bluuebunny/crossref_metadata_2025 huggingface dataset:
A Multicenter Phase I Gene Therapy Clinical Trial Involving Intraperitoneal Administration of E1A-Lipid Complex in Patients with Recurrent Epithelial Ovarian Cancer Overexpressing HER-2/
neu
Oncogene
Authors: Madhusudan Srinivasan , Tamir Ayala , Bates Nicholas , Flanagan Elizabeth , Gore Martin E. , Barton Desmond P. J. , Harper Peter , Seckl Michael , Thomas Hilary , Lemoine Nicholas R. , Charnock Mark , Habib Nagy A. , Lechler Robert , Nicholls Joanna , Pignatelli Massimo , Ganesan Trivadi S.
Published: September 2005

Purpose: HER-2/neu oncogene is overexpressed in 10–30% of epithelial ovarian cancers and is associated with a poor prognosis. The E1A gene product of adenovirus type 5 down-regulates HER-2/neu and causes tumor regression in animal models. In the current study, we sought to determine the toxicity and biological activity of E1A-lipid complex in ovarian cancer patients.

Experimental Design: A Phase I trial involving intraperitoneal (i.p.) administration of E1A-lipid complex was initiated in ovarian cancer patients to assess biological activity (E1A gene transfer/transcription/translation and HER-2/neu expression) and to determine the maximum tolerated dose. Successive cohorts received E1A-lipid complex at doses of 1.8, 3.6, and 7.2 mg DNA/m2, given as weekly i.p. infusions for 3 of 4 weeks (each cycle) up to a maximum of six cycles. Peritoneal fluid was sampled at baseline and twice monthly for cellularity, cytology, CA-125, and biological activity

Results: Fifteen patients, with a median age of 57 years (range, 43–81) were recruited. Three (1.8 mg DNA/m2), 4 (3.6 mg DNA/m2), and 8 patients (7.2 mg DNA/m2) received i.p. E1A. A total of 91 infusions (range, 1–18) was administered. Abdominal pain was the dose-limiting toxicity, and the maximum-tolerated dose was 3.6 mg DNA/m2. E1A gene transfer and expression was observed in all of the patients and at all of the dose levels. HER-2/neu down-regulation could be demonstrated in the tumor cells of 2 patients (18%). There was no correlation between dose and biological activity.

Conclusions: I.P. EIA-lipid complex gene therapy is feasible and safe. Future studies, either alone or in combination with chemotherapy, particularly in patients with minimal residual disease, should be evaluated.
A Phase I and Pharmacological Study of the Platinum Polymer AP5280 Given as an Intravenous Infusion Once Every 3 Weeks in Patients with Solid Tumors
Authors: Rademaker-Lakhai Jeany M. , Terret Catherine , Howell Stephen B. , Baud Christiane M. , de Boer Richard F. , Pluim Dick , Beijnen Jos H. , Schellens Jan H. M. , Droz Jean-Pierre
Published: September 2005

Purpose: This Phase I study was designed to determine the maximum tolerated dose, profile of adverse events, and dose-limiting toxicity of AP5280 in patients with solid tumors. Furthermore, the platinum (Pt) pharmacokinetics after AP5280 administration and preliminary antitumor activity were evaluated. AP5280 is a Pt agent linked to the water-soluble, biocompatible copolymer N-(2-hydroxypropyl)methacrylamide, which potentially increases Pt accumulation in tumors via the enhanced permeability and retention effect. In this way, it is anticipated that a higher activity of therapeutic Pt can be reached. The pharmaceutical product contains approximately 8.5% of Pt by weight and has a molecular weight of approximately 25,000.

Experimental Design: Adult patients with solid tumors received AP5280 as a 1-h i.v. infusion every 21 days. Pharmacokinetics of total and unbound Pt were determined during the first treatment course and before the start of each new cycle using noncompartmental pharmacokinetic analysis. Pt-DNA adduct concentrations in WBCs and, if available, in tumor tissue were quantified using a sensitive 32P postlabeling assay.

Results: Twenty-nine patients were treated at eight dose levels (90–4500 mg Pt/m2). The dose-limiting toxicity was Common Toxicity Criteria grade 3 vomiting and was experienced at 4500 mg Pt/m2 in two of six patients. The maximum tolerated dose on this schedule was therefore 4500 mg Pt/m2, and the recommended dose for a Phase II study is 3300 mg Pt/m2. Renal toxicity and myelosuppression, toxicities typically observed with cisplatin and carboplatin, were minimal for AP5280. The area under the curve of total Pt increased with increasing AP5280 dose. Plasma clearance of total Pt was 644 ± 266 ml/h, and the terminal half-life was 116 ± 46.2 h. After AP5280 administration, Pt-guanine-guanine DNA adduct concentrations in WBCs ranged from 70 to 1848 amol/μg DNA, concentrations that were substantially lower than concentrations measured after administration of therapeutic doses of cisplatin.

Conclusions: AP5280 can be administered safely as a 1-h i.v. infusion at a dose of 3300 mg Pt/m2 once every 3 weeks and produces prolonged plasma exposure compared with any of the free Pt-containing drugs. However, it remains to be determined whether AP5280 can actually increase Pt delivery to the DNA of tumor cells in man as has been shown in experimental models.
A Phase I and Pharmacokinetic Study of Weekly Paclitaxel and Carboplatin in Patients with Metastatic Esophageal Cancer
Authors: Polee Marco B. , Sparreboom Alex , Eskens Ferry A. L. M. , Hoekstra Ronald , van de Schaaf Jacqueline , Verweij Jaap , Stoter Gerrit , van der Gaast Ate
Published: September 2005

Purpose: To determine the maximum-tolerated dose, toxicity profile, and pharmacokinetics of a fixed dose of paclitaxel followed by increasing doses of carboplatin, given weekly to patients with advanced esophageal or gastric junction cancer.

