bluuebunny/crossref_metadata_2025 huggingface dataset:
Primary Acute Lymphoblastic Leukemia Cells Use a Novel Promoter and 5′Noncoding Exon for the Human Reduced Folate Carrier That Encodes a Modified Carrier Translated from an Upstream Translational Start
Authors: Flatley Robin M. , Payton Scott G. , Taub Jeffrey W. , Matherly Larry H.
Published: September 2005

The human reduced folate carrier (hRFC) is reported to be regulated by up to seven alternatively spliced noncoding exons (A1, A2, A, B, C, D, and E). Noncoding exon and promoter usage was analyzed in RNAs from 27 childhood acute lymphoblastic leukemia (ALL) specimens by real-time PCR and/or 5′ rapid amplification of cDNA ends (5′ RACE) assay. By real-time PCR, total hRFC transcripts in ALL spanned a 289-fold range. Over 90% of hRFC transcripts were transcribed with A1, A2, and B 5′ untranslated regions (UTRs). Analysis of 5′ RACE clones showed that the A1 + A2 5′UTRs contained A1 sequence alone or a fusion of A1 and A2, implying the existence of a single, alternatively spliced 1021-bp A1/A2 noncoding region. High frequency sequence polymorphisms (AGG deletion, C/T transition) identified in the A1/A2 region by 5′RACE were confirmed in normal DNAs. By reporter assays in HepG2 hepatoma and Jurkat leukemia cells, A1/A2 promoter activity was localized to a 134-bp minimal region. Translation from an upstream AUG in the A1/A2 noncoding region in-frame with the normal translation start resulted in synthesis of a larger (∼7 kDa) hRFC protein with transport properties altered from those for wild-type hRFC. Although there was no effect on transcript or protein stabilities, in vitro translation from A1/A2 transcripts was decreased compared with those with the B 5′UTR. Our results document the importance of the hRFC A1/A2 upstream region in childhood ALL and an intricate transcriptional and posttranscriptional regulation of hRFC-A1/A2 mRNAs. Furthermore, they suggest that use of the A1/A2 5′UTR may confer a transport phenotype distinct from the other 5′UTRs due to altered translation efficiency and transport properties.
Prognostic Analysis of Early Lymphocyte Recovery in Patients with Advanced Breast Cancer Receiving High-Dose Chemotherapy with an Autologous Hematopoietic Progenitor Cell Transplant
Authors: Nieto Yago , Shpall Elizabeth J. , McNiece Ian K. , Nawaz Samia , Beaudet Julie , Rosinski Steve , Pellom Julie , Slat-Vasquez Victoria , McSweeney Peter A. , Bearman Scott I. , Murphy James , Jones Roy B.
Published: September 2005

Purpose: The purpose of this study was to evaluate the prognostic effect of early posttransplant lymphocyte recovery in patients with advanced breast cancer receiving high-dose chemotherapy with autologous hematopoietic progenitor cell transplantation.

Experimental Design: We analyzed the effect of the absolute lymphocyte count on day +15 posttransplant on freedom from relapse and overall survival in patients with high-risk primary breast cancer or metastatic breast cancer, enrolled between 1990 and 2001 in prospective high-dose chemotherapy trials, using a uniform regimen of cyclophosphamide, cisplatin, and 1,3-bis(2-chloroethyl)-1-nitrosourea.

Results: Four hundred and seventy-six patients (264 high-risk primary breast cancer and 212 metastatic breast cancer patients) were evaluated at median follow-up of 8 years (range, 1.5–11 years). The disease-free survival and overall survival rates in the high-risk primary breast cancer group were 67% and 70%, respectively. Patients with metastatic breast cancer patients had 21.8% disease-free survival and 31.5% overall survival rates. Day +15 absolute lymphocyte count correlated with freedom from relapse (P = 0.007) and overall survival (P = 0.04) in the metastatic breast cancer group, but not in the high-risk primary breast cancer group (P = 0.5 and 0.8, respectively). The prognostic effect of absolute lymphocyte count in metastatic breast cancer was restricted to those patients receiving unmanipulated peripheral blood progenitor cells (P = 0.04). In contrast, absolute lymphocyte count had no significant effect in those metastatic breast cancer patients receiving bone marrow or a CD34-selected product. In multivariate analyses, the prognostic effect of day +15 absolute lymphocyte count in metastatic breast cancer was independent of other predictors, such as disease status, pre-high-dose chemotherapy treatment, number of tumor sites, or HER2.

Conclusions: Early lymphocyte recovery is an independent outcome predictor in metastatic breast cancer patients receiving high-dose chemotherapy and an autologous peripheral blood progenitor cell transplant. These observations suggest that immune strategies targeting minimal posttransplant residual disease may prove worthwhile.
Antitumor Activity of Systemically Delivered Ribozymes Targeting Murine Telomerase RNA
Authors: Nosrati Mehdi , Li Shang , Bagheri Sepideh , Ginzinger David , Blackburn Elizabeth H. , Debs Robert J. , Kashani-Sabet Mohammed
Published: September 2005

Purpose: To test ribozymes targeting mouse telomerase RNA (mTER) for suppression of the progression of B16-F10 murine melanoma metastases in vivo.

Experimental Design: Hammerhead ribozymes were designed to target mTER. The ribozyme sequences were cloned into a plasmid expression vector containing EBV genomic elements that substantially prolong expression of genes delivered in vivo. The activity of various antitelomerase ribozymes or control constructs was examined after i.v. injection of cationic liposome:DNA complexes containing control or ribozyme constructs. Expression of ribozymes and mTER at various time points were evaluated by quantitative real-time PCR. Telomerase activity was examined using the telomeric repeat amplification protocol.

