bluuebunny/crossref_metadata_2025 huggingface dataset:
Selective Modulation of the Therapeutic Efficacy of Anticancer Drugs by Selenium Containing Compounds against Human Tumor Xenografts
Authors: Cao Shousong , Durrani Farukh A. , Rustum Youcef M.
Published: September 2005

Purpose: Studies were carried out in athymic nude mice bearing human squamous cell carcinoma of the head and neck (FaDu and A253) and colon carcinoma (HCT-8 and HT-29) xenografts to evaluate the potential role of selenium-containing compounds as selective modulators of the toxicity and antitumor activity of selected anticancer drugs with particular emphasis on irinotecan, a topoisomerase I poison.

Experimental Design: Antitumor activity and toxicity were evaluated using nontoxic doses (0.2 mg/mouse/day) and schedule (14–28 days) of the selenium-containing compounds, 5-methylselenocysteine and seleno-l-methionine, administered orally to nude mice daily for 7 days before i.v. administration of anticancer drugs, with continued selenium treatment for 7–21 days, depending on anticancer drugs under evaluation. Several doses of anticancer drugs were used, including the maximum tolerated dose (MTD) and toxic doses. Although many chemotherapeutic agents were evaluated for toxicity protection by selenium, data on antitumor activity were primarily obtained using the MTD, 2 × MTD, and 3 × MTD of weekly ×4 schedule of irinotecan.

Results: Selenium was highly protective against toxicity induced by a variety of chemotherapeutic agents. Furthermore, selenium increased significantly the cure rate of xenografts bearing human tumors that are sensitive (HCT-8 and FaDu) and resistant (HT-29 and A253) to irinotecan. The high cure rate (100%) was achieved in nude mice bearing HCT-8 and FaDu xenografts treated with the MTD of irinotecan (100 mg/kg/week × 4) when combined with selenium. Administration of higher doses of irinotecan (200 and 300 mg/kg/week × 4) was required to achieve high cure rate for HT-29 and A253 xenografts. Administration of these higher doses was possible due to selective protection of normal tissues by selenium. Thus, the use of selenium as selective modulator of the therapeutic efficacy of anticancer drugs is new and novel.

Conclusions: We demonstrated that selenium is a highly effective modulator of the therapeutic efficacy and selectivity of anticancer drugs in nude mice bearing human tumor xenografts of colon carcinoma and squamous cell carcinoma of the head and neck. The observed in vivo synergic interaction is highly dependent on the schedule of selenium.
Zoledronic Acid Inhibits Visceral Metastases in the 4T1/luc Mouse Breast Cancer Model
Authors: Hiraga Toru , Williams Paul J. , Ueda Akimi , Tamura Daisuke , Yoneda Toshiyuki
Published: September 2005

Purpose: It is established that bisphosphonates (BPs), specific inhibitors of osteoclasts, have beneficial effects on bone metastases of breast cancer. In addition, recent studies have reported that BPs have anticancer effects and suppress visceral metastases, too. However, the results of clinical studies are still conflicting. In the present study, we examined the effects of the BP zoledronic acid (ZOL), one of the most potent BPs currently available, on visceral metastases of breast cancer using an animal model in which mouse breast cancer cells 4T1/luc implanted at the orthotopic mammary fat pad spontaneously metastasize to multiple organs including bone, lung, and liver in female BALB/c mice.

Experimental Design and Results: The 4T1/luc-bearing mice received single or four i.v. injections of ZOL (0.5 or 5 μg/mouse) during the whole experimental period. Bone metastases were reduced by the ZOL treatment. More importantly, ZOL significantly suppressed lung and liver metastases. Furthermore, ZOL prolonged overall survival of the tumor-bearing mice. Of interest, apoptosis in 4T1/luc cells colonized in bone was increased by ZOL; however, those in lung were not changed. In vitro studies demonstrated that ZOL inhibited cell migration and invasion and promoted apoptosis of 4T1/luc cells.

Conclusions: These results are consistent with the notion that ZOL affects breast cancer metastasis to visceral organs as well as bone. These effects of ZOL may be attributable to inhibition of migration and invasion of breast cancer cells. Clinical relevance of our experimental results needs to be determined in breast cancer patients with visceral metastases.
Gene Expression Profiling of Early- and Late-Relapse Nonseminomatous Germ Cell Tumor and Primitive Neuroectodermal Tumor of the Testis
Authors: Sugimura Jun , Foster Richard S. , Cummings Oscar W. , Kort Eric J. , Takahashi Masayuki , Lavery Todd T. , Furge Kyle A. , Einhorn Lawrence H. , Teh Bin Tean
Published: September 2005

Purpose: To better understand the molecular mechanisms that underlay the development and progression of nonseminomatous germ cell tumor of testis (NSGCTT) as well as malignant transformation of teratoma and primitive neuroectodermal tumor (PNET).

Experimental Design: We studied the gene expression profiles of 17 retroperitoneal NSGCTTs (10 yolk sac tumors, 3 embryonal carcinomas, 4 teratomas) and 2 PNETs obtained from patients with two clinical outcomes. Tissue samples were obtained from the Indiana University. One group of NSGCTT and PNET patients developed metastases within 2 years (early-relapse) of initial successful treatment, and the other group developed metastases after 2 years (late-relapse). Gene expression in these groups of patients was quantified using cDNA microarrays and real-time relative quantitative PCR.

Results: We demonstrate that the gene expression profiles of these tumors correlate with histological type. In addition, we identify type-specific genes that may serve as novel diagnostic markers. We also identify a gene set that can distinguish between early-relapse and late-relapse yolk sac tumors. The expression differences of these genes may underlie the differences in clinical outcome and drug response of these tumors.

