Yok-Lam Kwong

Hepatitis B Reactivation in Patients With Previous Hepatitis B Virus Exposure Undergoing Rituximab-Containing Chemotherapy for Lymphoma: A Prospective Study

PURPOSE Patterns of hepatitis B virus (HBV) reactivation in hepatitis B surface antigen (HBsAg) -negative, antihepatitis B core antigen antibody (anti-HBc) -positive patients with lymphoma receiving rituximab-containing chemotherapy have not been well described. PATIENTS AND METHODS HBsAg-negative, anti-HBc-positive Chinese patients with undetectable serum HBV DNA (< 10 IU/mL), diagnosed with hematologic malignancies and receiving rituximab-containing chemotherapy, were prospectively monitored every 4 weeks for up to 2 years. Entecavir was started when HBV reactivation (defined as detectable HBV DNA) was encountered. RESULTS Among 260 patients receiving rituximab-containing chemotherapy, 63 patients (24.2%) who were HBsAg negative and anti-HBc positive underwent follow-up for a median of 70 weeks (range, 6 to 104 weeks). The 2-year cumulative rate of HBV reactivation was 41.5%, occurring at a median of 23 weeks (range, 4 to 100 weeks) after rituximab treatment. The median HBV DNA level at reactivation was 43 IU/mL (range, 14 to 920 IU/mL). A baseline undetectable antibody to HBsAg (anti-HBs; < 10 mIU/mL) was the only significant risk factor that was positively associated with HBV reactivation (hazard ratio, 3.51; 95% CI, 1.37 to 8.98; P = .009). Patients with negative baseline anti-HBs, compared with those with positive anti-HBs, had a significantly higher 2-year cumulative rate of HBV reactivation (68.3% v 34.4%; P = .012). At HBV reactivation, all patients had normal ALT, and all patients but one were HBsAg negative. Entecavir successfully controlled HBV reactivation in all patients. CONCLUSION A high rate of HBV reactivation was observed in HBsAg-negative, anti-HBc-positive patients undergoing rituximab-containing chemotherapy, with the risk of reactivation significantly higher in anti-HBs-negative patients. Periodic HBV DNA monitoring was an effective strategy in preventing HBV-related complications.

Genetic and Functional Drivers of Diffuse Large B Cell Lymphoma

Diffuse large B cell lymphoma (DLBCL) is the most common form of blood cancer and is characterized by a striking degree of genetic and clinical heterogeneity. This heterogeneity poses a major barrier to understanding the genetic basis of the disease and its response to therapy. Here, we performed an integrative analysis of whole-exome sequencing and transcriptome sequencing in a cohort of 1,001 DLBCL patients to comprehensively define the landscape of 150 genetic drivers of the disease. We characterized the functional impact of these genes using an unbiased CRISPR screen of DLBCL cell lines to define oncogenes that promote cell growth. A prognostic model comprising these genetic alterations outperformed current established methods: cell of origin, the International Prognostic Index comprising clinical variables, and dual MYC and BCL2 expression. These results comprehensively define the genetic drivers and their functional roles in DLBCL to identify new therapeutic opportunities in the disease.

A novel tescalcin-sodium/hydrogen exchange axis underlying sorafenib resistance in FLT3-ITD+ AML

Internal tandem duplication (ITD) of fms-like tyrosine kinase 3 (FLT3) in acute myeloid leukemia (AML) is associated with inferior clinical prognosis. Sorafenib is effective in clearing leukemic blasts in chemorefractory FLT3-ITD(+) AML, but leukemia progression invariably occurs. Mechanisms of drug resistance are not completely understood. We hypothesized that a gene encoding tescalcin (TESC), known to be upregulated at leukemia progression during continuous sorafenib treatment and activate an Na(+)/H(+) exchanger type-1 (NHE1), may underlie tyrosine kinase inhibitor resistance. TESC was highly expressed in FLT3-ITD(+) AML lines MOLM-13 and MV4-11, and its knockdown by small-interfering RNA lowered intracellular pH (pHi) and induced apoptosis. The results were recapitulated by treatment with an NHE1 inhibitor, 5-(N,N-hexamethylene) amiloride (HMA). Induction of sorafenib resistance in the MOLM-13 cell line (M13-RE) significantly increased its sensitivity to HMA. The later also enhanced suppression of FLT3 signaling by sorafenib in otherwise resistant cell lines. HMA treatment of MOLM-13 and MV4-11 as well as primary FLT3-ITD(+) AML cells significantly reduced leukemia initiation in anti-CD122-primed NOD/SCID mouse xenotransplantation. These observations provided novel information about the pathogenetic role of a TESC-NHE1-pHi axis in mediating sorafenib resistance in AML.

