bluuebunny/crossref_metadata_2025 huggingface dataset:
Human T Cells Armed with Her2/neu Bispecific Antibodies Divide, Are Cytotoxic, and Secrete Cytokines with Repeated Stimulation
Authors: Grabert Ryan C. , Cousens Leslie P. , Smith Janelle A. , Olson Sarah , Gall Jonathan , Young Wendy B. , Davol Pamela A. , Lum Lawrence G.
Published: March 2006

Purpose: Cancer immunotherapy has been limited by anergy of patient T cells, inadequate numbers of precursor tumor-specific CTL, and difficulty in producing therapeutic doses of CTL. To overcome these limitations, bispecific antibodies have been used to create artificial antibody receptors that direct polyclonal activated T cells (ATC) to target tumor antigens. Studies reported herein were designed to characterize bispecific antibody–armed ATC functions during multiple rounds of targeted cell stimulation.

Experimental Design: ATCs were generated from human peripheral blood mononuclear cells (PBMC) by culture with anti-CD3 and interleukin 2 for 14 days and armed with anti-CD3 × anti-Her2 bispecific antibody (Her2Bi). In vitro, Her2Bi-armed ATC were examined for a range of functions after repeated stimulation with the Her2/neu-expressing breast cancer cell line SK-BR-3. PBMC isolated from cancer patients treated with Her2Bi-armed ATC were tested ex vivo for cytotoxicity against SK-BR-3.

Results: In vitro, armed ATC divided, maintained surface Her2Bi, and expressed a range of activities for extended periods of time. Perforin-mediated cytotoxic activity by armed ATC continued for at least 336 hours, and cytokines and chemokines (i.e., IFN-γ and regulated on activation, normal T-cell expressed and secreted protein [RANTES]) were secreted during successive rounds of stimulation. Furthermore, PBMC isolated from patients over their courses of immunotherapy exhibited significant cytolytic activity against SK-BR-3 as a function of Her2Bi-armed ATC infusions.

Conclusions: These studies show that armed ATC are specific, durable, and highly functional T-cell populations in vitro. These previously unappreciated broad and long-term functions of armed ATC are encouraging for their therapeutic use in treating cancer.
Gene Expression Differences Associated with Human Papillomavirus Status in Head and Neck Squamous Cell Carcinoma
Authors: Slebos Robbert J.C. , Yi Yajun , Ely Kim , Carter Jesse , Evjen Amy , Zhang Xueqiong , Shyr Yu , Murphy Barbara M. , Cmelak Anthony J. , Burkey Brian B. , Netterville James L. , Levy Shawn , Yarbrough Wendell G. , Chung Christine H.
Published: March 2006

Human papillomavirus (HPV) is associated with a subset of head and neck squamous cell carcinoma (HNSCC). Between 15% and 35% of HNSCCs harbor HPV DNA. Demographic and exposure differences between HPV-positive (HPV+) and negative (HPV−) HNSCCs suggest that HPV+ tumors may constitute a subclass with different biology, whereas clinical differences have also been observed. Gene expression profiles of HPV+ and HPV− tumors were compared with further exploration of the biological effect of HPV in HNSCC. Thirty-six HNSCC tumors were analyzed using Affymetrix Human 133U Plus 2.0 GeneChip and for HPV by PCR and real-time PCR. Eight of 36 (22%) tumors were positive for HPV subtype 16. Statistical analysis using Significance Analysis of Microarrays based on HPV status as a supervising variable resulted in a list of 91 genes that were differentially expressed with statistical significance. Results for a subset of these genes were verified by real-time PCR. Genes highly expressed in HPV+ samples included cell cycle regulators (p16INK4A, p18, and CDC7) and transcription factors (TAF7L, RFC4, RPA2, and TFDP2). The microarray data were also investigated by mapping genes by chromosomal location (DIGMAP). A large number of genes on chromosome 3q24-qter had high levels of expression in HPV+ tumors. Further investigation of differentially expressed genes may reveal the unique pathways in HPV+ tumors that may explain the different natural history and biological properties of these tumors. These properties may be exploited as a target of novel therapeutic agents in HNSCC treatment.
CR011, a fully human monoclonal antibody-auristatin E conjugate, for the treatment of melanoma.
Authors: Tse Kam Fai , Jeffers Michael , Pollack Vincent A , McCabe Denise A , Shadish Melanie L , Khramtsov Nikolai V , Hackett Craig S , Shenoy Suresh G , Kuang Bing , Boldog Ferenc L , MacDougall John R , Rastelli Luca , Herrmann John , Gallo Michael , Gazit-Bornstein Gadi , Senter Peter D , Meyer Damon L , Lichenstein Henri S , LaRochelle William J
Published: March 2006

PURPOSE: Advanced melanoma is a highly drug-refractory neoplasm representing a significant unmet medical need. We sought to identify melanoma-associated cell surface molecules and to develop as well as preclinically test immunotherapeutic reagents designed to exploit such targets. EXPERIMENTAL DESIGN AND RESULTS: By transcript profiling, we identified glycoprotein NMB (GPNMB) as a gene that is expressed by most metastatic melanoma samples examined. GPNMB is predicted to be a transmembrane protein, thus making it a potential immunotherapeutic target in the treatment of this disease. A fully human monoclonal antibody, designated CR011, was generated to the extracellular domain of GPNMB and characterized for growth-inhibitory activity against melanoma. The CR011 monoclonal antibody showed surface staining of most melanoma cell lines by flow cytometry and reacted with a majority of metastatic melanoma specimens by immunohistochemistry. CR011 alone did not inhibit the growth of melanoma cells. However, when linked to the cytotoxic agent monomethylauristatin E (MMAE) to generate the CR011-vcMMAE antibody-drug conjugate, this reagent now potently and specifically inhibited the growth of GPNMB-positive melanoma cells in vitro. Ectopic overexpression and small interfering RNA transfection studies showed that GPNMB expression is both necessary and sufficient for sensitivity to low concentrations of CR011-vcMMAE. In a melanoma xenograft model, CR011-vcMMAE induced significant dose-proportional antitumor effects, including complete regressions, at doses as low as 1.25 mg/kg. CONCLUSION: These preclinical results support the continued evaluation of CR011-vcMMAE for the treatment of melanoma.
A Synthetic Triterpenoid, CDDO-Me, Inhibits IκBα Kinase and Enhances Apoptosis Induced by TNF and Chemotherapeutic Agents through Down-Regulation of Expression of Nuclear Factor κB–Regulated Gene Products in Human Leukemic Cells
Authors: Shishodia Shishir , Sethi Gautam , Konopleva Marina , Andreeff Michael , Aggarwal Bharat B.
Published: March 2006

