Heterogeneity of lymphoblastic malignancies in children

Summary Lymphocyte surface markers show that lymphoblastic lymphomata in children are a heterogeneous but related group of diseases. Lymphoblastic lymphomas of T cell type fall into at least three subgroups: (1) HTLA positive, E rosette negative, TdT positive phenotype, with characteristically high levels of TdT and some associated expression of C-ALL antigen on a small proportion of cells. (2) E rosette positive lymphoblasts with intermediate range of TdT positivity, and which express antigens specific for thymic cortical lymphocytes. (3) E rosette positive and C3d positive T lymphoblasts with low levels of TdT enzyme.B lymphoblastic lymphomas show a major subgroup characterised by surface IgM expression, with or without detectable cytoplasmic IgM, and which may express ALL antigen. A minor subgroup of B lymphoblastic disease, of predominantly nodal presentation, expresses surface IgM with some expression of C3d receptors. Therefore at least 3 T cell, and 2 B cell subgroups of lymphoblastic lymphoma can be described.     

T-cell acute lymphoblastic leukemia

The T-cell variant accounts for about 15 to 25% of acute lymphoblastic leukemias in children and adults. In this perspective article. Drs. Chiaretti and Foà examine our present understanding of this disorder. See related article on page 224.T-cell acute lymphoblastic leukemia (T-ALL) accounts for about 15% and 25% of ALL in pediatric and adult cohorts respectively.¹ Patients usually have high white blood cell counts and may present with organomegaly, particularly mediastinal enlargement and CNS involvement. The biological knowledge of TALL has until recently been rather limited. Flow cytometry allowed the stage of differentiation of the leukemic clone to be defined, while cytogenetic analyses were often uninformative. The introduction of novel technologies has allowed an increasing number of alterations to be unraveled. The most relevant results have been obtained by using FISH (Fluorescent In Situ Hybridization), molecular biology and gene expression profiling, which have enabled five subgroups to be recognized, i.e. immature/LYL1, TAL1, HOX11, HOX11L2 and HOX.²The most important biological findings of T-ALL, their role in leukemia initiation and their possible therapeutic implications will be hereby revised.     

Heterogeneity of presenting features and their relation to treatment outcome in 120 children with T-cell acute lymphoblastic leukemia

Presenting features of 120 consecutive children with T-cell acute lymphoblastic leukemia (ALL), representing 15% of all patients diagnosed as having ALL during the study period, were analyzed to determine relationships with treatment outcome. Patients' ages ranged from 1.7 to 18.8 years (median, 10.3 years) and their leukocyte counts from 1.7 to 1,070 x 10(9)/L (median, 100 x 10(9)/L). Central nervous system (CNS) leukemia was present in 12.5% of the cases, a mediastinal mass in 61%, and L2 lymphoblast morphology in 32%. A relatively high proportion of cases, 26%, had normal karyotypes at presentation. Of the cases tested, membrane CD1 expression was found in 38% of cases, CD3 in 33%, CD4 in 50%, CD5 in 94%, CD8 in 55%, and CD10 in 35%. Four presenting features were found to confer an increased risk of treatment failure: age greater than or equal to 15 years, L2 lymphoblast morphology, abnormal karyotype, and membrane CD3 expression. This study illustrates the heterogeneity of presentations of childhood T-cell ALL and suggests that the relative importance of risk factors in ALL differs according to immunophenotype and treatment strategy.     

Adoptive T-cell therapy for B-cell malignancies

The success of allogeneic hematopoietic cell transplantation (HCT) for B-cell malignancies is evidence that these tumors can be eliminated by T lymphocytes. This has encouraged the development of specific adoptive T-cell therapy, both for augmenting the anti-tumor effect of HCT and for patients not undergoing HCT. T cells that are capable of recognizing antigens expressed on malignant B cells may be recruited from the endogenous repertoire or engineered to express tumor-targeting receptors. Critical insights into the qualities of T cells that enable their persistence and function in vivo have been derived, and obstacles to effective T-cell-mediated tumor eradication are being elucidated. These advances provide the tools to translate adoptive T-cell transfer into reliable clinical therapies.     

Epstein-Barr virus-associated T-cell acute lymphoblastic leukaemia

Epstein-Barr virus (EBV) is frequently detected in haematological malignancies, including Burkitt's lymphomas/leukaemias, Hodgkin's diseases and non-Hodgkin's lymphomas. However, immature T-cell malignancies associated with EBV have not been reported previously. We report a patient with T-cell acute lymphoblastic leukaemia (T-ALL), whose leukaemic cells had EBV, confirmed by Southern blotting and in situ hybridization. The EBV existed in episomal form and was detected in most leukaemic cells, but not in bystander normal B-cells. The leukaemic cells, massively infiltrated into the liver and spleen, were resistant to chemotherapy. EBV might be associated with tumorigenesis of T-ALL, and characteristic clinical features of the patient.     