Results: Systemic administration of cationic liposome:DNA complexes containing a plasmid-expressed ribozyme specifically targeting a cleavage site at mTER nucleotide 180 significantly reduced the metastatic progression of B16-F10 murine melanoma. The antitumor activity of the anti-TER 180 ribozyme in mice was abolished by a single inactivating base mutation in the ribozyme catalytic core. The EBV-based expression plasmid produced sustained levels of ribozyme expression for the full duration of the antitumor studies. In addition to antitumor activity, cationic liposome:DNA complex-based ribozyme treatment also produced reductions in both TER levels and telomerase enzymatic activity in tumor-bearing mice.

Conclusions: Systemic, plasmid-based ribozymes specifically targeting TER can reduce both telomerase activity and metastatic progression in tumor-bearing hosts. The work reported here demonstrates the potential utility of plasmid-based anti-TER ribozymes in the therapy of melanoma metastasis.
Elevated S-Phase Kinase-Associated Protein 2 Protein Expression in Acute Myelogenous Leukemia
Authors: Min Yoo Hong , Cheong June-Won , Lee Mark Hong , Kim Ji Yeon , Lee Seung Tae , Hahn Jee Sook , Ko Yun Woong
Published: September 2005

Purpose: The F-box protein S-phase kinase-associated protein 2 (Skp2) positively regulates the G1-S phase transition by controlling the stability of several G1 regulators, such as p27Kip1. However, the clinical significance of Skp2 in patients with acute myelogenous leukemia (AML) remains unknown.

Experimental Design: We examined the clinical and biological significance of Skp2 expression in AML and evaluated the relationship between Skp2 and p27Kip1 expression and phosphatase and tensin homologue (PTEN) phosphorylation.

Results: Western blot analysis showed that high Skp2 expression was observed in 57 (57.6%) cases and significantly correlated with unfavorable cytogenetics (P = 0.035) but not with age, white blood cell count, serum lactic dehydrogenase level, and the French-American-British subtype. An inverse correlation was not observed between Skp2 and p27Kip1 expression. However, p27Kip1 protein was preferentially localized to cytoplasm in the high-Skp2-expression group. The cytoplasmic to nuclear ratio of p27Kip1 expression was significantly correlated with the levels of Skp2 expression (P < 0.001). The frequency of PTEN phosphorylation was significantly higher in the high-Skp2-expression group compared with the low- Skp2-expression group (P = 0.035). The Skp2 overexpression was significantly associated with shorter disease-free survival and overall survival (P = 0.0386 and P = 0.0369, respectively). Multivariate analysis showed that Skp2 expression was an independent prognostic factor both in the disease-free survival and overall survival.

Conclusion: These findings suggest that Skp2 expression is an independent marker for a poor prognosis in AML. The presence of a positive correlation between Skp2 and phosphorylated PTEN suggests that an aberration in the PTEN/Skp2 signaling pathway might be operating in AML.
Cytochrome P450 and Glutathione Transferase Expression in Squamous Cell Cancer
Authors: Ali Shadan , El-Rayes Basil F. , Heilbrun Lance K. , Sarkar Fazlul H. , Ensley John F. , Kucuk Omar , Philip Philip A.
Published: September 2005

Purpose: The cytochrome P-450 (CYP) and glutathione S-transferase (GST) enzyme systems modulate the carcinogenic effects of tobacco. Therefore, the expression of these enzymes may be in part responsible for the observed interindividual and inter-racial differences in the risk of development of squamous cell carcinoma of the head and neck (SCCHN). The first aim of this study was to evaluate the feasibility of measuring the expression of the CYP and GST in target tissue from the head and neck. The second aim was to compare the expression of CYPs 1A1, 2E1, and 3A4 in squamous epithelium from African-American and Caucasian pediatric patients. The third aim was to compare the expression of CYPs 1A1, 2E1, 3A4, and GST-π on the p16 expression in patients with SCCHN.

Experimental Design: The expression of CYP 1A1, 2E1, 3A4, GST-π, and p16 was quantified by immunoblotting. Expression of CYPs 1A1, 2E1, and 3A4 was quantified in tissue from 160 pediatric patients undergoing tonsillectomy. Expression of CYPs 1A1, 2E1, 3A4, GST-π, and p16 was determined in 46 resected SCCHN patients.

Results: Large interindividual variability in the expression of these enzymes was observed in the pediatric and adult populations. No significant difference was observed in CYP 1A1, 2E1, and 3A4 expression of Caucasian and African-American patients. There was no correlation between p16 and enzyme expression in patients with SCCHN.

Conclusion: Evaluation of CYP expression in the target tissue of interest is feasible. The clinical significance of CYPs and GST-π alterations in the risk of developing SCCHN will need to be investigated in larger trials.
Phase II Study of the Antiangiogenic Agent SU5416 in Patients with Advanced Soft Tissue Sarcomas
Authors: Heymach John V. , Desai Jayesh , Manola Judith , Davis Darren W. , McConkey David J. , Harmon David , Ryan David P. , Goss Geraldine , Quigley Travis , Van den Abbeele Annick D. , Silverman Stuart G. , Connors Susan , Folkman Judah , Fletcher Christopher D. M. , Demetri George D.
Published: September 2005

Purpose: SU5416 (semaxanib) is a small molecule inhibitor of the vascular endothelial growth factor (VEGF) receptor-2 and KIT receptor tyrosine kinases. This Phase II study was conducted to investigate the safety and efficacy of SU5416 for patients with soft tissue sarcomas.

Experimental Design: Thirteen patients with locally advanced or metastatic soft tissue sarcomas were treated with SU5416 via intravenous infusion at a dose of 145 mg/m2 twice weekly. In selected cases tumor biopsies were taken before and after 2 months of treatment.