Experimental Design: Paclitaxel was administered on day 1 as a 1-h infusion at a fixed dose of 100 mg/m2 followed by a 1-h infusion of carboplatin targeting an area under the curve (AUC) of 2–5 mg × min/ml, with cycles repeated on days 8, 15, 29, 36, and 43.

Results: Forty patients [36 males; median (range) age, 57 (40–74) years] were enrolled. Dose-limiting toxicity was observed at a carboplatin AUC of 5 mg × min/ml and consisted of treatment delay attributable to myelosuppression. No grade 3/4 treatment-related nonhematological toxicity was observed. The highest dose intensity (>95% of the planned dose over time) was achieved with a carboplatin AUC of 4 mg × min/ml. The mean (±SD) AUCs of unbound (Cu) and total paclitaxel were 0.662 ± 0.186 and 7.37 ± 1.33 μm × h, respectively. Clearance of Cu was 188 ± 44.6 liter/h/m2, which is not significantly different from historical data (P = 0.52). Cremophor EL clearance was 123 ± 23 ml/h/m2, similar to previous findings. Of 37 patients evaluable for response, 1 had complete response, 19 had partial response, and 10 had stable disease, accounting for an overall response rate of 54%.

Conclusions: This regimen is very tolerable and effective, and the recommended doses for additional studies are paclitaxel (100 mg/m2), with carboplatin targeting an AUC of 4 mg × min/ml.
Prognostic Analysis of E-Cadherin Gene Promoter Hypermethylation in Patients with Surgically Resected, Node-Positive, Diffuse Gastric Cancer
Authors: Graziano Francesco , Arduini Federica , Ruzzo Annamaria , Bearzi Italo , Humar Bostjan , More Helen , Silva Rosarita , Muretto Pietro , Guilford Parry , Testa Enrica , Mari Davide , Magnani Mauro , Cascinu Stefano
Published: September 2005

Purpose: Recent investigations have demonstrated that hypermethylation is a frequent mechanism for silencing tumor suppressor genes. This is a potentially reversible epigenetic change, and it is the target of a novel class of anticancer compounds with demethylating activity. Better understanding of the clinical implications of hypermethylation will allow the optimal planning of future trials with demethylating drugs. In this perspective, we investigated whether hypermethylation in the CDH1 promoter region is correlated with poor prognosis of patients with surgically resected, node-positive, diffuse gastric cancer.

Experimental Design: Consecutive cases of diffuse gastric cancer were considered eligible for study entry. Additional inclusion criteria were radical surgery with a minimum of D1 lymphadenectomy, complete follow-up information, and availability of tumor specimens for methylation-specific PCR and immunohistochemistry analyses.

Results: CDH1 promoter hypermethylation was found in 40 of 73 cases (54%), and it was significantly associated with worse prognosis. In patients with and without hypermethylation, the 5-year event-free survival rate was 30% and 62%, respectively, and the 5-year overall survival rate was 35% and 67%, respectively. CDH1 promoter hypermethylation retained its prognostic role for disease-free survival (P < 0.001) and overall survival (P < 0.001) in multivariate analysis. Immunohistochemistry showed a significant association between CDH1 methylation and E-cadherin expression (P < 0.001).

Conclusions: This study shows adverse prognostic effect of CDH1 promoter hypermethylation in patients with diffuse gastric cancer. This form of cancer, and other types with frequent hypermethylation and silencing of critical tumor suppressor genes, would make appropriate targets for the testing of novel compounds with demethylating activity.
A Phase I Pharmacokinetic and Pharmacodynamic Study of the Farnesyl Transferase Inhibitor BMS-214662 in Combination with Cisplatin in Patients with Advanced Solid Tumors
Authors: Mackay Helen J. , Hoekstra Ronald , Eskens Ferry A. L. M. , Loos Walker J. , Crawford Donna , Voi Maurizio , Van Vreckem Anne , Evans T. R. Jeffrey , Verweij Jaap
Published: September 2005

Purpose: BMS-214662 is a potent and selective inhibitor of the farnesyl transferase enzyme with in vitro and in vivo antitumor activity. The aims of this study were to characterize the toxicities and to determine the pharmacokinetic profiles of BMS-214662 when administered in combination with cisplatin, and to determine the constitutive farnesyltransferase activity as a surrogate pharmacodynamic end point.

Experimental Design: Twenty-nine patients with advanced solid malignancy, refractory to conventional therapy, and with adequate hematological, renal, and hepatic function were treated with escalating doses of BMS-214662 administered as a 1-h infusion, followed after an interval of 30 min by 75 mg/m2 cisplatin administered as a 4-h infusion and repeated every 21 days. Blood and urine samples for pharmacokinetic and pharmacodynamic analyses were collected during the first cycle of treatment only.

Results: Dose-limiting toxicities occurred in 4 of 9 patients enrolled at the 225 mg/m2 BMS-214662 dose cohort, and included elevation of hepatic transaminases, nausea, vomiting, diarrhea, and renal failure. There was no apparent pharmacokinetic interaction between the two drugs at the recommended dose levels, and a dose-dependent inhibition of farnesyltransferase activity was observed, which returned to control levels within 24 h of drug administration. There were no objective responses, but disease stabilization was observed in 15 patients, including 4 patients with stable disease after 6 cycles of treatment.

Conclusions: A dose of 200 mg/m2 of BMS-214662 administered as a 1-h infusion with 75 mg/m2 cisplatin over 4 h is the recommended dose for additional studies.
Improved Targeting of Pancreatic Cancer
Authors: Cardillo Thomas M. , Karacay Habibe , Goldenberg David M. , Yeldell Dion , Chang Chien-Hsing , Modrak David E. , Sharkey Robert M. , Gold David V.
Published: September 2005

Purpose: The early detection and diagnosis of pancreatic cancer remains a major clinical challenge in which imaging procedures have a central role. The purpose of this study was to evaluate a pretargeting method with a bispecific PAM4 (bsPAM4; anti-MUC1) antibody for radioimmunoscintigraphy of experimental human pancreatic cancer.