Conclusion: This is the first study that used gene expression profiling to examine the molecular characteristics of the NSGCTTs and drug response in early- and late-relapse tumors. These results suggest that two molecularly distinct forms of NSGCTTs exist and that the integration of expression profile data with clinical parameters could enhance the diagnosis and prognosis of NSGCTTs. More importantly, the identified genes provide insight into the molecular mechanisms of aggressive NSGCTTs and suggest intervention strategies.
First 18F-Labeled Tracer Suitable for Routine Clinical Imaging of sst Receptor-Expressing Tumors Using Positron Emission Tomography
Authors: Schottelius Margret , Poethko Thorsten , Herz Michael , Reubi Jean-Claude , Kessler Horst , Schwaiger Markus , Wester Hans-Jürgen
Published: September 2005

Purpose: Despite excellent radionuclide characteristics, no 18F-labeled peptides are available for quantitative peptide receptor mapping using positron emission tomography (PET) so far, mainly due to time-consuming multistep radiosyntheses with limited overall yields. A newly developed two-step chemoselective conjugation method allows rapid and high-yield [18F]fluorination of peptides via oxime formation and was applied for the synthesis of new 18F-labeled carbohydrated Tyr3-octreotate (TOCA) analogs with optimized pharmacokinetics suitable for clinical routine somatostatin-receptor (sst) imaging.

Experimental Design: 18F-labeled glucose (Gluc-S-) and cellobiose (Cel-S-) derivatives of aminooxy-functionalized TOCA were synthesized via oxime formation with 4-[18F]fluorobenzaldehyde ([18F]FBOA-peptides). Both the in vitro internalization profile of Gluc-S-Dpr([18F]FBOA)TOCA and Cel-S-Dpr([18F]FBOA)TOCA in hsst2-expressing Chinese hamster ovary cells (dual tracer protocol) and their biodistribution in AR42J tumor-bearing mice were investigated and compared with two [18F]fluoropropionylated ([18F]FP) analogs, Gluc-Lys([18F]FP)TOCA and Gluc-S-Dpr([18F]FP)TOCA.

Results: In contrast to [18F]FP-labeling (3 h), chemo-selective [18F]FBOA-formation (50 min) afforded the respective radiopeptides in high yields (65–85%). In vitro, Gluc-S-Dpr([18F]FBOA)TOCA and Cel-S-Dpr([18F]FBOA)-TOCA showed high internalization (139 ± 2 and 163 ± 8 of the reference [125I]Tyr3-octreotide, respectively), which was reflected by high tumor accumulation in vivo [21.8 ± 1.4 and 24.0 ± 2.5% of injected dose/g (1 h), respectively]. How-ever, only Cel-S-Dpr([18F]FBOA)TOCA and Gluc-S-Dpr([18F]FP)TOCA (tumor: 15.1 ± 1.5% of injected dose/g) with its very low accumulation in all of the nontarget organs showed improved tumor:organ ratios compared with Gluc-Lys([18F]FP)TOCA. For Cel-S-Dpr([18F]FBOA)TOCA,tumor:organ ratios (1 h) were 42:1, 27:1, 15:1, 3:1, and 208:1 for blood, liver, intestine, kidney, and muscle, respectively.

Conclusion: Due to the fast and high-yield chemoselective radiofluorination strategy and to its excellent pharmacokinetics, Cel-S-Dpr([18F]FBOA)TOCA represents the first tracer suitable for routine clinical application in PET somatostatin receptor imaging.
Characterization of p53 Wild-Type and Null Isogenic Colorectal Cancer Cell Lines Resistant to 5-Fluorouracil, Oxaliplatin, and Irinotecan
Authors: Boyer John , McLean Estelle G. , Aroori Somaiah , Wilson Peter , McCulla Andrea , Carey P. Declan , Longley Daniel B. , Johnston Patrick G.
Published: September 2005

To elucidate mechanisms of resistance to chemotherapies currently used in the first-line treatment of advanced colorectal cancer, we have developed a panel of HCT116 p53 wild-type (p53+/+) and null (p53−/−) isogenic colorectal cancer cell lines resistant to the antimetabolite 5-fluorouracil (5-FU), topoisomerase I inhibitor irinotecan (CPT-11), and DNA-damaging agent oxaliplatin. These cell lines were generated by repeated exposure to stepwise increasing concentrations of each drug over a period of several months. We have demonstrated a significant decrease in sensitivity to 5-FU, CPT-11, and oxaliplatin in each respective resistant cell line relative to the parental line as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analysis, with increases in IC50 (72 h) concentrations ranging from 3- to 65-fold. Using flow cytometry, we have also demonstrated compromised apoptosis and cell cycle arrest in 5-FU-, oxaliplatin-, and CPT-11-resistant cell lines compared with the parental lines after exposure to each drug. In addition, we found that resistance to 5-FU and oxaliplatin was higher in parental p53−/− cells compared with parental p53+/+ cells, with an ∼5-fold increase in IC50 (72 h) for each drug. In contrast, the IC50 (72 h) doses for CPT-11 were identical in the p53 wild-type and null cell lines. Furthermore, apoptosis after treatment with 5-FU and oxaliplatin, but not CPT-11, was significantly reduced in parental p53−/− cells compared with parental p53+/+ cells. These data suggest that p53 may be an important determinant of sensitivity to 5-FU and oxaliplatin but not CPT-11. Using semiquantitative reverse transcription-PCR, we have demonstrated down-regulation of thymidine phosphorylase mRNA in both p53+/+ and p53−/− 5-FU-resistant cells, suggesting that decreased production of 5-FU active metabolites may be an important resistance mechanism in these lines. In oxaliplatin-resistant cells, we noted increased mRNA levels of the nucleotide excision repair gene ERCC1 and ATP-binding cassette transporter breast cancer resistance protein. In CPT-11-resistant cells, we found reduced mRNA levels of carboxylesterase, the enzyme responsible for converting CPT-11 to its active metabolite SN-38, and topoisomerase I, the SN-38 target enzyme. In addition, we noted overexpression of breast cancer resistance protein in the CPT-11-resistant lines. These cell lines are ideal tools with which to identify novel determinants of drug resistance in both the presence and absence of wild-type p53.
Osteopontin as an Adjunct to CA125 in Detecting Recurrent Ovarian Cancer
Authors: Schorge John O. , Drake Richard D. , Lee Hang , Skates Steven J. , Rajanbabu Ramababu , Miller David S. , Kim Jae-Hoon , Cramer Daniel W. , Berkowitz Ross S. , Mok Samuel C.
Published: September 2005

Purpose: CA125 is currently the only tumor marker to have a validated role in the postoperative monitoring of ovarian cancer. Osteopontin (OPN) is a putative plasma biomarker that was recently identified using high-throughput cDNA microarray technology. The purpose of this study was to test the hypothesis that OPN is a clinically useful adjunct to CA125 in detecting recurrent ovarian cancer.

