Recognition of a Caucasoid HLA-B locus allele, B*44:55, in a Taiwanese/Chinese bone marrow stem cell donor.

We detected a Caucasoid HLA-B allele, HLA-B*44:55, in a potential Taiwanese/Chinese bone marrow hematopoietic stem cell donor during our routine HLA SBT (sequence-based typing) practice. The sequence of B*44:55 varies with B*44:02:01:01 with one nucleotide in exon 2 at position 97 (T->C), while it differs from B*44:03:01 with one nucleotide in exon 2 at position 97 (T->C) and three nucleotides in exon 3 at residues 538-540 (CTG->GAC). The nucleotide replacements caused one amino acid variation with B*44:02:01:01 at residue 9 (Y->H) and two amino acid variations with B*44:03:01 at residue 9 (Y->H) and residue 156 (L->D). The formation of B*44:55 is probably the result of a nucleotide substitution involving B*44:02:01:01 at position 97 (T->C). The Taiwanese/Chinese donor with B*44:55 claims having no kinship with Caucasian. Our speculations on the origin of the Taiwanese/Chinese B*44:55 will be presented.

Discovery of the novel HLA-DRB1*09:01:08 allele in a Taiwanese volunteer bone marrow donor and identification of the probable HLA-A, HLA-B and HLA-DRB1 haplotype in association with DRB1*09:01:08.

We report here the novel variant of HLA-DRB1*09:01, DRB1*09:01:08, discovered in a Taiwanese volunteer bone marrow donor by a sequence-based typing (SBT) method. The DNA sequence of DRB1*09:01:08 is identical to the sequence of DRB1*09:01:02 in exon 2 except a silent mutation at nucleotide position 261(C→T) (GCC→GCT at codon 58). We hypothesize DRB1*09:01:08 was probably derived from DRB1*09:01:02 via a nucleotide point mutation event. The plausible HLA-A, HLA-B and HLA-DRB1 haplotype in association with DRB1*09:01:08 was deduced as A*02:07-B*46:01-DRB1*09:01:08.

Discovery of HLA-DRB1*03:20 allele in a Taiwanese volunteer hematopoietic stem cell donor and the probable HLA-A, -B, -C and -DRB1 haplotype in association with DRB1*03:20.

The allele HLA-DRB1*03:20, a variant of DRB1*03, was first reported to the IMGT HLA database in April 2001 without indication on the ethnicity of the blood donor (Cell ID: HC 125775). We found a Taiwanese volunteer hematopoietic stem cell donor carries DRB1*03:20 by a sequence-based typing (SBT) method. The DNA sequence of DRB1*03:20 is identical to the sequence of DRB1*03:01:01 in exon 2, except a nucleotide substitution at position 341(T→C) (GTT→GCT at codon 85). The nucleotide replacement produced an amino acid variation at residue 85 (V→A). We hypothesize that DRB1*03:20 was probably derived from DRB1*03:01:01 via a nucleotide point mutation event. The probable HLA haplotype in association with DRB1*03:20 was deduced as A*11:02-B*58:01-C*07:02-DRB1*03:20. We here report the Taiwanese/Chinese ethnicity of DRB1*03:20.

A novel HLA-B allele, B*58:01:12, detected in a Taiwanese volunteer bone marrow stem cell donor using sequence-based typing method.

Human leukocyte antigen-B*58:01:12, a novel rare allele of HLA-B*58:01 variant, was found in a Taiwanese volunteer bone marrow donor by SBT (sequence-based typing) method. The DNA sequence of B*58:01:12 is identical to the sequence of B*58:01:01 in exons 2, 3 and 4 except at nucleotide position 483 where nucleotide C is substituted by T (at codon 137; GAC GAT). Due to the silent point mutation, the amino acid sequence of B*58:01:12 is identical to the sequence of B*58:01:01. The HLA haplotype in association with B*58:01:12 may be deduced as A*33:03-B*58:01:12-DRB1*03:01. The discovery of B*58:01:12 adds further polymorphism of B*58:01 in Taiwanese population.

Discovery of the novel HLA-DRB1*04:05:14 allele in a Taiwanese unrelated haematopoietic stem cell donor by a sequence-based typing method.