Experimental Design: A bispecific F(ab′)2 antibody was generated from chimeric PAM4 Fab′ and murine 734 (anti-indium-diethylenetriaminepentaacetic acid) Fab′ fragments and then used in conjunction with 2 peptide haptens (111In-IMP-156 and 99mTc-IMP-192). Biodistribution studies and radioimmunoscintigraphic imaging properties of the radiolabeled bsPAM4, and pretargeted, radiolabeled peptides were examined in the CaPan1 human pancreatic tumor grown as s.c. xenografts in athymic nude mice. Tumor uptake and tumor:nontumor ratios were compared with a nontargeting irrelevant anti-CD20, bispecific rituximab, radiolabeled peptides alone, and with directly labeled PAM4.

Results: Biodistribution results indicated significantly greater tumor uptake of radiolabeled peptides at 3 h after injection when pretargeting was performed with bsPAM4 as compared with the bispecific rituximab [20.2 ± 5.5 percentage of injected dose per gram of tissue (%ID/g) versus 0.9 ± 0.1%ID/g, respectively, for 111In-IMP-156, and 16.8 ± 4.8%ID/g versus 1.1 ± 0.2%ID/g, respectively, for 99mTc-IMP-192]. Similar results were obtained at the 24-h time point. Tumor:nontumor ratios were >30 for all of the tissues except the kidneys, where a ratio of 7.8 ± 2.8 was observed. By immunoscintigraphy, tumors could be visualized as early as 30 min after injection of the radiolabeled peptide.

Conclusions: These studies demonstrate the feasibility of using the pretargeted, bispecific antibody technology for nuclear imaging of pancreatic cancer. The advantage of pretargeted bsPAM4 antibody as an imaging platform is the high specificity for pancreatic cancer as compared with the physicochemical parameters identified by current imaging technologies.
Tumor Tissue Levels of Tissue Inhibitor of Metalloproteinase-1 as a Prognostic Marker in Primary Breast Cancer
Authors: Schrohl Anne-Sofie , Holten-Andersen Mads N. , Peters Harry A. , Look Maxine P. , Meijer-van Gelder Marion E. , Klijn Jan G. M. , Brünner Nils , Foekens John A.
Published: September 2005

Purpose: In the present study, we investigated the association between tumor tissue levels of tissue inhibitor of metalloproteinase-1 (TIMP-1) and prognosis in patients with primary breast cancer and analyzed whether TIMP-1 may be useful as a prognostic marker in combination with urokinase plasminogen activator (uPA) and plasminogen activator inhibitor type-1 (PAI-1).

Experimental Design: In cytosolic extracts of 2984 primary breast tumors, total levels of TIMP-1 were determined using an established, validated ELISA. Levels of uPA and PAI-1 have previously been determined in the extracts.

Results: Univariate survival analysis showed a significant relationship between higher levels of TIMP-1 (continuous log-transformed variable) and poor prognosis [recurrence-free survival (RFS), overall survival (OS); P < 0.001]. Performing isotonic regression analysis, we identified a cut point to classify tumors as TIMP-1-low or TIMP-1-high. Using this cut point, high levels of TIMP-1 were significantly associated with shorter survival in univariate analysis, both in the total patient group (RFS, OS; P < 0.001), in the node-negative subgroup (RFS, hazard ratio = 1.28, P = 0.006), and in the node-positive subgroup (RFS, hazard ratio = 1.43, P < 0.001). In multivariate analysis, including uPA and PAI-1, TIMP-1 was significantly associated with shorter RFS, both when included as a continuous log-transformed (P = 0.03) and as a dichotomized variable (P = 0.002).

Conclusions: This study validates previous findings that tumor tissue levels of TIMP-1 are associated with prognosis in patients with primary breast cancer. It confirms that TIMP-1 may be useful as a prognostic marker in combination with uPA/PAI-1 and adds substantial positive information on the use of TIMP-1 as a prognostic marker in breast cancer.
Polymorphisms in Genes Involved in Homologous Recombination Repair Interact to Increase the Risk of Developing Acute Myeloid Leukemia
Authors: Seedhouse Claire , Faulkner Rowena , Ashraf Nadia , Das-Gupta Emma , Russell Nigel
Published: September 2005

Purpose: Double-strand break repair via homologous recombination is essential in maintaining genetic integrity. RAD51 and XRCC3 are involved in the repair of DNA by this pathway, and polymorphisms have been identified in both the RAD51 (RAD51-G135C) and XRCC3 (XRCC3-Thr241Met) genes. The object of this study was to examine whether these polymorphisms may modulate susceptibility to the development of acute myeloid leukemia (AML), a disease that is characterized by genetic instability.

Experimental Design: We studied the distribution of polymorphisms in RAD51 and XRCC3 in 216 cases of de novo AML, 51 cases of therapy-related AML (t-AML), and 186 control subjects using PCR followed by restriction enzyme digestion. The polymorphic deletion of the detoxification gene glutathione S-transferase M1 (GSTM1) was also examined by PCR.

Results: The risk of the development of AML was found to be significantly increased when both variant RAD51-135C and XRCC3-241Met alleles are present [odds ratio (OR), 3.77; 95% confidence interval (CI), 1.39–10.24], whereas the risk of t-AML development is even higher (OR, 8.11; 95% CI, 2.22–29.68), presumably because of the large genotoxic insult these patients receive after their exposure to radiotherapy or chemotherapy. If we further divide the AML group into patients in which the burden of DNA damage is increased, because of the deletion of the GSTM1 gene, the risk of development of AML is further increased (OR, 15.26; 95% CI, 1.83–127.27).

