Kevin Barton

Autologous Transplantation as Consolidation for Aggressive Non-Hodgkin's Lymphoma

BACKGROUND The efficacy of autologous stem-cell transplantation during the first remission in patients with diffuse, aggressive non-Hodgkins lymphoma classified as high-intermediate risk or high risk on the International Prognostic Index remains controversial and is untested in the rituximab era. METHODS We treated 397 patients who had disease with an age-adjusted classification of high risk or high-intermediate risk with five cycles of cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) or CHOP plus rituximab. Patients with a response were randomly assigned to receive three additional cycles of induction chemotherapy (control group) or one additional cycle of induction chemotherapy followed by autologous stem-cell transplantation (transplantation group). The primary efficacy end points were 2-year progression-free survival and overall survival. RESULTS Of 370 induction-eligible patients, 253 were randomly assigned to the transplantation group (125) or the control group (128). Forty-six patients in the transplantation group and 68 in the control group had disease progression or died, with 2-year progression-free survival rates of 69 and 55%, respectively (hazard ratio in the control group vs. the transplantation group, 1.72; 95% confidence interval [CI], 1.18 to 2.51; P=0.005). Thirty-seven patients in the transplantation group and 47 in the control group died, with 2-year overall survival rates of 74 and 71%, respectively (hazard ratio, 1.26; 95% CI, 0.82 to 1.94; P=0.30). Exploratory analyses showed a differential treatment effect according to risk level for both progression-free survival (P=0.04 for interaction) and overall survival (P=0.01 for interaction). Among high-risk patients, the 2-year overall survival rate was 82% in the transplantation group and 64% in the control group. CONCLUSIONS Early autologous stem-cell transplantation improved progression-free survival among patients with high-intermediate-risk or high-risk disease who had a response to induction therapy. Overall survival after transplantation was not improved, probably because of the effectiveness of salvage transplantation. (Funded by the National Cancer Institute, Department of Health and Human Services, and others; SWOG-9704 ClinicalTrials.gov number, NCT00004031.).

Analysis of expression of nuclear factor κB (NF-κB) in multiple myeloma: Downregulation of NF-κB induces apoptosis

Nuclear factor‐κB (NF‐κB) is an important transcription factor that regulates survival in many cells. Activated NF‐κB has been shown to protect some haematopoietic neoplastic cells from apoptosis. In the present study, we analysed NF‐κB status in 13 primary samples from patients with multiple myeloma (MM) and in four myeloma cell lines including U266, RPMI 8226, HS‐Sultan and K620. Constitutive activation of NF‐κB was evaluated by either immunohistochemistry or immunofluorescence using a monoclonal mouse anti‐human p65 (Rel A) antibody, which recognizes the unbound, active form of p65 (Rel A). Constitutively active NF‐κB was present in all MM patient samples as well as in all four myeloma cell lines. Inhibition of constitutively active NF‐κB, by either proteasome inhibitors (MG132, gliotoxin) or inhibitors of IκB phosphorylation (Bay117082, and Bay117085), induced apoptosis as demonstrated by both flow cytometric analysis and light microscopic morphological evaluation. This chemically induced apoptosis was associated with decreased DNA binding of nuclear NF‐κB as determined by the electrophoretic mobility shift assay. In addition, adenovirus vector with dominant negative IκBα (Ad5IκB) was used for inhibition of NF‐κB in the U266 cell line. Compared with wild‐type, super‐repressor‐treated cells showed an increased level of apoptosis. These results suggest that constitutive expression of NF‐κB plays an important role in plasma cell survival in MM.

Analysis of histone deacetylase inhibitor, depsipeptide (FR901228), effect on multiple myeloma.

Multiple myeloma (MM) is a neoplastic proliferation of plasma cells and remains an incurable disease because of the development of drug resistance. Histone deacytylase (HDAC) inhibitors are a new class of chemotherapeutic reagents that cause growth arrest and apoptosis of neoplastic cells. Depsipeptide, a new member of the HDAC inhibitors, was found to be safe in humans and has been shown to induce apoptosis in various cancers. In order to evaluate the effects of depsipeptide, a MM cell line, U266 [interleukin (IL)‐6 dependent], was analysed for viability and apoptosis. The combined effect of depsipeptide with melphalan and changes in BCL‐2 family proteins (BCL‐2, BCL‐XL, BAX and MCL‐1) were also investigated. In addition, the RPMI 8226 cell line (IL‐6 independent), and primary patient myeloma cells were also analysed for apoptosis after depsipeptide treatment. Depsipeptide induced apoptosis in both U266 and RPMI 8226 cell lines in a time‐ and dose‐dependent fashion, and in primary patient myeloma cells. We also demonstrated that depsipeptide had an additive effect with melphalan (10 μmol/l). BCL‐2, BCL‐XL and MCL‐1 showed decreased expression in depsipeptide‐treated samples. Based on recent clinical trials demonstrating minimal clinical toxicity, our study supports the future clinical utilization of depsipeptide in the management of MM.

