transfected cells were stimulated with anti-IgM and calcium flux and inositol triphosphate (IP3) production were measured. Cells bearing the mutant protein demonstrated consistently a 15-20% decrease in both calcium flux and IP3 production. These findings indicate that even a modest decrease in Btk function can impair B cell proliferation or survival. However, a mutation in Btk and reduced numbers of B cells are not always associated with clinical disease.

Pediatric acute myeloid leukemia with NPM1 mutations is characterized by a gene expression profile with dysregulated HOX gene expression distinct from MLL-rearranged leukemias.

Somatic mutations in nucleophosmin (NPM1) occur in approximately 35% of adult acute myeloid leukemia (AML). To assess the frequency of NPM1 mutations in pediatric AML, we sequenced NPM1 in the diagnostic blasts from 93 pediatric AML patients. Six cases harbored NPM1 mutations, with each case lacking common cytogenetic abnormalities. To explore the phenotype of the AMLs with NPM1 mutations, gene expression profiles were obtained using Affymetrix U133A microarrays. NPM1 mutations were associated with increased expression of multiple homeobox genes including HOXA9, A10, B2, B6 and MEIS1. As dysregulated homeobox gene expression is also a feature of MLL-rearranged leukemia, the gene expression signatures of NPM1-mutated and MLL-rearranged leukemias were compared. Significant differences were identified between these leukemia subtypes including the expression of different HOX genes, with NPM1-mutated AML showing higher levels of expression of HOXB2, B3, B6 and D4. These results confirm recent reports of perturbed HOX expression in NPM1-mutated adult AML, and provide the first evidence that the NPM1-mutated signature is distinct from MLL-rearranged AML. These findings suggest that mutated NPM1 leads to dysregulated HOX expression via a different mechanism than MLL rearrangement.

Novel cryptic, complex rearrangements involving ETV6-CBFA2 (TEL-AML1) genes identified by fluorescence in situ hybridization in pediatric patients with acute lymphoblastic leukemia.

In childhood B-lineage acute lymphoblastic leukemia (ALL), the most common genetic change, the ETV6-CBFA2 (TEL-AML1) fusion resulting from the cryptic t(12;21)(p13;q22) is associated with a favorable outcome. Therefore, it is important to identify patients with this translocation so that they can receive appropriate treatment. To identify new partner breakpoints for ETV6 and CBFA2, we selected 30 patients with childhood ALL in whose leukemic cells a t(12;21) had been detected by RT-PCR. Conventional cytogenetics revealed that 12p abnormalities were present in 10 patients and that other random abnormalities were present in another 15, including 9 with a numerical or structural abnormality of chromosome 21. Normal karyotypes were observed in the leukemic blasts of five patients. Interphase fluorescence in situ hybridization (FISH) confirmed the RT-PCR finding of the t(12;21) in each patient and detected the loss of the wild-type ETV6 allele in 14 (47%) patients. Metaphase cells from only 20 patients were available for additional FISH analysis. In 13 patients, the expected fusion signal of t(12;21) was observed on der(21)t(12;21), and the reciprocal CBFA2 signal was observed on der(12)t(12;21). However, in six patients with the ETV6-CBFA2 fusion on chromosome 21, the reciprocal CBFA2 signal was observed not on 12p13 but on 4q21, 4q27, 8q24, 11q24, 14q11.2, or 16p13.1. In four of these six patients, we found interstitial insertions of part of CBFA2. In another patient, the ETV6-CBFA2 fusion was observed on 4q21 rather than on 21q. Thus, seven (35%) of the 20 patients with a t(12;21) revealed complex rearrangements. Our findings also indicate the importance of analyzing metaphase chromosomes in identifying cryptic and complex rearrangements involving ETV6 and CBFA2.

Lack of BCL10 mutations in multiple myeloma and plasma cell leukemia.