Treatment of refractory T-cell malignancies using gemcitabine

We have treated 10 patients with relapsed or refractory T-cell malignancies using gemcitabine. The drug was administered intravenously over 30 min at a dose of 1200 mg/m2 on d 1, 8 and 15 of each 28-d cycle. The mean age of the patients was 62 years and the mean number of administered cycles was three. Of the 10 patients, two achieved a complete response and four a partial response, for an overall response rate of 60% (95% confidence interval 20--35%). The toxicity of chemotherapy was mild and manageable in all patients. These encouraging results warrant further investigation of gemcitabine either as a single agent or in combination regimens as early salvage treatment for patients with refractory T-cell haematological neoplasms.     

Somatic rearrangement of T-cell antigen receptor gene in human T-cell malignancies.

A cDNA clone representing the gene encoding the beta chain of the human T-cell antigen receptor has been isolated recently. By using fragments of this cDNA as hybridization probes in Southern blot analysis of restriction endonuclease-digested genomic DNA, we have now examined the structure of the gene in DNA from 26 patients with acute leukemia and from 23 normal individuals. We have found that the T-cell antigen receptor gene has undergone somatic rearrangement in 14 of 14 patients with the phenotypic diagnosis of T-cell acute lymphoblastic leukemia. In this group of patients, similar patterns of rearrangement appear to occur in different patients. This finding suggests that there is either a limited repertoire of possible rearrangements or an association between the development of leukemia and specific patterns of rearrangement. DNA from 6 patients with acute myeloblastic leukemia, 6 patients with non-B, non-T acute lymphoblastic leukemia, and 23 nonleukemic individuals showed no rearrangement or polymorphism. One case of T-cell acute lymphoblastic leukemia, however, showed rearrangement of both the T-cell receptor beta chain and the constant region of the immunoglobulin gene. Studies with mixtures of DNAs from leukemic bone marrow cells and cultured skin fibroblasts, as well as with remission and relapse marrow DNAs from the same patients, indicate that this technique can detect 1% leukemic cells in a mixed population. In addition, DNA from the marrow of a patient in relapse contains a similar rearrangement to that found in the marrow sample taken at the time of diagnosis, which suggests that the original clone of leukemic cells was responsible for relapse. Our results indicate that assessment of rearrangement of the T-cell antigen receptor gene will be valuable in the diagnosis and management of leukemia and can be used to evaluate clonality in T-cell neoplasia.     

Stages of T-cell receptor protein expression in T-cell acute lymphoblastic leukemia.

In this study five monoclonal antibodies (MoAbs) to T-cell receptor (TCR) proteins (WT31, alpha F1, beta F1, TCR delta-1 and delta TCS-1) were used to identify discrete maturative stages in 40 cases of T-cell acute lymphoblastic leukemia (T-ALL). These MoAbs reacted exclusively with CD3+ T cells and did not label B-lineage and myeloid cells. In 17 of the 40 T-ALL cases studied the leukemic blasts lacked membrane and cytoplasmic TCR chains (group I). In 12 cases cells did not have membrane CD3/TCR but expressed cytoplasmic TCR proteins heterogenously: nine cases had cytoplasmic TCR beta chains (beta F1+, alpha F1-; group II), one case had cytoplasmic TCR alpha chains (alpha F1+, beta F1-; group III), and two cases were labeled by both alpha F1 and beta F1 MoAbs (group IV). The remaining 11 cases were mCD3+: nine were TCR alpha beta+ (group Va) and two exhibited TCR gamma delta (TCR delta-1+, delta TCS-1+; group Vb). The analysis of the TCR beta, -gamma, and -delta gene configurations in 23 of the 40 T-ALLs showed that: (1) the lack of TCR protein expression was due to the lack of TCR gene rearrangements only in one of nine cases; (2) five of five TCR beta+, TCR alpha- cases studied had germline TCR alpha genes (ie, no detectable TCR delta gene deletions); (3) seven of eight cases with TCR delta gene deletions expressed TCR alpha proteins, whereas in 12 of 20 of the T-ALLs with TCR beta gene rearrangements the synthesis of the corresponding protein occurred; only 2 of 16 cases with rearranged TCR delta genes expressed TCR delta chains. The T-ALL categories identified with anti-TCR MoAbs did not have additional characteristic phenotypic patterns and may correspond to the normal stages of T-cell development more precisely than those defined by other differentiation antigens.     

