Eun-Yup Lee

Quercetin enhances susceptibility to NK cell-mediated lysis of tumor cells through induction of NKG2D ligands and suppression of HSP70.

It is known that treatments with heat shock, some anticancer drugs, and ionizing radiation increase the expression of heat-shock proteins (HSPs) and natural killer group 2D (NKG2D) ligands in tumor cells. The increased HSPs may make the tumor cells resistant to apoptosis and reduction of HSPs may make the tumor cells more susceptible to natural killer (NK)-cell mediated lysis of tumor cells. In this study, we investigated whether quercetin which has inhibitory activities against heat-shock factor, protein kinase C, nuclear factor-κB, and phosphatidyl inositol 3-kinase, can modulate the expression of NKG2D ligands and suppress the HSPs in tumor cells. The results of this study showed that quercetin significantly induced the expression of several NKG2D ligands including major histocompatibility complex class I-related chain B, UL16-binding protein 1, and UL16-binding protein 2 in K562, SNU1, and SNU-C4 cells. The quercetin-treated K562, SNU1, and SNU-C4 cells showed an enhanced susceptibility to NK-92 cells through induction of NKG2D ligands. This increased expression of NKG2D ligands seemed to be due to the inhibition of the nuclear factor-κB and phosphatidyl inositol 3-kinase pathways. The findings of this study suggest that the induced NKG2D ligands with the decrease of HSP70 protein by quercetin may provide an attractive strategy to improve the effectiveness of NK cell-based cancer immunotherapy.

Genetic correlation of community-associated methicillin-resistant Staphylococcus aureus strains from carriers and from patients with clinical infection in one region of Korea.

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is an increasingly common worldwide and colonizing S. aureus strains may serve as the causative pathogen for overt clinical infections. This study was performed to determine whether the pathogenic CA-MRSA isolate in clinical infections was genetically related to the MRSA isolates in community carriers. We prospectively collected a total of 42 CA-MRSA isolates (23 clinical infection isolates and 19 colonization isolates) in a local region of Korea. Antimicrobial susceptibility tests, staphylococcal toxin assays, SCCmec typing, multilocus sequence typing (MLST), and spa (staphylococcal protein A) typing were performed with all isolates. Thirty-four (81%) of 42 CA-MRSA isolates belonged to sequence type (ST) 72 in the MLST analysis. The distribution of STs did not differ significantly between colonization and clinical infection isolates (89.5% [17/19] vs. 73.9% [17/23], P=0.26). Among the ST72-MRSA isolates, spa type t664 (18, 52.9%) and t324 (8, 23.5%) were common in both groups. This study demonstrates that the community-associated MRSA strains from patients with clinical infections are closely related to the strains found in carriers from one local community.

Induction of NKG2D ligands and subsequent enhancement of NK cell-mediated lysis of cancer cells by arsenic trioxide.

Natural killer (NK) cells are important effector cells in immune responses to tumor cells and the activation of NK cells is mediated through specific interactions between activating receptors and their cognate ligands. Recently, it has been demonstrated that induction of NKG2D ligands on tumor cells by various stresses render them more sensitive to NK cell-mediated killing. Therefore, in this study, it was investigated whether arsenic trioxide (ATO) could up-regulate NKG2D ligands on tumor cells and increase the susceptibility of cancer cells against NK cells. ATO increased transcription of NKG2D ligands, predominantly ULBP1, in various cancer cell lines, such as K562 chronic myelogenous leukemic cells, NB4 acute promyelocytic leukemic cells, and MCF7 breast cancer cells, and subsequently the surface expression of NKG2D ligands. These results were followed by increased susceptibility of cancer cells to NK cell-mediated cytotoxicity after treatment with ATO. This increase in cytotoxicity was abolished by addition of a blocking NKG2D monoclonal antibody, indicating that the increased susceptibility of ATO-treated cancer cells to cytotoxicity of NK cells was mediated through up-regulation of NKG2D ligands. In addition, abrogation of heat shock proteins induction with KNK437 would sensitize the ATO-treated MCF-7 cells to NK cell-mediated killing. This study suggests that the immunomodulatory property of ATO would be an attractive strategy to improve the effectiveness of NK cell-based cancer immunotherapy.

