Jui-Feng Hsu

Altered expression of circadian clock genes in human chronic myeloid leukemia.

Circadian clock genes use transcriptional-translational feedback loops to control circadian rhythms. Recent studies have demonstrated that expression of some circadian clock genes displays daily oscillation in peripheral tissues including peripheral blood and bone marrow. Circadian rhythms regulate various functions of human body, and the disruption of circadian rhythm has been associated with cancer development and tumor progression. However, the direct links between aberrant circadian clock gene expression and human disorders remain largely unknown. In this study, comparisons were made between the expression profiles of 9 circadian clock genes from peripheral blood mononuclear cells (PBMCs) and polymorphonuclear cells (PMNs) from 18 healthy volunteers. Peripheral blood (PB) total leukocytes from 54 healthy volunteers and 95 patients with chronic myeloid leukemia (CML) were also investigated. Similar expression profiles of all 9 circadian clock genes were observed in PBMCs and PMNs of healthy individuals. In PB total leukocytes of healthy individuals, the daily pattern of PER1, PER2, PER3, CRY1, CRY2, and CKIε expression level peaked at 0800 h, and BMAL1 peaked at 2000 h. Daily pattern expression of these 7 genes was disrupted in newly diagnosed pre—imatinib mesylate—treated and blast crisis—phase patients with CML. Partial daily pattern gene expression recoveries were observed in patients with CML with complete cytogenetic response and major molecular response. The expression of CLOCK and TIM did not show a time-dependent variation among the healthy and patients with CML. These results indicate a possible association of the disrupted daily patterns of circadian clock gene expression with the pathogenesis of CML.

Induction of Cellular Senescence by Doxorubicin Is Associated with Upregulated miR-375 and Induction of Autophagy in K562 Cells

Background Cellular senescence is a specialized form of growth arrest that is generally irreversible. Upregulated p16, p53, and p21 expression and silencing of E2F target genes have been characterized to promote the establishment of senescence. It can be further aided by the transcriptional repression of proliferation-associated genes by the action of HP1γ, HMGA, and DNMT proteins to produce a repressive chromatin environment. Therefore, senescence has been suggested to functions as a natural brake for tumor development and plays a critical role in tumor suppression and aging. Methodology/Principal Findings An in vitro senescence model has been established by using K562 cells treated with 50 nM doxorubicin (DOX). Since p53 and p16 are homozygously deleted in the K562 cells, the DOX-induced senescence in K562 cells ought to be independent of p53 and p16-pRb pathways. Indeed, no change in the expression of the typical senescence-associated premalignant cell markers in the DOX-induced senescent K562 cells was found. MicroRNA profiling revealed upregulated miR-375 in DOX-induced senescent K562 cells. Treatment with miR-375 inhibitor was able to reverse the proliferation ability suppressed by DOX (p<0.05) and overexpression of miR-375 suppressed the normal proliferation of K562 cells. Upregulated miR-375 expression was associated with downregulated expression of 14-3-3zeta and SP1 genes. Autophagy was also investigated since DOX treatment was able to induce cells entering senescence and eventually lead to cell death. Among the 24 human autophagy-related genes examined, a 12-fold increase of ATG9B at day 4 and a 20-fold increase of ATG18 at day 2 after DOX treatment were noted. Conclusions/Significance This study has demonstrated that in the absence of p53 and p16, the induction of senescence by DOX was associated with upregulation of miR-375 and autophagy initiation. The anti-proliferative function of miR-375 is possibly exerted, at least in part, by targeting 14-3-3zeta and SP1 genes.

Herpes zoster is associated with an increased risk of subsequent lymphoid malignancies - A nationwide population-based matched-control study in Taiwan