Results: The median progression-free survival was 1.8 months. Median overall survival was 22.8 months. No objective tumor responses were observed. There was evidence of shorter survival among patients with high baseline urine VEGF levels (P = 0.04). No grade 4 toxicities were observed. The most common grade 3 toxicities were headache and thrombosis. Other less serious toxicities included fatigue, nausea, and abdominal pain. The median systolic blood pressure increased from 118 mmHg at baseline to 133 after 1 month of treatment (P = 0.01). Post-treatment tumor biopsies showed no significant decreases in VEGF receptor phosphorylation compared with baseline in 3 evaluable patients. One patient with gastrointestinal stromal tumor who had rapid progression during SU5416 treatment was subsequently treated with another KIT inhibitor, imatinib mesylate, and had a partial response lasting >36 months.

Conclusions: SU5416 was relatively well tolerated but did not demonstrate significant antitumor activity against advanced soft tissue sarcoma. Correlative studies suggest that VEGF receptor or KIT inhibition was incomplete in at least some cases, providing a possible explanation for the observed lack of activity.
Spontaneous Tumor-Specific Humoral and Cellular Immune Responses to NY-ESO-1 in Hepatocellular Carcinoma
Authors: Korangy Firouzeh , Ormandy Lars A. , Bleck Jörg S. , Klempnauer Jürgen , Wilkens Ludwig , Manns Michael P. , Greten Tim F.
Published: September 2005

Purpose: Hepatocellular carcinoma (HCC) is the fifth most common cancer around the world. Although several therapeutic approaches for treatment of HCC are available, survival rates for HCC patients are still very poor because of inefficient treatment options. For HCC, as well as other tumors, antigen-specific immunotherapy remains a viable approach that is dependent on the definition of tumor-associated antigens. NY-ESO-1, a member of the cancer testis antigen family, is one possible candidate for a tumor-specific antigen in HCC. The aim of this study was to show the relevance of NY-ESO-1 in hepatocellular carcinoma.

Experimental Design: Sera samples from 189 HCC patients were analyzed for NY-ESO-1-specific antibodies. Forty-nine HCC patients were screened for NY-ESO-1 mRNA expression in HCC tissue. Selected patients were followed for up to 3 years to correlate their immune response with their clinical course of events. NY-ESO-1-specific CD4+ and CD8+ T-cell responses from NY-ESO-1 seropositive patients were analyzed and a NY-ESO-1+ specific cytotoxic T-cell line was generated.

Results: Twelve of 49 analyzed tumor samples expressed NY-ESO-1 mRNA and 23 of 189 patients showed NY-ESO-1-specific antibody responses. These humoral immune responses were accompanied by NY-ESO-1-specific functional CD4+ and CD8+ T-cell responses. Finally, NY-ESO-1 humoral responses were dependent on the presence of NY-ESO-1-expressing tumors.

Conclusions: This is the first report of a spontaneous immune response in HCC patients to a known tumor-specific antigen, NY-ESO-1 protein. Our data favor the possibility of immunotherapeutic strategies for the treatment of HCC.
Gefitinib (ZD1839) Monotherapy as a Salvage Regimen for Previously Treated Advanced Non-Small Cell Lung Cancer
Authors: Park Jinny , Park Byung Bae , Kim Jee Youn , Lee Se-Hoon , Lee Soon Il , Kim Ho Young , Kim Jung Han , Park Se Hoon , Lee Kyung-Eun , Park Joon Oh , Kim Kihyun , Jung Chul Won , Park Young Suk , Im Young-Hyuck , Kang Won Ki , Lee Mark H. , Park Keunchil
Published: September 2005

Purpose: A worldwide compassionate-use program has enabled >42,000 patients with advanced non-small cell lung cancer (NSCLC) to receive gefitinib treatment. Here we report the outcome of gefitinib therapy in patients who enrolled in the “Iressa” Expanded Access Program at the Samsung Medical Center.

Experimental Design: Patients with advanced or metastatic NSCLC who had progressed after prior systemic chemotherapy and for whom no other treatment option was available were eligible to receive gefitinib treatment as part of the Expanded Access Program. A post hoc assessment of potential prognostic factors for response and survival was performed by multivariate analysis.

Results: All 111 evaluable patients had stage IV disease; most patients had a baseline performance status of 2 [n = 52 (47%)] or 3 [n = 18 (16%)] and had received ≥2 prior chemotherapy regimens (56%). The objective response rate was 26%, the disease control rate (measured over ≥8 weeks) was 40%, and the 1-year survival rate was 44%. Adenocarcinoma histology was associated with better response and disease control rates, and a performance status of 0–2 was also associated with a better disease control rate. Both of these factors, as well as female gender, were significantly associated with longer survival. Gefitinib was well tolerated; the most common adverse event was grade 1 skin rash.

Conclusions: Gefitinib demonstrated significant antitumor activity and a favorable tolerability profile in this series of NSCLC patients with poor prognosis.
Phospho-Serine-118 Estrogen Receptor-α Expression Is Associated with Better Disease Outcome in Women Treated with Tamoxifen
Authors: Murphy Leigh C. , Niu Yulian , Snell Linda , Watson Peter
Published: September 2005

Purpose: The purpose of this research was to determine whether estrogen receptor α specifically phosphorylated at Ser118 is associated with clinical outcome in primary breast tumors from estrogen receptor-positive and node-negative breast cancer patients.

Experimental Design: Estrogen receptor α specifically phosphorylated at Ser118 was determined by immunohistochemistry in 117 primary breast tumors from node-negative patients who were subsequently treated with adjuvant tamoxifen. The relationship of estrogen receptor α specifically phosphorylated at Ser118 expression to disease-free survival and overall survival was determined.