We report here a de novo HLA-DRB1*04 allele, DRB1*04:05:14, discovered in a Taiwanese unrelated volunteer bone marrow stem cell donor by a sequence-based typing method. In exon 2, the DNA sequence of DRB1*04:05:14 is identical to the sequence of DRB1*04:05:01 except the nucleotide at positions 321 where C is replaced by T (at codon 78; TAC→TAT). Due to the silent mutation, the nucleotide substitution produced no amino acid variation in comparison with DRB1*04:05:01. We assume DRB1*04:05:14 was derived from DRB1*04:05:01 via a point mutation. The probable HLA-A, -B and -DRB1 haplotype in association with DRB1*04:05:14 may be deduced as A*11-B*55-DRB1*04:05:14. We here report the Taiwanese ethnicity of DRB1*04:05:14.

Discovery of the novel HLA-DRB1*10:04 allele in a Taiwanese volunteer bone marrow donor and identification of the probable HLA-A, -B, -C and -DRB1 haplotype in association with DRB1*10:04.

We report here a novel variant of HLA-DRB1*10, DRB1*10:04, discovered in a Taiwanese volunteer bone marrow donor by a sequence-based typing (SBT) method. The DNA sequence of DRB1*10:04 differs from DRB1*10:01:01, in exon 2, at nucleotide positions 296 (G fi A) and 303 (T fi G). The nucleotide changes caused an amino acid substitution at amino acid residue 70 (R fi Q). We hypothesize that the formation of DRB1*10:04 was probably the result of a gene recombination event where DRB1*10:01:01 received a minimum length of DNA sequence from DRB1*04:05:01, as the sequence of DRB1*10:04 is identical to DRB1*10:01:01 in exon 2 except the sequence from nucleotide 296 to nucleotide 303, which is identical to DRB1*04:05:01. The plausible HLA-A, -B, -C and - DRB1 haplotypes in association with DRB1*10:04 was deduced as A*01:01-B*37:01-C*06:02-DRB1*10:04.

Detection of the rare HLA-B*40:97 allele in an unrelated Taiwanese bone marrow donor.

We detected a rare HLA-B locus allele, B*40:97, in a Taiwanese unrelated donor in our routine HLA SBT (sequence-based typing) exercise for a possible hematopoietic stem cell donation. In exons 2, 3 and 4, the sequence of B*40:97 is identical to the sequence of B*40:02:01 except one nucleotide at nucleotide position 760 (C->T) in exon 4. The nucleotide variation caused one amino acid alteration at residue 230 (L->F). B*40:97 was probably derived from a nucleotide substitution event where C was replaced by T at nucleotide 760 involving B*40:02:01. The HLA-A, HLA-B, HLA-C, HLA-DRB1 and HLA-DQB1 haplotype in association with B*40:97 may be deduced as A*26:01-B*40:97-C*03:03-DRB1*11:01-DQB1*03:03. Our recognition of B*40:97 in Taiwanese helps to fill the void of ethnic information for the allele B*40:97 reported to the IMGT/HLA Database.

Recognition of HLA-A*24:137 allele in a Taiwanese unrelated bone marrow stem cell donor and the plausible HLA haplotype associated with A*24:137.

We detected a rare HLA-A*24:137 allele in an unrelated Taiwanese haematopoietic stem cell donor during a routine SBT (sequence-based typing) HLA typing exercise. The DNA sequence of A*24:137 is identical to the sequence of A*24:02:01:01 in exons 2 and 3 except at codon 21 where CGC was replaced with CAA. The DNA variation caused an amino acid alteration at amino acid residue 21 (R->Q). The HLA haplotype in association with A*24:137 may be deduced as A*24:137-B*15-DRB1*14. The formation of A*24:137 was probably the result of a nucleotide point mutation involving A*24:02:01:01. It remains to be determined whether A*24:137 is restricted to Taiwanese/Chinese ethnicity.

Discovery of the novel HLA-DRB1*03:77 allele in a Taiwanese unrelated hematopoietic stem cell donor by a sequence-based typing method and identification of the probable HLA haplotype in association with DRB1*03:77.

