Application of droplet digital PCR in minimal residual disease monitoring of rare fusion transcripts and mutations in haematological malignancies.

Leukaemia of various subtypes are driven by distinct chromosomal rearrangement or genetic abnormalities. The leukaemogenic fusion transcripts or genetic mutations serve as molecular markers for minimal residual disease (MRD) monitoring. The current study evaluated the applicability of several droplet digital PCR assays for the detection of these targets at RNA and DNA levels (atypical BCR::ABL1 e19a2, e23a2ins52, e13a2ins74, rare types of CBFB::MYH11 (G and I), PCM1::JAK2, KMT2A::ELL2, PICALM::MLLT10 fusion transcripts and CEBPA frame-shift and insertion/duplication mutations) with high sensitivity. The analytical performances were assessed by the limit of blanks, limit of detection, limit of quantification and linear regression. Our data demonstrated serial MRD monitoring for patients at molecular level could become "digitalized", which was deemed important to guide clinicians in treatment decision for better patient care.

Next-generation sequencing and molecular cytogenetic characterization of ETV6-LYN fusion due to chromosomes 1, 8 and 12 rearrangement in acute myeloid leukemia.

In a newly diagnosed patient with acute myeloid leukemia (AML) and complex cytogenetics and negative for gene mutations associated with myeloid neoplasms, RNA sequencing by next-generation sequencing (NGS) through a large cancer-related gene panel showed ETV6-LYN leukemic fusion transcript. Breakpoint analysis of the NGS reads showed fusion of exon 5 of the ETV6 gene to exon 8 of the LYN gene. Metaphase fluorescence in situ hybridization (FISH) inferred a four-break rearrangement of three chromosomes, namely 1, 8 and 12. First, there was a balanced translocation t(1;12)(p13;p13.2) in which the ETV6 was split between der(1) and der(12). Second, an inverted insertion of 8q12.1~q24.21 into 1p13 occurred, thus bringing ETV6 and LYN into juxtaposition in the correct 5' to 3' orientation to produce an in-frame chimeric fusion gene on der(1). Notwithstanding two previous reports of ETV6-LYN fusion in myeloproliferative neoplasms (MPN), we report the first case of this fusion in AML and hence broaden its disease association. We also illustrate the clinical utility of NGS based detection of gene fusion in the setting of complex karyotype or cryptic aberration, since this method does not require a priori knowledge of the translocation partner and exact breakpoints to guide the application of appropriate primers or probes.

Effectiveness of prophylactic Anti-HBV therapy in allogeneic hematopoietic stem cell transplantation with HBsAg positive donors.

Use of hepatitis B surface antigen (HBsAg) positive donors for allogeneic hematopoietic stem cell transplantation (HSCT) causes serious hepatitis B virus (HBV)-related liver morbidity and mortality in the recipient. We compared the effectiveness of anti-HBV therapy in 29 recipients who underwent HSCT using HBsAg positive marrow (group I) against a historical control group of 25 patients who received HBsAg positive marrow without pre-HSCT prophylaxis (group II). Anti-HBV therapy consisted of lamivudine for HBsAg-positive donors and all recipients (n = 29) as well as HBV vaccination to all HBsAg-negative recipients (n = 10) before HSCT. After transplantation, HBV-related hepatitis was significantly higher in group II than group I recipients [12 of 25 recipients (48%) vs. 2 of 29 recipients (6.9%), p = 0.002] and in recipients whose donors had detectable serum HBV DNA by Digene Hybrid Capture II assay [8 of 14 recipients (57.1%) vs. 6 of 40 recipients (15.0%), p = 0.02]. Six recipients in group II and none in group I died of HBV-related hepatic failure (24.0% vs. 0%, p = 0.01). By multivariate Cox analysis, anti-HBV therapy effectively reduces post-HSCT HBV-related hepatitis (p = 0.01, adjusted hazards ratio 7.27, 95%CI 1.62-32.58). Our data support the use of prophylactic therapy in preventing HBV-related hepatitis after allogeneic HSCT from HBsAg-positive donor.