The C-28 methyl ester of 2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid (CDDO-Me), a synthetic triterpenoid based on naturally occurring ursolic and oleanolic acids, induces apoptosis in tumor cells, induces differentiation, and inhibits inflammatory response through a poorly understood mechanism. Because the nuclear transcription factor nuclear factor κB (NF-κB) has been shown to suppress apoptosis and promote proliferation and is linked with inflammation and differentiation, we postulated that CDDO-Me modulates NF-κB activity and NF-κB-regulated gene expression. Using human leukemia cell lines and patient samples, we show that CDDO-Me potently inhibits both constitutive and inducible NF-κB activated by tumor necrosis factor (TNF), interleukin (IL)-1β, phorbol ester, okadaic acid, hydrogen peroxide, lipopolysaccharide, and cigarette smoke. CDDO-Me was more potent than CDDO and its imidazole derivative. NF-κB suppression occurred through inhibition of IκBα kinase activation, IκBα phosphorylation, IκBα degradation, p65 phosphorylation, p65 nuclear translocation, and NF-κB-mediated reporter gene transcription. This inhibition correlated with suppression of NF-κB-dependent genes involved in antiapoptosis (IAP2, cFLIP, TRAF1, survivin, and bcl-2), proliferation (cyclin d1 and c-myc), and angiogenesis (VEGF, cox-2, and mmp-9). CDDO-Me also potentiated the cytotoxic effects of TNF and chemotherapeutic agents. Overall, our results suggest that CDDO-Me inhibits NF-κB through inhibition of IκBα kinase, leading to the suppression of expression of NF-κB-regulated gene products and enhancement of apoptosis induced by TNF and chemotherapeutic agents.
A Rapid and Sensitive Enzymatic Method for Epidermal Growth Factor Receptor Mutation Screening
Authors: Jänne Pasi A. , Borras Ana M. , Kuang Yanan , Rogers Andrew M. , Joshi Victoria A. , Liyanage Hema , Lindeman Neal , Lee Jeffrey C. , Halmos Balazs , Maher Elizabeth A. , Distel Robert J. , Meyerson Matthew , Johnson Bruce E.
Published: March 2006

Purpose: Mutations in the epidermal growth factor receptor (EGFR) are associated with clinical and radiographic responses to EGFR tyrosine kinase inhibitors gefitinib and erlotinib. Currently available methods of EGFR mutation detection rely on direct DNA sequencing, which requires isolation of DNA from a relatively pure population of tumor cells, cannot be done on small diagnostic specimens, and lack sensitivity. Here we describe the use of a sensitive screening method that overcomes many of these limitations.

Experimental Design: We screened 178 non–small cell lung cancer specimens for mutations in exons 18 to 21 of EGFR using a DNA endonuclease, SURVEYOR, which cleaves mismatched heteroduplexed DNA. Samples were analyzed by high-performance liquid chromatography on the Transgenomic WAVE HS system. Selected specimens that produced digestion products using SURVEYOR were subsequently reanalyzed by size separation or under partially denaturing conditions, followed by fractionation and sequencing. The specimens included DNA isolated from frozen tumor specimens, dissected formalin-fixed, paraffin-embedded tumor specimens undergoing clinical sequencing, and undissected formalin-fixed, paraffin-embedded specimens. One hundred sixty specimens were independently analyzed using direct DNA sequencing in a blinded fashion.

Results: EGFR mutations were detected in 16 of 61 fresh frozen tumor specimens, 24 of 91 dissected formalin-fixed, paraffin-embedded tumor specimens, and 11 of 26 undissected formalin-fixed, paraffin-embedded tumor specimens. Compared with sequencing, the sensitivity and specificity of the present method were 100% and 87%. The positive and negative predictive values were 74% and 100%, respectively. SURVEYOR analysis detected 7 (4%) mutations that were not previously detected by direct sequencing.

Conclusions: SURVEYOR analysis provides a rapid method for EGFR mutation screening with 100% sensitivity and negative predictive value. This unbiased scanning technique is superior to direct sequencing when used with undissected formalin-fixed, paraffin-embedded specimens.
A randomized phase II study of concurrent docetaxel plus vaccine versus vaccine alone in metastatic androgen-independent prostate cancer.
Authors: Arlen Philip M , Gulley James L , Parker Catherine , Skarupa Lisa , Pazdur Mary , Panicali Dennis , Beetham Patricia , Tsang Kwong Y , Grosenbach Douglas W , Feldman Jarett , Steinberg Seth M , Jones Elizabeth , Chen Clara , Marte Jennifer , Schlom Jeffrey , Dahut William
Published: March 2006