ABCG2 Q141K polymorphism is associated with chemotherapy-induced diarrhea in patients with diffuse large B-cell lymphoma who received frontline rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone chemotherapy.

ATP‐binding cassette transporter G2 (ABCG2) is the most recently described transporter of the multidrug‐resistance pump and it promotes resistance to anticancer drugs such as doxorubicin, mitoxantrone, topotecan, and SN‐38. Of the ABCG2 polymorphisms, V12M and Q141K alter the functional activity of the ABCG2 transporter and influence the drug response and various toxicities to chemotherapeutic agents. We therefore evaluated the impact of the ABCG2 V12M and Q141K polymorphisms on the therapeutic outcomes and toxicities of primary rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone (R‐CHOP) therapy in 145 Korean patients with diffuse large B‐cell lymphoma (DLBCL). ABCG2 V12M and Q141K genotyping was carried out by pyrosequencing of polymerase chain reaction products. The clinical characteristics, treatment outcomes, toxicities of the patients, and the predictive value of the polymorphisms on response, survival, and adverse events to R‐CHOP for 145 patients were analyzed according to the ABCG2 V12M and Q141K polymorphisms. No differences were observed according to ABCG2 Q141K and V12M genotype in patient characteristics, disease characteristics, response, survival, or hematology toxicity profiles in patients with DLBCL who received frontline R‐CHOP chemotherapy. On multivariate analysis, grade I–IV diarrhea was statistically significant according to ABCG2 Q141K polymorphism (the QQ genotype vs the QK or KK genotypes; hazard ratio 2.835; 95% confidence interval 1.432–5.613; P = 0.003). This study demonstrates that the ABCG2 Q141K polymorphism may correlate with chemotherapy‐induced diarrhea in patients with DLBCL who have received frontline R‐CHOP chemotherapy, and this has implications for optimizing treatment with such agents. (Cancer Sci 2008; 99: 2496–2501)

Phase I study of autologous dendritic cell tumor vaccine in patients with non-small cell lung cancer

BACKGROUND A dendritic cell vaccine has been developed as a novel strategy for generating antitumor immunity in the treatment of cancer. The purpose of this study was to assess the maximal tolerated dose, safety, and immunologic response of a new dendritic cell vaccine (DC-Vac) into which tumor lysate was loaded by electroporation and pulse in patients with advanced non-small cell lung cancer (NSCLC). PATIENTS AND METHODS Fifteen patients with inoperable stage III or IV NSCLC were assigned to cohorts that received 3, 6, or 12 × 10(6) DC-Vac intradermally 3 times at 2 week intervals. We also evaluated immunologic and tumor responses. RESULTS The maximum dose of DC-Vac (12 × 10(6)) was shown to be safe. In 5 of 9 patients, the vaccine resulted in increased interferon (IFN)-γ production by CD8+ cells after exposure to tumor lysate. Additionally, there were mixed responses which do fulfill progressive disease definition but demonstrate some clinical benefit in two patients. CONCLUSION The administration of tumor lysate-loaded autologous dendritic cells by electroporation and pulse was non-toxic and induced immunologic responses to tumor antigens. The two mixed tumor responses which were achieved may represent a potential benefit of this new DC-Vac.

DNA repair gene XRCC1 polymorphisms and haplotypes in diffuse large B-cell lymphoma in a Korean population

DNA repair gene XRCC1 polymorphisms could lead to defective DNA repair and increased risk of lymphoma. This study was performed to evaluate the effect of polymorphisms and haplotypes of the XRCC1 gene on the risk of diffuse large B-cell lymphoma (DLBCL) and treatment outcomes after rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone (R-CHOP) chemotherapy in a Korean population. Carriers of XRCC1 194 variant genotypes had a significantly increased risk of DLBCL [adjusted odds ratio (OR), 1.57; 95% confidence interval (95% CI), 1.06-2.32; P = 0.028] among three polymorphisms of XRCC1 Arg194Trp, Arg280His, and Arg399Gln in 145 patients with DLBCL and in 515 healthy controls. Three polymorphisms of XRCC1 showed very strong linkage disequilibrium (LD) and consisted of one haploblock. The frequency of XRCC1 haplotype A (194Arg-280Arg-399Arg) was significantly lower in DLBCL patients compared to controls (OR, 0.60; 95% CI, 0.15-0.81; P = 0.001). The frequency of XRCC1 haplotype B (194Arg-280Arg-399Gln) was significantly higher in DLBCL patients compared to controls (OR, 1.38; 95% CI, 1.05-1.80; P = 0.019). The association between haplotype A and decreased risk of DLBCL was stable on permutation testing (P = 0.038). However, no relation was noted between these variant genotypes and treatment outcomes in DLBCL patients treated with R-CHOP chemotherapy. These findings suggest that haplotype A of XRCC1 plays a protective role against development of this disease and the haplotype estimation is advantageous for association studies of various cancers showing strong LD.