Experimental Design: Thirty-eight ovarian cancer patients had a single pretreatment blood sample and 200 postoperative specimens were prospectively collected during chemotherapy and follow-up. OPN measurements were performed using an enzyme-linked immunoassay, and CA125 levels were concurrently obtained. Wilcoxon’s signed rank-sum test was used to perform paired comparisons between pretreatment and postoperative OPN and CA125 measurements. Longitudinal mixed effects polynomial models were used to determine whether OPN and CA125 levels correlated with the development of recurrent ovarian cancer.

Results: The median pretreatment OPN level was 178 ng/ml (range, 12–3468) and the median CA125 measurement was 812 units/ml (range, 12–81,500). There was a trend for OPN levels to decline after treatment was initiated (P = 0.07), but decreasing CA125 measurements were more consistently observed (P = 0.0009). The quadratic functional trends of OPN and CA125 were each highly significant (P < 0.0001). Although inferior to CA125 in predicting clinical response to therapy, OPN rose earlier in 90% (95% confidence interval, 56–100%) of the patients developing recurrent disease (median lead time, 3 months).

Conclusions: OPN may be a clinically useful adjunct to CA125 in detecting recurrent ovarian cancer.
Expression of Thyrotropin-Releasing Hormone by Human Melanoma and Nevi
Authors: Ellerhorst Julie A. , Naderi Aresu A. , Johnson Marilyn K. , Pelletier Pauline , Prieto Victor G. , Diwan A. Hafeez , Johnson Marcella M. , Gunn Debra C. , Yekell Sandra , Grimm Elizabeth A.
Published: September 2005

Purpose: Thyrotropin-releasing hormone (TRH) is a tripeptide hormone produced by the hypothalamus in response to hypothyroidism. RNA transcripts for the TRH prohormone have recently been described in melanoma cell lines. To expand these findings, we have examined cultured melanoma cells and melanocytes, human melanoma tumors, and nevi for the expression of TRH.

Experimental Design: Five melanoma cell lines were analyzed by reverse transcription-PCR/Southern blotting for preproTRH message. The same melanoma lines and two melanocyte lines were examined by immunocytochemistry for TRH protein expression and for growth response to exogenous TRH. Immunohistochemistry was used to test for TRH protein in sections of 19 melanomas, 33 dysplastic nevi, and 27 benign nevi.

Results: TRH message and protein were detected in all melanoma cell lines examined. Melanocytes were also found to express TRH protein. Four of the five melanoma cell lines but neither melanocyte line responded with a increase in proliferation to low concentrations of exogenous TRH. TRH immunoreactivity was observed in 12 of 19 melanomas (63%), 23 of 33 (69.7%) dysplastic nevi, and 14 of 27 (51.9%) benign nevi. Expression in dysplastic nevi was significantly greater than in benign nevi. Upon separate analysis of nevi from melanoma patients, the difference between dysplastic and benign nevi was even more significant. However, in healthy individuals, no difference between dysplastic and benign nevi was observed. Furthermore, dysplastic nevi from melanoma patients had a significantly higher percentage of TRH-positive cells when compared with healthy individuals.

Conclusions: TRH is commonly expressed by melanomas and dysplastic nevi and may function as a melanoma autocrine growth factor. The presence of TRH in dysplastic nevi may be predictive for the development of melanoma. Our findings have significant clinical and biological implications for future research into the early stages of melanoma initiation and progression.
Clinical Trial Substantiates the Predictive Value of
O-6-Methylguanine-DNA Methyltransferase
Promoter Methylation in Glioblastoma Patients Treated with Temozolomide
Authors: Hegi Monika E. , Diserens Annie-Claire , Godard Sophie , Dietrich Pierre-Yves , Regli Luca , Ostermann Sandrine , Otten Philippe , Van Melle Guy , de Tribolet Nicolas , Stupp Roger
Published: September 2005

Purpose: In the setting of a prospective clinical trial, we determined the predictive value of the methylation status of the O-6-methylguanine-DNA methyltransferase (MGMT) promoter for outcome in glioblastoma patients treated with the alkylating agent temozolomide. Expression of this excision repair enzyme has been associated with resistance to alkylating chemotherapy.

Experimental Design: The methylation status of MGMT in the tumor biopsies was evaluated in 38 patients undergoing resection for newly diagnosed glioblastoma and enrolled in a Phase II trial testing concomitant and adjuvant temozolomide and radiation. The epigenetic silencing of the MGMT gene was determined using methylation-specific PCR.

Results: Inactivation of the MGMT gene by promoter methylation was associated with longer survival (P = 0.0051; Log-rank test). At 18 months, survival was 62% (16 of 26) for patients testing positive for a methylated MGMT promoter but reached only 8% (1 of 12) in absence of methylation (P = 0.002; Fisher’s exact test). In the presence of other clinically relevant factors, methylation of the MGMT promoter remains the only significant predictor (P = 0.017; Cox regression).