Results: Estrogen receptor α specifically phosphorylated at Ser118 was limited to estrogen receptor α ligand binding assay-positive tumors and among this subset was expressed in 70 (62%) of these tumors. Estrogen receptor α specifically phosphorylated at Ser118 expression was more frequently observed in progesterone receptor-positive tumors compared with progesterone receptor-negative tumors (χ2 test, P = 0.012, n = 113). A significant correlation was also seen between estrogen receptor α specifically phosphorylated at Ser118 and progesterone receptor levels (Spearman r = 0.236, P = 0.0118, n = 113). Kaplan-Meier outcome analysis showed that patients whose primary tumors expressed estrogen receptor α specifically phosphorylated at Ser118 had a longer disease-free survival (P = 0.0018, n = 113) and a trend toward better overall survival, but this was not statistically significant. Among the subset of progesterone receptor-positive tumors, progesterone receptor-positive/estrogen receptor α specifically phosphorylated at Ser118-positive patients had a significantly longer disease-free survival that progesterone receptor-positive/estrogen receptor α specifically phosphorylated at Ser118-negative patients (P = 0.0041).

Conclusions: Our data suggest that estrogen receptor α specifically phosphorylated at Ser118 is a marker of a functional, intact ligand-dependent estrogen receptor signaling pathway in breast cancer and that estrogen receptor α specifically phosphorylated at Ser118 status has the potential to provide a more precise biomarker of responsiveness to endocrine therapy in conjunction with estrogen receptor α and progesterone receptor status.
Loss of Fhit Expression in Head and Neck Squamous Cell Carcinoma and Its Potential Clinical Implication
Authors: Tai Shyh-Kuan , Lee Janet I. , Ang K. Kian , El-Naggar Adel K. , Hassan Khaled A. , Liu Diane , Lee J. Jack , Ren Hening , Hong Waun K. , Mao Li
Published: September 2005

Purpose: Abnormalities of FHIT, a candidate tumor suppressor gene, have frequently been found in multiple malignancies, including head and neck squamous cell carcinoma (HNSCC). To define its role in HNSCC treated with surgery and postoperative radiotherapy (PORT), the Fhit protein expression status was investigated in 80 patients enrolled in a prospective Phase III clinical trial addressing the dose and fractionation regimen of PORT.

Experimental Design: Immunohistochemical staining of HNSCC tissue sections for Fhit expression was performed. The Fhit expression status was correlated with the clinicopathological characteristics and clinical course. The median follow-up duration was 4.9 years.

Results: Loss of Fhit expression was found in 52 of the 80 study patients (65%). There was not a significant association between Fhit expression and clinical characteristics. Patients whose tumor exhibited negative Fhit expression had a significantly worse 5-year overall survival duration [hazard ratio = 0.49; 95% confidence interval, 0.23–1.03; P = 0.05 (log-rank test)] than did those whose tumor exhibited positive Fhit expression. One third of the patients with a Fhit-negative tumor had distant metastasis during the follow-up period. Paradoxically, patients classified as high risk who had a Fhit-negative tumor experienced locoregional recurrence less often (18%) than did high-risk patients who had a Fhit-positive tumor (33%).

Conclusions: Loss of Fhit expression is a poor prognostic indicator in patients with HNSCC. However, tumors lacking Fhit expression may be more sensitive to PORT and therefore more susceptible to locoregional control.
Superior Activity of the Combination of Histone Deacetylase Inhibitor LAQ824 and the FLT-3 Kinase Inhibitor PKC412 against Human Acute Myelogenous Leukemia Cells with Mutant FLT-3
Authors: Bali Purva , George Prince , Cohen Pamela , Tao Jianguo , Guo Fei , Sigua Celia , Vishvanath Anasuya , Scuto Anna , Annavarapu Srinivas , Fiskus Warren , Moscinski Lynn , Atadja Peter , Bhalla Kapil
Published: September 2005

Purpose: Mutant FLT-3 receptor tyrosine kinase is a client protein of the molecular chaperone heat shock protein 90 and is commonly present and contributes to the leukemia phenotype in acute myelogenous leukemia (AML). LAQ824, a cinnamyl hydroxamate histone deacetylase inhibitor, is known to induce acetylation and inhibition of heat shock protein 90. Here, we determined the effects of LAQ824 and/or PKC412 (a FLT-3 kinase inhibitor) on the levels of mutant FLT-3 and its downstream signaling, as well as growth arrest and cell-death of cultured and primary human AML cells.

Experimental Design: The effect of LAQ824 and/or PKC412 treatment was determined on the levels of FLT-3 and phosphorylated (p)-FLT-3, on downstream pro-growth and pro-survival effectors, e.g., p-STAT5, p-AKT, and p-extracellular signal-regulated kinase (ERK) 1/2, and on the cell cycle status and apoptosis in the cultured MV4–11 and primary AML cells with mutant FLT-3.

Results: Treatment with LAQ824 promoted proteasomal degradation and attenuation of the levels of FLT-3 and p-FLT-3, associated with cell cycle G1-phase accumulation and apoptosis of MV4–11 cells. This was accompanied by attenuation of p-STAT5, p-AKT, and p-ERK1/2 levels. STAT-5 DNA-binding activity and the levels of c-Myc and oncostatin M were also down-regulated. Cotreatment with LAQ824 and PKC412 synergistically induced apoptosis of MV4–11 cells and induced more apoptosis of the primary AML cells expressing mutant FLT-3. This was also associated with more attenuation of p-FLT-3, p-AKT, p-ERK1/2, and p-STAT5.

Conclusions: The combination of LAQ824 and PKC412 is highly active against human AML cells with mutant FLT-3, which merits in vivo studies of the combination against human AML.
Breast Cancer Resistance Protein in Drug Resistance of Primitive CD34+38− Cells in Acute Myeloid Leukemia
Authors: Raaijmakers Marc H.G.P. , de Grouw Elke P.L.M. , Heuver Leonie H.H. , van der Reijden Bert A. , Jansen Joop H. , Scheper Rik J. , Scheffer George L. , de Witte Theo J.M. , Raymakers Reinier A.P.
Published: September 2005

Purpose: Acute myeloid leukemia (AML) is considered a stem cell disease. Incomplete chemotherapeutic eradication of leukemic CD34+38− stem cells is likely to result in disease relapse. The purpose of this study was to investigate the role of the breast cancer resistance protein (BCRP/ATP-binding cassette, subfamily G, member 2) in drug resistance of leukemic stem cells and the effect of its modulation on stem cell eradication in AML.