PURPOSE: Docetaxel has activity against androgen-independent prostate cancer and preclinical studies have shown that taxane-based chemotherapy can enhance antitumor response of vaccines. The primary objective of this study was to determine if concurrent docetaxel (with dexamethasone) had any effect on generating an immune response to the vaccine. Secondary end points were whether vaccine could be given safely with docetaxel and the clinical outcome of the treatment regimen. EXPERIMENTAL DESIGN: The vaccination regimen was composed of (a) recombinant vaccinia virus (rV) that expresses the prostate-specific antigen gene (rV-PSA) admixed with (b) rV that expresses the B7.1 costimulatory gene (rV-B7.1), and (c) sequential booster vaccinations with recombinant fowlpox virus (rF-) containing the PSA gene (rF-PSA). Patients received granulocyte macrophage colony-stimulating factor with each vaccination. Twenty-eight patients with metastatic androgen-independent prostate cancer were randomized to receive either vaccine and weekly docetaxel or vaccine alone. Patients on the vaccine alone arm were allowed to cross over to receive docetaxel alone at time of disease progression. The ELISPOT assay was used to monitor immune responses for PSA-specific T cells. RESULTS: The median increase in these T-cell precursors to PSA was 3.33-fold in both arms following 3 months of therapy. In addition, immune responses to other prostate cancer-associated tumor antigens were also detected postvaccination. Eleven patients who progressed on vaccine alone crossed over to receive docetaxel at time of progression. Median progression-free survival on docetaxel was 6.1 months after receiving vaccine compared with 3.7 months with the same regimen in a historical control. CONCLUSION: This is the first clinical trial to show that docetaxel can be administered safely with immunotherapy without inhibiting vaccine specific T-cell responses. Furthermore, patients previously vaccinated with an anticancer vaccine may respond longer to docetaxel compared with a historical control of patients receiving docetaxel alone. Larger prospective clinical studies will be required to validate these findings.
Human melanoma cells selected for resistance to apoptosis by prolonged exposure to tumor necrosis factor-related apoptosis-inducing ligand are more vulnerable to necrotic cell death induced by cisplatin.
Authors: Zhang Xu Dong , Wu Jing Jing , Gillespie Susan , Borrow Jodie , Hersey Peter
Published: March 2006

PURPOSE: Heterogeneous sensitivity of melanoma cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis may lead to outgrowth of TRAIL-resistant cells and limit successful treatment by TRAIL. The present study aims to better understand the biological characteristics of melanoma cells resistant to TRAIL-induced apoptosis. EXPERIMENTAL DESIGN: We generated TRAIL-resistant melanoma cells by prolonged exposure to TRAIL and characterized the cells in terms of their sensitivity to killing induced by a panel of cytotoxic agents using biological and biochemical methods. RESULTS: TRAIL-resistant melanoma cells are cross-resistant to apoptosis induced by another death ligand FasL, the DNA-damaging agent cisplatin, the histone deacetylase inhibitor suberic bishydroxamate, and the antimicrotubule Vinca alkaloid, vincristine. The apoptotic signaling seemed to be inhibited upstream of mitochondrial apoptotic events and was associated with decreased expression of multiple apoptotic mediators, including pro-caspase-8, Fas-associated death domain, Bid, Bim, p53, and the products of its proapoptotic target genes. Despite being resistant to apoptosis, TRAIL-resistant melanoma cells were more vulnerable to cisplatin-induced nonapoptotic cell death. This was characterized by lack of DNA fragmentation, delayed externalization of phosphatidylserine, caspase and p53 independence, and severe mitochondrial disruption, and was preceded by poly(ADP)ribose polymerase (PARP) activation and depletion of intracellular ATP, indicative of necrotic cell death. Inhibition of PARP activity partially converted the mode of cell death from necrosis to apoptosis. CONCLUSIONS: TRAIL-resistant melanoma cells are cross-resistant to apoptosis induced by various apoptotic stimuli but are more sensitive to nonapoptotic cell death induced by cisplatin. Exploration of chemotherapy-induced nonapoptotic cell death may provide an alternative strategy in overcoming resistance of melanoma cells to TRAIL-induced apoptosis.
HER2-Positive Circulating Tumor Cells Indicate Poor Clinical Outcome in Stage I to III Breast Cancer Patients
Authors: Wülfing Pia , Borchard Julia , Buerger Horst , Heidl Stefan , Zänker Kurt S. , Kiesel Ludwig , Brandt Burkhard
Published: March 2006

Purpose: Early metastasis in node-negative breast cancer indicates that breast cancer cells obviously can bypass the lymph nodes and disseminate directly hematogenous to distant organs. For this purpose, we evaluated the prognostic value of blood-borne, HER2-positive circulating tumor cells (CTC) in the peripheral blood from 42 breast cancer patients with a median follow-up of 95 months.

Experimental Design: Cells were isolated by the patented combined buoyant density gradient and immunomagnetic separation procedure and analyzed by immunocytochemistry.

Results: We detected one to eight CTCs in the peripheral blood of 17 of 35 patients (48.6%) presenting no overt metastasis. As a positive control, 7 of 7 (100%) patients with metastatic disease presented positive. Healthy persons and patients (n = 32) operated for nonmalignant diseases presented negative for CTCs. The presence and frequency of HER2-positive CTCs correlated with a significantly decreased disease-free survival (P < 0.005) and overall survival (P < 0.05). Interestingly, in 12 patients with HER2-positive CTCs, the primary tumor was negative for HER2 as assessed by immunohistochemical score and fluorescence in situ hybridization.

Conclusions: This study provides some evidence of a prognostic effect of HER2-positive CTCs in stage I to III breast cancer. Future studies have to determine the outcome of patients treated with HER2-targeting therapies with respect to HER2-positive CTC levels because it is not unlikely that high levels of HER2-positive CTCs reflect the activity of the tumor and may predict response to trastuzumab.
Phase I Trial with BMS-275183, a Novel Oral Taxane with Promising Antitumor Activity
Authors: Bröker Linda E. , de Vos Filip Y.F.L. , van Groeningen Cees J. , Kuenen Bart C. , Gall Helen E. , Woo Michael H. , Voi Maurizio , Gietema Jourik A. , de Vries Elisabeth G.E. , Giaccone Giuseppe
Published: March 2006

Purpose: BMS-275183 is an orally administered C-4 methyl carbonate analogue of paclitaxel. We did a dose-escalating phase I study to investigate its safety, tolerability, pharmacokinetics, and possible antitumor activity.