Conclusions: This prospective clinical trial identifies MGMT-methylation status as an independent predictor for glioblastoma patients treated with a methylating agent. The association of the epigenetic inactivation of the DNA repair gene MGMT with better outcome in this homogenous cohort may have important implications for the design of future trials and supports efforts to deplete MGMT by O-6-benzylguanine, a noncytotoxic substrate of this enzyme.
In vitro
and
In vivo
Activity of the Nuclear Factor-κB Inhibitor Sulfasalazine in Human Glioblastomas
Authors: Robe Pierre A. , Bentires-Alj Mohamed , Bonif Marianne , Rogister Bernard , Deprez Manuel , Haddada Heddi , Khac Minh-Tuan Nguyen , Jolois Olivier , Erkmen Kadir , Merville Marie-Paule , Black Peter M. , Bours Vincent
Published: September 2005

Glioblastomas, the most common primary brain cancers, respond poorly to current treatment modalities and carry a dismal prognosis. In this study, we demonstrated that the transcription factor nuclear factor (NF)-κB is constitutively activated in glioblastoma surgical samples, primary cultures, and cell lines and promotes their growth and survival. Sulfasalazine, an anti-inflammatory drug that specifically inhibits the activation of NF-κB, blocked the cell cycle and induced apoptosis in several glioblastoma cell lines and primary cultures, as did gene therapy with a vector encoding a super-repressor of NF-κB. In vivo, sulfasalazine also significantly inhibited the growth of experimental human glioblastomas in nude mice brains. Given the documented safety of sulfasalazine in humans, these results may lead the way to a new class of glioma treatment.
Amplification of the BRCA2 Pathway Gene
EMSY
in Sporadic Breast Cancer Is Related to Negative Outcome
Authors: Rodriguez Carmen , Hughes-Davies Luke , Vallès Hélène , Orsetti Béatrice , Cuny Marguerite , Ursule Lisa , Kouzarides Tony , Theillet Charles
Published: September 2005

DNA amplification at band q13 of chromosome 11 is common in breast cancer, and CCND1 and EMS1 remain the strongest candidate genes. However, amplification patterns are consistent with the existence of four cores of amplification, suggesting the involvement of additional genes. Here we present evidence strongly suggesting the involvement of the recently characterized EMSY gene in the formation of the telomeric amplicon. EMSY maps at 11q13.5, 100 kb centromeric to the GARP gene, which has been mapped within the core of the distal amplicon. The EMSY protein was shown to interact with BRCA2 and has a role in chromatin remodeling. This makes EMSY a strong candidate oncogene for the 11q13.5 amplicon. DNA amplification was studied in a total of 940 primary breast tumors and 39 breast cancer cell lines. Amplification profiles were consistent with the EMSY-GARP locus being amplified independently of CCND1 and/or EMS1. EMSY RNA expression levels were studied along with those of five other genes located at 11q13.5 by real-time quantitative PCR in the 39 cell lines and a subset of 65 tumors. EMSY overexpression correlated strongly with DNA amplification in both primary tumors and cell lines. In a subset of 296 patients, EMSY amplification was found by both uni- and multivariate analyses to correlate with shortened disease-free survival. These data indicate that EMSY is a strong candidate oncogene for the 11q13.5 amplicon.
Antisense Protein Kinase A RIα Inhibits 7,12-Dimethylbenz(
a
)anthracene-Induction of Mammary Cancer
Authors: Nesterova Maria V. , Cho-Chung Yoon S.
Published: September 2005

Purpose: There are two types of cyclic AMP (cAMP)-dependent protein kinase (PKA), type I (PKA-I) and type II (PKA-II), which share a common catalytic (C) subunit but contain distinct regulatory (R) subunits, RI versus RII, respectively. Evidence suggests that increased expression of PKA-I and its regulatory subunit (RIα) correlates with tumorigenesis and tumor growth. We investigated the effect of sequence-specific inhibition of RIα gene expression at the initial phase of 7,12-dimethylbenz(αa)anthracene (DMBA)-induced mammary carcinogenesis.

Experimental Design: Antisense RIα oligodeoxynucleotide (ODN) targeted against PKA RIα was administered (0.1 mg/day/rat, i.p.) 1 day before DMBA intubation and during the first 9 days post-DMBA intubation to determine the anticarcinogenic effects.

Results: Antisense RIα, in a sequence-specific manner, inhibited the tumor production. At 90 days after DMBA intubation, untreated controls and RIα-antisense-treated rats exhibited an average mean number of tumors per rat of 4.2 and 1.8, respectively, and 90% of control and 45% of antisense-treated animals had tumors. The antisense also delayed the first tumor appearance. An increase in RIα and PKA-I levels in the mammary gland and liver preceded DMBA-induced tumor production, and antisense down-regulation of RIα restored normal levels of PKA-I and PKA-II in these tissues. Antisense RIα in the liver induced the phase II enzymes, glutathione S-transferase and quinone oxidoreductase, c-fos protein, and activator protein 1 (AP-1)- and cAMP response element (CRE)-directed transcription. In the mammary glands, antisense RIα promoted DNA repair processes. In contrast, the CRE transcription-factor decoy could not mimic these effects of antisense RIα.

Conclusions: The results demonstrate that RIα antisense produces dual anticarcinogenic effects: (a) increasing DMBA detoxification in the liver by increasing phase II enzyme activities, increasing CRE-binding-protein phosphorylation and enhancing CRE- and Ap-1-directed transcription; and (b) activating DNA repair processes in the mammary gland by down-regulating PKA-I.
Epidermal Growth Factor Receptor, c-MET, β-Catenin, and p53 Expression as Prognostic Indicators in Stage II Colon Cancer
Authors: Resnick Murray B. , Routhier Justin , Konkin Tamako , Sabo Edmond , Pricolo Victor E.
Published: September 2005

Purpose: Through the use of molecular markers, it may be possible to identify aggressive tumor phenotypes and tailor therapies to treat them. This approach would be particularly useful for stage II colon cancer. The purpose of this study was to define the prognostic value of epidermal growth factor receptor (EGFR), c-MET, β-catenin, and p53 protein expression in TNM stage II colon cancer using tissue microarray technology.

Experimental Design: In this study, we retrospectively analyzed, resected, and otherwise untreated paraffin-embedded specimens from 134 consecutive patients with Tumor-Node-Metastasis stage II colonic carcinomas for EGFR, c-MET, β-catenin, and p53 protein expression by immunohistochemistry.