Experimental Design: BCRP expression (measured flow-cytometrically using the BXP21 monoclonal antibody) and the effect of its modulation (using the novel fumitremorgin C analogue KO143) on intracellular mitoxantrone accumulation and in vitro chemosensitivity were assessed in leukemic CD34+38− cells.

Results: BCRP was preferentially expressed in leukemic CD34+38− cells and blockage of BCRP-mediated drug extrusion by the novel fumitremorgin C analogue KO143 resulted in increased intracellular mitoxantrone accumulation in these cells in the majority of patients. This increase, however, was much lower than in the mitoxantrone-resistant breast cancer cell line MCF7-MR and significant drug extrusion occurred in the presence of BCRP blockage due to the presence of additional drug transport mechanisms, among which ABCB1 and multiple drug resistance protein. In line with these findings, selective blockage of BCRP by KO143 did not enhance in vitro chemosensitivity of leukemic CD34+38− cells.

Conclusions: These results show that drug extrusion from leukemic stem cells is mediated by the promiscuous action of BCRP and additional transporters. Broad-spectrum inhibition, rather than modulation of single mechanisms, is therefore likely to be required to circumvent drug resistance and eradicate leukemic stem cells in AML.
The Absence of Human Equilibrative Nucleoside Transporter 1 Is Associated with Reduced Survival in Patients With Gemcitabine-Treated Pancreas Adenocarcinoma
Authors: Spratlin Jennifer , Sangha Randeep , Glubrecht Darryl , Dabbagh Laith , Young James D. , Dumontet Charles , Cass Carol , Lai Raymond , Mackey John R.
Published: September 2005

Purpose: Gemcitabine monotherapy is the standard palliative chemotherapy for pancreatic adenocarcinoma. Gemcitabine requires plasma membrane nucleoside transporter proteins to efficiently enter cells and exert it cytotoxicity. In vitro studies have demonstrated that deficiency of human equilibrative nucleoside transporter 1 (hENT1), the most widely abundant and distributed nucleoside transporter in human cells, confers resistance to gemcitabine toxicity, but the distribution and abundance of nucleoside transporters in normal and malignant pancreatic tissue is unknown.

Experimental Design: We studied tumor blocks from normal pancreas and 21 Alberta patients with gemcitabine-treated pancreatic cancer. Immunohistochemistry on the formalin-fixed, paraffin-embedded tissues was performed with specific hENT1 and human Concentrative Nucleoside Transporter 3 monoclonal antibodies and scored by a pathologist blinded to clinical outcomes.

Results: hENT1 was detected in normal Langerhan cells and lymphocytes but not in normal glandular elements. Patients in whom all adenocarcinoma cells had detectable hENT1 had significantly longer median survivals from gemcitabine initiation than those for whom hENT1 was absent in a proportion (10 to 100%) of adenocarcinoma cells (median survival, 13 versus 4 months, P = 0.01). Immunohistochemistry for human Concentrative Nucleoside Transporter 3 revealed moderate to high-intensity staining in all adenocarcinoma tissue samples.

Conclusions: Patients with pancreatic adenocarcinoma with uniformly detectable hENT1 immunostaining have a significantly longer survival after gemcitabine chemotherapy than tumors without detectable hENT1. Immunohistochemistry for hENT1 shows promise as a molecular predictive assay to appropriately select patients for palliative gemcitabine chemotherapy but requires formal validation in prospective, randomized trials.
Molecular Prediction of Response to 5-Fluorouracil and Interferon-α Combination Chemotherapy in Advanced Hepatocellular Carcinoma
Authors: Kurokawa Yukinori , Matoba Ryo , Nagano Hiroaki , Sakon Masato , Takemasa Ichiro , Nakamori Shoji , Dono Keizo , Umeshita Koji , Ueno Noriko , Ishii Shin , Kato Kikuya , Monden Morito
Published: September 2005

Purpose: The prognosis of hepatocellular carcinoma (HCC) is very poor, particularly in patients with tumors that have invaded the major branches of the portal vein. Combination chemotherapy with intra-arterial 5-fluorouracil and subcutaneous interferon-α has shown promising results for such advanced HCC, but it is important to develop the ability to accurately predict chemotherapeutic responses.

Experimental Design: We analyzed the expression of 3,080 genes using a polymerase chain reaction-based array in 20 HCC patients who were treated with combination chemotherapy after reduction surgery. After unsupervised analyses, a supervised classification method for predicting chemotherapeutic responses was constructed. To minimize the number of predictive genes, we used a random permutation test to select only significant (P < 0.01) genes. A leave-one-out cross-validation confirmed the gene selection. We also prepared an additional 11 cases for validation of predictive performance.

Results: Hierarchical clustering analysis and principal component analysis with all 3,080 genes revealed distinct gene expression patterns in responders (those with complete response or partial response) and nonresponders (those with stable disease or progressive disease) to the combination chemotherapy. Using a weighted-voting classification method with either all genes or only significant genes as assessed by permutation testing, the objective responses to treatment were correctly predicted in 17 of 20 cases (accuracy, 85%; positive predictive value, 100%; negative predictive value, 80%). Moreover, patients in the validation dataset could be classified into two distinct prognostic groups using 63 predictive genes.

Conclusions: Molecular analysis of 63 genes can predict the response of patients with advanced HCC and major portal vein tumor thrombi to combination chemotherapy with 5-fluorouracil and interferon-α.
Prognostic Significance of
TP53
Tumor Suppressor Gene Expression and Mutations in Human Osteosarcoma
Authors: Pakos Emilios E. , Kyzas Panayiotis A. , Ioannidis John P. A.
Published: September 2005

Purpose: Various studies examining the relationship between tumor suppressor protein TP53 overexpression and/or TP53 gene mutations and the response to chemotherapy and clinical outcome in patients with osteosarcoma have yielded inconclusive results. The purpose of the current study was to evaluate the relation of TP53 status with response to chemotherapy and/or clinical outcome in osteosarcoma.