To determine whether the BCL10 mutation plays a role in the oncogenesis of plasma cell dyscrasias, we used polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) and direct sequencing analysis and examined the genomic BCL10 mutations in 57 patients with multiple myeloma or plasma cell leukemia and 52 normal bone marrow samples. We found three polymorphic sequence variants, either alone or in combination, at codons 5 and 8, and in intron 1 at base 58 of the BCL10 gene in 37 patients with plasma cell dyscrasia. Identical aberrant band shifts were also observed in 34 normal marrow samples. No polymorphic variants were identified in exon 2 or 3 in either patient or control samples, and no pathogenic mutations were detected. Patients with plasma cell dyscrasias in Taiwan appeared to have a higher frequency of polymorphisms at codon 5 and intron 1 at base 58, and a lower frequency at codons 8 and 213. Our results suggest that BCL10 is not involved in the oncogenesis of plasma cell dyscrasias.

Use of reverse transcriptase polymerase chain reaction for diagnosis and staging of alveolar rhabdomyosarcoma, Ewing sarcoma family of tumors, and desmoplastic small round cell tumor.

PURPOSE: To compare the use of reverse transcriptase polymerase chain reaction (RT-PCR) with that of morphology-based methods for diagnosis, staging, and detection of metastatic disease in pediatric alveolar rhabdomyosarcoma (ARMS), Ewing sarcoma family of tumors (ESFT), and desmoplastic small round cell tumors (DSRCT). MATERIALS AND METHODS: RT-PCR assays for the EWS-FLII, EWS-ERG, PAX3-FKHR, PAX7-FKHR, and EWS-WTI fusion transcripts were performed on RNA extracted from the primary tumor tissue, bone marrow, and body fluids obtained at initial presentation and relapse. Molecular findings were compared with original histologic diagnoses and results of staging procedures. RESULTS: Eighty-eight samples from 47 patients with ARMS (n = 13), ESFT (n = 31), or DSRCT (n = 3) were analyzed. The detection rate of metastatic disease was significantly higher with RT-PCR (95%) as compared with the morphologic methods (70%) for the three pediatric sarcomas studied. In primary tumors with characteristic fusion transcript, RT-PCR was positive in all cases with morphologic evidence of metastatic disease. Moreover, in six patients (3 with ARMS, 2 with DSRCT, and 1 with ESFT) with metastatic disease, micrometastases in bone marrow (4) and other sites (2) were detected by RT-PCR alone. Importantly, none of the patients with localized disease diagnosed had micrometastases detected by RT-PCR in bone marrow. CONCLUSIONS: The high sensitivity and specificity of RT-PCR for the characteristic fusion transcripts of pediatric sarcomas make it an ideal method to aid in the routine staging of these patients. In addition, the 100% sensitivity of RT-PCR in detection of micrometastasis makes it useful for follow-up and detection of minimal residual disease. However, the clinical significance of molecularly-detectable disease remains unknown. Further studies should aim to elucidate the therapeutic and prognostic implications of micrometastases detected by RT-PCR alone.

NPM-ALK Reverse Transcriptase-Polymerase Chain Reaction for Detecting the t(2;5) of Non-Hodgkin's Lymphoma.

The development of modern molecular biology techniques and their use in characterizing the genetic abnormalities that are of pathogenic significance in non-Hodgkin's lymphoma (NHL) now provides a means to diagnose and rationally subcategorize these neoplasms in addition to the more traditional use of morphologic and immunophenotypic criteria. The shortcomings of traditional methods of NHL diagnosis and classification have been especially evident within the morphological subset commonly referred to as the large-cell lymphomas, which comprise approx 25% and 40% of NHL in children and adults, respectively (1). The marked cytological, immunological, and clinical heterogeneity of this group of tumors suggests that it is comprised of several biologically different neoplasms, but morphological and immunophenotypic subclassification schema have failed to identify meaningful subsets within the large-cell lymphomas. Recently, however, two recurrent genetic abnormalities in large-cell NHL-activation of the BCL6/LAZ3 zinc finger gene located at 3q27 (altered in approx 30% of large-cell lymphomas [2-4]) and the NPM-ALK fusion gene (5-7) produced by the t(2;5)-have been analyzed to now permit the identification of patient subsets that have reproducibly different therapeutic responses and survival rates in most studies, both of these molecular genetic subtypes appearing to have a superior prognosis compared to those cases lacking these gene abnormalities.