Results: Thirty-five percent, 77, and 65% of tumors exhibited strong (+2 and +3 immunopositivity) expression of EGFR, c-MET, and β-catenin, respectively. Fifty-four percent exhibited nuclear staining for p53 in >10% of the tumor cells. Univariate analysis revealed that increased nuclear p53 expression (P = 0.001), strong membranous EGFR expression (P = 0.04), and lymphovascular invasion (P = 0.01) were significantly related to disease recurrence and that p53 (P = 0.02) and EGFR (P = 0.05) expression were associated with decreased survival. Increased nuclear p53 expression also correlated with the presence of distal metastasis (P = 0.027). No significant association was seen between c-MET expression and prognosis, whereas a strong trend was detected between loss of β-catenin (P = 0.065) expression and poor outcome. Multivariate analysis indicated that p53 (P = 0.04), EGFR (P = 0.05), and lymphovascular invasion (P = 0.03) were independent predictors of recurrence and that p53 (P = 0.02) and EGFR (P = 0.01) expression were both associated with poor survival.

Conclusions: This retrospective tumor microarray study, restricted to Tumor-Node-Metastasis stage II colon cancer patients who did not undergo adjuvant therapy, supports the use of EGFR and p53 as biological markers, which may assist in predicting disease recurrence and survival.
In vitro
Toxicity of A-431 Carcinoma Cells with Antibodies to Epidermal Growth Factor Receptor and Epithelial Glycoprotein-1 Conjugated to Radionuclides Emitting Low-Energy Electrons
Authors: Michel Rosana B. , Castillo Mary Ellen , Andrews Philip M. , Mattes M. Jules
Published: September 2005

Purpose: The ability of antibodies (Abs) conjugated to radionuclides emitting low-energy electrons to specifically kill nonadherent lymphoma target cells in vitro was demonstrated previously. This study extends this work to adherent carcinoma cells. The fact that these cells are spread out on plastic can potentially make it more difficult to deliver radiation to the nucleus from decays in the cytoplasm or on the cell surface.

Experimental Design: The Abs tested were anti-epidermal growth factor receptor and anti-epithelial glycoprotein-1, conjugated to indium-111 or iodine-125, which emit low-energy Auger and conversion electrons. Conjugates of the β-particle emitter, iodine-131, also were tested, for comparison. Abs were incubated with the cells for 2 days, and then the treated cells were assayed for colony-forming units. The radiation dose delivered to the nucleus was calculated from the cumulative decays per cell.

Results: With conjugates of 111In, very potent killing was obtained with both of the Abs, with 100% kill (approximately 4–5 logs) even at subsaturating Ab concentrations. Lower levels of kill were obtained with 125I or 131I conjugates. Conjugates with 131I, a β-particle emitter, produced greater nonspecific toxicity. The greater potency of 111In could be attributed to the higher specific activity that was obtained routinely with this radiolabel, up to 70 mCi/mg. Uptake of radioactivity peaked at approximately 200 cpm per cell. Dosimetry calculations, using subcellular S values, demonstrated that the toxicity observed was consistent with the amount of radiation delivered to the nucleus.

Conclusions: These results are similar to previous results obtained with B lymphoma cells and indicate that this approach is applicable to a wide range of tumor types. Radionuclides emitting low-energy electrons are effective at killing target cells with relatively little nonspecific toxicity, if sufficient activity is delivered to the cell. Most Abs to high-density cell surface antigens would probably be effective.
Overexpressed eIF4E Is Functionally Active in Surgical Margins of Head and Neck Cancer Patients via Activation of the Akt/Mammalian Target of Rapamycin Pathway
Authors: Nathan Cherie-Ann O. , Amirghahari Nazanin , Abreo Fleurette , Rong Xiaohua , Caldito Gloria , Jones M. Lamar , Zhou Huijuan , Smith Melanie , Kimberly Donnellan , Glass Jonathan
Published: September 2005

Purpose: Overexpression of eIF4E in surgical margins of head and neck cancer patients is an independent risk factor for recurrence. We hypothesize that overexpressed eIF4E is functionally active in tumor margins through activation of the Akt/mammalian target of rapamycin (mTOR) pathway

Experimental Design: Western blots and/or immunohistochemistry were performed to determine whether phosphorylation of mTOR and activation of its downstream molecules eIF4E-binding protein-1 (4E-BP1) and p70 S6 kinase and the upstream modulator of mTOR, Akt, were expressed in margins overexpressing eIF4E.

Results: There was a significant association between phospho-4E-BP1 and eIF4E expression of a margin or a significant difference in phospho-4E-BP1 expression between the eIF4E-positive and -negative margins (P < 0.01). A significant association between eIF4E and phospho-p70 S6 kinase as well as eIF4E and phospho-mTOR was also noted (P < 0.05). Western blot analysis indicated a highly significant difference in the phosphorylation status of 4E-BP1 between tumors and resection margins. A total of 89% of the 4E-BP1-expressing margins expressed more of the phosphorylated (β, γ, and δ) isoforms, whereas 81% of the 4E-BP1-expressing tumors expressed more of the unphosphorylated α isoform. A similar difference in Akt activation was noted between eIF4E-positive margins and tumors (P < 0.05).

Conclusions: Overexpression of eIF4E is functionally active in tumor margins through activation of the Akt/mTOR signaling pathway. The greater degree of expression of downstream targets and upstream regulators of mTOR in margins compared with the tumors indicates preferential activation of the Akt/mTOR signaling pathway in margins overexpressing eIF4E. Rapamycin analogs can potentially be used as adjuvant therapy for patients with eIF4E-positive margins.
Nuclear Localization of KLF4 Is Associated with an Aggressive Phenotype in Early-Stage Breast Cancer
Authors: Pandya Ashka Y. , Talley Lynya I. , Frost Andra R. , Fitzgerald Thomas J. , Trivedi Vivek , Chakravarthy Mithun , Chhieng David C. , Grizzle William E. , Engler Jeffrey A. , Krontiras Helen , Bland Kirby I. , LoBuglio Albert F. , Lobo-Ruppert Susan M. , Ruppert J. Michael
Published: September 2005

Purpose: The Krüppel-like transcription factor KLF4/GKLF induces both malignant transformation and a slow-growth phenotype in vitro. Although KLF4 expression is increased in most cases of breast cancer, it was unknown whether these cases represent a distinct subtype with a different clinical outcome.