Experimental Design: We conducted a meta-analysis of 16 studies (n = 499 patients) that evaluated the correlation between TP53 status and histologic response to chemotherapy and 2-year survival. Data were synthesized in summary receiver operating characteristic curves and with summary likelihood ratios (LRs) and risk ratios.

Results: The quantitative synthesis showed that TP53 status is not a prognostic factor for the response to chemotherapy. The positive LR was 1.21 (95% confidence interval, 0.86–1.71), and the negative LR was 0.91 (95% confidence interval, 0.77–1.07). There was no significant between-study heterogeneity. TP53-positive status tended to be associated with a worse 2-year survival, but the overall results were not formally statistically significant. The association was formally significant in studies that clearly stated that measurements were blinded to outcomes (risk ratio, 2.05; 95% confidence interval, 1.23–3.44), and in studies using reverse transcription-PCR for evaluating TP53 alterations (risk ratio, 1.76; 95% confidence interval, 1.07–2.91).

Conclusions: TP53 status is not associated with the histologic response to chemotherapy in patients with osteosarcoma, whereas TP53 gene alterations may be associated with decreased survival.
Effects of the Antiestrogen Tamoxifen and the Aromatase Inhibitor Letrozole on Serum Hormones and Bone Characteristics in a Preclinical Tumor Model for Breast Cancer
Authors: Núñez Nomelí P. , Jelovac Danijela , Macedo Luciana , Berrigan David , Perkins Susan N. , Hursting Stephen D. , Barrett J. Carl , Brodie Angela
Published: September 2005

Purpose: The purpose of this study was to evaluate and compare the effects of the antiestrogen tamoxifen and the aromatase inhibitor letrozole on tumor growth, serum hormones, uterine weight, body composition, and bone characteristics in mice.

Experimental Design: Human estrogen-dependent breast cancer cells stably transfected with the aromatase gene (MCF-7CA cells) were inoculated in Matrigel subcutaneously into ovariectomized nude mice. This model represents postmenopausal breast cancer in many respects, including the fact that estrogen is no longer produced by the ovaries and is not under feedback regulation by gonadotropins. Mice that received subcutaneously implanted MCF-7CA cancer cells were then treated with tamoxifen or letrozole for 7 weeks.

Results: As reported previously, tumor growth was markedly inhibited by both tamoxifen (100 μg/day) and letrozole (10 μg/day). Tamoxifen treatment led to increased bone mineral density (BMD) and hyperplastic uteri. Mice treated with letrozole had significantly smaller uteri than the controls and tamoxifen-treated mice. Letrozole did not affect BMD. There was no significant difference in systemic leptin and insulin-like growth factor I levels as a result of tamoxifen or letrozole treatment.

Conclusions: Tamoxifen treatment inhibited breast cancer cell growth and increased BMD but caused uterine hypertrophy in this preclinical model of postmenopausal breast cancer. Letrozole inhibited tumor growth without inducing uterine hypertrophy. In addition, letrozole had no effect on BMD. These findings provide experimental evidence that letrozole is an effective and safe (in terms of risk of endometrial cancer risk and osteoporosis) alternative or complement to tamoxifen treatment for breast cancer.
Epratuzumab, a Humanized Anti-CD22 Antibody, in Aggressive Non-Hodgkin’s Lymphoma
Authors: Leonard John P. , Coleman Morton , Ketas Jamie C. , Chadburn Amy , Furman Richard , Schuster Michael W. , Feldman Eric J. , Ashe Michelle , Schuster Stephen J. , Wegener William A. , Hansen Hans J. , Ziccardi Heather , Eschenberg Michael , Gayko Urte , Fields Scott Z. , Cesano Alessandra , Goldenberg David M.
Published: September 2005

Purpose: We conducted a single-center, dose-escalation study evaluating the safety, pharmacokinetics, and efficacy of epratuzumab, an anti-CD22 humanized monoclonal antibody, in patients with aggressive non-Hodgkin’s lymphoma.

Experimental Design: Epratuzumab was administered once weekly for 4 weeks at 120-1000-mg/m2 doses to 56 patients [most (n = 35) with diffuse large B-cell lymphoma].

Results: Patients were heavily pretreated (median, 4 prior therapies), 25% received prior high-dose chemotherapy with stem cell transplant, and 84% had bulky disease (≥5 cm). Epratuzumab was well tolerated, with no dose-limiting toxicity. Most (95%) infusions were completed within 1 h. The mean serum half-life was 23.9 days. Across all dose levels and histologies, objective responses (ORs) were observed in five patients (10%; 95% confidence interval, 3–21%), including three complete responses. In patients with diffuse large B-cell lymphoma, 15% had ORs. Overall, 11 (20%) patients experienced some tumor mass reduction. Median duration of OR was 26.3 weeks, and median time to progression for responders was 35 weeks. Two responses are ongoing at ≥34 months, including one rituximab-refractory patient.

Conclusions: These data demonstrate that epratuzumab has a good safety profile and exerts antitumor activity in aggressive non-Hodgkin’s lymphoma at doses of ≥240 mg/m2, thus warranting further evaluation in this clinical setting.
Proteomic Analysis of Human Acute Leukemia Cells
Authors: Cui Jiu-Wei , Wang Jie , He Kun , Jin Bao-Feng , Wang Hong-Xia , Li Wei , Kang Li-Hua , Hu Mei-Ru , Li Hui-Yan , Yu Ming , Shen Bei-fen , Wang Guan-Jun , Zhang Xue-Min
Published: September 2005

Purpose: French-American-British (FAB) classification of acute leukemia with genetic heterogeneity is important for treatment and prognosis. However, the distinct protein profiles that contribute to the subtypes and facilitate molecular definition of acute leukemia classification are still unclear.

