Novel molecular mechanism of cellular transformation by a mutant molecular chaperone in myeloproliferative neoplasms

Deregulation of the cytokine‐receptor signaling pathway plays a significant role in tumorigenesis. Such deregulation is frequently caused by alterations in the genes involved in the signaling pathway. At the end of 2013, recurrent somatic mutations in the calreticulin (CALR) gene that encodes a molecular chaperone were identified in a subset of patients with Philadelphia‐chromosome negative myeloproliferative neoplasms (MPN). The present review focuses on the role of CALR mutations in the oncogenic transformations observed in MPN. All the CALR mutations were found to generate a + 1 frameshift in the reading frame on exon 9, which encodes the carboxy (C)‐terminus end of CALR, and thus conferred a common mutant‐specific sequence in all the CALR mutants. The mutant CALR (but not the wild‐type) constitutively activates the thrombopoietin (TPO) receptor, myeloproliferative leukemia protein (MPL), even in the absence of TPO to induce cellular transformation. Preferential interaction between the mutant CALR and MPL is achieved by a presumptive conformational change induced by the mutant‐specific C‐terminus domain, which allows N‐domain binding to MPL. Even though mutant CALR is expressed on the cell surface and is secreted out of cells, it only presents autocrine capacity for MPL activation. These findings define a novel molecular mechanism by which the mutant molecular chaperone constitutively activates the cytokine receptor to induce cellular transformation.     

Spleen deflation and beyond: The pros and cons of Janus kinase 2 inhibitor therapy for patients with myeloproliferative neoplasms

The myeloproliferative neoplasms (MPNs) essential thrombocythemia (ET), polycythemia vera (PV), and primary myelofibrosis (MF) are malignancies that frequently harbor the recurrent somatic point mutation JAK2^V617F. The discovery of this mutation has fueled the development of Janus kinase 2 (JAK2) inhibitors. Available results have indicated that JAK2 inhibitors are particularly effective at reducing spleen size. However, the activity of these agents is multifaceted and also involves a marked improvement of systemic symptoms and, for those agents with dual JAK1 and JAK2 inhibitory activity, a marked reduction in the levels of circulating cytokines involved in the pathogenesis of the disease. Because JAK2 inhibitors are not specific for JAK2^V617F, responses have also been observed in JAK2^V617F‐negative MPNs because of the inhibition of wild‐type JAK2, which is also likely responsible for the induction of cytopenias in patients with MF and for the normalization of peripheral blood counts observed in patients with ET or PV. Given the distinct mortality and morbidity associated with ET, PV, and MF, the use of JAK2 inhibitors appears reasonable for patients with MF as well as for those with ET or PV who have become resistant or intolerant to hydroxyurea. Ongoing randomized, placebo‐controlled, phase 3 trials will further delineate the role of these agents in the management of patients with MPNs. The pros and cons of JAK2 kinase inhibitor therapy are herein discussed. Cancer 2012;. © 2011 American Cancer Society.     

The molecular landscape of childhood myeloproliferative neoplasms

The classical myeloproliferative neoplasms (MPN) are comparatively uncommon in children and display a degree of mutational naivety if considering the high frequency of known MPN driver events observed in the corresponding adult diseases. Whole exome sequencing has unravelled much of the underlying molecular complexity of MPN in adult patients yet less is known of the pathogenetic mechanisms when these diseases occur in childhood. It is proposed that such methodological approaches will contribute significant insights into the molecular landscape of childhood MPN that may in turn impact on understanding the pathophysiology of disease in their adult counterparts.     