Experimental Design: A cycle consisted of four weekly doses of BMS-275183. The starting dose was 5 mg, which was increased by 100% increments (i.e., 5, 10, 20 mg/m2, etc.) in each new cohort consisting of one patient. Cohorts were expanded when toxicity was encountered, and 20 patients were treated at the maximum tolerated dose (MTD). Plasma pharmacokinetics were done on days 1 and 15.

Results: A total of 48 patients were enrolled in this trial. Dose-limiting toxicities consisted of neuropathy, fatigue, diarrhea, and neutropenia. First cycle severe neuropathy was reported in four patients treated at 320 (n = 1), 240 (n = 2), and 160 mg/m2 (n = 1), whereas eight patients treated at dose levels ranging from 160 to 320 mg/m2 experienced grade 2 neuropathy in cycle one. The MTD was 200 mg/m2, as 3 of 20 patients experienced grade 3 or 4 toxicity in cycle one [fatigue (n = 2), and neutropenia/diarrhea (n = 1)]. BMS-275183 was rapidly absorbed with a mean plasma half-life of 22 hours. We observed a significant correlation between drug-exposure and toxicity. Tumor responses were observed in 9 of 38 evaluable patients with non–small cell lung cancer, prostate carcinoma, and other tumor types.

Conclusions: BMS-275183 is generally well tolerated on a weekly schedule. The main toxicity is peripheral neuropathy, and the MTD is 200 mg/m2. Promising activity was observed in several tumor types, and a phase II trial in non–small cell lung cancer has been initiated.
High Levels of Aberrant DNA Methylation in Helicobacter pylori–Infected Gastric Mucosae and its Possible Association with Gastric Cancer Risk
Authors: Maekita Takao , Nakazawa Kazuyuki , Mihara Mami , Nakajima Takeshi , Yanaoka Kimihiko , Iguchi Mikitaka , Arii Kenji , Kaneda Atsushi , Tsukamoto Tetsuya , Tatematsu Masae , Tamura Gen , Saito Daizo , Sugimura Takashi , Ichinose Masao , Ushijima Toshikazu
Published: March 2006

Introduction: Risk prediction of gastric cancers is important to implement appropriate screening procedures. Although aberrant DNA methylation is deeply involved in gastric carcinogenesis, its induction by Helicobacter pylori, a strong gastric carcinogen, is unclear. Here, we analyzed the effect of H. pylori infection on the quantity of methylated DNA molecules in noncancerous gastric mucosae and examined its association with gastric cancer risk.

Experimental Design: Gastric mucosae were collected from 154 healthy volunteers (56 H. pylori negative and 98 H. pylori positive) and 72 cases with differentiated-type gastric cancers (29 H. pylori negative and 43 H. pylori positive) by endoscopy. The numbers of DNA molecules methylated and unmethylated for eight regions of seven CpG islands (CGI) were quantified by quantitative PCR after bisulfite modification, and fractions of methylated molecules (methylation levels) were calculated.

Results: Among healthy volunteers, methylation levels of all the eight regions were 5.4- to 303-fold higher in H. pylori positives than in H. pylori negatives (P < 0.0001). Methylation levels of the LOX, HAND1, and THBD promoter CGIs and p41ARC exonic CGI were as high as 7.4% or more in H. pylori–positive individuals. Among H. pylori–negative individuals, methylation levels of all the eight regions were 2.2- to 32-fold higher in gastric cancer cases than in age-matched healthy volunteers (P ≤ 0.01). Among H. pylori–positive individuals, methylation levels were highly variable, and that of only HAND1 was significantly increased in gastric cancer cases (1.4-fold, P = 0.02).

Conclusions: It was indicated that H. pylori infection potently induces methylation of CGIs to various degrees. Methylation levels of specific CGIs seemed to reflect gastric cancer risk in H. pylori–negative individuals.
Expression of DNA Double-Strand Break Repair Proteins ATM and BRCA1 Predicts Survival in Colorectal Cancer
Authors: Grabsch Heike , Dattani Mit , Barker Lisa , Maughan Nicola , Maude Karen , Hansen Olaf , Gabbert Helmut E. , Quirke Phil , Mueller Wolfram
Published: March 2006

Purpose: The double-strand break (DSB) is the major DNA lesion leading to chromosomal aberrations and faithful repair is crucial for maintaining genomic instability. Very little is known about the expression of DNA DSB repair proteins in colorectal cancer. To address this issue, we examined the expression pattern of DSB repair key proteins ATM, BRCA1, BRCA2, Ku70, and Ku80 and their putative role in patients survival in a large series of colorectal cancer.

Experimental Design: 342 sporadic colorectal cancer were subjected to immunohistochemistry by using specific antibodies for the various proteins investigated. Staining results were compared with clinicopathologic data, patient survival, as well as expression of mismatch repair proteins MLH1 and MSH2.

Results: The expression pattern of both ATM and BRCA1 predicted survival in all colorectal cancer patients as well as in the small subgroup of patients that received adjuvant therapy. Low expression of ATM and BRCA1 was associated with loss of MLH1 or MSH2 expression.