BackgroundInfectious agents have been shown to contribute to the development of lymphoid malignancies. The different distribution of lymphoid malignancies in Asian and Western populations suggests possibly different etiologies in Asian populations. Herpes zoster infection, commonly seen in immunocompromised persons, has been reported to be associated with lymphoid malignancies in retrospective case–control studies from Western populations, but the results are controversial and large-scale prospective studies from Asian populations are lacking.MethodsA nationwide population-based matched-controlled prospective study on Taiwanese patients was performed using the National Health Insurance Research Database from 1996 to 2007. Herpes zoster and malignancies were defined by compatible ICD-9-CM (International Classification of Disease, 9th Revision, Clinical Modification) codes. Patients who had been diagnosed with any malignancies before herpes zoster, with known viral infections including human immunodeficiency virus, and duration from herpes zoster to diagnosis of malignancies less than 6 months were excluded.ResultsOf 42,498 patients with herpes zoster prior to the diagnosis of any malignancies, the cumulative incidence for lymphoid malignancies was 0.11% (n = 48), compared with 0.06% (n = 106) in 169,983 age- and gender-matched controls (univariate hazard ratio (HR): 1.82, 95%CI: 1.29-2.55). The most common lymphoid malignancy was non-Hodgkin’s lymphoma (60.4%, n = 29), followed by multiple myeloma (27.1%, n = 13). Risk for developing lymphoid malignancies is significantly higher in herpes zoster patients (log rank P = 0.005). After adjusting for presence of any comorbidities in Charlson comorbidity index, time-dependent covariate for herpes group, and income category using Cox proportional hazard regressions, herpes zoster patients had an increased risk of developing lymphoid malignancies (adjusted HR: 1.68, 95%CI: 1.35-2.42, P = 0.0026), but did not have an increased risk of developing non-lymphoid malignancies (adjusted HR: 1.00, 95%CI: 0.91-1.05, P = 0.872).ConclusionPreceding herpes zoster infection is an independent risk marker for subsequent lymphoid malignancies in Taiwanese subjects. Further studies are warranted for pathogenesis exploration and preventive strategies in Asian populations.

Additional chromosome abnormalities in chronic myeloid leukemia

The Philadelphia (Ph) chromosome and/or Breakpoint cluster region‐Abelson leukemia virus oncogene transcript are unique markers for chronic myeloid leukemia (CML). However, CML demonstrates heterogeneous presentations and outcomes. We analyzed the cytogenetic and molecular results of CML patients to evaluate their correlation with clinical presentations and outcome. A total of 84 newly diagnosed CML patients were enrolled in the study. Patients were treated according to disease status. Bone marrow samples were obtained to perform cytogenetic and molecular studies. Clinical presentations, treatment courses, and survival were reviewed retrospectively. Among 84 patients, 72 had chronic phase and 12 had accelerated phase CML. Cytogenetic study showed 69 (82.1%) with the classic Ph chromosome, 6 (7.2%) with a variant Ph chromosome, and 9 (10.7%) with additional chromosome abnormalities. Fifty‐four (64.3%) cases harbored b3a2 transcripts, 29 (34.5%) had b2a2 transcript, and 1 had e19a2 transcript. There was no difference in clinical presentations between different cytogenetic and molecular groups; however, additional chromosome abnormalities were significantly associated with the accelerated phase. Imatinib therapy was an effective treatment, as measured by cytogenetic response, when administered as first‐ and second‐line therapy in chronic phase patients. Survival analysis showed that old age, additional chromosome abnormalities, high Sokal score, and no cytogenetic response in second‐line therapy had a significant poor impact (p < 0.05). In conclusion, we presented the cytogenetic and molecular pattern of CML patients and demonstrated that the additional chromosome abnormality was associated with poor outcome.

Breakthrough Fusarium solani infection in a patient with acute myeloid leukemia receiving posaconazole prophylaxis.