Acute lymphoblastic leukemia in a developing country: preliminary results of a nonrandomized clinical trial in El Salvador.

PURPOSE: To improve outcome and study biology of childhood acute lymphoblastic leukemia (ALL) in El Salvador. PATIENTS AND METHODS: Between January 1994 and December 1996, 153 children of El Salvador had newly diagnosed ALL treated in a collaborative program between Hospital Benjamin Bloom and St. Jude Children's Research Hospital (SJCRH). Therapy was based on a modified SJCRH protocol, with uniform remission induction (prednisone, vincristine, L-asparaginase) followed-up by consolidation with teniposide/cytarabine and/or high-dose methotrexate. Continuation treatment was risk-stratified: 123 patients assigned to the high-risk group received weekly rotational drug pairs, and 16 assigned to the standard-risk group received daily 6-mercaptopurine, weekly methotrexate, and monthly pulses of vincristine plus dexamethasone. High risk was defined as: DNA index < 1.16, age 12 months or younger, white blood cell count > or = 50 x 10(9)/L, T-cell immunophenotype, anterior mediastinal mass, central nervous system leukemia at diagnosis, or t(4;11), t(1;19), or t(9;22). Duration of the continuation treatment was 2.5 years in both groups. The median age at diagnosis of all patients was 4.8 (range I d-17 yrs), median leukocyte count was 15 (range 1-766) x 10(9)/L, and sex distribution was equal. RESULTS: Immunophenotypes were early beta-progenitor in 79%, T-cell in 3.9%, and inconclusive in 17% of cases. DNA index was <1.16 in 80.5% and was > or = 1.16 in 19.5% of the 123 known cases. For the analyzes, patients who refused therapy (abandoned treatment) were considered to have treatment failure as of their last follow-up dates. Complete remission was achieved in 126 of 151 (82.4%) patients (11 abandoned therapy during induction). The overall 4-year event-free survival (EFS) rate +/- 1 standard error was 48 +/- 6%. The 4-year EFS rates in patients at high-risk and standard-risk were 46 +/- 7% (n = 121) and 69 +/- 15% (n = 16), respectively (P = 0.20). When patients who refused further treatment are censored, the corresponding 4-year estimates of EFS are 51 +/- 8% and 75 +/- 14%, respectively. CONCLUSIONS: These results suggest that the biology of childhood ALL in El Salvador appears to be similar to that seen in the United States. Risk-directed chemotherapy can successfully be used in developing countries, but risk factors must be carefully determined and applied.

Cytogenetic abnormalities associated with the t(12;21): a collaborative study of 169 children with t(12;21)-positive acute lymphoblastic leukemia.

The t(12;21)(p13;q22) is a cryptic abnormality observed in 25% of children with B-lineage acute lymphoblastic leukemia (ALL), associated with a favorable prognosis. To determine whether specific cytogenetic abnormalities accompany the t(12;21), we analyzed the cytogenetic profiles of blast cells from 169 ALL cases positive for the t(12;21), previously identified by molecular methods. Only 13.6% of samples had normal karyotypes. Structural changes were detected in 89.7% of abnormal karyotypes, and numerical abnormalities in 47%. Rearrangements of 12p were the most frequent structural aberration (57 out of 146 patients with chromosomal abnormalities). Nonspecific deletions of chromosomes 6 and 9 were also found. The most frequent numerical abnormalities was trisomy for chromosomes 21. Blast cells were pseudodiploid (45.6%), hyperdiploid with 47 to 51 chromosomes (24.3%), hypodiploid with 44 to 45 chromosomes (10%), near-triploid (0.6%), or near-tetraploid (5.9%). Our results show that the t(12;21) is not associated with hyperdiploidy of 52 to 68 chromosomes or with the prognostic t(1;19), t(4;11) or t(9;22). Only children with B-lineage ALL who lack these abnormalities detected by conventional cytogenetics will probably benefit from additional testing by molecular methods to detect the t(12;21).