Conclusions: This is the first study to show a relationship between the expression of DNA DSB repair proteins ATM and BRCA1 and survival in colorectal cancer patients. Studies in tumors from large randomized trials are now necessary to validate our pilot data and establish the clinical usefulness of the immunohistochemical assay in predicting response to a particular adjuvant therapy regimen. Furthermore, our results indicate a possible link between expression of DNA mismatch repair and DNA DSB repair proteins in sporadic colorectal cancer, which warrants further investigation.
Clinical Efforts to Combine Endocrine Agents with Targeted Therapies against Epidermal Growth Factor Receptor/Human Epidermal Growth Factor Receptor 2 and Mammalian Target of Rapamycin in Breast Cancer
Authors: Johnston Stephen R.D.
Published: March 2006

Enhancing the benefit of endocrine therapy by overcoming de novo or acquired resistance remains an important goal in systemic breast cancer therapy. Progress continues to be made in elucidating the molecular pathways by which estrogen receptor–positive breast cancer cells escape from endocrine therapy. The increasing recognition of the roles of epidermal growth factor receptor (EGFR) and human EGFR2 in cross-talk activation of estrogen receptor signaling has led to studies aimed at identifying whether small-molecule tyrosine kinase inhibitors targeted against these receptors give additive or synergistic effects when combined with endocrine agents. Activation of the phosphatidylinositol-3-OH kinase/Akt pathway has also been associated with resistance to either tamoxifen or estrogen deprivation, and preclinical studies have shown that the mammalian target of rapamycin antagonist temsirolimus can restore endocrine sensitivity in breast cancer cells. Randomized phase II trials of aromatase inhibitors combined with EGFR/human EGFR2 tyrosine kinase inhibitors or mammalian target of rapamycin antagonists have been completed in both the neoadjuvant and advanced breast cancer settings. Larger phase III trials with both approaches are now in progress and have been powered to detect whether either strategy can significantly prolong time to disease progression compared with endocrine therapy alone. The correlation of molecular and clinical results from these ongoing studies will be important to establish appropriate biological variables for selecting those patients who may benefit most from this combined approach.
Ingenuity Network-Assisted Transcription Profiling: Identification of a New Pharmacologic Mechanism for MK886
Authors: Mayburd Anatoly L. , Martlínez Alfredo , Sackett Daniel , Liu Huaitian , Shih Joanna , Tauler Jordy , Avis Ingalill , Mulshine James L.
Published: March 2006

The small molecular inhibitor MK886 is known to block 5-lipoxygenase-activating protein ALOX5AP and shows antitumor activity in multiple human cell lines. The broad antitumor therapeutic window reported in vivo for MK886 in rodents supports further consideration of this structural class. Better understanding of the mode of action of the drug is important for application in humans to take place. Affymetrix microarray study was conducted to explore MK886 pharmacologic mechanism. Ingenuity Pathway Analysis software was applied to validate the results at the transcriptional level by putting them in the context of an experimental proteomic network. Genes most affected by MK886 included actin B and focal adhesion components. A subsequent National Cancer Institute-60 panel study, RT-PCR validation followed by confocal microscopy, and Western blotting also pointed to actin B down-regulation, filamentous actin loss, and disorganization of the transcription machinery. In agreement with these observations, MK886 was found to enhance the effect of UV radiation in H720 lung cancer cell line. In light of the modification of cytoskeleton and cell motility by lipid phosphoinositide 3-kinase products, MK886 interaction with actin B might be biologically important. The low toxicity of MK886 in vivo was modeled and explained by binding and transport by dietary lipids. The rate of lipid absorbance is generally higher for tumors, suggesting a promise of a targeted liposome-based delivery system for this drug. These results suggest a novel antitumor pharmacologic mechanism.
PIK3CA Mutations in Head and Neck Squamous Cell Carcinoma
Authors: Qiu Wanglong , Schönleben Frank , Li Xiaojun , Ho Daniel J. , Close Lanny G. , Manolidis Spiros , Bennett Boyce P. , Su Gloria H.
Published: March 2006

Purpose: Recent studies have reported high frequencies of somatic mutations in the phosphoinositide-3-kinase catalytic α (PIK3CA) gene in several human solid tumors. Although gene amplifications of PIK3CA have been reported in head and neck squamous cell carcinoma (HNSCC), small mutation of the gene has not been evaluated in HNSCC previously. In this study, we examined the mutation frequency of PIK3CA in HNSCC.

Experimental Design: More than 75% of the somatic mutations of PIK3CA are clustered in the helical (exon 9) and kinase domains (exon 20). To investigate the possible role of PIK3CA in HNSCC tumorigenesis, exons 1, 4, 5, 6, 7, 9, and 20 of the gene were analyzed by direct genomic DNA sequencing in 38 HNSCC specimens.

Results: We identified four missense mutations in the seven exons of PIK3CA from 38 HNSCC specimens (11%). Three of the four mutations (i.e., H1047R, E542K, and E545K) have been previously reported as hotspot mutations. The remaining novel mutation, Y343C, is identified at exon 4 nucleotide 1028 A → G. Three of the four mutations were shown to be somatic, whereas the fourth mutation (H1047R) was identified in a cell line. Interestingly, three of the four mutations identified were in pharyngeal cancer samples.

Conclusions: These data provide evidence that oncogenic properties of PIK3CA contribute to the carcinogenesis of human head and neck cancers, especially in pharyngeal cancer. A specific kinase inhibitor to PIK3CA may potentially be an effective therapeutic reagent against HNSCC or pharyngeal cancer in particular.
Managing Patients on Endocrine Therapy: Focus on Quality-of-Life Issues
Authors: Whelan Timothy J. , Pritchard Kathleen I.
Published: March 2006

Purpose: To review the health-related quality of life (QOL) of women treated with adjuvant hormonal therapy.

Experimental Design: To review the limited QOL data from randomized trials of tamoxifen versus placebo and ovarian ablation versus none. To discuss QOL results from randomized trials of aromatase inhibitors compared with tamoxifen or placebo for adjuvant therapy of postmenopausal women with estrogen receptor–positive and/or progesterone receptor–positive breast cancer.

