Acute lymphoblastic leukemia of childhood: identification of two distinct regions of deletion on the short arm of chromosome 12 in the region of TEL and KIP1

Cytogenetic analysis of acute lymphoblastic leukemia (ALL) of childhood identified nonrandom chromosomal abnormalities of the short arm of chromosome 12. The alterations include deletions that are thought to be indicative of the presence of a tumor suppressor gene that is mutated on the remaining allele. To refine further the chromosomal localization of this gene, we analyzed the loss of heterozygosity (LOH) of chromosome 12 in 100 primary ALL samples using 22 polymorphic markers and identified two distinct smallest common deleted regions on chromosome 12p13. One region is flanked by D12S77 and D12S98 and has a size of 4 cM. Twenty-six percent of informative patients showed LOH in this region. This region may contain the TEL gene. The other region is flanked by D12S269 and D12S308 including the KIP1 gene. Forty-four percent of informative patients showed LOH in this second region. Mutational analysis of KIP1 using polymerase chain reaction-single- strand conformation polymorphism analysis and Southern blot analysis showed no homozygous deletions and point mutations suggesting that the altered gene in this second region is not the KIP1. Clinical data showed that LOH of 12p was demonstrated more frequently in precursor-B ALLs (32 of 80; 40%) than in T-ALLs (1 of 20; 5%) (P = .0027). Furthermore, patients with 12p LOH were younger (P = .013), with a lower DNA index (P = .046), but they had the same survival rates at 3 years. In summary, these data suggest that two different tumor suppressor genes are on chromosome arm 12p, which act separately in the development of childhood precursor-B ALLs. One of the tumor suppressor genes is in the region the KIP1 gene, but our data suggest this gene is not abnormal. The other target is in the region of the TEL gene; and this candidate deserves further study.     

Anti-graft-versus-host disease effect of DT390-anti-CD3sFv, a single- chain Fv fusion immunotoxin specifically targeting the CD3 epsilon moiety of the T-cell receptor

In a recent study, we showed that an immunotoxin (IT) made with a conventional monoclonal antibody targeting the CD3 epsilon moiety of the T-cell receptor (TCR) had a potent, but partial, graft-versus-host disease (GVHD) effect (Vallera et al, Blood 86:4367, 1995). Therefore, in this current study, we determined whether a fusion immunotoxin made with anti-CD3 single-chain Fv (sFv), the smallest unit of antibody recognizing antigen, would have anti-GVHD activity. A fusion protein was synthesized from a construct made by splicing sFv cDNA from the hybridoma 145-2C11 to a truncated form of the diphtheria toxin (DT390) gene. DT390 encodes a molecule that retains full enzymatic activity, but excludes the native DT binding domain. The DT390-anti-CD3sFv hybrid gene was cloned into a vector under the control of an inducible promoter. The protein was expressed in Escherichia coli and then purified from inclusion bodies. The DT390 moiety of the protein had full enzymatic activity compared with native DT and DT390-anti-CD3sFv, with an IC50 of 1 to 2 nmol/L against phytohemagglutinin-stimulated and alloantigen-stimulated T cells. Specificity was shown (1) by blocking the IT with parental anti-CD3 antibody, but not with a control antibody; (2) by failure of DT390-anti-CD3sFv to inhibit lipopolysaccharide-stimulated murine B cells; (3) by failure of an Ig control fusion protein, DT390-Fc, to inhibit T-cell responses; and (4) with in vivo immunohistochemisty studies. GVHD was studied in a model in which C57BL/6 (H-2b)-purified lymph node T cells were administered to major histocompatibility complex (MHC) antigen disparate unirradiated C.B.-17 scid (H-2d) mice to assess GVHD effects in the absence of irradiation toxicity. Flow cytometry studies showed that donor T cells were expanded 57-fold and histopathologic analysis showed the hallmarks of a lethal model of GVHD. Control mice receiving phosphate-buffered saline showed 17% survival on day 80 after bone marrow transplantation, and mice receiving 2 micrograms DT390-Fc fusion toxin control administered in 2 daily doses for 6 days (days 0 through 5) had a 43% survival rate. In contrast, 86% of mice receiving the same dose of DT390-anti-CD3sFv were survivors on day 80, a significant improvement, although survivors still showed histopathologic signs of GVHD. These findings suggest that new anti-GVHD agents can be genetically engineered and warrant further investigation of fusion proteins for GVHD treatment.     