Homoharringtonine (omacetaxine mepesuccinate) as an adjunct for FLT3-ITD acute myeloid leukemia.

An in vitro drug screening using primary AML samples identified homoharringtonine (omacetaxine mepesuccinate) as an effective adjunct for treatment of FLT3-ITD AML. More than a FLT-ing success in leukemia Acute myeloid leukemia is a difficult disease to treat under the best of circumstances, and the subtype containing internal tandem duplication of fms-like tyrosine kinase 3 (FLT3-ITD) tends to be particularly challenging. Lam et al. performed a high-throughput drug screen and identified homoharringtonine as a candidate treatment for this type of leukemia and then confirmed its effectiveness in cancer cells, in mouse models, and in patients. The treatment showed promising results in a phase 2 clinical trial, which included elderly patients and those who have failed all previous treatments, paving the way for further development of this drug. An in vitro drug-screening platform on patient samples was developed and validated to design personalized treatment for relapsed/refractory acute myeloid leukemia (AML). Unbiased clustering and correlation showed that homoharringtonine (HHT), also known as omacetaxine mepesuccinate, exhibited preferential antileukemia effect against AML carrying internal tandem duplication of fms-like tyrosine kinase 3 (FLT3-ITD). It worked synergistically with FLT3 inhibitors to suppress leukemia growth in vitro and in xenograft mouse models. Mechanistically, the effect was mediated by protein synthesis inhibition and reduction of short-lived proteins, including total and phosphorylated forms of FLT3 and its downstream signaling proteins. A phase 2 clinical trial of sorafenib and HHT combination treatment in FLT3-ITD AML patients resulted in complete remission (true or with insufficient hematological recovery) in 20 of 24 patients (83.3%), reduction of ITD allelic burden, and median leukemia-free and overall survivals of 12 and 33 weeks. The regimen has successfully bridged five patients to allogeneic hematopoietic stem cell transplantation and was well tolerated in patients unfit for conventional chemotherapy, including elderly and heavily pretreated patients. This study validated the principle and clinical relevance of in vitro drug testing and identified an improved treatment for FLT3-ITD AML. The results provided the foundation for phase 2/3 clinical trials to ascertain the clinical efficacy of FLT3 inhibitors and HHT in combination.

Molecular and Cellular Mechanisms of Myelodysplastic Syndrome: Implications on Targeted Therapy

Myelodysplastic syndrome (MDS) is a group of heterogeneous clonal hematopoietic stem cell disorders characterized by cytopenia, ineffective hematopoiesis, and progression to secondary acute myeloid leukemia in high-risk cases. Conventional prognostication relies on clinicopathological parameters supplemented by cytogenetic information. However, recent studies have shown that genetic aberrations also have critical impacts on treatment outcome. Moreover, these genetic alterations may themselves be a target for treatment. The mutation landscape in MDS is shaped by gene aberrations involved in DNA methylation (TET2, DNMT3A, IDH1/2), histone modification (ASXL1, EZH2), the RNA splicing machinery (SF3B1, SRSF2, ZRSR2, U2AF1/2), transcription (RUNX1, TP53, BCOR, PHF6, NCOR, CEBPA, GATA2), tyrosine kinase receptor signaling (JAK2, MPL, FLT3, GNAS, KIT), RAS pathways (KRAS, NRAS, CBL, NF1, PTPN11), DNA repair (ATM, BRCC3, DLRE1C, FANCL), and cohesion complexes (STAG2, CTCF, SMC1A, RAD21). A detailed understanding of the pathogenetic mechanisms leading to transformation is critical for designing single-agent or combinatorial approaches in target therapy of MDS.

Next-generation sequencing with a myeloid gene panel in core-binding factor AML showed KIT activation loop and TET2 mutations predictive of outcome.