Experimental Design: We examined expression of KLF4 by immunostaining 146 cases of human primary infiltrating ductal carcinoma of the breast. Staining patterns were correlated with clinical outcome and with established prognostic factors.

Results: Subcellular localization exhibited case-to-case variation. Tumors with high nuclear staining and low cytoplasmic staining were termed type 1. For patients with early-stage disease (i.e., stage I or IIA), type 1 staining was associated with eventual death because of breast cancer (hazard ratio, 2.8; 95% confidence interval, 1.23–6.58; P = 0.011). The association was stronger in patients with early-stage cancer and small primary tumors (i.e., ≤2.0 cm in diameter; hazard ratio, 4.3; 95% confidence interval, 1.75–10.62; P < 0.001). For patients with early-stage disease, multivariate analysis indicated that type 1 staining was independently associated with outcome (adjusted hazard ratio 2.6; 95% confidence interval, 1.10–6.05; P = 0.029). Type 1 staining was also associated with high histological grade (P = 0.032), increased expression of Ki67 (P = 0.016), and reduced expression of BCL2 (P = 0.032). In vitro, KLF4 was localized within the nucleus of transformed RK3E epithelial cells, consistent with a nuclear function of this transcription factor during induction of malignant transformation.

Conclusions: The results suggest that localization of KLF4 in the nucleus of breast cancer cells is a prognostic factor and identify KLF4 as a marker of an aggressive phenotype in early-stage infiltrating ductal carcinoma.
Decreased Expression of 14-3-3σ Is Associated with Advanced Disease in Human Epithelial Ovarian Cancer
Authors: Akahira Jun-ichi , Sugihashi Youko , Suzuki Takashi , Ito Kiyoshi , Niikura Hitoshi , Moriya Takuya , Nitta Makoto , Okamura Hitoshi , Inoue Satoshi , Sasano Hironobu , Okamura Kunihiro , Yaegashi Nobuo
Published: September 2005

Purpose: In this study, we examined the promoter methylation status and expression of 14-3-3σ and evaluated its clinical significance in epithelial ovarian cancer.

Experimental Design: Twelve ovarian cancer cell lines; 2 ovarian surface epithelial cell lines; and 8 normal, 8 benign, 12 borderline, and 102 ovarian cancer tissues were examined. Methylation-specific PCR, quantitative reverse transcription-PCR, and immunohistochemistry were used to evaluate methylation status and expression of 14-3-3σ gene and protein.

Results: Among the 12 ovarian cancer cell lines, the presence of a methylated band was detected in seven cell lines. Median values of relative 14-3-3σ gene expression in cancers with methylation (3.27) were significantly lower than those without methylation (16.4; P < 0.001). Treatment of 5-aza-2′-deoxycitidine resulted in the demethylation of the promoter CpG islands and reexpression. All of the normal, benign, and borderline tissues were positive for 14-3-3σ protein, and in ovarian cancer tissues, 73.5% (75 of 102) were positive for 14-3-3σ protein and was almost consistent with methylation status. Negative immunoreactivity of 14-3-3σ was significantly correlated with high age and serous histology, high-grade, advanced-stage residual tumor of >2 cm, high serum CA125, high Ki-67 labeling index, and positive p53 immunoreactivity. 14-3-3σ immunoreactivity was significantly associated with overall survival (P = 0.0058).

Conclusions: Our findings suggest that 14-3-3σ is inactivated mainly by aberrant DNA methylation and that it may play an important role in the pathogenesis of epithelial ovarian cancer.
The Roles of Thymidylate Synthase and p53 in Regulating Fas-Mediated Apoptosis in Response to Antimetabolites
Authors: Longley Daniel Broderick , Allen Wendy Louise , McDermott Ultan , Wilson Timothy Richard , Latif Tariq , Boyer John , Lynch Maria , Johnston Patrick Gerard
Published: September 2005

Fas (CD95/Apo-1) is a member of the tumor necrosis factor receptor family. Receptor binding results in activation of caspase 8, leading to activation of proapoptotic downstream molecules. We found that expression of Fas was up-regulated >10-fold in MCF-7 breast and HCT116 and RKO colon cancer cell lines after treatment with IC60 doses of 5-fluorouracil (5-FU) and raltitrexed (RTX). Combined treatment with the agonistic Fas antibody CH-11 and either 5-FU or RTX resulted in a highly synergistic induction of apoptosis in these cell lines. Similar results were obtained for another antifolate, Alimta. Induction of thymidylate synthase expression inhibited Fas induction in response to RTX and Alimta, but not in response to 5-FU. Furthermore, thymidylate synthase induction abrogated the synergy between CH-11 and both antifolates but had no effect on the synergistic interaction between 5-FU and CH-11. Inactivation of p53 in MCF-7 and HCT116 cell lines blocked 5-FU- and antifolate-mediated up-regulation of Fas. Furthermore, Fas was not up-regulated in response to 5-FU or antifolates in the p53-mutant H630 colon cancer cell line. Lack of Fas up-regulation in the p53-null and -mutant lines abolished the synergistic interaction between 5-FU and CH-11. Interestingly, synergy was still observed between the antifolates and CH-11 in the p53-null HCT116 and p53-mutant H630 cell lines, although this was significantly reduced compared with the p53 wild-type cell lines. Our results indicate that Fas is an important mediator of apoptosis in response to both 5-FU and antifolates.
Patient Characteristics Compete with Dose as Predictors of Acute Treatment Toxicity in Early Phase Clinical Trials
Authors: Rogatko André , Babb James S. , Wang Hao , Slifker Michael J. , Hudes Gary R.
Published: September 2005

Purpose: The purpose of this study was to identify patient characteristics that may be risk factors or markers of susceptibility to adverse treatment effects in cancer Phase I and II clinical trials.