Experimental Design: The proteins of leukemic cells from 61 cases of acute leukemia characterized by FAB classification were separated by two-dimensional electrophoresis, and the differentially expressed protein spots were identified by both matrix-assisted laser desorption/ionization–time-of-flight–mass spectrometry (MALDI-TOF-MS) and tandem electrospray ionization MS (ESI-MS/MS).

Results: The distinct protein profiles of acute leukemia FAB types or subtypes were successfully explored, including acute myeloid leukemia (AML), its subtypes (M2, M3, and M5) and acute lymphoid leukemia (ALL), which were homogeneous within substantial samples of the respective subgroups but clearly differed from all other subgroups. We found a group of proteins that were highly expressed in M2 and M3, rather than other subtypes. Among them, myeloid-related proteins 8 and 14 were first reported to mark AML differentiation and to differentiate AML from ALL. Heat shock 27 kDa protein 1 and other proteins that are highly expressed in ALL may play important roles in clinically distinguishing AML from ALL. Another set of proteins up-regulated was restricted to granulocytic lineage leukemia. High-level expression of NM23-H1 was found in all but the M3a subtype, with favorable prognosis.

Conclusions: These data have implications in delineating the pathways of aberrant gene expression underlying the pathogenesis of acute leukemia and could facilitate molecular definition of FAB classification. The extension of the present analysis to currently less well-defined acute leukemias will identify additional subgroups.
Nitric Oxide Accelerates Interleukin-13 Cytotoxin-Mediated Regression in Head and Neck Cancer Animal Model
Authors: Kawakami Koji , Kawakami Mariko , Puri Raj K.
Published: September 2005

Receptors for interleukin-13 (IL-13R) are overexpressed on several types of solid cancers including gliobastoma, renal cell carcinoma, AIDS Kaposi’s sarcoma, and head and neck cancer. Recombinant fusion proteins IL-13 cytotoxin (IL13-PE38QQR or IL13-PE38) have been developed to directly target IL-13R-expressing cancer cells. Although it has been found that IL-13 cytotoxin has a direct potent antitumor activity in vivo in nude mice models of human cancers, the involvement of indirect antitumor effecter molecules such as nitric oxide (NO) is unknown. To address this issue, we assessed the effect of NO inhibiter Nω-monomethyl-l-arginine on IL-13 cytotoxin-mediated cytotoxicity and NO2/NO3 production in HN12 head and neck cancer cells. In addition, antitumor effects and NO levels in HN12 and KCCT873 head and neck tumors xenografted s.c. in nude mice when treated with IL-13 cytotoxin were evaluated by tumor measurement, Western blot, and immunohistochemistry analyses. Pretreatment of animals with Nω-monomethyl-l-arginine significantly decreased the NO levels and IL-13 cytotoxin-mediated antitumor effects. In addition, depletion of macrophages, known to produce NO, also decreased antitumor activity of IL-13 cytotoxin. Based on these studies, we concluded that NO accelerates antitumor effect of IL-13 cytotoxin on head and neck tumor cells. Because IL-13 cytotoxin is currently being tested in the clinic for the treatment of patients with recurrent glioblastoma maltiforme, our current findings suggest maintaining macrophage and NO-producing cellular function for optimal therapeutic effect of this targeted agent.
Immunization of Patients with the hTERT:540-548 Peptide Induces Peptide-Reactive T Lymphocytes That Do Not Recognize Tumors Endogenously Expressing Telomerase
Authors: Parkhurst Maria R. , Riley John P. , Igarashi Takehito , Li Yong , Robbins Paul F. , Rosenberg Steven A.
Published: September 2005

Purpose: Telomerase is an attractive target antigen for cancer immunotherapies because it is expressed in >85% of human tumors but is rarely found in normal tissues. A HLA-A*0201-restricted T-cell epitope was previously identified within telomerase reverse transcriptase hTERT:540-548. This peptide was reported to induce CTL that recognized tumor cells and transfectants that endogenously expressed telomerase. Therefore, we initiated a clinical protocol to evaluate the therapeutic and immunological efficacy of this peptide.

Experimental Design: Fourteen patients with metastatic cancers were vaccinated with hTERT:540-548 emulsified in incomplete Freund’s adjuvant.

Results: In 7 patients, peripheral blood mononuclear cells collected after immunization recognized hTERT:540-548, whereas those collected before vaccination did not. However, none of these CTLs recognized tumors that endogenously expressed telomerase, and none of the patients had an objective clinical response. Several highly avid T-cell clones were generated that recognized T2 cells pulsed with ≤1 nm hTERT:540-548, but none of these recognized HLA-A*0201+ hTERT+ tumors or cells transduced with the human telomerase reverse transcriptase (hTERT) gene. Also, an antibody specific for hTERT:540-548/HLA-A*0201 complexes stained peptide-pulsed cells but not telomerase+ tumors.

Conclusions: Our results are discordant with previous studies and those of a clinical trial that claimed peripheral blood mononuclear cells from patients vaccinated with peptide-pulsed dendritic cells lysed hTERT+ tumors. However, our findings are consistent with a previous study that demonstrated that the hTERT:540-548 peptide is cleaved in the proteasome. These results suggest that hTERT:540-548 is not presented on the surfaces of tumor cells in the context of HLA-A*0201 and will not be useful for the immunotherapy of patients with cancer.
Identification of Glypican-3 as a Novel Tumor Marker for Melanoma
Authors: Nakatsura Tetsuya , Kageshita Toshiro , Ito Shosuke , Wakamatsu Kazumasa , Monji Mikio , Ikuta Yoshiaki , Senju Satoru , Ono Tomomichi , Nishimura Yasuharu
Published: September 2005

Purpose: We reported recently the novel tumor marker glypican-3 (GPC3) for hepatocellular carcinoma. In the present study, we investigated the expression of GPC3 in human melanoma cell lines and tissues and asked whether GPC3 could be a novel tumor marker for melanoma.

