Yok L. Kwong

FLT-3 aberrations in acute promyelocytic leukaemia: clinicopathological associations and prognostic impact.

FLT‐3 aberrations that occur as an internal tandem duplication (ITD) or a mutation at the activation‐loop position 835, D835, are common in acute promyelocytic leukaemia (APL). We investigated the clinicopathological associations and prognostic impact of FLT‐3 aberrations in a cohort of APL patients. FLT‐3 exons 11 and 12 were amplified by polymerase chain reaction (PCR), and the ITD was recognized as an increase in the size of the PCR product. FLT‐3 exon 17 was amplified, and D835 mutation was identified by loss of an EcoRV site, followed by DNA sequencing. Of 82 patients studied, FLT‐3 aberrations were detected in 35 cases (43%) at diagnosis (ITD: 16; D835 mutation: 18; ITD + D835 mutation: 1). FLT‐3 ITD, but not D835 mutations, was significantly associated with higher presentation white blood cell count (WBC) and microgranular morphology. Early/induction deaths were related to male sex and high presentation WBC. There was a trend for FLT‐3 ITD to be associated with non‐remission (P = 0·06). For disease‐free survival, high WBC was the only significant adverse factor. Male sex, high WBC and FLT‐3 ITD were significant adverse factors for overall survival. These findings have important implications on the possible use of FLT‐3 inhibitors in the treatment of APL.

Aberrant promoter CpG methylation as a molecular marker for disease monitoring in natural killer cell lymphomas

Summary. Natural killer (NK) cell lymphomas lack suitable clonal markers for tumour cell detection, making the monitoring of minimal residual lymphoma difficult. Aberrant promoter CpG methylation occurs frequently in NK cell lymphomas. The objective of this study was to assess the potential of aberrant methylation as a surrogate tumour marker. Twenty‐five primary tumours and 105 serial biopsies taken at various time points after treatment were examined using a methylation‐specific polymerase chain reaction (MSP) for a panel of genes, comprising p73, p16, hMLH1, RARβ and p15, previously shown to be methylated in NK cell lymphomas. All samples underwent independent morphological examination, supplemented by immunostaining for CD56 and in‐situ hybridization for Epstein–Barr‐virus‐encoded RNA. Primary tumours showed the frequent methylation of the genes p73 (92%), p16 (71%), hMLH1 (61%), RARβ (56%) and p15 (48%). MSP results in serial post‐treatment biopsies were correlated with clinicopathological findings. Results were concordant in 89 follow‐up samples (18 samples, histology positive/MSP positive; 71 samples, histology negative/MSP negative) and discordant in 16. Fifteen samples were histology negative/MSP positive, and tumour involvement was subsequently confirmed (positive re‐biopsies or relapses at the same sites), indicating that MSP was more sensitive for minimal lymphoma detection. One sample was histology positive/MSP negative; a subsequent histological review and continuous clinical remission of the patient did not support tumour involvement. Our findings suggest that MSP for aberrantly methylated genes is a potentially valuable molecular marker for detecting either residual or relapsed disease in NK cell lymphoma patients.

Aberrant gene promoter methylation in acute promyelocytic leukaemia: profile and prognostic significance.

Summary. Acute promyelocytic leukaemia (APL) has distinct clinicopathological and molecular features. However, the profile of aberrant gene promoter methylation is undefined. In this study, methylation‐specific polymerase chain reaction (MSP) was used to define the methylation status of a panel of nine genes, comprising p15, p16, RARβ, oestrogen receptor (ER), E‐cadherin (E‐CAD), p73, caspase 8 (CASP8), VHL and MGMT, in 29 patients with APL. Aberrant methylation of p15, ER, RARβ, p16 and E‐CAD occurred, respectively, in 23 (79%), 14 (48%), six (21%), six (21%) and two (7%) patients at diagnosis, but p73, VHL, CASP8 and MGMT were not methylated in any patients. There was methylation of one gene in 13 patients (45%), two genes in four patients (14%), three genes in six patients (21%) and four genes in three patients (10%). Concurrent methylation of two or more genes occurred in 13 patients (45%). No association was identified between gene methylation and presenting clinicopathological features. However, p15 methylation was significantly associated with an inferior disease‐free survival (DFS, P = 0·008), and remained the only poor prognostic factor in multivariate analysis (P = 0·019). In APL, p15, p16, ER and RARβ were most frequently methylated. This profile is distinct from other types of myeloid leukaemias. p15 methylation has a poor prognostic impact on DFS.