Clinical outcome and mutations of 96 core-binding factor acute myeloid leukemia (AML) patients 18–60 years old were examined. Complete remission (CR) after induction was 94.6%. There was no significant difference in CR, leukemia-free-survival (LFS) and overall survival (OS) between t(8;21) (N=67) and inv(16) patients (N=29). Univariate analysis showed hematopoietic stem cell transplantation at CR1 as the only clinical parameter associated with superior LFS. Next-generation sequencing based on a myeloid gene panel was performed in 72 patients. Mutations in genes involved in cell signaling were associated with inferior LFS and OS, whereas those in genes involved in DNA methylation were associated with inferior LFS. KIT activation loop (AL) mutations occurred in 25 patients, and were associated with inferior LFS (P=0.003) and OS (P=0.001). TET2 mutations occurred in 8 patients, and were associated with significantly shorter LFS (P=0.015) but not OS. Patients negative for KIT-AL and TET2 mutations (N=41) had significantly better LFS (P<0.001) and OS (P=0.012) than those positive for both or either mutation. Multivariate analysis showed that KIT-AL and TET2 mutations were associated with inferior LFS, whereas age ⩾40 years and marrow blast ⩾70% were associated with inferior OS. These observations provide new insights that may guide better treatment for this AML subtype.

T-cell lymphoma: Microenvironment-related biomarkers.

Mature T-cell lymphomas are a heterogeneous group of lymphoid malignancies that occur in nodal and extranodal sites. The diverse tissues involved and the highly variable clinicopathologic features are related to an interaction between the neoplastic lymphoid cells and the tissues that they infiltrate. When different subpopulations of T-cells undergo transformation, they retain some of their inherent characteristics, including tissue tropisms, gene expression profiles and cytokine secretion patterns; which collaborate to impact on the cellular composition and structure of the lymphoma. Non-neoplastic cellular components of the lymphoma, including normal T and B lymphocytes, macrophages and eosinophils are altered by the neoplastic lymphoid cells, leading to changes in patterns of gene and protein expressions. These changes in turn modify the microenvironment of the lymphoma, inducing blood vessel formation and providing survival and proliferative signals to the neoplastic lymphoid cells. Tumor-infiltrating macrophages undergo M2 polarization as a result of the cytokines produced by the neoplastic T-cells. These macrophages in turn promote angiogenesis and inhibit anti-tumor cellular immunity. The macrophage content has been shown to correlate with treatment outcome in clinical studies. The effects of normal lymphocytes on T-cell lymphoma biology are more conflicting, with both stimulatory and inhibitory effects on the neoplastic cells demonstrated in vitro. Endothelial cells from micro-vessels interact with neoplastic T-cells mainly through the production of vascular endothelial growth factor. The prognostic and therapeutic significance of microvasculature have also been demonstrated in some clinical trials. This complex network of cellular, immunological and molecular interactions underlies the pathologies, clinical manifestations and treatment outcome of T-cell lymphomas. The understanding of the impact of the microenvironment on neoplastic T-cells should be further exploited in translational research to generate biomarkers that may be applicable to disease classification, treatment and prognostication.

Sequential chemotherapy/radiotherapy was comparable with concurrent chemoradiotherapy for stage I/II NK/T-cell lymphoma

Background In stage I/II natural killer (NK)/T-cell lymphoma, concurrent chemoradiotherapy (CCRT) had previously been shown to result in superior outcome compared with anthracycline-containing regimens, which have since been considered ineffective. The role of CCRT in comparison with approaches employing nonanthracycline-containing chemotherapy (CT) and sequential radiotherapy (RT) in such patients remains to be defined. Patients and methods Three hundred and three untreated patients (207 men, 96 women; median age: 51, 18-86 years) with stage I/II NK/T-cell lymphoma who had received nonanthracycline-containing regimens were collected from an international consortium and retrospectively analyzed. Treatment included single modality (CT and RT), sequential modalities (CT + RT; RT + CT) and concurrent modalities (CCRT; CCRT + CT). The impact of clinicopathologic parameters and types of treatment on complete response (CR) rate, progression-free-survival (PFS) and overall-survival (OS) was evaluated. Results For CR, stage (P = 0.027), prognostic index for NK/T-cell lymphoma (PINK) (P = 0.026) and types of initial treatment (P = 0.011) were significant prognostic factors on multivariate analysis. On Cox regression analysis, ECOG performance score (P = 0.021) and PINK-EBV DNA (PINK-E) (P = 0.002) significantly impacted on PFS; whereas ECOG performance score (P = 0.008) and stage (P < 0.001) significantly impacted on OS. For comparing CCRT ± CT and sequential CT + RT, CCRT ± CT patients (n = 190) were similar to sequential CT + RT patients (n = 54) in all evaluated clinicopathologic parameters except two significantly superior features (higher proportion of undetectable circulating EBV DNA on diagnosis and lower PINK-E scores). Despite more favorable pre-treatment characteristics, CCRT ± CT patients had CR rate, PFS and OS comparable with sequential CT + RT patients on multivariate and Cox regression analyses. Conclusions In stage I/II NK/T-cell lymphomas, when effective chemotherapeutic regimens were used, CCRT and sequential CT + RT gave similar outcome.