Lymphokine-induced phagocytosis in angiocentric immunoproliferative lesions (AIL) and malignant lymphoma arising in AIL

A factor that augmented the phagocytosis of IgG-coated ox red blood cells by the human monocyte/macrophage line U937 was identified in cell culture supernatants from two of two patients with angiocentric peripheral T cell lymphomas, three of three patients with angiocentric immunoproliferative lesions that were not frankly malignant, and one of two patients with T lymphoblastic malignancies. The factor was not present in supernatants derived from 14 nonangiocentric peripheral T cell lymphomas of other histologic types nor in ten cases of B cell lymphoma and two cases of Hodgkin's disease. A similar factor was present in the supernatants of concanavalin A (Con A)-stimulated normal peripheral blood mononuclear cells and in the supernatants of IL-2- dependent T cell lines derived from normal peripheral blood. The factor had an apparent mol wt of greater than 50,000 daltons, was heat labile (100 degrees C for two minutes), and stable at pH 2.0. Its stimulation of phagocytosis was independent of any increase in number of Fc receptors. Thus, this factor is probably not gamma-interferon. This factor may play a pathogenetic role in the hemophagocytic syndromes associated with certain T cell malignancies and immunodeficient states.     

Immune responses to major histocompatibility complex homozygous lymphoid cells in murine F1 hybrid recipients: implications for transfusion-associated graft-versus-host disease

Graft-versus-host disease (GVHD) is currently encountered after bone marrow transplantation and transfusion. GVHD associated with transfusion (TA-GVHD) in apparently immunocompetent recipients has been recently reported with increasing frequency. A consistent finding in many of these cases is that the recipient received blood from a donor homozygous for one of the recipient's HLA haplotypes. However, the observed frequency of TA-GVHD is much lower than the estimated probability of this donor/recipient combination. The potential role of recipient immune responses in controlling TA-GVHD was investigated using an analogous murine model in which GVHD is induced by the injection of parental lymphoid cells into unirradiated F1 hybrid recipients. The effect of various immune manipulations of the recipient of GVHD induction was assessed by determining the number of donor lymphoid cells required to induce GVHD responses. Whereas depletion of recipient CD4+ cells increased the number of donor cells needed to induce GVHD, depletion of recipient CD8+ and natural killer cells resulted in fewer donor cells being needed to induce a GVHD response. These studies suggest a central role for functioning recipient CD8 and natural killer cells in the down-regulation of TA-GVHD development in recipients.     

Case Report: Congenital Radioulnar Synostosis and Its Embryological Correlation and Functional Assessment

Congenital radioulnar synostosis(CRS) is a rare anomaly and approximately 400 cases were reported worldwide so far. CRS is the failure of the longitudinal segmentation and the persistence of the cartilaginous anlage between the radius and ulna during the seventh week of development that results in a persistent bridge of tissue. Here we are discussing on a case of 25yrs old, male patient with bilateral congenital synostosis. On the left hand the pronation and supination movements are restricted completely where as on right side 10 degree of supination and 20 degree of pronation is possible. Radiologically in our patient synostosis is classified as type II variety by Wilkie(1914)1 classification and type IV by the Cleary and Omer classification(1985). The position of forearm was not found to be related to subjective functional limitation, or employment status. Main line of treatment is surgical mainly rotational osteotomy but is rarely indicated. Our patient is not able to rotate his forearm especially on the left side still he has no functional limitation so he has refused for the operative treatment. No study has objectively compared the preoperative functional limitation of the patients with their postoperative functional improvement in order to justify surgical intervention In the authors opinion the only major factor that is to be taken into consideration of operative treatment is functional limitation to the patient.     

Diabetes Patients' Experiences With the Implementation of Insulin Therapy and Their Perceptions of Computer-Assisted Self-Management Systems for Insulin Therapy

Background

Computer-assisted decision support is an emerging modality to assist patients with type 2 diabetes mellitus (T2DM) in insulin self-titration (ie, self-adjusting insulin dose according to daily blood glucose levels). Computer-assisted insulin self-titration systems mainly focus on helping patients overcome barriers related to the cognitive components of insulin titration. Yet other (eg, psychological or physical) barriers could still impede effective use of such systems.