T-cell receptor delta gene rearrangement in childhood T-cell acute lymphoblastic leukemia

During the development of functional T lymphocytes, a variety of genes involved in antigen recognition undergo somatic rearrangement. These include the alpha, beta, and gamma chain genes. Recently a fourth rearranging gene, the delta chain gene, embedded in the alpha chain locus, has been described. We have determined the structure of the beta, gamma, and delta chain genes in 15 cases of T-cell acute lymphoblastic leukemia (T-ALL) representing stage I (CD7+, CD1-, CD3-) and stage II (CD7+, CD1+, CD3-) of intrathymic T-cell development. The alpha-delta locus was rearranged in 14 of the 15 cases. In three cases the delta constant region was deleted on both chromosomes, suggesting biallelic V-J alpha rearrangement. A limited pattern of rearrangement of the delta locus was observed in the remaining 11 cases. When the alpha-delta region was rearranged, there was rearrangement of the beta and gamma TcR in all cases except two; in these cases the beta chain was in the germline configuration. These findings support the hypothesis that delta chain gene rearrangement is an early event in T- cell development, possibly contemporary to gamma gene rearrangement, and that the delta locus has a limited repertoire.     

T-cell regulation of erythropoiesis during acute lymphoblastic leukemia

How and where erythropoiesis is maintained during advanced leukemic disease is an important and, as yet, unresolved question in hematology. To address the potential role of T-lymphocytes as cells that regulate CFU-E differentiation during leukemogenesis, an experimental model of disease has been developed in inbred Balb/c mice. Specifically, three-week-old Balb/c By mice were injected with murine sarcoma virus-murine leukemia virus-Moloney (MSV-MuLV-M), which resulted 6-8 months later in the development of immunoblastic T-cell sarcomas with a leukemic phase. Splenic T cells from either normal or tumor-bearing mice were assessed for their relative ability to modulate erythroid differentiation. Quantitatively, T cells, Ly1 or Ly 2,3 T-cell subsets isolated from tumor-bearing animals significantly enhanced erythropoiesis when compared with comparable normal T-cell subsets. These data suggest that the compensatory shift of erythropoiesis from the bone marrow to the spleen observed during leukemogenesis was facilitated by splenic T cells. In this circumstance, the enhanced erythropoietic function may be mediated by splenic T cells, which are selectively activated by virus.     

Molecular-genetic insights in paediatric T-cell acute lymphoblastic leukaemia

Paediatric T-cell acute lymphoblastic leukaemia (T-ALL) is an aggressive malignancy of thymocytes that accounts for about 15% of ALL cases and for which treatment outcome remains inferior compared to B-lineage acute leukaemias. In T-ALL, leukemic transformation of maturating thymocytes is caused by a multistep pathogenesis involving numerous genetic abnormalities that drive normal T-cells into uncontrolled cell growth and clonal expansion. This review provides an overview of the current knowledge on onco- and tumor suppressor genes in T-ALL and suggests a classification of these genetic defects into type A and type B abnormalities. Type A abnormalities may delineate distinct molecular-cytogenetic T-ALL subgroups, whereas type B abnormalities are found in all major T-ALL subgroups and synergize with these type A mutations during T-cell pathogenesis.     

Aberrant T-cell antigen expression in B lymphoblastic leukaemia

B lymphoblastic leukaemia (B-ALL) cells are characterized by the expression of various B-cell antigens. Expression of T/Natural Killer-cell antigens, however, has rarely been reported in B-ALL (TAg+ B-ALL), and the significance of this aberrant antigen expression is unclear. We thus analysed the frequency of TAg+ B-ALL at our institution and investigated its significance in the context of immunophenotypes, cytogenetic/molecular findings, and prognosis. We reviewed 134 consecutive cases of B-ALL and found 18 cases (13·4%) of TAg+ B-ALL. The most common aberrant T-cell antigens expressed were CD2, CD5, and CD7 at equivalent rates (each in six cases), CD4 (two cases), and CD56 (three cases). Adverse cytogenetic abnormalities were seen in a significantly larger proportion of the TAg+ cases (72·2%) than the TAg- cases (32·2%; P = 0·003). Multivariate Cox-regression analysis showed that the risk of relapse over time was higher in the TAg+ cases, independent of high risk status (based on age and white blood cell count) and the presence of adverse cytogenetic abnormalities (hazard ratio = 2·256, P = 0·065). These findings suggest that T-cell antigen expression in B-ALL may be an independent predictor of poor prognosis, and a useful marker to identify patients at increased risk for relapse and for harbouring adverse cytogenetic abnormalities.     