A multiplex RT-PCR assay for the detection of chimeric transcripts encoded by the risk-stratifying translocations of pediatric acute lymphoblastic leukemia.

Modern therapy for pediatric acute lymphoblastic leukemia (ALL) is based on the principle of risk stratification. One of the most important laboratory features used to accurately risk stratify patients is the presence of specific chromosomal translocation within the leukemic blasts. In this paper, we describe a multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) assay for the accurate, sensitive, and rapid identification of chimeric transcripts encoded by the major risk-stratifying translocations of pediatric ALL. This assay will identify both the CML- and ALL-type BCR-ABL transcripts encoded by the t(9;22), all described variants of the E2A-PBX1 transcripts encoded by the t(1;19), the MLL-AF4 transcripts encoded by the t(4;11), and all variants of TEL-AML1 encoded by the t(12;21). In addition, we have developed a reverse dot-blot detection system as an alternative to traditional post-PCR Southern blot analysis. Application of this combined assay to the analysis of 70 leukemic samples and five cell lines resulted in a complete concordance between this multiplex assay and individual PCR reactions. The characteristics of the multiplex assay suggest that its application to routine clinical screening will significantly improve the ability of clinical laboratories to accurate risk stratify pediatric ALL patients.

12p abnormalities and the TEL gene (ETV6) in childhood acute lymphoblastic leukemia.

Although abnormalities involving the short arm of chromosome 12 (12p) are one of the most frequently observed rearrangements in childhood acute lymphoblastic leukemia (ALL), little is known about the frequency of different structural abnormalities and their relationship to the status of the ETV6 (also named TEL) gene in this region. Of 815 children with newly diagnosed ALL, 94 (11.5%) had a total of 104 cytogenetic 12p abnormalities. Loss of genetic material was observed in 67 (64%) of these abnormalities. Cases with 12p alterations had a much lower frequency of hyperdiploidy greater than 50 (7%) than did the ALL population in general, but these cases had a similar distribution of immunophenotype and similar 5-year event-free survival (70% +/- 5% SE v 64% +/- 2%, P = .64). Rearrangement of the ETV6 gene was identified in 36 (56%) of 64 cases evaluated. The ETV6-CBFA2 (TEL-AML1) fusion transcript was found in 25 (66%) of 38 cases evaluated, and all but one of these showed ETV6 rearrangement. Importantly, ETV6 rearrangement was associated with a favorable prognosis (5-year event-free survival: 89% +/- 6% v 60% +/- 1%, P < .01). We conclude that most but not all 12p cytogenetic abnormalities in childhood ALL involve ETV6, and that rearrangement of ETV6 is associated with a favorable treatment outcome.

Primitive neuroectodermal tumor of bone as a second malignant neoplasm in a child previously treated for acute lymphoblastic leukemia.

PURPOSE: Although rare, second malignant neoplasms (SMNs) are a devastating consequence of successful treatment of childhood cancer. The 15-year estimated risk of developing a second malignant neoplasm after treatment of childhood acute lymphoblastic leukemia (ALL) is 2.5%. Most of these neoplasms are central nervous system tumors. The risk of secondary acute myeloid leukemia has been negligible in most treatment regimens. Here, we report the first case of a primitive neuroectodermal tumor (PNET) in a patient treated for ALL. PATIENTS AND METHODS: A 15.7-year-old girl developed pain in her left leg 7 years after diagnosis of low-risk ALL. Imaging studies revealed lytic lesions in her left proximal tibia and several vertebra as well as metastatic nodules in both lungs. RESULTS: Immunocytochemical and molecular analyses led to the diagnosis of PNET. The treatment of this SMN was composed of combination chemotherapy with hematopoietic growth factor support and radiotherapy to the primary lesion and affected spine. The tumor recurred 5 months after the completion of treatment, and the patient is now undergoing salvage therapy composed of chemotherapy and radiotherapy. CONCLUSIONS: To our knowledge, this is the first report of PNET as an SMN after successful treatment of ALL.

TEL gene rearrangement in acute lymphoblastic leukemia: a new genetic marker with prognostic significance.

