bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5506: Trabedersen (AP 12009) for the treatment of patients with advanced tumors: A phase I/II study
Authors: Oettle Helmut , Hilbig Andreas , Seufferlein Thomas , Luger Thomas , Schmid Roland , von Wichert Götz , Endlicher Esther , Garbe Claus , Kaehler Katharina C. , Hauschild Axel , Kiessling Peter , Schmaus Susanne , Heinrichs Hubert , Schlingensiepen Karl-Hermann
Published: June 2012

Background: Transforming growth factor-beta 2 (TGF-β2) regulates key mechanisms of carcinogenesis, in particular immunosuppression and metastasis. The antisense oligonucleotide trabedersen (AP 12009) specifically inhibits TGF-β2 expression. A randomized and active-controlled phase IIb study in high-grade glioma patients showed a clear survival benefit for intratumoral treatment with trabedersen over standard chemotherapy. In this study we evaluate the maximum tolerated dose (MTD), safety, pharmakokinetics, and efficacy of intravenous trabedersen treatment in patients with advanced solid tumors.

Methods: In the open label, multicenter, Phase I/II study, 33 patients with advanced pancreatic carcinoma (PancCa, stage III/IV, N=23), malignant melanoma (stage III/IV, N=5), or colorectal carcinoma (stage III/IV, N=5) were enrolled in cohorts for dose-escalation. Patients were treated intravenously with trabedersen as 2nd to 4th-line therapy as monotherapy with escalating doses in 2 treatment schedules (1st schedule: 7d on, 7d off; 2nd schedule: 4d on, 10d off; up to 10 cycles). After completion of dose-escalation, further patients were enrolled for the treatment with one defined dose in the Phase II part of the study.

Results: Trabedersen showed excellent safety and tolerability. The only expected adverse reaction identified is non-serious, transient and moderate thrombocytopenia. Within the 1st schedule, MTD was established at a dose of 160 mg/m2/d. In the 2nd schedule (4d on, 10d off) a well tolerated dose (140 mg/m2/d) was identified and subsequently further 14 PancCa patients were treated with this regimen. The median overall survival of PancCa patients treated 2nd-line with the 2nd schedule-140 mg/m2/d regimen (N=9) has not yet been reached and currently is 9.2 months (status Oct 2010). Further promising efficacy data include: one PancCa patient had a complete, sustained response of liver metastases and is alive 61 months after monotherapy with trabedersen; another patient is alive 31 months after treatment start (status Oct 2010). Promising efficacy was also observed in malignant melanoma patients with a current median overall survival of 13.8 months.

Conclusions: Currently the study continues with the treatment of 12 patients with malignant melanoma with the 2nd schedule-140 mg/m2/d regimen. A randomized, active-controlled study in PancCA patients is in preparation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5506. doi:10.1158/1538-7445.AM2011-5506
Abstract 5507: Immunological findings in a phase II immunotherapy study using allogeneic lung cancer cells modified to express alpha(1,3)galactosyltransferase in advanced non-small cell lung cancer (NSCLC)
Authors: Rossi Gabriela R. , Vahanian Nicholas N. , Morris John C. , Ramsey Jay W. , Tennant Lucinda , Link Charles J.
Published: June 2012