Efficacy of NS-018, a potent and selective JAK2/Src inhibitor, in primary cells and mouse models of myeloproliferative neoplasms

Aberrant activation of Janus kinase 2 (JAK2) caused by somatic mutation of JAK2 (JAK2V617F) or the thrombopoietin receptor (MPLW515L) plays an essential role in the pathogenesis of myeloproliferative neoplasms (MPNs), suggesting that inhibition of aberrant JAK2 activation would have a therapeutic benefit. Our novel JAK2 inhibitor, NS-018, was highly active against JAK2 with a 50% inhibition (IC₅₀) of <1 n, and had 30–50-fold greater selectivity for JAK2 over other JAK-family kinases, such as JAK1, JAK3 and tyrosine kinase 2. In addition to JAK2, NS-018 inhibited Src-family kinases. NS-018 showed potent antiproliferative activity against cell lines expressing a constitutively activated JAK2 (the JAK2V617F or MPLW515L mutations or the TEL–JAK2 fusion gene; IC₅₀=11–120 n), but showed only minimal cytotoxicity against most other hematopoietic cell lines without a constitutively activated JAK2. Furthermore, NS-018 preferentially suppressed in vitro erythropoietin-independent endogenous colony formation from polycythemia vera patients. NS-018 also markedly reduced splenomegaly and prolonged the survival of mice inoculated with Ba/F3 cells harboring JAK2V617F. In addition, NS-018 significantly reduced leukocytosis, hepatosplenomegaly and extramedullary hematopoiesis, improved nutritional status, and prolonged survival in JAK2V617F transgenic mice. These results suggest that NS-018 will be a promising candidate for the treatment of MPNs.     

费城染色体阴性骨髓增殖性肿瘤的诊断与治疗:JAK2 V617F基因突变评价

简要回顾了近百余年人们对骨髓增殖性肿瘤(MPN)的认识过程,重点讨论这一类疾病的诊断与治疗.JAK2 V617F基因突变的发现将费城染色体阴性(Ph-)MPN带入分子生物学时代,为临床提供了重要的诊断手段和依据,指导、研发了芦可替尼(ruxolitinib)等一批靶向药物.但是,与慢性粒细胞白血病(CML)中的bcr-abl不同,JAK2 V617F突变不是MPN诊断的“金标准”,其他辅助检查和鉴别诊断仍不可少.目前,JAK抑制剂开始用于Ph-MPN患者,有一定的适应证,远期疗效正在观察,目前还不能替代有效的常规治疗,如羟基脲、阿司匹林等.     

8p11骨髓增殖综合征的研究进展

8p11骨髓增殖综合征是一种少见的以嗜酸粒细胞增多及合并T淋巴母细胞淋巴瘤／白血病为主要特点的骨髓增殖性疾病,所有患者都有包括发生于8p11染色体上的成纤维细胞生长因子受体1（FGFR1）基因突变,造血干细胞移植是唯一有希望能治愈此病的手段.     

Development of ZMYM2‐FGFR1 driven AML in human CD34+ cells in immunocompromised mice

Acute myelogenous leukemia (AML) has an overall poor survival rate and shows considerable molecular heterogeneity in its etiology. In the WHO classification there are >50 cytogenetic subgroups of AML, many showing highly specific chromosome translocations that lead to constitutive activation of individual kinases. In a rare stem cell leukemia/lymphoma syndrome, translocations involving 8p11 lead to constitutive activation of the fibroblast growth factor receptor 1 (FGFR1) kinase. This disorder shows myeloproliferative disease with almost invariable progresses to AML and conventional therapeutic strategies are largely unsuccessful. Because of the rare nature of this syndrome, models that faithfully recapitulate the human disease are needed to evaluate therapeutic strategies. The t(8;13)(p11;q12) chromosome translocation is most common rearrangement seen in this syndrome and creates a ZMYM2‐FGFR1 chimeric kinase. To understand more about the molecular etiology of AML induced by this particular rearrangement, we have created a model human CD34+ cells transplanted into immunocompromized mice which develop myeloproliferative disease that progresses to AML with a long (>12 months) latency period. As in humans, these mice show hepatospenomegaly, hypercellular bone marrow and a CD45 + CD34 + CD13+ immunophenotype. Molecular studies demonstrate upregulation of genes such as KLF4 and FLT3 that promote stemness, and overexpression of MYC, which is associated with suppression of myeloid cell differentiation. This murine model, therefore, provides an opportunity to develop therapeutic strategies against the most common subtype within these FGFR1 driven neoplasms and study the molecular etiology in more depth.     