A long-term follow-up study on hepatitis B surface antigen-positive patients undergoing allogeneic hematopoietic stem cell transplantation.

The long-term hepatic complications after allogeneic hematopoietic stem cell transplantation (HSCT) in hepatitis B virus (HBV) endemic area are unknown. We examined the serological and liver-related outcome of 803 consecutive patients who received allogeneic HSCTs, with a median follow-up period of 83 months (range, 0.5-155 months). Late HBV-related hepatitis occurred in 2 of the 721 hepatitis B surface antigen-negative (HBsAg-) recipients compared with 16 of the 82 HBsAg+ recipients after HSCT (0.3% vs 19.5%; P < .001 by log-rank). Liver cirrhosis developed in 8 of the 82 HBsAg+ recipients compared with none of the 721 HBsAg- recipients (9.8% vs 0%; P < .001 by log-rank). Twenty of the 31 (64.5%) HBsAg+ recipients of hematopoietic stem cells from donors with natural immunity to HBV had sustained serologic clearance of HBsAg after HSCT. Eight of the 62 recipients without sustained HBsAg clearance compared with none of the 20 recipients with sustained HBsAg clearance developed liver cirrhosis (12.9% vs 0%; P = .02 by log-rank). Our study showed that long-term hepatic complications occur in a significant proportion of HBsAg+ patients after HSCT and the incidence of liver cirrhosis is reduced in those with sustained serologic clearance of HBsAg after HSCT.

Primary CD30+ve cutaneous T-cell lymphoma associated with chronic burn injury in a patient with longstanding psoriasis.

An increased incidence of lymphoma has been reported in psoriatic patients, but most cases are nodal B-cell lymphoma. We report a unique case of CD30-expressing cutaneous T-cell lymphoma arising from underlying psoriatic plaque after intractable caustic burns and cellulitis in the palm of a patient with generalized chronic plaque psoriasis. Molecular studies confirmed a localized clonal T-cell expansion, and the lesion responded dramatically to multiagent chemotherapy. The case highlighted the possible role of chronic systemic and local T-cell activation in the pathogenesis of primary CD30+ve cutaneous T-cell lymphoma, and the importance of histologic assessment in chronic nonhealing skin lesions.

Primary nasal natural killer cell lymphoma: long-term treatment outcome and relationship with the International Prognostic Index.

Nasal natural killer (NK) cell lymphoma is rare, so that its optimal therapy, long-term outcome, and prognostic factors are unclear. Data on 52 men and 15 women with well-characterized nasal NK cell lymphomas were analyzed retrospectively to define the impact of primary therapy on remission and long-term outcome and the validity of the International Prognostic Index (IPI). Most (84%) had stage I/II disease with an IPI score of 1 or less (52%). Seven patients received radiotherapy only; 47 patients received anthracycline-containing chemotherapy plus consolidation radiotherapy; and 12 patients received nonanthracycline-containing chemotherapy plus radiotherapy. The overall complete remission (CR) rate was 64.2%; the 20-year overall survival (OS) and disease-free survival (DFS) rates were 37.1% and 33.5%, respectively. Front-line radiotherapy was apparently better than chemotherapy for CR (100% versus 59%, P =.04) and OS (83.3% versus 32.0%, P =.03). Relapses occurred in 4 radiotherapy-treated (all local) and 14 chemotherapy-treated patients (9 local, 4 systemic). Among these, 5 late relapses (4 local, 1 systemic) occurred at 170 months (range, 92-348 months) from CR. The IPI score was of prognostic significance for the whole group (IPI or= 2 for 20-year OS: 57.4% versus 27.6%, P = 0.012), as well as for patients treated with chemotherapy/radiotherapy (IPI or= 2 for CR: 76.7% versus 35.7%, P =.017; and 10-year OS: 63.8% versus 26.8%, P =.003).

Hepatitis B reactivation in patients positive for hepatitis B surface antigen undergoing autologous hematopoietic cell transplantation.