Patients and Methods: A total of 459 patients enrolled in 23 therapeutic Phase I and II studies at the Fox Chase Cancer Center were included in the analysis. Patient-specific characteristics, medical and treatment history, doses of experimental agents, and graded toxicities were extracted from case report forms. We developed a novel summary measure, the toxicity index (TI), to better discriminate patients on the basis of their overall toxicity experiences. Mixed model ANOVA was used to model TI on the basis of data from all trials using a specific agent. Generalized estimating equations in the context of binary logistic regression were used to model dose-limiting toxicity.

Results: Seventeen pretreatment factors, including performance status, alkaline phosphatase, total bilirubin, serum creatinine, and tobacco use, emerged as significant predictors of toxicity as defined by dose-limiting toxicity or TI. Unexpectedly, dose was not always a predictor of toxicity. Even for values within the normal range, the TI identified serum bilirubin and alkaline phosphatase as predictors of toxicity after treatment with docetaxel and alkaline phosphatase as a predictor for toxicity after treatment with irinotecan.

Conclusions: Independent of dose, certain pretreatment characteristics, including measures of organ function that are in the normal range, were found to be predictors of treatment toxicity. Because of its sensitivity to differences in overall toxicity, the TI should prove to be a useful tool for identifying predictors of chemotherapy-related toxicity.
High Level of Glutathione-
S
-Transferase π Expression in Mantle Cell Lymphomas
Authors: Bennaceur-Griscelli Annelise , Bosq Jacques , Koscielny Serge , Lefrère François , Turhan Ali , Brousse Nicole , Hermine Olivier , Ribrag Vincent
Published: September 2005

Purpose: Prognosis of mantle cell lymphoma (MCL) remains poor. Patients who achieve a response to first line therapy usually relapse, and the probability of cure remains low. Glutathione-S-transferase π (GST-π) overexpression has been associated with alkylating agents and anthracycline resistance. GST-π gene is located in 11q13 and is coamplified along with CCND1 gene in some human solid tumors.

Experimental Design: We performed immunohistochemical analysis of GST-π expression in 24 consecutive MCLs, 12 follicular lymphomas (FLs), and 69 diffuse large B-cell lymphomas (DLBCLs). Cases were classified in three groups: high GST-π expression (> 50% of cells were stained), moderate (5 to 50% cells were stained), or absent (< 5% cells were stained). GST-π and CCND1 mRNA levels were also assessed by real-time reverse transcription-PCR analysis.

Results: All MCLs exhibit high GST-π protein expression, compared with 29% of the DLBCLs and none of the FLs. MCLs expressed high levels of GST-π and CCND1 mRNAs compared with DLBCLs and FLs. There was a strong relation between GST-π and CCND1 mRNAs transcript levels in MCLs but not in DLBCLs. In conclusion, protein and mRNA GST-π expression is high in MCL compared with FL and DLBCL.

Conclusions: Overexpression of CCND1 in MCL is associated with a transcriptional up-regulation of the GST-π gene. Our results suggest that the glutathione system could play a role in drug resistance in MCL.
trans-
3,4,5′-Trihydroxystibene Inhibits Hypoxia-Inducible Factor 1α and Vascular Endothelial Growth Factor Expression in Human Ovarian Cancer Cells
Authors: Cao Zongxian , Fang Jing , Xia Chang , Shi Xianglin , Jiang Bing-Hua
Published: September 2005

trans-3,4,5′-Trihydroxystibene (resveratrol) is a natural product commonly found in the human diet and has been shown recently to have anticancer effects on various human cancer cells. However, the molecular basis for its anticancer action remains to be elucidated. In this study, we investigated the effect of resveratrol on hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) expression in human ovarian cancer cells A2780/CP70 and OVCAR-3. We found that although resveratrol did not affect HIF-1α mRNA levels, it did dramatically inhibit both basal-level and growth factor-induced HIF-1α protein expression in the cells. Resveratrol also greatly inhibited VEGF expression. Mechanistically, we demonstrated that resveratrol inhibited HIF-1α and VEGF expression through multiple mechanisms. First, resveratrol inhibited AKT and mitogen-activated protein kinase activation, which played a partial role in the down-regulation of HIF-1α expression. Second, resveratrol inhibited insulin-like growth factor 1-induced HIF-1α expression through the inhibition of protein translational regulators, including Mr 70,000 ribosomal protein S6 kinase 1, S6 ribosomal protein, eukaryotic initiation factor 4E-binding protein 1, and eukaryotic initiation factor 4E. Finally, we showed that resveratrol substantially induced HIF-1α protein degradation through the proteasome pathway. Our data suggested that resveratrol may inhibit human ovarian cancer progression and angiogenesis by inhibiting HIF-1α and VEGF expression and thus provide a novel potential mechanism for the anticancer action of resveratrol.
EphA2 Expression Is Associated with Aggressive Features in Ovarian Carcinoma
Authors: Thaker Premal H. , Deavers Michael , Celestino Joseph , Thornton Angela , Fletcher Mavis S. , Landen Charles N. , Kinch Michael S. , Kiener Peter A. , Sood Anil K.
Published: September 2005

Purpose: EphA2 (epithelial cell kinase) is a transmembrane receptor tyrosine kinase that has been implicated in oncogenesis. There are no published data regarding the role of EphA2 in ovarian carcinoma, which is the focus of the present study.

Experimental Design: Nontransformed (HIO-180) and ovarian cancer (EG, 222, SKOV3, and A2780-PAR) cell lines were evaluated for EphA2 by Western blot analysis. Five benign ovarian masses, 10 ovarian tumors of low malignant potential, and 79 invasive ovarian carcinomas were also evaluated for EphA2 expression by immunohistochemistry. All samples were scored in a blinded fashion. Univariate and multivariate analyses were used to determine significant associations between EphA2 expression and clinicopathological variables.