Here, we report a novel human leucocyte antigen (HLA)-DRB1 allele, DRB1*03:77, discovered in a Taiwanese unrelated volunteer hematopoietic stem cell donor by a sequence-based typing (SBT) method. The DNA sequence of DRB1*03:77 is identical to the DNA sequence of DRB1*03:01:01 in exon 2 except one nucleotide at position 223 (G→C). The nucleotide substitution caused an amino acid replacement at residue 46 (E→Q). The formation of DRB1*03:77 was thought as the result of a nucleotide point mutation. The probable HLA-A, HLA-B and HLA-DRB1 haplotype in association with DRB1*03:77 may be deduced as A*33-B*58-DRB1*03:77. The donor was a Minna Taiwanese whose ancestors came from mainland China.

Discovery of the novel HLA-DRB1*16:16 allele in a Taiwanese unrelated bone marrow stem cell donor by a sequence-based typing method and the probable haplotype associated with DRB1*16:16.

We report here a de novo HLA-DRB1 allele, DRB1*16:16, discovered in a Taiwanese unrelated volunteer bone marrow stem cell donor by a sequence-based typing (SBT) method. In exon 2, the DNA sequence of DRB1*16:16 is identical to the sequence of DRB1*16:02:01 except the nucleotides at positions 258 (C→T), 260 (C→A) and 261 (T→G). The nucleotide substitution produced an amino acid replacement at residue 58 (A→E). The formation of DRB1*16:16 was probably generated by a DNA sequence recombination event involving DRB1*11:01:01 and DRB1*16:02:01. The probable HLA-A, -B and -DRB1 haplotype in association with DRB1*16:16 may be deduced as A*02-B*38-DRB1*16:16.

Identification of two novel HLA-A*02 variants, A*02:319 and A*02:01:64, in two Taiwanese marrow stem cell donors by sequence-based typing.

We report here two novel variants of HLA-A*02 allele, A*02:319 and A*02:01:64, discovered in two Taiwanese unrelated volunteer bone marrow donors by sequence-based typing (SBT) method. The DNA sequence of A*02:319 is identical to A*02:07 in exons 2 and 3 but varies with one nucleotide at codon 9 (TTC->TCC). The variation caused one amino acid substitution at residue 9 (F->S). On the other hand, the DNA sequence of A*02:01:64 is identical to the sequence of A*02:01:01:01 in exons 2 and 3 except a silent mutation at codon 114 (CAC->CAT). The probable HLA-A, HLA-B and HLA-DRB1 haplotypes in association with A*02:319 and A*02:01:64 were deduced as A*02:319-B*46:01-DRB1*04 and A*02:01:64-B*38:02-DRB1*16:02, respectively.

Overexpressed hPTTG1 promotes breast cancer cell invasion and metastasis by regulating GEF-H1/RhoA signalling.

Human pituitary tumour-transforming gene 1 (hPTTG1) is an oncogenic transcription factor that is overexpressed in many tumour types, especially tumours with metastatic abilities. However, how hPTTG1 overexpression drives metastasis is not yet clear. As a transcription factor, hPTTG1 may promote metastasis by activating target genes that are involved in the metastatic process. Here, we showed that Rho guanine nucleotide exchange factor-H1 (GEF-H1) was transcriptionally activated by hPTTG1, thereby promoting breast cancer metastasis. Luciferase reporter analyses and chromatin immunoprecipitation (ChIP) assays showed that hPTTG1 directly bound and activated the GEF-H1 gene promoter. In this study, RNA interference-mediated knockdown of hPTTG1 in highly metastatic breast tumour cells decreased GEF-H1 expression and RhoA activation, thereby reducing cell motility and invasion, and interfering with cytoskeletal remodelling in vitro, and impairing the tumour metastasis in vivo. The restoration of GEF-H1 expression in hPTTG1-knockdown cells rescued the hPTTG1-knockdown effects on cytoskeletal changes in vitro and tumour metastasis in vivo. Conversely, ectopic expression of hPTTG1 in non-metastatic breast tumour cells induced cytoskeletal rearrangements, and allowed these cells to metastasise in a mouse model by orthotopic implantation. In human tumour samples, hPTTG1 expression was also correlated to GEF-H1 expression in aggressive breast carcinoma. Altogether, these findings definitively establish a role for hPTTG1 in activating the GEF-H1/RhoA pathway as a newly identified mechanism in breast cancer metastasis.