PURPOSE: TEL gene rearrangements due to the 12;21 chromosomal translocation are the most common molecular genetic abnormality in childhood acute lymphoblastic leukemia (ALL), occurring in approximately 25% of cases with a B-precursor immunophenotype. The limited number of clinically useful genetic markers in this leukemia subtype prompted us to assess TEL status as a predictor of treatment outcome. PATIENTS AND METHODS: We determined the status of the TEL gene (rearranged or germline) in 188 cases of B-precursor acute leukemia using Southern blot analysis and related the findings to event-free survival. All comparisons of outcome were stratified by treatment regimen, risk classification, age, and leukocyte count. RESULTS: Forty-eight patients (26%) had a rearranged TEL gene. At 5 years of follow-up, an estimated 91% +/- 5% (SE) of this group were event-free survivors, compared with only 65% +/- 5% of the group with germline TEL (stratified log-rank P = .011). For nonhyperdiploid patients, the odds ratio of an adverse event in the germline TEL group to that for the rearranged TEL group was 4.06 (95% confidence interval, 1.86 to 8.84). The relationship of TEL rearrangement to a favorable prognosis was independent of recognized good-risk features in B-precursor leukemia, including age, initial leukocyte count, and hyperdiploidy. CONCLUSION: Rearrangement of the TEL gene distinguishes a large subset of children with favorable-prognosis B-precursor leukemia who cannot be identified by standard prognostic features. It may be possible to treat these patients less aggressively without loss of therapeutic efficacy.

Disseminated intrathoracic desmoplastic small round-cell tumor: a case report.

PURPOSE: Recently recognized as a distinct clinicopathologic entity, desmoplastic small round-cell tumors typically affect young men. These aggressive tumors usually arise in the abdomen; other sites of primary disease have been described only rarely. We report the case of an extraabdominal primary tumor with widespread dissemination, including the subcutaneous tissue, a previously unrecognized metastatic site. PATIENT AND METHODS: We describe the case of a 16-year-old boy with a primary extraabdominal metastatic desmoplastic small round-cell tumor. RESULTS: Our patient had a primary intrathoracic desmoplastic small round-cell tumor and widespread dissemination involving the subcutaneous tissue, kidney, liver, bone, and lymph nodes. Histopathologic analysis found intense desmoplasia and polyphenotypic expression of neural, muscle, and epithelial markers. Reverse transcriptase-polymerase chain reaction analysis of fresh tumor tissue confirmed the characteristic EWS-WT1 transcript. CONCLUSIONS: Broader than originally anticipated, the clinical spectrum of desmoplastic small round-cell tumors continues to evolve. Primary intrathoracic tumors with soft-tissue dissemination and polyphenotypic expression should prompt suspicion of this malignancy. Molecular analysis of fresh tumor tissue is an important adjunct to diagnosing this rare neoplasm.

Molecular analysis of the PML/RAR alpha chimeric gene in pediatric acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is characterized cytogenetically by the t(15;17)(q22;q11-21) translocation. To compare molecular events among pediatric and adult APL cases, we designed two sets of oligonucleotide primers using published cDNA sequence for PML/RAR alpha fusion transcripts, and undertook reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of 22 US pediatric cases of APL. PML/RAR alpha fusion transcripts were detected in all APL cases, including two cases lacking cytogenetic evidence of t(15;17). Breakpoint usage in PML was determined using a combination of PCR amplification with differing 5' primers, junction-specific probes, and sequence analysis in selected cases. Consistent with previously published data, case analysis demonstrated fusion products resulting from three breakpoint cluster regions (bcr) in PML, and a single breakpoint region in intron 2 of RAR alpha. Transcripts resulting from breakpoints in bcr1 were detected in 59 percent of cases, bcr2 in 27 percent and bcr3 in 14 percent. This distribution is dissimilar to that observed in adults, where bcr2 comprises a lesser and bcr3 a greater portion of cases. These results suggest that the pathogenesis of the t(15;17) in APL may differ among patient sets. RT-PCR with these primer sets is a reliable method for detecting PML/RAR alpha chimeric transcript in t(15; 17)-containing APL.