Results: By Western blot analysis, EG, 222, and SKOV3 cell lines overexpressed EphA2, whereas A2780-PAR and HIO-180 had low to absent EphA2 expression. All of the benign tumors had low or absent EphA2 expression. Among the invasive ovarian carcinomas examined (mean age of patients was 59.2 years), 60 (75.9%) tumors overexpressed EphA2 and the other 19 tumors had negative or minimal EphA2 expression. There was no association of EphA2 overexpression with ascites, likelihood of nodal positivity, pathological subtype, and optimum surgical cytoreduction (residual tumor <1 cm). However, EphA2 overexpression was significantly associated with higher tumor grade (P = 0.02) and advanced stage of disease (P = 0.001). The median survival for patients with tumor EphA2 overexpression was significantly shorter (median, 3.1 years; P = 0.004); the median survival for patients with low or absent EphA2 tumor expression was at least 12 years and has not yet been reached. In multivariate analysis using the Cox proportional hazards model, only volume of residual disease (P < 0.04) and EphA2 overexpression (P < 0.01) were significant and independent predictors of survival.

Conclusions: EphA2 overexpression is predictive of aggressive ovarian cancer behavior and may be an important therapeutic target.
Phase I Study of Concomitant Chemoradiotherapy with Paclitaxel, Fluorouracil, Gemcitabine, and Twice-Daily Radiation in Patients with Poor-Prognosis Cancer of the Head and Neck
Authors: Milano Michael T. , Haraf Daniel J. , Stenson Kerstin M. , Witt Mary Ellyn , Eng Cathy , Mittal Bharat B. , Argiris Athanassios , Pelzer Harold , Kozloff Mark F. , Vokes Everett E.
Published: September 2005

Purpose: We previously demonstrated high locoregional control, in patients with poor-prognosis head and neck cancer (HNC), using paclitaxel, 5-fluorouracil, hydroxyurea, and concomitant hyperfractionated radiotherapy. In the present phase I trial, gemcitabine, a novel antimetabolite with strong radiation-enhancing activity, replaces hydroxyurea. We sought to determine the recommended phase II dose and clinical efficacy in poor-prognosis HNC patients.

Experimental Design: Seventy-two patients enrolled. Eligibility criteria included recurrent or second primary HNC, metastases or expected 2-year survival <20%. Chemoradiotherapy consisted of 5-fluorouracil, 600 mg/m2/d, for 5 days; paclitaxel, 100 mg/m2 on Day 1; and concurrent 1.5 Gy twice-daily radiation for 5 days. Gemcitabine was dose escalated, 50–300 mg/m2 on day 1. Cycles repeated every 14 days until the completion of chemoradiation. Dose-limiting toxicities (DLTs) included: neutropenic fever; grade ≥4 neutropenia or thrombocytopenia for >4 days; grade ≥4 mucositis or dermatitis for >7 days; or grade 3 toxicity necessitating chemotherapy dose reductions. Non-DLT dose reductions in 5-fluorouracil and/or paclitaxel were allowed.

Results: Seventy-nine percent of assessable patients experienced a clinical response. Five-year actuarial survival is 33.0%, and locoregional control is 61.4%. The recommended phase II dose of gemcitabine in this regimen is 100 mg/m2 during cycles 1–5 (1 of 7 patients with DLT) or 200 mg/m2 delivered only during cycles 3–5 (3 of 19 with DLT). Grades 3 and 4 mucositis (56 and 21%, respectively) and dermatitis (25 and 21%, respectively) were common.

Conclusions: Gemcitabine, 5-fluorouracil, paclitaxel, and twice-daily radiation, delivered on alternating weeks, is active in patients with poor-prognosis HNC, although severe mucositis limits the clinical applicability of this regimen. Refinements in radiotherapy, including intensity-modulated radiation therapy, may improve the tolerance for this regimen.
Phase I and Pharmacokinetic Study of Genexol-PM, a Cremophor-Free, Polymeric Micelle-Formulated Paclitaxel, in Patients with Advanced Malignancies
Authors: Kim Tae-You , Kim Dong-Wan , Chung Jae-Yong , Shin Sang Goo , Kim Sung-Chul , Heo Dae Seog , Kim Noe Kyeong , Bang Yung-Jue
Published: September 2005

Purpose: The rationale for developing an alternative paclitaxel formulation concerns Cremophor EL-related side effects, and a novel paclitaxel delivery system might augment its therapeutic efficacy. Genexol-PM is a polymeric micelle formulated paclitaxel free of Cremophor EL. A phase I study was performed to determine the maximum tolerated dosage, dose-limiting toxicities, and the pharmacokinetic profile of Genexol-PM in patients with advanced, refractory malignancies.

Experimental Design: Twenty-one patients were entered into the study. Genexol-PM was i.v. administered over 3 h every 3 weeks without premedication. The Genexol-PM dose was escalated from 135 mg/m2 to 390 mg/m2.

Results: All of the patients were evaluable for toxicity and response. Acute hypersensitivity reactions were not observed. Neuropathy and myalgia were the most common toxicities. During cycle 1, grade 3 myalgia occurred in 1 patient at 230 and 300 mg/m2, respectively. At 390 mg/m2, 2 of 3 patients developed grade 4 neutropenia or grade 3 polyneuropathy. Therefore, the maximum tolerated dosage was determined to be 390 mg/m2. There were 3 partial responses (14%) among the 21 patients. Of the 3 responders, 2 were refractory to prior taxane therapy. The paclitaxel area under the curve from time 0 to infinity and peak or maximum paclitaxel concentration seemed to increase with escalating dose, except at 230 mg/m2, which suggests that Genexol-PM has linear pharmacokinetics.

Conclusion: The main dose-limiting toxicities were neuropathy, myalgia, and neutropenia, and the recommended dosage for a phase II study is 300 mg/m2. Genexol-PM is believed to be superior to conventional paclitaxel in terms of the obviation of premedication and the delivery of higher paclitaxel doses without additional toxicity.