Translocation between chromosome 5q35 and chromosome 11q13-- an unusual cytogenetic finding in a primary refractory acute myeloid leukemia.

Cytogenetic abnormalities are observed in approximately two-thirds of patients with acute myeloid leukemia (AML). Chromosome rearrangements are associated with specific subtypes of AML and associated prognosis. We report a patient with AML, M2, who was primarily refractory to standard induction chemotherapy with idarubicin and cytarabine. Flow cytometry of a bone marrow aspirate showed aberrant expression of B-cell markers including CD19. Cytogenetic studies disclosed a translocation between 5q35 and 11q13. Fluorescence in situ hybridization analyses demonstrated that neither the NSD1 nor MLL genes were involved in this case. Further study is required to define conclusively the genes involved and their contribution to pathogenesis in this case.

BAG-i expression in human breast cancer: interrelationship between BAG-1 RNA, protein, HSC70 expression and clinico-pathological data.

BAG-1 (BCL-2 athanogene-1), a multifunctional protein which associates with steroid hormone receptors (including the oestrogen receptor) and the anti-apoptotic BCL-2 protein, regulates steroid hormone-dependent transcription and apoptosis. Direct interaction with 70 kD heat-shock proteins, HSC70 and HSP70, may mediate the diverse functions of BAG-1. Immunohistochemistry was used to examine the expression of BAG-1 and HSC70 in 160 cases of invasive breast cancer. BAG-1 was expressed in 92% of cases; most tumours exhibited cytoplasmic BAG-1, while a smaller proportion also had nuclear immunostaining. There was a significant inverse correlation between histological grade and nuclear BAG-1 expression, with higher-grade tumours tending to have reduced nuclear BAG-1 expression, but there was no association with cytoplasmic BAG-1. There was also no significant correlation between nuclear or cytoplasmic BAG-1 expression and oestrogen receptor positivity. Since BAG-1 may be influenced by hormonal background, the relationship between grade and oestrogen receptor was examined separately in pre-menopausal and post-menopausal women. The statistically significant correlation between nuclear BAG-1 expression and low tumour grade was strong in pre-menopausal, but not apparent in postmenopausal women. A statistically significant correlation was observed between cytoplasmic, but not nuclear, BAG-1 expression and oestrogen receptor status in pre-menopausal, but not postmenopausal, women. There was no correlation between BAG-1 protein expression and RNA, suggesting that important post-transcriptional mechanisms control BAG-1 expression in vivo. HSC70 was also detected in the majority (97%) of cases, although expression was not correlated with BAG-1 levels, oestrogen receptor status or tumour grade. Overall survival in cases with high levels of nuclear BAG-1 expression was improved, though not significantly. These results are consistent with the hypothesis that BAG-1 plays an important but variable role in breast cancers developing in pre-menopausal and post-menopausal women.

Identification of changes in gene expression associated with minimal residual disease.

Minimal residual disease (MRD), the tumour burden which remains after a course of treatment that has resulted in clinical remission [1], appears to differ in certain characteristics from the primary tumour population. Certainly the cells which comprise MRD have had to escape from the constraints of the primary tumour mass, invade normal tissue and penetrate small vessels in order to enter the circulation in which they then have had to survive. Such activities are the consequence of the expression of specific proteins and these may well be a reflection of alterations in DNA or RNA levels. Identifying the changes in RNA expression levels between related cell groups exhibiting different phenotypes recently has become a great deal easier as a consequence of developments in analytical procedures such as Differential Display (DD) and Serial Analysis of Gene Expression (SAGE). Application of these procedures to MRD cells recovered from blood, bone marrow or lymph node, should identify novel sequences associated with tumour progression and the development of disseminated disease.

Myocardial tumor necrosis factor-alpha secretion in hypertensive and heart failure-prone rats.