High incidence of TEL/AML1 fusion resulting from a cryptic t(12;21) in childhood B-lineage acute lymphoblastic leukemia in Taiwan.

Despite its rarity by routine karyotypic analysis, cryptic t(12;21)(p12-13;q22) translocation leading to TEL/AML1 fusion has been recognized as the most frequent genetic rearrangement in childhood acute lymphoblastic leukemia (ALL) in two recent studies, one from France and the other from the United States. To estimate the frequency of this abnormality in the Chinese population, we studied 41 children with ALL and 17 with acute myeloid leukemia (AML) in two medical centers in Taiwan, using the reverse transcriptase polymerase chain reaction (RT-PCR) assay. Results of this analysis demonstrated a 17% frequency of this translocation in the ALL population overall and 19% in patients with B-lineage ALL, similar to previous findings in Caucasian children. None of the patients with AML had TEL/AML1 fusion transcripts. In addition to its association with the B-lineage immunophenotype, TEL/AML1 was also correlated with a low presenting leukocyte count and favorable age (1-10 years). These findings, combined with earlier reports, indicate that TEL/AML1 fusion is the most frequent genetic abnormality in childhood ALL, regardless of race. Molecular diagnosis of t(12;21)-positive ALL may identify a subgroup of patients who do not require intensive treatment for cure.

Molecular analysis of t(11;19) breakpoints in childhood acute leukemias.

MLL is fused to ENL or ELL in acute leukemias that contain t(ll;19)(q23;p13). Although ENL and ELL localize to chromosome 19, bands p13.3 and p13.1, respectively, these breakpoints are not always readily distinguished by standard cytogenetics. We therefore used reverse transcriptase-polymerase chain reaction (RT-PCR) assays to analyze 26 cases of childhood acute leukemia containing t(11;19) to determine the frequencies of ENL and ELL involvement. All 17 cases of acute lymphoblastic leukemia (ALL) had MLL/ENL fusion transcripts. By contrast, of the 9 cases of acute myeloid leukemia (AML) analyzed, 6 had MLL/ENL fusions, 2 had MLL/ELL fusions, and 1 case had no RT-PCR-detectable MLL fusion mRNA. These data suggest that the majority of 11;19 translocations involve ENL, whereas involvement of ELL is relatively uncommon in childhood acute leukemia and may be restricted to AML.

NPM-ALK Reverse Transcriptase-Polymerase Chain Reaction Analysis for Detection of the t(2;5) Translocation of Non-Hodgkin's Lymphoma.

The diagnosis and classification of non-Hodgkin's lymphoma (NHL) has traditionally been made based on morphologic and mununophenotypic criteria. Unfortunately, because of the diverse nature of this group of diseases, rehante solely on these criteria has frequently resulted in misdiagnosis. In addition, investigators have been limited m their ability to define biologically and clinically relevant non-Hodgkin's lymphoma subgroups using this diagnostic approach. The development of modern molecular biological techniques and their use to characterize the genetic abnormalities that are of pathogenic significance in NHL now provides an additional means to identify and to rationally subcategorize these neoplasms. The shortcomings of traditional methods of NHL diagnosis and classification have been especially evident within the morphological subset commonly referred to as the large-cell lymphomas, which comprise approx 25 and 40% of NHL m children and adults, respec tively (1). The marked cytological, immunological, and clinical heterogeneity of this group of tumors suggests that it is comprised of several biologically different neoplasms. However, morphological and immunophenotypic subclassification of the large-cell lymphomas has failed to identify a subset of tumors having a reproducibly different therapeutic response or patient survival rate. Until recently, recurrent genetic abnormalities characteristic of the large-cell lymphomas had not been identified.However, the characterization during the past 2 yr of chromosomal rearrangements mvolvmg the BCL6/LAZ3 zmc finger gene located at 3q27 (which is altered m approx 30% of these tumors [2,3]), and the cloning by our group (4) of the NPM-ALK fusion gene produced by the t(2;5) may now permrt the identrficatton of molecular genetic subtypes of largecell lymphoma that possess unique btological and/or clinical features.