Dear Editor, A 23-year-old woman presented with fever, exertional dyspnea, and bruised limbs was diagnosed as having acute myeloid leukemia (AML; French–American–British classification type M0) with normal metaphase cytogenetics and negative NPM1 , FLT3-ITD , and CEBPA gene mutations. Laboratory evaluations revealed a leukocyte count of 65,800/μL (blasts 83.5 %), hemoglobin of 3.8 g/dL, and platelets of 9,000/μL. She received induction chemotherapy with idarubicin and cytarabine (I3A7) along with cefepime. The fever subsided without identified organisms. Posaconazole (200 mg three times daily) was started as antifungal prophylaxis on day 1 of chemotherapy. The fever recurred on day 9, and the antibiotics were shifted to imipenam/cilastatin and vancomycin. However, the fever persisted, and multiple tender erythematous papules with central necrosis developed over lower limbs on day 13, rapidly extended to trunk, upper limbs, and scalp areas. Scattered whitish plaques were found on soft palate and conjunctiva (Fig. 1a–d). Skin biopsy revealed numerous fungal hyphae highlighted by Gomori methenamine silver stain (Fig. 1e, f). Whole-body imaging studies showed maxillary sinusitis without pulmonary, intracranial, or intraabdominal lesions. Cultures from blood and skin lesions yielded Fusarium species. Amphotericin B (1 mg kg day) was used instead of posaconazole. Fusarium solani was identified by microscopic morphology and confirmed by polymerase chain reactionbased methods targeting elongation factor alpha and internal transcribed spacers [1, 2]. The minimal inhibitory concentration study revealed 1 μg/mL for amphotericin B, 8 μg/mL for voriconazole, >8 μg/mL for posaconazole, caspofungin, micafungin, and anidulafungin, and >16 μg/ mL for itraconazole. The fever subsided following amphotericin B; the skin lesions resolved gradually and turned to crust-like appearance without recurrence. Subsequent cultures were negative for Fusarium species. Despite the improvement in disseminated fusariosis, failure to achieve remission and increasing blasts refractory to further antileukemia therapies indicated a primary refractory disease. The patient eventually expired on day 78 due to refractory AML. Posaconazole is effective as primary antifungal prophylaxis for invasive fungal diseases (IFDs) in AML patients [3]. To date, only two AML cases of breakthrough fusariosis were reported from a single institution without identification of the species [4]. The incidence of proven and probable IFDs following posaconazole prophylaxis varied C.<H. Wu :H.<H. Hsiao : T.<C. Liu : S.<F. Lin : C.<S. Chang : J.<F. Hsu :Y.<C. Liu (*) Division of Hematology–Oncology, Department of Internal Medicine, Kaohsiung Medical University Hospital, 100 Tzyou 1st Road, 807, Kaohsiung, Taiwan e-mail: ycliu@kmu.edu.tw

Higher lipocalin 2 expression may represent an independent favorable prognostic factor in cytogenetically normal acute myeloid leukemia

Abstract Several molecular markers, such as NPM1, FLT3 and CEBPA, have been incorporated into both the World Health Organization and European LeukemiaNet classifications as routine assessments for the diagnosis and evaluation of prognostic significance in acute myeloid leukemia (AML). Lipocalin 2 (LCN2) is related to cancer development and is believed to be associated with the outcome of cytogenetically normal (CN)-AML. In the present study, we analyzed the prognostic effects and interactions of LCN2 expression (by molecular analysis, quantitative real-time polymerase chain reaction [qRT-PCR]) with neucleophosmin 1, fms-related tyrosine kinase 3 (FLT3) and CCAAT/enhancer-binding protein alpha mutations in 85 patients with CN-AML receiving intensive induction chemotherapy. Our results indicate that patients with higher LCN2 mRNA expression in the bone marrow (LCN2high), especially in combination with wild type FLT3-ITD, had better prognoses. FLT3-ITD compensated LCN2-overexpression-enhanced oxidative stress-induced apoptosis in cell line studies. In conclusion, LCN2high was associated with better prognosis, and FLT3 status had an adjuvant effect on overall survival.

MDM2 promoter polymorphism and p53 codon 72 polymorphism in chronic myeloid leukemia: The association between MDM2 promoter genotype and disease susceptibility, age of onset, and blast-free survival in chronic phase patients receiving imatinib

The genetic or functional inactivation of the p53 pathway plays an important role with regards to disease progression from the chronic phase (CP) to blast phase (BP) and imatinib treatment response in chronic myeloid leukemia (CML). Two functional single nucleotide polymorphisms (SNPs), p53 R72P and MDM2 SNP309, are associated with alternation of p53 activity, however the association regarding CML susceptibility and BP transformation under imatinib treatment is unclear. The MDM2 SNP309 genotype was determined by polymerase chain reaction–restriction fragment length polymorphism and confirmed by direct sequencing from 116 CML patients, including 104 in the CP at diagnosis, and 162 healthy Taiwanese controls. The p53 R72P polymorphism was examined in all CML patients. The SNP309 G/G genotype was associated with an increased risk of CML susceptibility (OR: 1.82, 95% CI: 1.03–3.22, P = 0.037), and an earlier age of disease onset (log‐rank P = 0.005) compared with the T/T + T/G genotypes. Higher MDM2 mRNA expression was found in G/G genotype compared with T/T (P = 0.034) and T/T + T/G (P = 0.056) genotypes. No associations were found between the p53 R72P genotypes and clinical parameters and survival outcomes. Among 62 CP patients receiving imatinib as first‐line therapy, the G/G genotype was associated with a shorter blast‐free survival (log‐rank P = 0.048) and more clonal evolution compared with the T/T + T/G genotypes. In patients with advanced diseases at diagnosis, the G/G genotype was associated with a poor overall survival (log‐rank P = 0.006). Closely monitoring CML patients harboring the G/G genotype and further large‐scale studies are warranted.