Results: QOL is generally good in up to 3 years of follow-up with either tamoxifen or aromatase inhibitors. Vasomotor and sexual complaints remain problematic, however, in only a small proportion of women. There are fewer data regarding the QOL effects of ovarian ablation, which may nonetheless be more substantial.

Conclusion: Tamoxifen and aromatase inhibitors cause specific vasomotor or gynecologic symptoms, which may affect sexual function. However, clinical benefits of these agents are generally achieved without major detrimental effect on overall QOL.
Phase 1 Trial of Gefitinib Plus Sirolimus in Adults with Recurrent Malignant Glioma
Authors: Reardon David A. , Quinn Jennifer A. , Vredenburgh James J. , Gururangan Sridharan , Friedman Allan H. , Desjardins Annick , Sathornsumetee Sith , Herndon James E. , Dowell Jeannette M. , McLendon Roger E. , Provenzale James M. , Sampson John H. , Smith Robert P. , Swaisland Alan J. , Ochs Judith S. , Lyons Peggy , Tourt-Uhlig Sandy , Bigner Darell D. , Friedman Henry S. , Rich Jeremy N.
Published: March 2006

Purpose: To determine the maximum tolerated dose (MTD) and dose-limiting toxicity (DLT) of gefitinib, a receptor tyrosine kinase inhibitor of the epidermal growth factor receptor, plus sirolimus, an inhibitor of the mammalian target of rapamycin, among patients with recurrent malignant glioma.

Patients and Methods: Gefitinib and sirolimus were administered on a continuous daily dosing schedule at dose levels that were escalated in successive cohorts of malignant glioma patients at any recurrence who were stratified based on concurrent use of CYP3A-inducing anticonvulsants [enzyme-inducing antiepileptic drugs, (EIAED)]. Pharmacokinetic and archival tumor biomarker data were also assessed.

Results: Thirty-four patients with progressive disease after prior radiation therapy and chemotherapy were enrolled, including 29 (85%) with glioblastoma multiforme and 5 (15%) with anaplastic glioma. The MTD was 500 mg of gefitinib plus 5 mg of sirolimus for patients not on EIAEDs and 1,000 mg of gefitinib plus 10 mg of sirolimus for patients on EIAEDs. DLTs included mucositis, diarrhea, rash, thrombocytopenia, and hypertriglyceridemia. Gefitinib exposure was not affected by sirolimus administration but was significantly lowered by concurrent EIAED use. Two patients (6%) achieved a partial radiographic response, and 13 patients (38%) achieved stable disease.

Conclusion: We show that gefitinib plus sirolimus can be safely coadministered on a continuous, daily dosing schedule, and established the recommended dose level of these agents in combination for future phase 2 clinical trials.
Characterization of Familial Non-BRCA1/2Breast Tumors by Loss of Heterozygosity and Immunophenotyping
Authors: Oldenburg Rogier A. , Kroeze-Jansema Karin , Meijers-Heijboer Hanne , van Asperen Cristi J. , Hoogerbrugge Nicoline , van Leeuwen Inge , Vasen Hans F.A. , Cleton-Jansen Anne-Marie , Kraan Jaennelle , Houwing-Duistermaat Jeanine J. , Morreau Hans , Cornelisse Cees J. , Devilee Peter
Published: March 2006

Purpose: Since the identification of BRCA1 and BRCA2, there has been no major breast cancer susceptibility gene discovered by linkage analysis in breast cancer families. This has been attributed to the heterogeneous genetic basis for the families under study. Recent studies have indicated that breast tumors arising in women carrying a BRCA1 mutation have distinct histopathologic, immunophenotypic, and genetic features. To a lesser extent, this is also true for breast tumors from BRCA2 carriers. This indicates that it might be possible to decrease the genetic heterogeneity among families in which BRCA1 and BRCA2 have been excluded with high certainty (BRCAx families) if distinct subgroups of BRCAx-related breast tumors could be identified.

Experimental Design: Loss of heterozygosity (LOH) analysis with at least one marker per chromosomal arm (65 markers) was used to characterize 100 breast tumors derived from 92 patients from 42 selected BRCAx families. In addition, the immunophenotype of 10 markers was compared with that of 31 BRCA1- and 21 BRCA2-related breast tumors.

Results and Conclusions: The BRCAx-related tumors were characterized by more frequent LOH at 22q relative to sporadic breast cancer (P < 0.02), and differed significantly from BRCA1- and BRCA2-related tumors in their positivity for Bcl2. However, cluster analyses of the combined data (LOH and immunohistochemistry) did not result in subgroups that would allow meaningful subclassification of the families. On chromosomes 2, 3, 6, 12, 13, 21, and 22, we found markers at which LOH occurred significantly more frequent among the tumors from patients belonging to a single family than expected on the basis of overall LOH frequencies. Nonetheless, linkage analysis with markers for the corresponding regions on chromosomes 12, 21, and 22 did not reveal significant logarithm of the odds.
Defective Antitumor Function of Monocyte-Derived Macrophages from Epithelial Ovarian Cancer Patients
Authors: Gordon Ilyssa O. , Freedman Ralph S.
Published: March 2006

Purpose: Monocytes/macrophages (MO/MA) are an important but heterogeneous population of immune inflammatory cells that have diverse effector functions. We examined and compared these differences in peripheral blood and ascites of epithelial ovarian cancer patients with peripheral blood of normal donors.

Experimental Design: Comparisons were made of cell surface subsets, cytokine production, and FcR-dependent cytotoxicity of CD14+ MO/MA and the CD14brightCD16−HLA-DR+ MO/MA subset in normal donor peripheral blood and peripheral blood and ascites from epithelial ovarian cancer patients. Studies were done on monocyte-derived macrophages cultured with macrophage colony-stimulating factor and activated with lipopolysaccharide or a combination of lipopolysaccharide plus recombinant IFN-γ.