Background: Alpha(1,3)galactose carbohydrate (αGal) epitopes are highly immunogenic in man. In animal models, vaccination with αGal expressing tumor cells induced rejection of tumor and improved survival. In a phase II anti-tumor immunotherapy study, we evaluated the safety, anticancer activity and immunogenicity of allogeneic human lung cancer cells expressing αGal epitopes HyperAcute-Lung (HAL) vaccines. Methods: Patients with advanced NSCLC received intradermal injections of 300 million HAL vaccine cells every 2 weeks for up to eight doses. Anti-αGal (IgG) and anti-CEA antibodies (Ab) were monitored by ELISA. Cellular immune responses were evaluated by ELISPOT assay (IFN-γ and IL-5). Serum was collected before and at several time points post-immunization. PBMC were collected prior to immunization, after the 4th vaccination and after 8 vaccinations. PBMC were cultured with autologous dendritic cells (DC) pulsed with one of 4 irradiated NSCLC cell lines, three of which were the parental (wild type) cell lines used to generate the vaccine. The fourth was a NSCLC line that was not a component of the vaccine. We compared the reactivity of PBMC stimulated with DC pulsed with these tumor cells pre- and post-immunization in 18 patients with available samples. Results: All patients demonstrated anti-αGal Ab prior to immunization and all responded to vaccination by increasing anti-αGal Ab 2 to 100-fold. Thirty-two patients were tested for anti-CEA Ab. Twenty patients showed significant increases in anti-CEA Ab after immunization. IFN-γ release was detected in 10 of 18 patients evaluated. Patients demonstrating IFN-γ release by ELISPOT after immunization had a median overall survival of 92 weeks (range 40 to 157) comparing favorably with 33.5 weeks (range, 12 to 200) in patients that showed little IFN-γ response. The majority of patients tested showed IL-5 induction after vaccination. In both assays, the HAL1 and HAL2 vaccine cell lines induced greater reactivity on ELISPOT. Patients responding with higher levels of IFN-γ and IL-5 also showed reactivity to the NSCLC cell line that was not a component of the vaccine suggesting that reactivity and possibly cross-priming to shared tumor antigens is induced after vaccination with HyperAcute immunotherapy. Conclusion: Hyperacute anti-tumor immunotherapy induced humoral and cellular responses. Cytokine expression, especially IFN-γ, might be predictive for increased survival.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5507. doi:10.1158/1538-7445.AM2011-5507
Abstract 5513: An immune-active tumor microenvironment favors clinical response to ipilimumab
Authors: Ji Rui-Ru , Hamid Omid , Chasalow Scott D. , Cogswell John , Alaparthy Suresh , Berman David , Shahabi Vafa
Published: June 2012

Ipilimumab, a fully human monoclonal antibody specific to CTLA-4, has shown improvement in overall survival in previously treated advance melanoma patients. As a consequence of blockade of the T cell regulatory molecule CTLA-4, ipilimumab treatment is associated with proliferation and activation of peripheral T cells. An important factor for impacting tumor growth however, is whether these T cells can successfully migrate into the tumor. In order to gain a better understanding of the various tumor-associated components that determine the clinical activity of ipilimumab in melanoma tumors, Affymetrix gene expression profiling and pathway analysis were retrospectively performed on 46 primary or metastatic melanoma tumors, collected in a complete phase II clinical trial in advanced melanoma patients. In biopsies obtained at baseline (before treatment), there were 361 differentially genes expressed (≥ 1.5 fold difference in expression, p ≤ 0.01); most over-expressed transcripts and pathways associated with favorable clinical activity were immune-related. Furthermore, differential analysis of mRNA from pre- and post-treatment tumor biopsies pointed to additional increase in the expression of these genes after treatment. Interestingly, ipilimumab treatment was associated with down-regulation of a number of melanoma-associated transcripts in the tumors within a short time (3 weeks after the first dose). Our preliminary results suggest that high pre-existing immune-activity favors clinical response to ipilimumab. These analyses also provide a list of candidate biomarkers with potential predictive value for response to ipilimumab, to be confirmed in a larger, placebo controlled clinical trial.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5513. doi:10.1158/1538-7445.AM2011-5513
Abstract 5520: Therapeutic vaccination with epitope-enhanced and wild-type TARP peptides in stage D0 prostate cancer
Authors: Wood Lauren V. , Dahut William , Gulley James L. , Madan Ravi A. , Arlen Philip , Parnes Howard L. , Roberson Brenda D. , Steinberg Seth M. , Terabe Masaki , Berzofsky Jay A.
Published: June 2012

Background: T-cell receptor alternate reading frame protein (TARP) is immunogenic, is expressed in ∼95% of prostate cancer specimens and hence is a good candidate for therapeutic vaccination. The immunogenicity of HLA-A*0201 binding TARP peptides can be improved through epitope-enhancement achieved by amino acid substitutions that result in increased HLA binding affinity and immunogenicity at least in mice. We conducted a pilot study to determine the safety and immunogenicity of TARP peptide vaccination and its impact on PSA doubling time (PSADT) in men with Stage D0 prostate cancer.