Hepatitis C transmission from viremic donors in hematopoietic stem cell transplant.

Transmission of hepatitis C virus (HCV) to recipients of hematopoietic stem cell transplant (HSCT) occurs frequently from HCV viremic donors and causes complications. Here, we report the outcomes of 3 cases from our 265 allogeneic HSCTs, whose donors had HCV infections. Successful prevention of HCV transmission was noted in 1 recipient by pretreatment of the donor with peginterferon/ribavirin to undetectable levels of HCV viremia before stem cell harvest. This case stressed the important role of effective antiviral therapy and HCV RNA seronegativity before cell harvest for prevention of HCV transmission in HSCT.

JAK2V617F mutation is associated with special alleles in essential thrombocythemia.

Janus kinase 2 mutation (JAK2V617F) has been identified in myeloproliferative neoplasms. Furthermore, special single nucleoside polymorphisms (SNPs) have been found to be associated with the JAK2V617F mutation. Therefore, the associations among JAK2V617F and special SNPs and the allelic location between them were investigated in patients with essential thrombocythemia (ET). A total of 61 patients with ET and 106 healthy individuals were enrolled. The PCR-RFLP method was applied to investigate the pattern of three SNPs, rs10974944, rs12343867, and rs12340895. Allele-specific PCR was used to examine the allelic location between rs10974944 and JAK2V617F. Among the patients with ET, 34 (55.7%, 34/61) were JAK2V617F positive (heterozygous) while the other 27 (44.3%, 27/61) were negative, and there were no MPLW515L/K mutations noted. The pattern of special SNPs in JAK2V617F(+) was significantly different from that in normal individuals (p <0.05), while there was no difference between JAK2V617F(−) patients and normal individuals. Allele-specific PCR showed high association of a cis-location between the special G-allele of rs10974944 and JAK2V617F(+). Based on this small numbered study, the results show the association between special SNPs and JAK2V617F mutation and a cis-location between the special G-allelic form of rs10974944 and the JAK2V617F mutation. These data highlight a close relationship between them in patients with ET.

Recurrent local advanced hypopharyngeal squamous cell carcinoma with isolated quadriceps femoralis metastasis

We report the case of a 43-year-old male with a history of hypopharyngeal squamous cell carcinoma (SCC) (T1N1M0, Stage III) in February 2010, who was treated with wide excision and concurrent chemoradiation therapy. No evidence of recurrence was found during 16 months of followup. He had been suffering from left knee pain while walking or at rest since June 2011. Four months later, one palpable, fixed, aggregately painful mass over the medial site of the left distal thigh was noted, and then he visited our orthopedic outpatient department. The physical examination revealed swelling of the left knee, and one nonmovable, firm, tender mass measuring about 6 5 cm over the distal, medial part of the left quadriceps femoris. No other abnormalities were found. A left femur X-ray revealed soft-tissue calcification at the medial aspect of the distal thigh medial to the femur without remarkable bony destruction (Fig. 1A). The magnetic resonance tomography scan of low limb revealed an infiltrative fusiform soft-tissue tumor (5.2 4.9 cm) with central necrosis adjacent to the medial aspect of the distal femur (Fig. 1B). A complete surgical excision of the mass was performed, and the pathological study revealed that the soft tissue was infiltrated by neoplastic cells exhibiting hyperchromatic, pleomorphic nuclei and distinct nucleoli; keratin pearls were also found (Fig. 1C). The result was compatible with the morphology of the previous hypopharyngeal SCC. No secondary metastatic lesions were found via whole body imaging using 18F-fluoro-deoxy-glucose positron emission