Diagnostic application of next-generation sequencing in ZMYM2-FGFR1 8p11 myeloproliferative syndrome: A case report

The 8p11 myeloproliferative syndrome (EMS), also known as 8p11 myeloproliferative neoplasm (8p11 MPN), is a collection of rare hematologic malignancies that are associated with fusion genes involving the tyrosine kinase receptor gene FGFR1 in chromosome 8p11. The entity is an aggressive disease with a high rate of transformation to acute myeloid leukemia (AML) and pathologically characterized by its associated eosinophilia. In this study, we reported a distinctive EMS case featuring an in-frame ZMYM2-FGFR1 fusion gene identified by next-generation sequencing technology (NGS). This patient exhibited not only typical EMS signs including elevated white blood cells in peripheral blood and hypercellular bone marrow with marked leukocytosis, but also exceptional characteristics including erythrocytosis in blood and bone marrow basophilia. Moreover, we detected 2 novel genomic mutations in 2 known leukemogenic genes, IKZF1 and ASXL1. Whether these 2 mutations play a part in EMS pathogenesis or contribute to its specific presentations clinically remain to be determined. In summary, we present a unique EMS case involving a ZMYM2-FGFR1 fusion with distinctive hematologic characteristics.     

Common features of myeloproliferative disorders with t(8;9)(p12;q33) and CEP110-FGFR1 fusion: report of a new case and review of the literature

The 8p12 myeloproliferative syndrome is a rare, generally aggressive chronic myeloproliferative disorder (MPD). The hallmark of this MPD is the disruption of the FGFR1 gene, which encodes a tyrosine kinase receptor for members of the fibroblast growth factor family. In MPD cells FGFR1 is fused to several partners. The most frequent partner genes are BCR, CEP110, FOP, and ZNF198, localized on 22q11, 9q33, 6q27, and 13q12, respectively. We report here the tenth case of translocation (8;9)(p12;q33) in an acute myelomonocytic leukemia and provide a review of the literature that points to common syndrome features: the t(8;9)(p11;q33) MPD transforms rapidly, and always in myelomonocytic leukemia, with a possible B- or T-lymphoid involvement, which may include tonsil invasion. The FGFR1-MPD seems refractory to current chemotherapies and is not sensitive to imatinib. Currently, only the patients with bone marrow transplantation stand a chance of survival.     

胶质瘤中FGFR1OP和p57/Kip2蛋白的临床意义

目的:探讨FGFR1OP和p57/Kip2在胶质瘤发生、发展中的作用和临床意义.方法:对中国医科大学附属盛京医院和北京通州区潞河医院的54例胶质瘤手术切除标本,采用SP法进行免疫组织化学染色,检测FGFR1OP和p57/Kip2的表达情况,分析其与胶质瘤临床病理各参数的相关性,并评价其临床意义.结果:FGFR1OP和p57/Kip2在胶质瘤中的阳性表达率分别为66.7%和44.4%,且随着胶质瘤的恶性程度不同表现出显著性差异(P<0.05),FGFR1OP在高度恶性胶质瘤(Ⅲ～Ⅳ级)中的OD值为0.131±0.010,高于在低度恶性胶质瘤(Ⅰ～Ⅱ级)中的OD值(0.118±0.010),两者有显著统计学差异(P=0.000).p57/Kip2在高度恶性胶质瘤(Ⅲ～Ⅳ级)中的OD值(0.156±0.008),低于在低度恶性胶质瘤(Ⅰ～Ⅱ级)中的OD值(0.165±0.006),两者有显著统计学差异(P=0.014).Pearson相关系数检验分析提示FGFR10P和p57/Kip2之间呈负直线相关(r=-0.732,P<0.01).结论:FGFR1OP的高表达与p57/Kip2的低表达可能参与胶质瘤的生长、分化和进展,提示预后不良.     