miR-874-3p is down-regulated in hepatocellular carcinoma and negatively regulates PIN1 expression

PIN1 is a peptidyl-prolyl cis/trans isomerase (PPIase) that regulates multiple signaling pathways to control cell fate and is found to be over-expressed in cancers, including hepatocellular carcinoma (HCC). However, the regulation of PIN1 in HCC remains poorly defined. Micro-RNAs (miRNAs) have been reported to play a pivotal role in oncogenesis by targeting the 3′-untranslated region (UTR) of mRNAs encoded by oncogenes and tumour suppressor genes, thereby suppressing the levels of both oncoproteins and tumour suppressors. In this report, we aimed to identify miRNAs that suppress PIN1 expression and to determine their role in HCC. By searching the TargetScan database, miR-874-3p was identified as a potential negative regulator of PIN1. miR-874-3p was demonstrated to bind the 3′UTR of PIN1 mRNA directly to suppress expression of PIN1. Functionally, over-expression of miR-874-3p in HCC cell line PLC/PRF/5 inhibited cell growth and colony formation in-vitro, and promoted cellular apoptosis. Furthermore, these tumour suppressive functions conferred by miR-874-3p were abrogated by over-expression of PIN1. Similarly, expression of miR-874-3p in PLC/PRF/5 with PIN1 knocked-down did not further suppress cellular proliferation, suggesting that PIN1 was a major target of miR-874-3p. More importantly, miR-874-3p was found to be down-regulated in HCC tissues and its expression was negatively correlated with that of PIN1. Down-regulation of miR-874-3p was also associated with poorly differentiated tumour cells, more advanced staging, and inferior patient outcomes. In addition, over-expression of miR-874-3p suppressed tumour growth in vivo. Taken together, our data suggested that miR-874-3p plays a tumour suppressive role in HCC through down-regulation of PIN1.

From chronic immune thrombocytopenia to severe aplastic anemia: recent insights into the evolution of eltrombopag:

Thrombopoietin (TPO) is the most potent cytokine stimulating thrombopoiesis. Therapy with exogenous TPO is limited by the formation of antibodies cross-reacting with endogenous TPO. Mimetics of TPO are compounds with no antigenic similarity to TPO. Eltrombopag is an orally-active nonpeptide small molecule that binds to the transmembrane portion of the TPO receptor MPL. Initial trials of eltrombopag have centered on immune thrombocytopenia (ITP), which is due to both increased destruction and decreased production of platelets. Eltrombopag at 25–75 mg/day has been shown to be highly effective in raising the platelet count in ITP with suboptimal response to immunosuppression and splenectomy. These successful results led to the exploration of eltrombopag in other thrombocytopenic disorders. In hepatitis C viral infection, eltrombopag raises the platelet count sufficiently enough to allow treatment with ribavirin and pegylated interferon. Because MPL is expressed on hematopoietic cells, eltrombopag use in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) might enhance leukemic proliferation. Clinical trials of eltrombopag in MDS and AML, however, have shown amelioration of thrombocytopenia without promoting disease progression. In severe aplastic anemia (SAA) not responding to immunosuppression with anti-thymocyte globulin (ATG) and cyclosporine, eltrombopag as a single agent at 150–300 mg/day results in an overall response rate of 40–70%. At high doses, adverse effects including pigmentation, gastrointestinal upset and hepatic derangement have become evident. Current studies have examined the first-line use of eltrombopag in combination with ATG in SAA. In a recent study, eltrombopag used at 150 mg/day with horse ATG resulted in an overall response rate of 90% in newly diagnosed SAA patients, with a complete response rate of about 50%. Clonal karyotypic aberrations are, however, found in 10–20% of SAA patients treated with eltrombopag. The safety and efficacy of eltrombopag in SAA require further evaluation, particularly when it is used with less intensive immunosuppression.