Acute increases in blood pressure (BP) increase myocardial tumor necrosis factor (TNF)-alpha production, but it is not known whether chronic hypertensive stress elevates myocardial TNF-alpha production, possibly contributing to cardiac remodeling, decreased cardiac function, and faster progression to heart failure. BP, cardiac function, and size were evaluated in normotensive [Sprague-Dawley (SD)], spontaneously hypertensive (SHR), and spontaneously hypertensive heart failure-prone (SHHF) rats at 6, 12, 15, and 18 mo of age and in failing SHHF. Left ventricular tissues were evaluated for secretion of bioactive TNF-alpha and inhibition of TNF-alpha secretion by phosphodiesterase inhibitors. All ventricles secreted bioactive and immunoreactive TNF-alpha, but secretion decreased with age. SHR and SHHF rats secreted more TNF-alpha than SD rats at 6 mo of age, but only failing SHHF rats secreted significantly more TNF-alpha at 18 mo. Amrinone inhibited TNF-alpha secretion in all rats and was less potent but more efficacious than RO-201724 in all strains. TNF-alpha secretion correlated with BP and left ventricular mass in 6-mo-old rats, but this relationship disappeared with age. Results suggest that hypertension and/or cardiac remodeling is associated with elevated myocardial TNF-alpha, and, although hypertension, per se, did not maintain elevated cardiac TNF-alpha levels, SHHF rats increase TNF-alpha production during the end stages of failure.

Age-dependent augmentation of cardiac endothelial NOS in a genetic rat model of heart failure.

Alterations in nitric oxide (NO) biosynthesis in the heart have been implicated in the pathophysiology of heart failure. We compared changes in cardiac nitric oxide synthase (NOS) activity and expression in genetically heart failure-prone (SHHF) rats at 6, 12, and 18 mo of age with those in age-matched spontaneously hypertensive (SHR) and Sprague-Dawley (SD) rats. Systolic blood pressure was significantly higher in SHHF and SHR rats compared with SD rats; however, it declined with age in SHHF rats only. Left ventricular mass increased with age in SHR and SHHF, but not in SD rats. Plasma nitrate and nitrite level was elevated in SHHF and SHR rats at 18 mo only. In left ventricular homogenates from SHHF rats, Ca(2+)-dependent NOS activity increased markedly with age and was accompanied by enhanced expression of endothelial NOS (eNOS). In contrast, SHR rats showed a much smaller increase in Ca(2+)-dependent NOS activity over time without changes in eNOS expression; neither parameter was altered with age in SD rats. Ca(2+)-independent NOS activity was not detected in any heart. This is the first report of a unique alteration in myocardial NOS activity in hypertensive rats genetically prone to heart failure.

Characteristics of malignant lymphoma in eastern Taiwan: comparison between aborigines and nonaborigines.

To study the characteristics of malignant lymphoma in aboriginal and nonaboriginal patients in eastern Taiwan, the records of 90 patients treated from July 1986 to September 1994 were reviewed. Immunohistologic staining with UCHL-1 and L-26 was used to determine immunophenotype. There were 20 aboriginal and 70 nonaboriginal patients (22% vs 78%). Among them, three had Hodgkin's diseases (HD) and 87 had non-Hodgkin's lymphomas (NHL). Comparing the histology of malignant lymphoma in aboriginal and nonaboriginal patients, there was no case of HD or follicular lymphoma in aborigines. Concerning primary extranodal lymphoma, no aboriginal patients were found to have gastric lymphoma, while eight nonaboriginal patients did. Among patients with intermediate and high-grade NHL, aboriginal patients had a higher rate of B symptoms (weight loss, fever, night sweats) than did nonaboriginal patients. In T cell lymphoma, three out of four (75%) aboriginal patients had angioimmunoblastic lymphadenopathy with dysproteinemia as compared with three out of 20 (15%) nonaboriginal patients. There were no significant differences in demographic data, stage or distribution of immunophenotype between the two groups. Several combination chemotherapy regimens were used and only 31 patients were considered evaluable, of which 12 were aboriginal and 19 were nonaboriginal patients. No distinct difference was found between aboriginal and nonaboriginal patients in overall response rate, complete response rate, disease-free survival or overall survival. Comparison of our results with other lymphoma studies in Taiwan revealed that the frequencies of HD and follicular lymphoma were higher in northern Taiwan studies than in southern and eastern Taiwan studies, and the frequency of primary extranodal lymphoma was much higher in eastern Taiwan than in other areas. While the rate of T cell lymphoma in NHL for nonaboriginal patients was similar to that of northern Taiwan studies, it was closer to that of southern Taiwan studies in aboriginal patients.