Conclusions: These results strongly suggest that DNA double-strand breaks and their repair are important in the pathogenesis of both de novo and t-AML.
Improved Outcome When B-Cell Lymphoma Is Treated with Combinations of Immunoliposomal Anticancer Drugs Targeted to Both the CD19 and CD20 Epitopes
Authors: Sapra Puja , Allen Theresa M.
Published: September 2005

Purpose: We have reported previously that successful immunoliposomal drug therapy with liposomal doxorubicin (DXR) against xenograft B-lymphoma models required targeting against an internalizing B-cell antigen, CD19 (P. Sapra and T. M. Allen. Cancer Res 2002;62:7190–4.). Here we compare targeting of immunoliposomal formulations of DXR with vincristine (VCR) targeted against CD19 versus a noninternalizing (CD20) epitope. We also examine the effect of targeting immunoliposomes with antibody combinations in an attempt to increase the total number of binding sites (apparent antigen density) at the target cell surface.

Experimental Design: Cell association of immunoliposomes (CD19-targeted, CD20-targeted, or combinations of the two) with human B-cell lymphoma (Namalwa) cells were studied using radiolabeled liposomes. Therapeutic efficacy of the same formulations was determined in a severe combined immunodeficient murine model.

Results: Therapeutic results in severe combined immunodeficient mice bearing Namalwa cells administered anti-CD20-targeted liposomal DXR were barely improved over those found for nontargeted liposomal DXR or free DXR but, surprisingly, administration of anti-CD20-targeted liposomal VCR resulted in a significantly improved therapeutic outcome compared with nontargeted liposomal VCR, free VCR, or anti-CD20-targeted liposomal DXR.

Treatment of murine B lymphoma with single injections of combinations of anti-CD19- and anti-CD20-targeted liposomal VCR led to cures in 70% of mice. However, mice injected with similar combinations of liposomal DXR did not have improved survival rates over anti-CD19-targeted liposomal DXR by itself.

Conclusions: The success of immunoliposomal therapy in combination regimens varies with the type of encapsulated drug and the nature of the target epitopes.
Human Telomerase Reverse Transcriptase Gene Expression and the Surgical Management of Suspicious Thyroid Tumors
Authors: Umbricht Christopher B. , Conrad George T. , Clark Douglas P. , Westra William H. , Smith Dañelle C. , Zahurak Marianna , Saji Motoyasu , Smallridge Robert C. , Goodman Steven , Zeiger Martha A.
Published: September 2005

Purpose: Patients with a preoperative cytologic diagnosis of a suspicious thyroid nodule present a therapeutic dilemma because surgery differs for benign and malignant lesions. To address this issue, several molecular markers, including human telomerase reverse transcriptase (TERT), have been tested as markers of thyroid cancer. Because most studies select cases falling into well-defined categories to test new markers, they may overestimate their discriminatory power when applied to samples that are difficult to classify. Fine-needle aspirates (FNAs) of the thyroid with indeterminate cytology are an example of such cases.

Experimental Design: We examined whether assessing TERT mRNA by reverse transcription-PCR could have improved the surgical management in a cohort of 100 patients undergoing thyroidectomy for indeterminate FNA results.

Results: Ninety percent of 48 cancers were TERT positive, as were 35% of 52 benign lesions. When 10 cases with concomitant lymphocytic thyroiditis were excluded, the overall sensitivity of TERT was 91% (95% confidence interval, 80–98%) and specificity was 79% (64–90%). No clinical or tumor variable contributed to the predictive ability of TERT except for tumor size, which added only marginally. Basing the surgical approach on the TERT assay alone would have reduced lobectomies performed for malignant disease from 11 to 4 cases and reduced total thyroidectomies for benign lesions from to 15 to 9, an overall 50% reduction in suboptimal treatment.

Conclusions: The overall performance of preoperative differential diagnosis for thyroid tumors with indeterminate FNA results can be substantially improved by the inclusion of molecular markers such as TERT.
Mammaglobin Expression in Leukapheresis Products Is a Predictive Marker of Poor Prognosis in Women with High-Risk Breast Cancer
Authors: Ferrucci Pier Francesco , Rabascio Cristina , Mazzetta Chiara , Cocorocchio Emilia , Agazzi Alberto , Vanazzi Anna , Cinieri Saverio , Peccatori Fedro A. , Paolucci Mara , Bertolini Francesco , Martinelli Giovanni
Published: September 2005

Purpose: The purpose of this study was to investigate the incidence and prognostic relevance of tumor cell detection in granulocyte colony-stimulating factor–mobilized peripheral blood progenitor cell collections (PBPCCs) using cytokeratin (CK), maspin (MAS), and mammaglobin (MAM) genes as epithelial cell markers. The population on which the study was conducted was drawn from stage III breast cancer patients undergoing high-dose chemotherapy and autologous transplantation with PBPCCs.

Experimental Design: One hundred and ninety-four patients were enrolled in the study and analyzed for tumor cell detection on the basis of 481 PBPCCs gathered before administration of chemotherapy. CK, MAS, and MAM gene expressions were investigated by means of the reverse transcription nested polymerase chain reaction, and those samples expressing CK were further hybridized with a radiolabeled internal probe to reduce false-positive results. Sensitivity and specificity were assessed on 37 controls (12 cell lines, 12 healthy donors, and 13 nonepithelial malignancies). Each of the known prognostic variables (age, stage, lymph node status, receptor status, c-ErbB2 status, and Ki67 status) was then analyzed (both individually and together with CK, MAS, and MAM expression on PBPCCs) in relation to patient overall survival (OS) and relapse-free survival (RFS).