Activation of NF-κB mediates the PMA-induced differentiation of K562 cells

The role of NF-κB during the PMA-induced megakaryocytic differentiation of K562 cells was investigated using K562 cells transfected with each or both subunits of NF-κB. The NF-κB subunit-transfected cells have shown much higher sensitivity to PMA-induced differentiation than their parental cells. This result was consistent with the findings that PMA-stimulated activities of NF-κB were markedly increased in the NF-κB subunit-transfected cells in comparison with their parental cells and PMA-induced differentiation was enhanced by pretreatment with IκB-α antisense oligonucleotide in the NF-κB subunit-transfected cells. Meanwhile, there were basically no difference in the basal and PMA-stimulated MAP kinase activities among the parental and NF-κB subunit-transfected cells, respectively. However, PMA-induced differentiation was blocked by pretreatment with PD98059, a specific inhibitor of MEK, in both parental and NF-κB-transfected cells. Therefore, these results suggest that during the PMA-induced megakaryocytic differentiation of K562 cells, NF-κB works downstream of MAP kinase, or that activation of both NF-κB and MAP kinase pathways is involved.

Dendritic cells loaded with exogenous antigen by electroporation can enhance MHC class I-mediated antitumor immunity

To develop an efficient antitumor immunotherapy, we have examined if dendritic cells (DCs) loaded with soluble antigens by electroporation present more antigens via the MHC (major histocompatibility complex) class I pathway, which mediate a cytotoxic T-cell response. DCs loaded with ovalbumin (OVA) by electroporation presented more MHC class I–restricted determinants compared with DCs pulsed with OVA. When electroporated DCs were pulsed with OVA for additional times, both MHC class I– and II–restricted presentation of OVA were increased compared with each single procedure, including electroporation or simple pulse. Immunization with DCs loaded with OVA by electroporation induced higher cytotoxicity of splenocytes to E.G7 cells, a clone of EL4 cells transfected with an OVA cDNA, than immunization with DCs pulsed with OVA. In the animal study, immunization with DCs loaded with OVA or tumor cell lysates by electroporation induced an effective antitumor immunity against tumor of E.G7 cells or Lewis lung carcinoma cells, respectively. In addition, immunization with DCs loaded with antigen by combination of electroporation and pulse, completely protected mice from tumor formation, and prolonged survival, in both tumor models. These results demonstrated that electroporation would be a useful way to enhance MHC class I–mediated antitumor immunity without functional deterioration, and that the combination of electroporation and pulse could be a simple and efficient antigen-loading method and consequently lead to induction of strong antitumor immunity.

Meta-Analysis for the Pooled Sensitivity and Specificity of Hepatitis B Surface Antigen Rapid Tests