Hydroxyurea, azacitidine and gemtuzumab ozogamicin therapy in patients with previously untreated non-M3 acute myeloid leukemia and high-risk myelodysplastic syndromes in the elderly: results from a pilot trial.

Elderly patients with acute myeloid leukemia (AML) and high-risk myelodysplastic syndromes (MDS) have a poor prognosis due to low response rates (26–46%) to standard chemotherapy and high treatment-related mortality (11–31%). In this Phase II study, we used a combination of hydroxyurea (HU), azacitidine and low dose gemtuzumab ozogamicin (GO) to assess its efficacy and toxicity in this group of patients. Twenty patients with non-M3 AML and MDS were treated with this regimen. The treatment was begun with HU 1500 mg orally twice daily to lower white blood cell count below 10,000/μL, followed by azacitidine 75 mg/m2 subcutaneously for 7 days and GO 3 mg/m2 on day 8. Patients who achieved complete remission (CR) received a consolidation course. The median age of patients was 76 years. Eleven patients (55%) were treated in the outpatient setting. Fourteen (70%) achieved a CR, three of which were incomplete (CRi). The median duration of remission was 8 months and median survival was 10 months. Performance status of 0–1 was associated with high complete response rate. Overall toxicity was acceptable with only one (5%) early death due to disease progression. The combination of HU, azacitdine and GO appears to be a safe and effective regimen in the treatment of AML and high risk MDS in the elderly. These results need to be confirmed in a larger cohort of patients.

Expanding the Pathologic Spectrum of Immunoglobulin Light Chain Proximal Tubulopathy

CONTEXT In plasma cell dyscrasias, involvement of the distal tubules is frequent and well characterized. In contrast, proximal tubules have only rarely been reported to show diagnostic pathology such as intracytoplasmic crystals. OBJECTIVE To look for additional morphologic features that might be helpful in the diagnosis of proximal tubulopathy associated with an underlying plasma cell dyscrasia. DESIGN We examined patients presenting with nonspecific renal symptoms who were found to have light chain restriction limited to proximal tubular epithelium by immunofluorescence. We correlated these results with light microscopy, electron microscopy, and the clinical findings. RESULTS By immunofluorescence, 5 patients had light chain restriction in proximal tubular epithelium. By light microscopy, only 1 patient had focal rhomboid crystals in the proximal tubular epithelium; all other biopsies failed to show any discernible pathology within the proximal tubules or elsewhere in the kidney. By electron microscopy, proximal tubules from 2 patients showed crystals with a latticelike structure, whereas the remaining 3 patients had only prominent phagolysosomes. However, by immunoelectron microscopy, the lysosomal content showed light chain restriction (in 2 cases studied). Post-kidney biopsy, all patients were diagnosed with multiple myeloma or plasma cell dyscrasia. One patient developed renal failure and had recurrence of crystals in the allograft. CONCLUSIONS Light chain proximal tubulopathy may be associated with the presence of crystals or with the presence of phagolysosomes with light chain restriction as the sole abnormality. Both kappa and lambda light chains may be involved. The prognosis is variable and the pathology may recur in transplants.

Transcriptional regulation of CD1D1 by Ets family transcription factors

CD1 molecules are MHC class I-like glycoproteins specialized in presenting lipid/glycolipid Ags to T cells. The distinct cell-type specific expression of CD1D1 plays an important role in the development and function of NKT cells, a unique subset of immunoregulatory T cells. However, the mechanisms regulating CD1D1 expression are largely unknown. In this study, we have characterized the upstream region of the CD1D1 gene and identified a minimal promoter region within 200 bp from the translational start site of CD1D1 that exhibits cell-type specific promoter activity. Analysis of this region revealed an Ets binding site critical for CD1D1 promoter activity. Gel shift assays and chromatin immunoprecipitation experiments showed that Elf-1 and PU.1 bind to the CD1D1 promoter. Furthermore, we found that gene disruption of Elf-1 resulted in decreased CD1D1 expression on B cells but not other cell types, whereas conditional activation of PU.1 negatively regulated CD1D1 expression in PU.1-deficient myeloid cells. These findings are the first to demonstrate that Ets proteins are involved in the transcriptional regulation of CD1D1 and that they may function uniquely in different cell types.