Results: After a 3-year follow-up, an estimated 83% (95% confidence interval, 77.1–88.8%) of the patients were alive and an estimated 67% (95% confidence interval, 60.1–74.6%) were free of relapse. One hundred and seventy-six of the 194 patients (91%) had contaminated PBPCCs evidenced by at least one positive sample for any of the markers evaluated. The PBPCC frequency of CK, MAS, and MAM positivity (+) was 71%, 36%, and 16%, respectively. MAM expression on PBPCC was associated with an increased risk of relapse (P = 0.003), whereas CK and MAS expressions were not associated with changes in either RFS or OS.

Conclusions: MAM gene expression on leukapheresis products of high-risk breast cancer patients is an indicator of poor prognosis. The method of evaluation is simple and reproducible and provides new tools for evaluating the role played by tumor cells in apheresis products and their potential in causing metastasis.
Oral RDP58 Allows CPT-11 Dose Intensification for Enhanced Tumor Response by Decreasing Gastrointestinal Toxicity
Authors: Zhao Jingsong , Huang Lingyan , Belmar Nicole , Buelow Roland , Fong Timothy
Published: September 2005

Cancer patients undergoing triple therapy (CPT-11, 5-fluorouracil, and leucovorin) often present with severe delayed diarrhea as a result of chemotherapy-induced gastrointestinal (GI) toxicity and inflammation. RDP58 is a novel, anti-inflammatory, d-amino acid decapeptide that inhibits the production of tumor necrosis factor α, IFN-γ, and interleukin 12, and has been shown to effectively inhibit clinical symptoms and intestinal inflammation in several rodent models of chemically induced colitis, nonhuman primates with spontaneous colitis, and humans with mild to moderate ulcerative colitis. We evaluated RDP58 as a potential protective agent in chemotherapy-induced GI inflammation. Oral administration of RDP58 significantly decreased the incidence of diarrhea and improved the survival rates of mice treated with toxic doses of CPT-11 or 5-fluorouracil. Histological analysis showed that RDP58 significantly reduced the destruction of the intestinal mucosa by inhibiting local overproduction of tumor necrosis factor α, IFN-γ, and interleukin 12 in vivo. Furthermore, RDP58 administration allowed the maximum tolerated dose of CPT-11 to be doubled in tumor-bearing mice resulting in significantly enhanced primary tumor responses and prolongation of time to relapse without a concomitant increase in GI toxicity. Our results suggest that RDP58 may have clinical utility in cancer therapy by preventing treatment-associated GI toxicity and potentially increasing the effectiveness of chemotherapy.
Disease-Stage Variance in Functional CD4+ T-Cell Responses Against Novel Pan-Human Leukocyte Antigen-D Region Presented Human Papillomavirus-16 E7 Epitopes
Authors: Warrino Dominic E. , Olson Walter C. , Knapp William T. , Scarrow Meera I. , D’Ambrosio-Brennan Lori J. , Guido Richard S. , Edwards Robert P. , Kast W. Martin , Storkus Walter J.
Published: September 2005

Given the anticipated clinical importance of helper and regulatory CD4+ T cells reactive against human papillomavirus-16 E7 in the cervical carcinoma setting, we performed this study to identify novel E7-derived T helper (Th) epitopes and to characterize functional anti-E7 Th responses in normal donors and patients with cervical intraepithelial neoplasia I-III or cervical cancer.

Candidate pan-HLA-DR (D region) binding peptides were identified and synthesized based on results obtained using a predictive computer algorithm, then applied in short-term in vitro T-cell sensitization assays. Using IFN-γ/IL-5 (interleukin 5) enzyme-linked immunospot assays as readouts for Th1-type and Th2-type CD4+ T-cell responses, respectively, we identified three E7-derived T helper epitopes (E71–12, E748–62, and E762–75), two of which are novel.

Normal donor CD4+ T cells failed to react against these E7 peptides, whereas patients with premalignant cervical intraepithelial neoplasia I-III lesions displayed preferential Th1-type responses against all three E7 epitopes. Th1-type responses were still observed to the E748–62 but not to the E71–12 and E762–75 peptides in cancer patients, where these latter two epitopes evoked Th2-type responses. Notably all responders to the E71–12 and E762–75 peptides expressed the HLA-DR4 or -DR15 alleles, whereas all responders to the E748–62 peptide failed to express the HLA-DR4 allele.

Our results are consistent with a model in which cervical cancer progression is linked to an undesirable Th1- to Th2-type shift in functional CD4+ T cell responses to two novel E7-derived epitopes. These peptides may prove important in vaccines to promote and maintain protective Th1-type antihuman papillomavirus immunity and in the immune monitoring of treated patients harboring HPV-16+ malignancies.
Suppressing Orthotopic Pancreatic Tumor Growth with a Fiber-Modified Adenovector Expressing the
TRAIL
Gene from the Human Telomerase Reverse Transcriptase Promoter
Authors: Jacob Dietmar , Davis John , Zhu Hongbo , Zhang Lidong , Teraishi Fuminori , Wu Shuhong , Marini Frank C. , Fang Bingliang
Published: September 2005

An adenoviral vector with RGD-modified fibers and expressing the human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene from the human telomerase reverse transcriptase (hTERT) promoter (designated Ad/TRAIL-F/RGD) was constructed, and its antitumor activity was evaluated in vitro and in vivo. An in vitro study showed that treatment with Ad/TRAIL-F/RGD elicited a high rate of apoptosis in human pancreatic and colon cancer cell lines that were either susceptible or resistant to conventional adenovectors. In vivo study showed that direct administration of Ad/TRAIL-F/RGD to an orthotopic implantation tumor model established in the pancreatic tails of nu/nu mice significantly suppressed tumor growth: tumors in the animals treated with Ad/TRAIL-F/RGD were approximately eight times smaller than those in animals treated with a control vector. We also evaluated hTERT promoter activity and the effect of Ad/TRAIL-F/RGD on mesenchymal stem cells. Our results showed that transgene expression from the hTERT promoter in human bone marrow mesenchymal stem cells was minimal. No adverse effect was observed in mesenchymal stem cells treated with Ad/TRAIL-F/RGD. Together, our results suggest that Ad/TRAIL-F/RGD could become a potent therapeutic agent for the management of pancreatic cancer.
Activation of theMDR1Upstream Promoter in Breast Carcinoma as a Surrogate for Metastatic Invasion
Authors: Raguz Selina , Tamburo De Bella Manuela , Tripuraneni Gopichani , Slade Martin J. , Higgins Christopher F. , Coombes R. Charles , Yagüe Ernesto
Published: September 2005

Purpose: Activation of the MDR1 upstream promoter (USP) has been described previously in four lymphoblastic leukemia patients, where it is the major MDR1 promoter associated with P-glycoprotein overexpression. We asked whether MDR1 USP-derived transcripts were also present in breast carcinoma and assessed their potential as a biomarker.