BACKGROUND Although hepatitis B surface antigen (HBsAg) rapid test based on immunochromatographic assay (ICA) is now widely used, the test has not been evaluated sufficiently enough to validate its performance. Thus, it is important to summarize the clinical performance of the test kits. In this study, we performed meta-analysis for the performance of the HBsAg rapid tests. METHODS PubMed database was searched using keywords about the accuracy of diagnostic tests for hepatitis B virus (HBV) infection. Two investigators assessed methodological quality utilizing standards for reporting of diagnostic accuracy studies (STARD) checklist. After performing a heterogeneity test, we obtained pooled sensitivity and specificity. Positive and negative predictive values (PPV and NPV) were simulated according to HBV prevalence. RESULTS A total of 38 studies was selected from 10 papers. The quality scores ranged from 3 to 13 (median, 8). Kappa value was good (0.85). The performance of the 38 studies was heterogeneous. When 33 studies with better quality from 7 papers were re-selected, the pooled sensitivity and specificity were 98.07% (95% confidence interval, CI: 97.67-98.47%) and 99.56% (95% CI: 99.21-99.91%), respectively. With an HBV prevalence of 5%, PPV and NPV were predicted to be 92.14% and 99.90%, respectively. CONCLUSIONS In view of high HBV prevalence in Korea, it is thought that the HBsAg rapid test can be used for HBV screening in small-sized laboratories or for epidemiologic studies. This study should be helpful in establishing a guideline for the proper performance evaluation of the HBsAg rapid tests.

Nilotinib-induced bone marrow aplasia.

To the Editor: The pathogenetic mechanism in chronic myeloid leukemia (CML) is the presence of BCR-ABL tyrosine kinase, the product of a reciprocal (9; 22) chromosome translocation. Imatinib, and more recently, second generation BCR-ABL tyrosine kinase inhibitors (TKIs), have been developed for the treatment of patients with CML. Nilotinib is a second generation TKI that decreases the proliferation and variability of wild-type BCR-ABL and imatinib-resistant BCR-ABL mutant-expressing cells in vitro by selectively inhibiting BCR-ABL autophosphorylation. In a previous study, grade 3–4 neutropenia and thrombocytopenia was observed in 29% of nilotinib-treated CML patients following imatinib resistance and intolerance (1). Although several authors have reported on imatinib-associated bone marrow aplasia (BMA; 2–4), a case associated with nilotinib has not been reported. Herein, we report one case of nilotinib-induced BMA in an imatinib-intolerant CML patient. A 77-year-old man was admitted to our hospital due to general fatigue, lower abdominal pain and leukocytosis without fever in July 2007. The complete blood count (CBC) showed an abnormally elevated white blood cell count of 12.3 · 10 ⁄L, a hemoglobin (Hb) level of 12.6 g ⁄dL and a platelet (PLT) count of 563 · 10 ⁄L. He had no significant medical history or concomitant diseases. On physical examination, the spleen was palpable 3 cm below the costal margin. A computed tomography (CT) scan confirmed the splenomegaly (14 cm). A bone marrow (BM) biopsy was consistent with Philadelphia chromosome-positive CML in the chronic phase with a 45,-X,Y, t(9;22)(q34;q11.2) karyotype and cellularity of 100% (Fig. 1A). He was initially treated with imatinib at a dose of 400 mg ⁄d. After 3 months of imatinib therapy, he achieved a complete hematologic response. However, he developed a grade 1 skin rash and pruritus. After 9 months of therapy, a follow-up BM biopsy showed >50% cellularity and major cytogenetic remission (BCR-ABL by FISH on BM, positivity = 18%) was achieved. However, intolerance symptoms persisted despite a dose reduction to 300 mg and subsequent symptomatic treatment. In September 2008, the drug was discontinued and nilotinib (400 mg twice daily) was started. After 2 months of nilotinib therapy, he developed massive hematochezia with pancytopenia without fever. At that time, the CBC showed a leukocyte count of 0.3 · 10 ⁄L, a neutrophil count of 0.05 · 10 ⁄L, a Hb levelof 4.7 g ⁄dL and a PLT count of 3 · 10 ⁄L. Nilotinib was discontinued, and red blood cells and PLTs were transfused. A BM biopsy was performed because of persistent pancytopenia, and was diagnostic of BMA with <5% of normal cellularity and fatty tissue without evidence of myelofibrosis (MF) or adipocyte deposition, and the presence of an eosinophilic extracellular matrix (ECM; Fig. 1B). Cytogenetic analysis did not evidence additional chromosome abnormalities (46, XY). Serologic testing for human immunodificiency virus, hepatitis A, hepatits B, hepatitis C and parvovirus (IgM) were negative. As the patient had received only nilotinib without any side effects, it was considered that the cause of pancytopenia was nilotinib. Four months after discontinuing nilotinib, the patient remained in complete cytogenetic remission without