AML1 haploinsufficiency, gene dosage, and the predisposition to acute leukemia

Hematopoiesis is the complex developmental process through which undifferentiated, pluripotent, hematopoietic stem cells come to generate mature, functional blood cells. This process is regulated in large part by specific transcription factors that control expression of genes necessary for the developmental sequence. Leukemias represent one form of disruption of this normal developmental process, and studies over the past few years have shown that many of the genes that underlay leukemogenesis are also essential for normal hematopoiesis. In an interesting recent example, Song et al.((1)) demonstrate that haploinsufficiency of the AML1 gene is the genetic basis of a form of familial thrombocytopenia which predisposes the affected individuals to the development of acute myeloid leukemia. Here we summarize Songs paper and current information describing the interesting dosage effects of this gene and other members of its gene family.

Management of Diffuse Low-Grade Cerebral Gliomas

World Health Organization grade II gliomas (GIIG) are diffuse, slow-growing, primary neuroectodermal tumors that occur in the central nervous system. They are generally seen in young individuals and are slightly more common in Whites and males. Most patients present with seizures but neurologic deficits are rare. Magnetic resonance imaging best detects GIIG and they are most frequently located in the frontal and temporal lobes. An accurate pathologic diagnosis is essential because the natural history of a GIIG may be unpredictable. In recent years, the emphasis has been on surgically removing as much tumor as safely possible to obtain an accurate diagnosis, improve symptoms, reduce tumor burden, and determine the need for adjuvant therapies. Radiation and chemotherapy are integral to the management of GIIG but their efficacy varies by tumor histology and is balanced against complications associated with them. Genetic, histopathologic, clinical, and radiographic changes are noted as GIIG progress to malignant gliomas. The risk of malignant transformation and subsequent survival may be predicted by pretreatment and treatment-related factors.

Thrombotic microangiopathy associated with sunitinib, a VEGF inhibitor, in a patient with factor V Leiden mutation.

Haemolytic uraemic syndrome (HUS) and thrombotic thrombocytopenic purpura (TTP) are closely related disorders characterised by non-immune microangiopathic haemolytic anaemia and thrombocytopenia. Thrombotic microangiopathyistheunderlyingpathologiclesioninboth syndromes. Under physiological conditions, platelets bind to endothelium via von Willebrand Factor (vWF) and are released back into the circulation by ADAMTS13, a metalloprotease that cleaves ultra-large, newly synthesized vWF multimers. A deficiency of ADAMTS13 (due to either a decrease in its production or the presence of a circulating inhibitor) may thus result in microvascular thrombi. However, patients with thrombotic microangiopathy may have normal ADAMTS13 levels and no evidence of a circulating inhibitor [1]. Sunitinib malate, an oral multi-targeted tyrosine kinase inhibitor that blocks the receptors for VEGF, has utility in a variety of metastatic tumours [2]. We report a patient who developed proteinuria, hypertension and biopsy-proven thrombotic microangiopathy after treatment with sunitinib malate. This patient had normal ADAMTS13 levels but had factor V Leiden mutation that may have predisposed her to this complication.

Ibritumomab consolidation after 3 cycles of CHOP plus radiotherapy in high-risk limited-stage aggressive B-cell lymphoma: SWOG S0313

In the S0313 trial, we evaluated the impact of adding ibritumomab tiuxetan consolidation to 3 cycles of standard cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) chemotherapy plus involved field radiotherapy (IFRT) in patients with limited-stage aggressive B-cell non-Hodgkin lymphoma (LD-NHL). Patients with at least 1 stage-modified adverse risk factor (nonbulky stage II, age >60 years, elevated lactate dehydrogenase, or World Health Organization performance status of 2) were treated with CHOP on days 1, 22, and 43, followed 3 weeks later by 40 to 50 Gy of IFRT. An ibritumomab tiuxetan regimen was initiated 3 to 6 weeks following IFRT. Forty-six patients were registered and eligible, with median follow-up of 7.3 years. The progression-free survival estimate is 89% at 2 years, 82% at 5 years, and 75% at 7 years. The overall survival estimate is 91% at 2 years, 87% at 5 years, and 82% at 7 years. Grade 4 adverse events occurring more than once included neutropenia (8), leukopenia (5), and lymphopenia (2). Febrile neutropenia was observed in 4 patients. No cases of treatment-related myeloid neoplasms were noted. In conclusion, patients with high-risk LD-NHL treated with 3 cycles of CHOP plus IFRT followed by ibritumomab tiuxetan consolidation had outcomes that compare favorably to our historical experience. The clinical trial was registered at www.clinicaltrials.gov as #NCT00070018.