Experimental Design: Expression of GPC3 mRNA and protein was investigated in human melanoma cell lines and tissues using reverse transcription-PCR and immunohistochemical analysis. Secreted GPC3 protein was quantified using ELISA in culture supernatants of melanoma cell lines and in sera from 91 patients with melanoma and 28 disease-free patients after surgical removal of primary melanoma. All of the subjects were Japanese nationals.

Results: In >80% of melanoma and melanocytic nevus, there was evident expression of GPC3 mRNA and protein. Furthermore, GPC3 protein was evidenced in sera of 39.6% (36 of 91) of melanoma patients but not in sera from subjects with large congenital melanocytic nevus (0 of 5) and from healthy donors (0 of 60). Twenty-seven of 36 serum GPC3-positive patients were negative for both serum 5-S-cysteinyldopa and melanoma-inhibitory activity, well-known tumor markers for melanoma. The positive rate of serum GPC3 (39.6%) was significantly higher than that of 5-S-cysteinyldopa (26.7%) and of melanoma-inhibitory activity (20.9%). Surprisingly, we detected serum GPC3 even in patients with stage 0 in situ melanoma. The positive rate of serum GPC3 at stage 0, I, and II (44.4%, 40.0%, and 47.6%) was significantly higher than that of 5-S-cysteinyldopa (0.0%, 8.0%, and 10.0%). Also observed was the disappearance of GPC3 protein in sera from 11 patients after surgical removal of the melanoma.

Conclusions: GPC3 is apparently a novel tumor marker useful for the diagnosis of melanoma, especially in early stages of the disorder.
Hypoxia and Photofrin Uptake in the Intraperitoneal Carcinomatosis and Sarcomatosis of Photodynamic Therapy Patients
Authors: Busch Theresa M. , Hahn Stephen M. , Wileyto E. Paul , Koch Cameron J. , Fraker Douglas L. , Zhang Paul , Putt Mary , Gleason Kristen , Shin Daniel B. , Emanuele Michael J. , Jenkins Kevin , Glatstein Eli , Evans Sydney M.
Published: September 2005

Purpose: Response to photodynamic therapy depends on adequate tumor oxygenation as well as sufficient accumulation of photosensitizer in the tumor. The goal of this study was to investigate the presence of hypoxia and retention of the photosensitizer Photofrin in the tumors of patients with intra-abdominal carcinomatosis or sarcomatosis.

Experimental Design: Tumor nodules from 10 patients were studied. In nine of these patients, hypoxia was identified in histological sections of biopsied tumor after administration of the hypoxia marker 2-(2-nitroimidazol-1[H]-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide (EF5). In separate tumor nodules from 10 patients, Photofrin uptake was measured by fluorescence after tissue solubilization.

Results: Hypoxia existed in the tumors of five patients, with three of these patients demonstrating at least one severely hypoxic nodule. Physiological levels of oxygen were present in the tumors of four patients. An association between tumor size and hypoxia was not evident because some tumor nodules as small as ∼2 mm in diameter were severely hypoxic. However, even these tumor nodules contained vascular networks. Three patients with severely hypoxic tumor nodules exhibited moderate levels of Photofrin uptake of 3.9 ± 0.4 to 3.9 ± 0.5 ng/mg (mean ± SE). The four patients with tumors of physiological oxygenation did not consistently exhibit high tumor concentrations of Photofrin: mean ± SE drug uptake among these patients ranged from 0.6 ± 0.8 to 5.8 ± 0.5 ng/mg.

Conclusions: Carcinomatosis or sarcomatosis of the i.p. cavity may exhibit severe tumor hypoxia. Photofrin accumulation in tumors varied by a factor of ∼10× among all patients, and, on average, those with severe hypoxia in at least one nodule did not demonstrate poor Photofrin uptake in separate tumor samples. These data emphasize the need for reconsideration of the generally accepted paradigm of small tumor size, good oxygenation, and good drug delivery because this may vary on an individual tumor basis.
Noscapine Crosses the Blood-Brain Barrier and Inhibits Glioblastoma Growth
Authors: Landen Jaren W. , Hau Vincent , Wang Mingshen , Davis Thomas , Ciliax Brian , Wainer Bruce H. , Van Meir Erwin G. , Glass Johnathan D. , Joshi Harish C. , Archer David R.
Published: September 2005

The opium alkaloid noscapine is a commonly used antitussive agent available in Europe, Asia, and South America. Although the mechanism by which it suppresses coughing is currently unknown, it is presumed to involve the central nervous system. In addition to its antitussive action, noscapine also binds to tubulin and alters microtubule dynamics in vitro and in vivo. In this study, we show that noscapine inhibits the proliferation of rat C6 glioma cells in vitro (IC50 = 100 μm) and effectively crosses the blood-brain barrier at rates similar to the ones found for agents such as morphine and [Met]enkephalin that have potent central nervous system activity (P ≤ 0.05). Daily oral noscapine treatment (300 mg/kg) administered to immunodeficient mice having stereotactically implanted rat C6 glioblasoma into the striatum revealed a significant reduction of tumor volume (P ≤ 0.05). This was achieved with no identifiable toxicity to the duodenum, spleen, liver, or hematopoietic cells as determined by pathological microscopic examination of these tissues and flow cytometry. Furthermore, noscapine treatment resulted in little evidence of toxicity to dorsal root ganglia cultures as measured by inhibition of neurite outgrowth and yielded no evidence of peripheral neuropathy in animals. However, evidence of vasodilation was observed in noscapine-treated brain tissue. These unique properties of noscapine, including its ability to cross the blood-brain barrier, interfere with microtubule dynamics, arrest tumor cell division, reduce tumor growth, and minimally affect other dividing tissues and peripheral nerves, warrant additional investigation of its therapeutic potential.