Molecular mechanisms underlying IGF-I-induced attenuation of the growth-inhibitory activity of trastuzumab (Herceptin) on SKBR3 breast cancer cells

The clinical usefulness of trastuzumab (Herceptin; Genentech, San Francisco, CA) in breast cancer treatment is limited by the rapid development of resistance. We previously reported that IGF-I signaling confers resistance to the growth-inhibitory actions of trastuzumab in a model system, but the underlying molecular mechanism remains unknown. We used SKBR3/neo cells (expressing few IGF-I receptors) and SKBR3/IGF-IR cells (overexpressing IGF-I receptor) as our experimental model. IGF-I antagonized the trastuzumab-induced increase in the level of the Cdk inhibitor p27(Kip1). This resulted in decreased association of p27(Kip1) with Cdk2, restoration of Cdk2 activity and attenuation of cell-cycle arrest in G(1) phase, all of which had been induced by trastuzumab treatment in SKBR3/IGF-IR cells. We also found that the decrease in p27(Kip1) induced by IGF-I was accompanied by an increase in expression of Skp2, which is a ubiquitin ligase for p27(Kip1), and by increased Skp2 association with p27(Kip1). A specific proteasome inhibitor (LLnL) completely blocked the ability of IGF-I to reduce the p27(Kip1) protein level, while IGF-I increased p27(Kip1) ubiquitination. This suggests that the action of IGF-I in conferring resistance to trastuzumab involves targeting of p27(Kip1) to the ubiquitin/proteasome degradation machinery. Finally, specific inhibitors of MAPK and PI3K suggest that the IGF-I-mediated reduction in p27(Kip1) protein level by increased degradation predominantly involves the PI3K pathway. Our results provide an example of resistance to an antineoplastic therapy that targets one tyrosine kinase receptor by increased signal transduction through an alternative pathway in a complex regulatory network.     

Systemic mastocytosis associated with myelodysplastic/myeloproliferative neoplasms with ring sideroblasts and thrombocytosis: Report of three cases

The association of systemic mastocytosis with another hematologic neoplasia of myeloid or lymphoid origin is recognized as an advanced subvariant of mastocytosis. Here, we report the association of indolent or smoldering systemic mastocytosis with three cases of myelodysplastic/myeloproliferative neoplasms with ring sideroblasts and thrombocytosis, a recently recognized disease characterized by SF3B1 mutations. The hierarchical pattern of KIT, SF3B1, JAK2, and additional mutations was studied in whole and fractionated subpopulations of peripheral blood cells and whole bone marrow. In two cases, we could demonstrate a multilineage D816V KIT mutation, involving all myeloid lineages in one patient and also the lymphoid series in the other. Two patients displaying both SF3B1 and V617F JAK2 mutations had a very poor prognosis. Another patient bearing SF3B1, but not V617F JAK2 mutation, had a favorable response to erythropoietin treatment and long survival.     

Association of lnc-LAMC2-1:1 rs2147578 and CASC8 rs10505477 Polymorphisms with Risk of Childhood Acute Lymphoblastic Leukemia

Long non-coding RNAs (lncRNAs) are a novel class of non-protein coding RNAs that are involved in a wide variety of biological processes. There are limited data regarding the impact of lnc-LAMC2-1:1 rs2147578 as well as CASC8 rs10505477 T>C polymorphisms on cancer development. Here we examined for the first time whether rs2147578 and rs10505477 polymorphisms are associated with childhood acute lymphoblastic leukemia (ALL) in a total of 110 cases and 120 healthy controls. Genotyping was achieved by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The rs2147578 variant increased the risk of ALL in codominant (OR=4.33, 95%CI=2.00-9.37, p<0.0001, CG vs CC, and OR=5.81, 95%CI=2.30-14.69, p=0.0002, GG vs CC), dominant (OR=4.63, 95%CI=2.18-9.86, p<0.0001, CG+GG vs CC), overdominant (OR=1.74, 95%CI=1.02-2.97, p=0.0444, CG vs CC+GG) and allele (OR=1.91, 95%CI=1.32-2.77, p=0.0008, G vs C) inheritance models tested. No significant association was found between the CASC8 rs10505477 T>C variant and risk of childhood ALL. In conclusion, the present study revealed that the lnc-LAMC2-1:1 rs2147578 polymorphism may be a risk factor for developing childhood ALL. Further studies with larger sample sizes with different ethnicities are now required to confirm our findings.     