Experimental Design: We developed a sensitive method for detecting transcripts derived from the MDR1 USP and used it to identify MDR1 USP-derived transcripts in cell model systems, in 61 breast carcinoma biopsies of the primary tumor, and in isolated malignant epithelial cells both from the primary tumor and from the associated invaded lymph nodes.

Results: The MDR1 USP was not active in several independent leukemic and breast cancer cell lines or nucleated peripheral blood cells (n = 9). However, transcripts derived from the MDR1 USP were detected in some drug-resistant cell lines and a high proportion of primary breast tumors (71.6%; n = 61), whereas they were present at low frequency in normal breast tissue (10%; n = 10). Activation of MDR1 USP was not due to chromosomal amplifications or rearrangements at the MDR1 locus. Transcription from the MDR1 USP correlated with metastatic node invasion [N = 0–3 versus N > 3 (N = number of lymph nodes invaded); Fisher’s exact test, P = 0.011] and was detected in malignant epithelial cells from the primary tumor and those that metastasized to the lymph nodes.

Conclusions: MDR1 USP activation is a surrogate marker for breast carcinoma progression and can be used as a marker to study breast cancer susceptibility.
Frequent Loss of pRb2/p130 in Human Ovarian Carcinoma
Authors: D’Andrilli Giuseppina , Masciullo Valeria , Bagella Luigi , Tonini Tiziana , Minimo Corrado , Zannoni Gian Franco , Giuntoli Robert L. , Carlson John A. , Soprano Dianne Robert , Soprano Kenneth J. , Scambia Giovanni , Giordano Antonio
Published: September 2005

Purpose: RB2/p130, a member of the retinoblastoma gene family, maps to human chromosome 16q12.2, a region in which deletions have been found in several human neoplasms including breast, prostatic, and ovarian carcinoma. We sought to evaluate pRb2/p130 protein expression and function in ovarian carcinoma.

Experimental Design: pRb2/p130 expression was detected by immunohistochemical and Western blot analyses in 45 primary ovarian carcinoma samples.

Results: Immunohistochemical analysis revealed loss or decrease of pRb2/p130 expression in 18 cases (40%). pRb2/p130 expression was mostly nuclear and inversely correlated to the tumor grade (P < 0.05). Western blot analysis correlated with immunohistochemical expression. Reverse transcription-PCR followed by Southern blot analysis was performed on a representative set of 20 ovarian carcinomas. RB2/p130 mRNA levels were consistent with protein expression. We found a significant increase in the percentage of G1-phase-arrested cells in CAOV3 and A2780 ovarian carcinoma cell lines after transduction with an adenovirus carrying the RB2/p130 gene (Ad-CMV-RB2/p130).

Conclusions: These data indicate that loss or decrease of pRb2/p130 expression is a frequent event in ovarian carcinoma and is regulated mostly at the transcriptional level. Moreover, pRb2/p130 overexpression is able to arrest cell growth in ovarian carcinoma cells, suggesting the putative role of pRb2/p130 as a tumor suppressor in this malignancy.
Expression of Matrix Metalloproteinase-9 in Head and Neck Squamous Cell Carcinoma
Authors: Ruokolainen Henni , Pääkkö Paavo , Turpeenniemi-Hujanen Taina
Published: September 2005

Purpose: Previous studies have shown that matrix metalloproteinase-9 (MMP-9) is expressed in malignant head and neck squamous cell carcinoma. The prognostic role of MMP-9 is still unclear. The aim of this study was to investigate the role of MMP-9 immunoreactive protein as a prognostic marker for survival in head and neck squamous cell carcinoma.

Experimental Design: Overexpression of the immunoreactive protein for MMP-9 was evaluated in tissue sections of 74 primary head and neck carcinomas with a monoclonal antibody using a biotin-streptavidin immunohistochemical staining method. The staining results were compared with the clinical data and to the patients’ outcome.

Results: Positive immunostaining for MMP-9 was observed in 82% of the head and neck carcinomas, 39% of the cases being extensively positive. MMP-9 protein expression was independent of the stage or the grade of the tumor. The expression of MMP-9 was prognostic for shortened survival, the 5-year cause-specific survival being 45% in MMP-9 positive cases, and 92% in cases negative for MMP-9 (P = 0.013). MMP-9 positivity also correlated to the relapse-free survival (P = 0.019). At the 5-year follow-up, the cumulative relapse-free survival rate was 79% for patients with MMP-9-negative tumor and 42% for the patients with positive immunostaining for MMP-9. High expression of MMP-9 seemed to be linked with more aggressive relapses, appearing in 33% of the cases in local relapses, in 52% of cases with lymph node relapses, and in 60% of the cases with hematogenic relapses.