Hepatitis due to reactivation of hepatitis B virus is an important cause of liver-related morbidity and mortality in hepatitis B surface antigen (HBsAg) positive patients undergoing autologous hematopoeitic cell transplantation. With the recent introduction of sensitive serum HBV DNA quantitation assay, the diagnosis of hepatitis B reactivation can now be made more reliably. As these hepatitis are driven by the host immune response to a surge of hepatitis B viral load, the availability of effective nucleoside analogues which can inhibit hepatitis B viral replication has opened up new approaches to this previously untreatable condition. Up till now, two such nucleoside analogues, lamivudine and adefovir dipivoxil, have been approved for the treatment of chronic hepatitis B infection. However, further studies are needed to determine which nucleoside analogues should be chosen in this transplant setting. Due to the high dose chemotherapy generally needed in autologous hematopoeitic cell transplantation, there is a high risk of post-transplant hepatitis B reactivation. Hence, all HBsAg positive patients undergoing autologous hematopoeitic cell transplantation should preferably be treated pre-emptively with nucleoside analogous. An alternative approach is to defer treatment with nucleoside analogous until there is evidence of hepatitis B virological reactivation. However, the latter approach would need the patient's hepatitis B viral load be monitored at a very close interval and might not be cost-effective.

Role of liver biopsy in the management of liver dysfunction after hematopoietic stem-cell transplantation in a hepatitis B virus-prevalent patient population.

BACKGROUND: Derangement of liver function tests (LFT) is common after hematopoietic stem-cell transplantation (HSCT). The role of liver biopsy in such cases has not been defined in hepatitis B virus (HBV)-prevalent patients. The impact of liver biopsy in the management of LFT derangement after HSCT in an HBV-prevalent population was examined. METHODS: Seventy-five liver biopsies, performed for 323 patients with LFT derangement post-HSCT (263 allogeneic, 60 autologous), were analyzed. The HBV carrier rate was 13.6%. RESULTS: Significantly more LFT derangements and therefore liver biopsies occurred in allogeneic versus autologous HSCT. Before biopsy, graft-versus-host disease (GVHD) and HBV reactivation were clinically diagnosed in 70.6% and 25.3% of cases, respectively. A definite histopathologic diagnosis was obtained after biopsy in 53 cases, with GVHD, HBV hepatitis, and concomitant GVHD-HBV hepatitis found in 33%, 21%, and 8% of cases, respectively. The clinical and histopathologic diagnoses were concordant in 43 cases and discordant in 9 cases. Clinical management was altered in six of nine discordant cases, five of which were caused by HBV or hepatitis C virus (HCV) reactivation. Twenty-two biopsy specimens showed nondiagnostic histopathologic features. Twenty of these cases were successfully managed on the basis of clinical diagnoses. The clinical-biochemical features of patients clinically diagnosed to have GVHD did not differ significantly whether or not they were HBV-HCV carriers. However, liver biopsies in HBV-HCV carriers resulted in significantly more treatment alterations as compared with noncarriers. CONCLUSIONS: Clinical diagnoses of LFT derangements post-HSCT might be adequate for initiation of treatment, but liver biopsies in HBV-HCV carriers were needed, as this might impact on management.

Aetiology in sixteen cases of toxic epidermal necrolysis and Stevens-Johnson syndrome admitted within eight months in a teaching hospital.

Toxic epidermal necrolysis and Stevens-Johnson syndrome are serious cutaneous reactions associated with significant mortality and morbidity. Eight patients with toxic epidermal necrolysis and eight patients with Stevens-Johnson syndrome were admitted consecutively to a single centre between August 2001 and March 2002. An aetiological study including viral serology and PCR was performed in view of the clustering of admissions related to these two conditions. The majority of cases were drug induced, the drug most commonly involved being allopurinol (toxic epidermal necrolysis, 50%; Stevens-Johnson syndrome, 13%). Two cases were related to drug abuse. Possible aetiological co-factors were cancers, radiotherapy and renal failure. No association with viral infection, including human herpesvirus-6 and parvovirus B19, was detected in the present series. Early diagnosis and prompt withdrawal of suspected drugs remain the most important measures in managing this condition. Further studies to identify the co-factors precipitating severe cutaneous drug reactions are warranted.