Methods: HLA-A*0201 positive men were randomized to receive epitope-enhanced (29-37-9V) or wild type (27-35) TARP peptides administered as a peptide emulsion with Montanide ISA 51VG plus GM-CSF (Arm A) or as an autologous peptide-pulsed dendritic cell vaccine (Arm B). Vaccines are administered q 3 weeks for a total of 5 vaccines with an optional 6th dose of vaccine at 36 weeks based on immune response or PSADT criteria and a booster dose of vaccine for all patients at 48 weeks.

Results: 26 patients have enrolled in the study to date (n = 14 Arm A, n = 12 Arm B) with a mean on study duration of 7.5 months. Mean baseline cohort characteristics: age 63 yrs, PSA 4.43, PSADT 7.3 months and Gleason Score 6.7 with 19 of 26 s/p radical prostatectomy. TARP vaccination was safe and well tolerated with adverse events limited to local injection site reactions < Grade 2. Nine of 18 patients (50%) reaching 24 weeks on study were documented to have an increase in their PSADT compared to baseline.

Mean Percent Increase/Prolongation in PSADT:

Surprisingly TARP-specific IFN-γ ELISPOT responses have been detected ex vivo in only 3 of 14 (21%) patients examined to date.

Conclusion: Vaccination with wild type and epitope-enhanced TARP peptides results in prolongation of PSADT in a substantial fraction of patients. The immunologic correlates responsible for PSADT lengthening are poorly understood and require additional study.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5520. doi:10.1158/1538-7445.AM2011-5520
Abstract 5533: A RT-PCR study of cytokine expression with macrophages and tumors
Authors: Syndergaard Chas , Knechtel Daniel C. , Clark Daniel N. , Keller Mary A. , O'Neill Kim , Robison Richard A.
Published: June 2012

Macrophages (MØ) are very important in initiating and directing an immune response. However, it has been noted that when MØ are in the presence of a tumor, they often undergo a change in phenotype. In this environment, MØ will be converted from an aggressive phenotype (M1) to one that promotes tissue remodeling and repair (M2) through a process that is not fully understood. The purpose of this study was to identify which cytokines, in both MØ and cancer cells, show altered expression when the two are in close proximity. To accomplish this, MCF-7 (breast cancer), MDA-MB-231 (breast cancer), MDA-MB-435 (melanoma), and HT29 (colon cancer) cell cultures were grown in prescribed media followed by a 48-hour co-incubation with whole blood MØ. Lipopolysaccharide (LPS) was used as a positive control of an M1-type stimulation. The cytokine levels of both the cancer cells and MØ were then analyzed by quantitative PCR (qPRC) for expression of interleukin-1beta (IL-1β), macrophage migration inhibition factor (MIF), and tumor necrosis factor-alpha (TNF-α). Mathematical calculations were analyzed using a Step One® comparative quantification program and the cytokine expression equation 2⁁-(Δct1- Δct2). MØ IL-1β showed a fold change of 2.76, 1.58, 0.95, 3.09, 10.11 when exposed to MCF-7, MDA-MB-231, MDA-MB-435, and HT29 cells and LPS respectively. These expressions correlated with the aggressiveness of the cancer. Aggressive MDA-MB-435's down regulated IL-1β. Fold change in MIF expression was 3.14, 1.33, 7.56, and 1.22 in MCF-7, MDA-MB-231, MDA-MB-435, and HT29 cells respectively. HT29's had the lowest expression levels suggesting one reason why it's been reported that infiltration of MØ happens properly in colon cancers. Fold expression in MØ expression of TNF-α was 1.34, 1.65, 1.10, and 0.85 with MCF-7, MDA-MB-231, MDA-MB-435, and HT29 cells respectively. An interesting finding that warrants further investigation was in regards to the colon cancer. We observed with interest that the colon cancer line, HT29, caused a decrease in expression of MØ TNF-α. These findings show that colon cancer HT29 decreases TNF-α expression whereas other tested cancer lines did not. This suggests that colon cancers might evade the immune system by regulating MØ TNF-α, an important factor in initiating the caspase-8 regulated apoptotic pathway. These findings suggest that each cancer has an individual method in managing it's microenvironment in respect to MØ.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5533. doi:10.1158/1538-7445.AM2011-5533
Abstract 5539: A predominant serrated pathway in hyperplastic polyposis syndrome generates microsatellite-stable and -unstable colorectal cancers
Authors: Noesel Carel V.
Published: June 2012