Conclusions: This is the first study with a long follow-up showing that the immunoreactive protein of MMP-9 in head and neck carcinoma is associated with shortened relapse-free and cause-specific survival, suggesting that MMP-9 has a role in tumor progression of head and neck carcinomas, as well as in estimation of the prognosis of these diseases.
Exon 15
BRAF
Mutations Are Uncommon in Melanomas Arising in Nonsun-Exposed Sites
Authors: Cohen Yoram , Rosenbaum Eli , Begum Shahnaz , Goldenberg David , Esche Clemens , Lavie Ofer , Sidransky David , Westra William H.
Published: September 2005

Purpose: An activating point mutation of the BRAF oncogene has been identified in a high proportion of cutaneous nevi and cutaneous melanomas, but its frequency in melanomas arising from the mucosa of head and neck is unknown.

Experimental Design: We tested 17 malignant mucosal melanomas of the head and neck for the thymine (T)→adenine (A) missense mutation at nucleotide 1796 in the BRAF gene using direct sequencing and a newly developed assay that uses a novel primer extension method (Mutector assay). We also tested 21 cutaneous melanomas, including 13 arising from sun-exposed sites and 8 from a nonsun-exposed site, the vulvar skin.

Results: The 1796T→A mutation was detected in only 1 (6%) of the sinonasal melanomas. As for cutaneous melanomas, a BRAF mutation was detected in 8 (62%) of the tumors arising in sun-exposed sites but in none (0%) of vulvar melanomas.

Conclusions: In contrast to cutaneous melanomas arising in sun-exposed sites, mucosal melanomas of the head and neck do not frequently harbor an activating mutation of BRAF. This finding additionally supports the view that the various subtypes of melanoma are not equivalent and that distinct genetic alterations may underlie well recognized differences in risk factors and behavioral patterns. Accordingly, patients with melanomas should not be collectively regarded as a uniform group as new strategies are developed that target specific genetic alterations.
DNA Aneuploidy and Integration of Human Papillomavirus Type 16 E6/E7 Oncogenes in Intraepithelial Neoplasia and Invasive Squamous Cell Carcinoma of the Cervix Uteri
Authors: Melsheimer Peter , Vinokurova Svetlana , Wentzensen Nicolas , Bastert Gunther , Doeberitz Magnus von Knebel
Published: September 2005

Purpose: Increasingly deregulated expression of the E6-E7 oncogenes of high-risk human papillomaviruses (HR-HPVs) has been identified as the major transforming factor in the pathogenesis of cervical dysplasia and derived cancers. The expression of these genes in epithelial stem cells first results in chromosomal instability and induces chromosomal aneuploidy. It is speculated that this subsequently favors integration of HR-HPV genomes into cellular chromosomes. This in turn leads to expression of viral cellular fusion transcripts and further enhanced expression of the E6-E7 oncoproteins. Chromosomal instability and aneuploidization thus seems to precede and favor integration of HR-HPV genomes.

Experimental Design: To prove this sequential concept, we analyzed here the sequence of events of DNA aneuploidization and integration in a series of HPV-16-positive cervical dysplastic lesions and carcinomas. Eighty-five punch biopsies of HPV-16-positive cervical lesions (20 CIN1/2, 50 CIN3, and 15 CxCa) were analyzed for DNA ploidy by DNA flow cytometry and for integration of HPV E6/E7 oncogenes using the amplification of papillomavirus oncogene transcripts assay, a reverse transcription-PCR method to detect integrate-derived human papillomavirus oncogene transcripts.

Results: DNA aneuploidy and viral genome integration were both associated with increasing dysplasia (P < 0.001, χ2 test for trend). In addition, DNA aneuploidy was associated with increased viral integration (P < 0.01, Fisher’s exact test). Nineteen of 20 (95%) lesions with integrated viral genomes had aneuploid cell lines; however, only 19 of 32 (59%) lesions with aneuploid cell lines had integrated viral genomes.

Conclusions: These data support the hypothesis that aneuploidization precedes integration of HR-HPV genomes in the progression of cervical dysplasia. Accordingly, deregulated viral oncogene expression appears to result first in chromosomal instability and aneuploidization and is subsequently followed by integration of HR-HPV genomes in the affected cell clones.
Changes in Gene Expression Induced by Phosphorothioate Oligodeoxynucleotides (Including G3139) in PC3 Prostate Carcinoma Cells Are Recapitulated at Least in Part by Treatment with Interferon-β and -γ
Authors: Benimetskaya Luba , Wittenberger Timo , Stein C. A. , Hofmann Hans-Peter , Weller Claudia , Lai Johnathan C. , Miller Paul , Gekeler Volker
Published: September 2005

Purpose: G3139 is an antisense bcl-2 phosphorothioate oligodeoxyribonucleotide that is currently being evaluated in Phase III clinical trials in several human cancers. The aim of the present work was to further identify the apparent non-bcl-2-dependent mechanism of this action of this compound in PC3 prostate cancer cells.

Experimental Design: We performed Affymetrix U95A oligonucleotide microarray studies on mRNA isolated from cells treated with G3139 and related oligonucleotides.

Results: Hierarchical clustering revealed the presence of a set of genes of which the expression was elevated on both 1 and 3 days after oligonucleotide treatment. Significantly, the persistence of expression of the up-regulation of these genes, many of which are members of the IFN cascade, was greater for G3139 than for any other oligomer evaluated. Furthermore, many of the genes with the greatest up-regulation of expression are also those of which the expression is up-regulated after treatment of cells with IFNs. Treatment of PC3 cells with either IFN-β or -γ recapitulated some of the aspects of the molecular and phenotypic changes observed after treatment with a G3139/Lipofectin complex. These include down-regulation of bcl-2 protein expression itself, down-regulation of protein kinase C α protein expression (but not that of other protein kinase C isoforms), alteration in p21/Waf1/Cip1 protein expression, up-regulation of MHC-I cell surface expression, and profound suppression of cell growth in the absence of a notable increase in cellular apoptosis. However, G3139 (when complexed with Lipofectin) did not induce the up-regulation of expression of either type I or type II IFNs, nor could IFNs be found in conditioned media from treated cells.