Objective

Our primary aim was to identify experiences with and barriers to self-monitoring of blood glucose, insulin injection, and insulin titration among patients with T2DM. Our research team developed a computer-assisted insulin self-titration system, called PANDIT. The secondary aim of this study was to evaluate patients’ perceptions of computer-assisted insulin self-titration. We included patients who used PANDIT in a 4-week pilot study as well as patients who had never used such a system.

Methods

In-depth, semi-structured interviews were conducted individually with patients on insulin therapy who were randomly recruited from a university hospital and surrounding general practices in the Netherlands. The interviews were transcribed verbatim and analyzed qualitatively. To classify the textual remarks, we created a codebook during the analysis, in a bottom-up and iterative fashion. To support examination of the final coded data, we used three theories from the field of health psychology and the integrated model of user satisfaction and technology acceptance by Wixom and Todd.

Results

When starting insulin therapy, some patients feared a lifelong commitment to insulin therapy and disease progression. Also, many barriers arose when implementing insulin therapy (eg, some patients were embarrassed to inject insulin in public). Furthermore, patients had difficulties increasing the insulin dose because they fear hypoglycemia, they associate higher insulin doses with disease progression, and some were ignorant of treatment targets. Patients who never used a computer-assisted insulin self-titration system felt they had enough knowledge to know when their insulin should be adjusted, but still believed that the system advice would be useful to confirm their reasoning. Furthermore, the time and effort saved with automated insulin advice was considered an advantage. Patients who had used PANDIT found the system useful if their glycemic regulation improved. Nevertheless, for some patients, the absence of personal contact with their caregiver was a drawback. While guidelines state that adjustment of basal insulin dose based on fasting plasma glucose values is sufficient, both patients who had and those who had not used PANDIT felt that such a system should take more patient data into consideration, such as lifestyle and diet factors.

Conclusions

Patients encounter multiple obstacles when implementing insulin therapy. Computer-assisted insulin self-titration can increase patient awareness of treatment targets and increase their confidence in self-adjusting the insulin dose. Nevertheless, some barriers may still exist when using computer-assisted titration systems and these systems could also introduce new barriers.     

Ambiguous phenotypes and genotypes in 16 children with acute leukemia as characterized by multiparameter analysis

Ambiguous phenotypes and genotypes were observed in 16 children with acute leukemia. Surface marker, cytogenetic, molecular genetic, and DNA flow cytometric analyses as well as standard morphologic and cytochemical studies were used to divide the patients into three groups. The first group comprised five children with acute leukemia whose blast cells were morphologically lymphoid, while immunophenotyping disclosed simultaneous expression of early pre-B cell and myeloid features. Molecular genetic studies showed evidence of heavy-chain immunoglobulin (Ig) gene rearrangements in all patients. Cytogenetic data, available in three of these children, revealed t(4;11). In five of the 16 patients, morphologic and surface marker analyses indicated the coexistence of two separate cell populations, one with myeloid and the other with early pre-B cell features. Further evidence of B cell commitment in these patients was provided by demonstration of Ig heavy-chain gene rearrangements in all five patients. Surprisingly, one of the five patients showed oligoclonal Ig heavy-chain as well as monoclonal gene rearrangement for the beta chain of the T cell receptor (beta-TCR). The last group consisted of four cases with otherwise typical acute lymphoblastic leukemia (ALL), early pre-B cell phenotype, and coexpression of myeloid or T cell-associated antigens, and two children with unequivocal acute myeloid leukemia (AML) and coexpression of T cell antigens. Gene rearrangement of Ig heavy-chain could be demonstrated in five of six patients, additional Ig light-chain gene rearrangement in two children with ALL, and bigenotypic features (Ig heavy-chain and beta-TCR gene rearrangement) in one patient. In none of the 16 patients did flow cytometry disclose clonal abnormalities of leukemic cell DNA content. Based on these findings, we suggest that malignant transformation in the first and second group of patients took place at a stage ontogenetically close to the pluripotent stem cell, whereas ambiguous phenotypes in the third group resulted from aberrant gene expression or insufficient reagent specificity.