Clinical characteristics of and response to combination chemotherapy and subsequent application of international prognostic index in non-Hodgkin's lymphoma--an experience from a medical center in Southern Taiwan.

A retrospective analysis was performed of 117 non-Hodgkin's lymphoma (NHL) patients (72 male and 45 female, mean age 55 years) treated at NCKUH between July 1988 and December 1993. Of the 115 patients who could be classified by Ann Arbor staging system, 26 patients (22.2%) were in stage 1; 23 (19.7%) in stage 2; 29 (24.8%) in stage 3; and 37 (31.6%) in stage 4. According to the International Working Formulation, three patients (2.6%) were low grade NHL, 90 (76.9%) were intermediate, and 8 (6.8%) were high grade NHL. Histologically, diffuse large cell NHL accounted for 52.1% of cases, followed by 16.2% of cases exhibiting diffuse mixed NHL. Immunophenotype analysis was available in 95 cases, which revealed 76 (80%) cases exhibiting B-cell origin, 17 (18%) cases exhibiting diffuse mixed NHL. Immunophenotype analysis was available in 95 cases, which revealed 76 (80%) cases exhibiting B-cell origin, 17 (18%) cases exhibiting T-cell origin and 2 (2%) cases were of null cell type. All patients underwent two groups of induction chemotherapy, either CHOP (Cyclophosphamide, Epirubicin, Oncovin, and Prednisolone), or "modified" COPBLAM (Cyclophosphamide, Epirubicin, Oncovin, and Prednisolone), or "modified" COPBLAM (Cyclophosphamide, Epirubicin, Oncovin, Vinblastine, Bleomycin, Procarbazine, and Prednisolone). Seventy-two cases treated through COPBLAM and 45 cases treated through CHOP were evaluated. The response rate (RR) to COPBLAM treatment was 72.2% and was 68.9% for the CHOP group (P = 0.51). The 5-year overall survival rate (OAS) was 44.1% for COPBLAM, versus 40% for CHOP (P = 0.15). The disease-free survival (DFS) was 72.6% at 63 months for COPBLAM and 58% at 51 months for CHOP (P = 0.16). Neither B cell nor T-cell lineages of NHL showed any statistical difference in RR (P = 0.53, DFS (P = 0.58) or OAS (P = 0.97) to the different treatments. Using multiple logistic analysis, two independent factors, high LDH and advanced stage, were found to adversely affect the rate of complete remission. The application of the International Prognostic Index to our patients needs modification, which suggests the necessity of more evaluation before it can accurately be applied to all international series of NHL.

Chemotherapy with cisplatin and continuous infusion of 5-fluorouracil and bleomycin for recurrent and metastatic nasopharyngeal carcinoma in Taiwan.

Twenty-five patients with metastatic and/or recurrent nasopharyngeal carcinoma were treated with cisplatin 20 mg/m2/day on days 1-5 i.v. with hydration; 5-fluorouracil (5-FU) 1,000 mg/m2/day by continuous infusion (CI); and bleomycin 15 mg/m2 on day 1 also by CI. These cycles were repeated every 4 weeks. Twenty-three (92%) had distant metastases. Bone was the most frequently involved site (72%), followed by lungs (44%) and liver (40%). More than half the patients (14/25) presented with at least 3 organ sites involved or had local T3/T4 or N3 lesions with a distant metastasis. The median time from relapse to start of chemotherapy was 7.5 months. We observed 1 (4%) complete response (CR), and 9 (36%) partial responses (PR). The objective response rate (CR + PR) was 40%. Hematologic toxicities were frequently encountered. Twenty (80%) patients experienced leukopenia during the treatment courses and 9 (36%) had severe (grade 3 or 4) leukopenia. Eight patients had grade 3 or 4 infections. Two of them died of sepsis and 1 succumbed to uncontrolled pneumonia. The objective response rate was inferior to other series. Possible explanation included longer delay before initiation of definitive treatment, larger tumor burdens, higher severe hematologic toxicity and lower dosage of bleomycin. The results suggested metastatic and/or recurrent nasopharyngeal carcinoma is chemosensitive, however, for patients with large tumor burdens, more intensive chemotherapy regimens with support of hematopoietic growth factors may be required to achieve a better control.