Conclusions: Oligonucleotide microarray experiments demonstrated that G3139 could induce elements of the IFN cascade in PC3 cells in vitro. In addition, the cellular phenotype obtained after treatment with exogenous IFN could, at least in part, recapitulate that obtained after G3139 treatment. Nevertheless, the oligonucleotide microarray experiments we performed also demonstrated that there are extremely large qualitative and quantitative differences between the two treatments.
Extended Follow-up of Patients Treated with Imatinib Mesylate (Gleevec) for Chronic Myelogenous Leukemia Relapse after Allogeneic Transplantation
Authors: DeAngelo Daniel J. , Hochberg Ephraim P. , Alyea Edwin P. , Longtine Janina , Lee Stephanie , Galinsky Ilene , Parekkedon Ben , Ritz Jerome , Antin Joseph H. , Stone Richard M. , Soiffer Robert J.
Published: September 2005

Purpose: Over the last several years, donor lymphocyte infusions have become the standard approach for patients with chronic myelogenous leukemia (CML) who relapse after allogeneic stem cell transplantation (SCT). Recent reports indicate that imatinib mesylate (Gleevec) can induce remissions in these patients as well. Less is known about the extent and durability of these responses.

Experimental Design: We studied 15 patients treated with imatinib for recurrent CML after SCT, 10 patients with stable phase CML (SP-CML), 1 with accelerated phase (AP-CML), and 4 with blast phase (BP-CML). The dose of imatinib was 600 mg (n = 10) or 400 mg (n = 5) daily. Patients were followed for hematological, cytogenetic, and molecular response. A susbset of responders was followed for changes in donor-derived hematopoietic chimerism.

Results: Of the 10 patients with SP-CML, all achieved a hematological response. Within 3 months, five of these patients had achieved a complete cytogenetic response (CCR). By six months, 9 of 10 patients had achieved CCR. The BCR-ABL transcript could not be detected by reverse transcription-PCR in 7 of these 9 patients. Patients who achieved CCR showed evidence of conversion to complete donor chimerism. No patient developed graft-versus-host disease. With a median follow up of 25 months, all patients are alive and 9 of 10 patients remain in CCR. Only one of the 5 patients with AP/BP-CML achieved a complete cytogenetic response.

Conclusions: We find that imatinib is well tolerated in patients with SP-CML who relapse after SCT. Responses were rapid, durable, and associated with conversion to full donor chimerism without graft-versus-host disease. Imantinib was far less effective in patients who relapsed with AP/BP-CML. Imatinib should be evaluated as either an alternative or as an adjunct to donor lymphocyte infusions for patients with SP-CML who relapse after SCT.
p53
Polymorphism and Age of Onset of Hereditary Nonpolyposis Colorectal Cancer in a Caucasian Population
Authors: Jones J. Shawn , Chi Xuedong , Gu Xiangjun , Lynch Patrick M. , Amos Christopher I. , Frazier Marsha L.
Published: September 2005

Purpose: Hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal dominant syndrome of familial malignancies. Colorectal and endometrial cancers are most frequently observed. The syndrome results mainly from germ-line mutations in DNA mismatch repair genes. A common G-to-C polymorphism at codon 72 in the p53 gene has been associated with increased risk for lung, nasopharyngeal, oral, prostate, and breast cancers and may be a marker for genetic susceptibility to colorectal cancer. We studied the influence of this p53 polymorphism on HNPCC age of onset.

Experimental Design: We determined the p53 genotype of 92 Caucasian mismatch repair mutation carriers, of which, 47 had colorectal cancer. The subjects were genotyped by single-strand conformational polymorphism analysis. We tested the association between age of onset and the p53 genotypes by comparing Kaplan-Meier survival curves, evaluating the homogeneity of the curves using the log-rank test and Wilcoxon’s test, and estimating the association using the Cox proportional hazards regression model to adjust for potential demographic confounding factors.

Results: The HNPCC patients who were heterozygous developed their colorectal cancer 13 years earlier than HNPCC patients who were homozygous for the wild-type allele.

Conclusions: Combining knowledge of an individual’s p53 genotype with information on other genetic and environmental risk factors may improve risk estimates and help to identify individuals who are genetically susceptible to developing HNPCC at an earlier age.
Flat-Fixed Dosing of Irinotecan
Authors: de Jong Floris A. , Mathijssen Ron H. J. , Xie Rujia , Verweij Jaap , Sparreboom Alex
Published: September 2005

Purpose: In a previous analysis, it was shown that body-surface area (BSA) is not a predictor of irinotecan pharmacokinetic parameters. Here, we prospectively evaluated the effects of administering a flat-fixed irinotecan dose to cancer patients, regardless of BSA.

Experimental Design: Twenty-six cancer patients (12 females) received a fixed irinotecan dose of 600 mg, given as a 90-min i.v. infusion. Plasma concentrations of irinotecan and its metabolites SN-38 (7-ethyl-10-hydroxycamptothecin) and SN-38G (SN-38 glucuronide) were measured during the first cycle and analyzed using nonlinear mixed-effect modeling. Data were compared with those obtained in 47 cancer patients (19 females) who received irinotecan at a BSA-normalized dose of 350 mg/m2.

Results: The interindividual variability in irinotecan clearance (25.9% versus 25.1%; P = 0.93), in relative extent of conversion to SN-38 (47.8% versus 42.7%; P = 0.24), and in relative extent of SN-38 glucuronidation (71.2% versus 72.4%; P = 0.95) were not significantly different between the two dose groups. Variance differences in irinotecan-mediated hematological side effects were also similar between the 600 mg and 350 mg/m2 groups (P > 0.14).

Conclusions: These findings suggest that flat-fixed dosing of irinotecan does not result in increased pharmacokinetic/pharmacodynamic variability and could be safely used to supplant current dosing strategies based on BSA.