Use of the polymerase chain reaction in the detection of AML1/ETO fusion transcript in t(8;21)

Background. t(8;21)(q22;q22), found in acute myeloid leukemia (AML) and occasionally in myelodysplasia (MDS), results in the fusion of the AML1 gene on 22q22 to the ETO gene on 8q22, generating a chimeric AML1/ETO transcript, which is a molecular marker of the translocation.

Fludarabine, mitoxantrone and dexamethasone in the treatment of indolent B- and T-cell lymphoid malignancies in Chinese patients

The treatment results of indolent lymphoid malignancies in Chinese are poorly reported. The efficacy of FND (fludarabine 25 mg/m2/d, ×3; mitoxantrone 10 mg/m2/d, ×1; dexamethasone 20 mg/d, ×5; monthly cycles, ×6) in 95 Chinese patients with indolent B‐cell malignancies (at diagnosis: 55, relapse/refractory disease: 40) and nine Chinese patients with T‐cell large granular lymphocyte leukaemia (T‐LGL leukaemia) (at diagnosis: two, refractory disease: seven) was evaluated. For B‐cell malignancies, the complete response (CR), partial response (PR) and overall response (OR) rates were 50·5%, 18% and 68·5% respectively. Better results were obtained for primary versus relapse/refractory disease (CR: 60% vs. 37·5%, P = 0·03; OR: 84% vs. 47·5%, P < 0·001; median progression‐free survival (PFS): 44 months vs. 22 months; 2‐year PFS: 66% vs. 47%, P = 0·039; overall survival (OS): not reached vs. 32%; 2‐year OS: 92% vs. 58%, P < 0·001). Responsive patients (CR/PR) had a better median PFS (44 months vs. 5 months, P < 0·001) and OS (67 months vs. 13 months, P < 0·001) than unresponsive patients. For T‐LGL leukaemia, the CR and molecular‐remission rates were 56% and 67% (median follow‐up: 23 months). FND is an active regimen for the treatment of indolent B‐ and T‐cell malignancies in Chinese patients, with results comparable with Western patients with similar indolent lymphomas.

T-Cell Large Granular Lymphocyte Leukemia of Donor Origin After Allogeneic Bone Marrow Transplantation

A 39-year-old man with chronic myeloid leukemia in accelerated phase underwent allogeneic bone marrow transplantation (BMT). At 6 months after BMT, lymphocytosis (WBC count, 23,100/microL [23.1 x 10(9)/L]; 80% (0.80) large granular lymphocytes [LGLs]) occurred. The LGLs were CD3+CD4-CD8+, with clonally rearranged T-cell receptor gamma gene, and of donor origin, as shown by analysis of polymorphic microsatellite markers. Epstein-Barr virus was not present. The diagnosis, therefore, was consistent with T-cell large granular lymphocytic (T-LGL) leukemia. Corticosteroids controlled the LGL count, but progressive pancytopenia led to death 4 months later. Retrospective analysis showed that the T-LGL leukemia apparently had arisen as early as 3 months after BMT. The distinguishing features of this case included donor origin, neoplastic nature, and the aggressive fatal outcome.

Evans' syndrome complicating chronic graft versus host disease after cadaveric liver transplantation

Acute graft versus host disease (GVHD) occurred in a patient after cadaveric liver transplantation from an HLA disparate donor. Immunosuppression resulted in a remission, but chronic GVHD with a scleroderma-like syndrome ensued. This was further complicated by immune hemolytic anemia and thrombocytopenia (Evans syndrome). Semi-quantitative microsatellite analysis of circulating lymphoid cells showed that T cells were predominantly of donor origin, thereby explaining the chronic GVHD. The marrow hematopoietic cells remained recipient, so that the immune cytopenias were expected to be alloimmune in nature. However, the red cell antibodies were shown to have anti-C and anti-e specificity, with both the donor (R1R1) and recipient (R1r) possessing the C and e antigens. Therefore, the immune hemolysis might be considered both alloimmune and autoimmune. The patient finally died of sepsis. This case illustrates that chronic GVHD due to stable donor T cell engraftment may rarely occur in liver transplantation despite HLA disparity. Immunosuppression may result in dysregulation of T cell functions, leading to alloimmune and autoimmune problems.