Background and aims: Hyperplastic polyposis syndrome (HPS) is characterized by the presence of multiple colorectal serrated polyps and is associated with an increased colorectal cancer (CRC) risk. The mixture of distinct precursor lesion types and malignancies provides a unique model to study the canonical pathway and a proposed serrated CRC-pathway in humans.

Methods: To establish which CRC-pathways play a role in HPS and particularly to obtain new support for the serrated CRC pathway, we analyzed the molecular characteristics of polyps (n=84) and CRCs (n=19) in 17 HPS patients as compared to control groups of various sporadic polyps(n=59) and sporadic microsatellite-stable CRCs(n=16).

Results: In both HPS and sporadic polyps, APC mutations were exclusively identified in adenomas whereas BRAF mutations were confined to serrated polyps. Six of 19(32%) HPS CRCs were identified within a serrated polyp. Mutation analysis performed in both the CRC and the serrated component of these lesions showed identical BRAF mutations. One HPS CRC was located within an adenoma, both components harboring an identical APC mutation. Overall, 10/19(53%) HPS CRCs carried a BRAF mutation, compared to none in control group CRCs (p=0.001). Six of the BRAF-mutated HPS CRCs were microsatellite-unstable (MSI-high) due to MLH1 methylation.

Conclusion: Our findings provide novel supporting evidence for the existence of a serrated CRC pathway which predominates over the classical Wnt pathway of carcinogenesis in HPS.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5539. doi:10.1158/1538-7445.AM2011-5539
Abstract 5552: New models for prostate and colon carcinogenesis in PhIP-treated hCYP1A mice
Authors: Li Guang Xun , Cheung Connie , Loy Shea , Liu Anna B. , Yang Chung S.
Published: June 2012