Results: We determined that MO/MA or its subset from epithelial ovarian cancer patients had altered morphology and significantly less antibody-dependent cell-mediated cytotoxicity and phagocytic activity than did MO/MA from normal donors. Our findings also showed that monocyte-derived macrophages from both epithelial ovarian cancer patients and normal donors produce macrophage colony-stimulating factor–stimulated cytokines, including interleukin-8, tumor necrosis factor-α, and interleukin-6.

Conclusions: These findings highlight for the first time the defective antibody-dependent cell-mediated cytotoxicity and phagocyte functions of epithelial ovarian cancer–associated MO/MA, which could have implications for immunobiotherapeutic strategies.
Prognostic Significance of Basal-Like Phenotype and Fascin Expression in Node-Negative Invasive Breast Carcinomas
Authors: Rodríguez-Pinilla Socorro María , Sarrió David , Honrado Emiliano , Hardisson David , Calero Francisco , Benitez Javier , Palacios José
Published: March 2006

Purpose: Basal-like phenotype tumors are frequently found among BRCA1 germ-line mutated breast carcinomas. They are biologically aggressive and have a tendency towards visceral metastasis when untreated. Nevertheless, it has been suggested that they respond to chemotherapy better than other types of tumors. Fascin expression has been associated with lung metastasis in breast cancer. The aim of this study was to determine whether basal-like phenotype and fascin were related in both sporadic and familial tumors and with prognosis in node-negative sporadic breast cancers.

Experimental design: 230 nonfamilial and 28 hereditary node-negative invasive breast carcinomas were immunohistochemically analyzed using tissue microarrays. Tumors that were estrogen receptor/HER2 negative and cytokeratin 5/6 and/or epidermal growth factor receptor positive were considered to have a basal-like phenotype.

Results: A basal-like phenotype was found in 11.9% of sporadic cancers. Among patients not receiving adjuvant chemotherapy, a basal-like phenotype was associated with poor prognosis (P = 0.001, log-rank test) whereas no such association was found in patients receiving it. Tumors with a basal-like phenotype showed local recurrence (17.4%) or visceral metastasis (13%) but not bone metastasis (P = 0.001). Fascin expression was observed in 25.1% of sporadic invasive breast carcinomas and was associated with the basal-like phenotype, but not with prognosis or recurrence pattern. Fascin was expressed in 83.3% and 16.7% BRCA1- and BRCA2-associated carcinomas, respectively (P = 0.048).

Conclusions: Basal-like tumors had a tendency towards visceral metastasis and their prognosis was dependent on the use of postoperative chemotherapy. Although fascin expression was associated with the basal-like phenotype, it was not associated with their metastatic behavior. Fascin expression is frequent in BRCA1-associated tumors.
ErbB Receptor Signaling and Therapeutic Resistance to Aromatase Inhibitors
Authors: Shin Incheol , Miller Todd , Arteaga Carlos L.
Published: March 2006

We have investigated the effect of HER-2 overexpression on resistance to the aromatase inhibitor letrozole in MCF-7 breast cancer cells stably expressing cellular aromatase (MCF-7/CA). MCF-7/CA cells overexpressing HER-2 showed a >2-fold increase in estrogen receptor (ER)–mediated transcriptional reporter activity upon treatment with androstenedione compared with vector-only control MCF-7/CA cells. Cotreatment with letrozole did not abrogate androstenedione-induced transcription and cell proliferation in HER-2-overexpressing cells. Chromatin immunoprecipitation assays using cross-linked protein-DNA from MCF-7/CA/HER-2 cells indicated ligand-independent association of the ERα coactivators AIB-1 and CBP to the promoter region of the estrogen-responsive pS2 gene. Upon treatment with androstenedione, there were increased associations of AIB1 and CBP with the pS2 promoter in the HER-2-overexpressing compared with control MCF-7/CA cells. These results suggest that ligand-independent recruitment of coactivator complexes to estrogen-responsive promoters as a result of HER-2 overexpression may play a role in the development of letrozole resistance.
Targeted Modulation of MGMT: Clinical Implications
Authors: Liu Lili , Gerson Stanton L.
Published: March 2006

O6-Methylguanine DNA methyltransferase (MGMT) has been studied for >20 years as a gene that is associated with the mutagenicity and cytotoxicity induced by either methylating carcinogens or alkylating (methylating and chloroethylating) therapeutic agents. Pioneering studies of alkylating agents identified alkylated guanine at the O6 position, the substrate of MGMT, as a potentially promutagenic and lethal toxic DNA lesion. MGMT plays a prominent role in DNA adduct repair that limits the mutagenic and cytotoxic effect of alkylating agents. Because of its role in cancer etiology and chemotherapy resistance, MGMT is of particular interest. In this article, the clinical effect of MGMT expression and targeted modulation of MGMT will be summarized.
Blood Pharmacokinetics of Various Monoclonal Antibodies Labeled with a New Trifunctional Chelating Reagent for Simultaneous Conjugation with 1,4,7,10-Tetraazacyclododecane-N,N′,N″,N‴-Tetraacetic Acid and Biotin before Radiolabeling
Authors: Wang Zhongmin , Mårtensson Linda , Nilsson Rune , Bendahl Pär-Ola , Lindgren Lars , Ohlsson Tomas , Sjögren Hans-Olov , Strand Sven-Erik , Tennvall Jan
Published: March 2006

Purpose: Knowledge of the blood pharmacokinetics of monoclonal antibodies is crucial in deciding the optimal time for starting the administration of a “clearing agent” or using a “clearing device.” The primary purpose was to investigate whether the pharmacokinetics of various antibodies labeled with the same chelator and 111In differed significantly after i.v. injection in immunocompetent rats. A new trifunctional chelator called “1033” containing a biotin and a radiometal chelation moiety is introduced, making it possible to use only one conjugation procedure for the antibody.