The activation of ERK1/2 via a tyrosine kinase pathway attenuates trail-induced apoptosis in HeLa cells

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) serves as an extracellular signal that triggers apoptosis in tumor cells. To characterize the molecular events involved in TRAIL-induced apoptotic signaling, we investigated the role of extracellular signal-regulated kinase 1/2 (ERK1/2) in HeLa cell death. Here we show that TRAIL-activated ERK1/2 through a tyrosine kinase-dependent pathway, subsequently elevated anti-apoptotic Bcl-2 protein levels. ERK1/2 inhibition with PD98059 promoted apoptotic cell death through the downregulation of ERK1/2 activity and Bcl-2 protein levels. Moreover, tyrosine kinase inhibition with Genistein in TRAIL-induced apoptosis effectively attenuated ERK1/2 activity and enhanced apoptotic cell death. Taken together, our results indicate that ERK1/2 activation via tyrosine kinase pathway plays a protective role as the cellular defense mechanism through the upregulation of Bcl-2 protein levels in TRAIL-induced apoptosis.     

BTK抑制剂治疗原发免疫豁免部位大B细胞淋巴瘤的研究进展

原发免疫豁免部位大B细胞淋巴瘤（primary immune-privileged site large B-cell lymphoma,IP-DLBCLs）是世界卫生组织（WHO）淋巴样肿瘤分类第5版新的类别总称,指一组原发于免疫功能正常患者的免疫屏障之后部位的侵袭性B细胞淋巴瘤,起源于各自的解剖结构（如血脑、血网膜和血睾丸屏障）和各自原发部位的免疫调节系统所形成的免疫庇护所,并具有相同的免疫表型和分子特征,目前包括原发性中枢神经系统大B细胞淋巴瘤（primary central nervous system lymphoma,PCNSL）、原发睾丸大B细胞淋巴瘤（primary testicular large B-cell lymphoma,PTL）和原发玻璃体视网膜大B细胞淋巴瘤（primary vitreoretinal large B-cell lymphoma,PVRL）。该类疾病预后相对较差,目前尚无标准的治疗方案。Toll样受体（TLR）信号（通过MYD88突变）和B细胞受体（BCR）信号（通过CD79B突变）通路介导的NF-κB激活是三者发病的核心机制。该共同属性为这一类疾病的治疗提供了通用的靶点。BTK（Bruton's tyrosine kinase,BTK）是上述信号通路的中心分子。因此,BTK抑制剂可能是这类疾病合理的治疗药物选择。本文将就BTK抑制剂治疗原发免疫豁免部位大B细胞淋巴瘤的作用机制、临床研究、不良反应及耐药问题进行综述。     

Herceptin: mechanisms of action and resistance

HER-2 is overexpressed in 20-25% of invasive breast cancers and is associated with an aggressive tumor phenotype and reduced survival rate. The HER-2 status of a tumor is the critical determinant of response to the HER-2-targeted antibody Herceptin. Thus, accurate assessment of HER-2 expression levels is essential for identifying breast cancer patients who will benefit from HER-2-targeted therapy. Herceptin combined with chemotherapy increases response rates, time to disease progression, and survival. However, the majority of cancers that initially respond to Herceptin begin to progress again within 1 year. This review describes mechanisms by which Herceptin inhibits cell growth in breast cancers that overexpress HER-2 and highlights possible mechanisms contributing to Herceptin resistance.