Gene expression profiling in T-cell acute lymphoblastic leukemia

T-cell acute lymphoblastic leukemia (T-ALL) presents a difficult medical problem. T-ALL's clinical features and the biological properties of the leukemia cells are not predictive of prognosis, and thus have not been useful for risk-specific adjustments in therapeutic intensity. Microarray gene expression analyses of T-cell leukemic lymphoblasts have not only improved our understanding of the biological heterogeneity of this disease but have revealed clinically relevant molecular subtypes. Five different multistep molecular pathways have been identified that lead to T-ALL, involving activation of different T-ALL oncogenes: (1) HOX11, (2) HOX11L2, (3) TAL1 plus LMO1/2, (4) LYL1 plus LMO2, and (5) MLL-ENL. Gene expression studies indicate activation of a subset of these genes-HOX11, TAL1, LYL1, LMO1, and LMO2-in a much larger fraction of T-ALL cases than those harboring activating chromosomal translocations. In many such cases, the abnormal expression of one or more of these oncogenes is biallelic, implicating upstream regulatory mechanisms. Among these molecular subtypes, overexpression of the HOX11 orphan homeobox gene occurs in approximately 5% to 10% of childhood and 30% of adult T-ALL cases. Patients with HOX11-positive lymphoblasts have an excellent prognosis when treated with modern combination chemotherapy, while cases at high risk of early failure are included largely in the TAL1- and LYL1-positive groups. Supervised learning approaches applied to microarray data have identified a group of genes whose expression is able to distinguish high-risk cases. Further analyses of gene expression signatures of T-ALL lymphoblasts are especially needed for patients treated on modern combination chemotherapy trials to clearly distinguish the 10% to 15% of patients who fail induction or relapse in the first year of treatment. These high-risk patients would be ideal candidates for more intensive therapies in first remission, such as myeloablative regimens with stem cell rescue. Based on the rapid pace of research in T-ALL, made possible in large part through microarray technology, deep analysis of molecular pathways should lead to new and much more specific targeted therapies.     

Defective expression of the T-cell receptor-CD3 zeta chain in T-cell acute lymphoblastic leukaemia

This study analysed the T-cell receptor (TCR)-CD3 zeta complex and the signal transduction apparatus of T-acute lymphoblastic leukaemia (T-ALL) blasts, and investigated the function of the ubiquitin-proteasome system. In all nine T-ALL samples studied, the leukaemic cells showed a marked reduction in the expression of the zeta chain, while a variety of tyrosine kinases (p56lck, ZAP70 and SYK) were normally present. There was no expression of the FcepsilonRIgamma chain. To confirm that this aberration was specific to immature T-ALL blasts, we investigated two patients with lymphoproliferative disorders of granular lymphocytes (LDGL), characterized by the expansion of mature T lymphocytes and found normal zeta chain expression. The reduction of the zeta chain protein was not reversible after 72 h stimulation with the anti-CD3 monoclonal antibody and interleukin 2, either alone or in combination. Northern blot analysis indicated that the reduced protein expression did not correspond to a defect at the mRNA level, nor were mutations in the coding region of the zeta chain found. We, therefore, hypothesized that the observed reduction of protein expression in T-ALL blasts could be secondary to an increased degradation at the proteasome level. Following selective inhibition of the proteasome, a marked increase of the zeta chain expression was observed. Moreover, an increase in the surface expression of CD3 was also documented. Taken together, these results indicate that the expression of the zeta subunit of the TCR-CD3 complex is consistently reduced in T-ALL blasts and that degradation of the protein is mediated by the proteasome system.     

Heterogeneity of immunologic markers and surface morphology in childhood lymphoblastic lymphoma

The neoplastic cells from seven patients with childhood lymphoblastic lymphoma were studied for cell surface markers and surface morphology in the scanning electron microscope (SEM). The cells were studied for surface immunoglobulin (Slg), complement receptors (EAC), receptors for cytophilic antibody (IgGEA), and nonimmune rosette formation with sheep red blood cells (E). In one patient the cells exclusively bound E, suggesting a T-lymphocytic origin. In two patients the cells bound EAC, but demonstrated no other B-lymphocytic markers. In two patients no markers were detected, and in two patients receptors for both E and EAC were demonstrated. Additional studies in one of these patients permitted simultaneous demonstration of both markers on the same neoplastic cells. The neoplastic cells were also examined by SEM after fixation and critical point dehydration. No consistent surface morphology was observed. In four patients the cells were predominately smooth, whereas in two patients variable numbers of surface microvilli were present. A correlation of the surface features with membrane markers could not be established. A comparison of the surface markers with clinical and cytologic features revealed clinical homogeneity in spite of the heterogeneous immunologic markers. This heterogeneity was most likely a reflection of neoplastic alteration and disordered differentiation of the cells. The observation of complement receptors on the cells of four cases is a feature not previously reported in this disease and should be investigated in other presumed T-cell malignancies.