Experimental Design: Sixty-five non–tumor-bearing rats were included and divided into four groups (I-IV). The blood pharmacokinetics was investigated for rituximab, BR96, and trastuzumab labeled with 1033 and 111In (I-III). The whole-body activity and activity uptake in muscle, liver, and kidney, which might explain differences in the early pharmacokinetics in blood, were also measured. hMN14 labeled with another chelator [1,4,7,10-tetraazacyclododecane-N,N′,N″,N‴-tetraacetic acid (DOTA)], but with the same radionuclide (111In-biotin-DOTA-hMN14), was studied (IV). The blood pharmacokinetics from another 15 tumor-bearing rats was compared with those of non–tumor-bearing rats (III) by injection of 111In-1033-BR96.

Results: No statistical difference was detected between the groups regarding the blood pharmacokinetics of rituximab, BR96, or trastuzumab. The pharmacokinetics and biodistribution of 111In-biotin-DOTA-hMN14 exhibited a clear difference compared with others. There were no significant differences in the blood pharmacokinetics of 111In-1033-BR96 between tumor-bearing rats and non–tumor-bearing rats.

Conclusions: Different antibodies labeled with the trifunctional chelator 1033 and 111In did not exhibit different blood pharmacokinetics, which means that the pharmacokinetics could be predicted irrespective of the IgG1 antibody chosen. A small tumor burden did not change the pharmacokinetics of the radioimmunoconjugates.
Improving the Delivery of Radionuclides for Imaging and Therapy of Cancer Using Pretargeting Methods
Authors: Sharkey Robert M. , Karacay Habibe , Cardillo Thomas M. , Chang Chien-Hsing , McBride William J. , Rossi Edmund A. , Horak Ivan D. , Goldenberg David M.
Published: March 2006

The article reviews the background and current status of pretargeting for cancer imaging and therapy with radionuclides. Pretargeting procedures were introduced ∼20 years ago as an alternative to directly radiolabeled antibodies. Because they were multistep processes, they were met with resistance but have since progressed to simple and improved procedures that could become the next generation of imaging and therapy with radionuclides. The separation of the radiolabeled compound from the antibody-targeting agent affords pretargeting procedures considerable flexibility in the radiolabeling process, providing opportunities for molecular imaging using γ- or positron-emitting radionuclides and a variety of β- and α-emitting radionuclides of therapeutic applications. Pretargeting methods improve tumor/nontumor ratios, exceeding that achieved with directly radiolabeled Fab′ fragments, particularly within just a few hours of the radionuclide injection. In addition, tumor uptake exceeds that of a Fab′ fragment by as much as 10-fold, giving pretargeting a greatly enhanced sensitivity for imaging. Advances in molecular biology have led to the development of novel binding proteins that have further improved radionuclide delivery in these systems. Studies in a variety of hematologic and solid tumor models have shown advantages of pretargeting compared with directly radiolabeled IgG for therapy, and there are several clinical studies under way that are also showing promising results. Thus, the next generation of targeting agents will likely employ pretargeting approaches to optimize radionuclide delivery for a wide range of applications.
Enhancement of the Therapeutic Index: From Nonmyeloablative and Myeloablative toward Pretargeted Radioimmunotherapy for Metastatic Prostate Cancer
Authors: DeNardo Sally J. , Richman Carol M. , Albrecht Huguette , Burke Patricia A. , Natarajan Arut , Yuan Aina , Gregg Jeff P. , O'Donnell R.T. , DeNardo Gerald L.
Published: March 2006

Purpose: New strategies that target selected molecular characteristics and result in an effective therapeutic index are needed for metastatic, hormone-refractory prostate cancer.

Experimental Design: A series of preclinical and clinical studies were designed to increase the therapeutic index of targeted radiation therapy for prostate cancer. 111In/90Y-monoclonal antibody (mAb), m170, which targets aberrant sugars on abnormal MUC1, was evaluated in androgen-independent prostate cancer patients to determine the maximum tolerated dose and efficacy of nonmyeloablative radioimmunotherapy and myeloablative combined modality radioimmunotherapy with paclitaxel. To enhance the tumor to liver therapeutic index, a cathepsin degradable mAb linkage (111In/90Y-peptide-m170) was used in the myeloablative combined modality radioimmunotherapy protocol. For tumor to marrow therapeutic index improvement in future studies, anti-MUC1 scFvs modules were developed for pretargeted radioimmunotherapy. Anti-MUC1 and anti-DOTA scFvs were conjugated to polyethylene glycol scaffolds tested on DU145 prostate cancer cells and prostate tissue arrays, along with mAbs against MUC1 epitopes.

Results: The nonmyeloablative maximum tolerated dose of 90Y-m170 was 0.74 GBq/m2 for patients with not more than 10% axial skeleton involvement. Metastatic prostate cancer was targeted in all 17 patients; mean radiation dose was 10.5 Gy/GBq and pain response occurred in 7 of 13 patients reporting pain. Myeloablative combined modality radioimmunotherapy with 0.4 GBq/m2 of 90Y-peptide-m170 and paclitaxel showed therapeutic effects in 4 of 6 patients and 30% less radiation to the liver per unit of activity. Neutropenia was dose limiting without marrow support and patient eligibility was a major limitation to dose escalation. Hypoglycosylated MUC1 epitopes were shown to be abundant in prostate cancer and to increase with disease grade. Anti-MUC1 scFvs binding to prostate cancer tissue and live cells were developed into di-scFv binding modules.

Conclusions: The therapeutic index enhancement for prostate radioimmunotherapy was achieved in clinical studies by the addition of cathepsin cleavable linkers to 90Y-conjugated mAbs and the use of paclitaxel. However, the need for marrow support in myeloablative combined modality radioimmunotherapy restricted eligible patients. Therefore, modular pretargeted radioimmunotherapy, aiming at improving the tumor to marrow therapeutic index, is being developed.