The food mutagen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine(PhIP), is mainly metabolized by human CYP1A2 through N-CYP1A2 through N2-hydroxylation (activation) and by mouse Cyp1a2 through 4′-hydroxylation (detoxification). In order to mimic human PhIP carcinogenesis in mice, we treated hCYP1A mice (which express human CYP1A but not mouse Cyp1a) with PhIP by oral gavage. Following a dose of PhIP (200 mg/kg b.w.), low-grade prostate intraepithelial neoplasia (PIN) were observed after 20 weeks, high-grade PIN were observed after 30 weeks, and microinvasive carcinomas and adenocarcinomas in ventral, dorsolateral, and coagulating gland were observed after 40 and 50 weeks. Similar to human prostate carcinogenesis, higher than normal GSTP1 levels were observed in proliferative inflammatory atrophic lesions, and GSTP1 expression was lost in high-grade PIN and carcinomas. Under these experimental conditions, colon tumors were not observed. However, in mice that were treated with PhIP (200 mg/kg) and, a week later, followed by treatment with 1.5% dextran sodium sulfate (DSS) in drinking fluid for 1 week, colon tumors were observed after 6 weeks (with 85% tumor incidence and 3.75±0.70 tumors/mouse) and >90% of the tumors were adenocarcinomas. The tumors had overexpressed β-catenin, c-myc, cyclinD1, iNOS and COX-2. In parallel studies with wild type mice treated with PhIP, or PhIP plus DSS, only low-grade PIN, but no colon tumors, were observed. Experiments are ongoing to study the effects of high-fat diet and other dietary factors in modulating carcinogenesis. These models, when fully characterized, could be very useful experimental systems to study diet and prostate/colon cancer (supported by NIH grants CA120915 and CA120915-S2).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5552. doi:10.1158/1538-7445.AM2011-5552
Abstract 5558: Persistent induction of cytochrome P450 (CYP)1A1 in human lung cell line, H358, by 3-methylcholanthrene: Implications for pulmonary carcinogenesis by polycyclic aromatic hydrocarbons
Authors: Moorthy Bhagavatula , Jiang Weiwu , Wang Lihua , Chu Chun
Published: June 2012

Humans are constantly exposed to environmental carcinogenic polycyclic aromatic hydrocarbons (PAHs) through cigarette smoke, diesel exhausts, charcoal-broiled meats, etc. Cytochrome P4501A (CYP1A) enzymes play important roles in the activation of PAHs such as 3-methylcholanthrene (MC) to carcinogenic DNA-binding metabolites, which could in turn lead to pulpmnary carcinogenesis. We recently reported that MC elicits persistent induction of hepatic and pulmonary CYP1A1 in mice by mechanisms other than persistence of the parent MC, and that this phenomenon contributes to pulmonary carcinogenesis. The regulation of CYP1A1 in human lung is not well understood as MC fails to induce CYP1A1 in many lung cells lines, including A549 cells. In this study, we tested the hypothesis that MC elicits persistent induction of CYP1A1 in the human bronchoalveolar cell line, H358. H358 cells were exposed to MC (2.5 µM) dissolved in DMSO, or DMSO alone as controls. At different time points, CYP1A1 activities were determined by measuring the activities of ethoxyresorufin O-dethylase (EROD). CYP1A1 apoprotein and mRNA expression were determined by Western blotting and real time RT-PCR, respectively. MC induced CYP1A1 activities by 8-fold at 8 h, and this effect was persistent even at 72 h. This was accompanied by persistent expression of CYP1A1 apoprotein. Real-time RT-PCR showed that 8 h treatment with 2.5 µM MC induced mRNA of CYP1A1 by 850-fold, and the effect was persistent up to 72 h. These results suggest that the H358 cell line could be an excellent model to study the molecular mechanisms of CYP1A1 regulation by MC in the human lung. Since lung is a target organ for pulmonary carcinogenesis in humans exposed to environmental pollutants such as cigarette smoke, further studies could lead to improved understanding of the mechanisms of pulmonary carcinogenesis, which could in turn lead to the development of rational strategies for the prevention/treatment of lung cancers in humans. (Supported by NIH grant ES009132.)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5558. doi:10.1158/1538-7445.AM2011-5558
Abstract 5567: Short-term pregnancy levels of estradiol delays Her-2 positive mammary tumor development by altering key signaling pathways
Authors: Parada Jacqueline , Arumugam Arunkumar , Myers Stephanie , Boopalan Thiyagarajan , Delgado Alejandro , Agullo Pamela , Lakshmanaswamy Rajkumar
Published: June 2012

Approximately 20-30% of all breast cancers overexpress the human epidermal growth factor receptor 2 (Her2). Breast cancers that overexpress Her2 have been associated with a more aggressive phenotype and decreased disease free survival. It has been demonstrated that uniparous Her2/neu transgenic mice developed smaller mammary tumors and these tumors had significantly decreased metastatic potential despite that this cancer type is phenotypically ER negative. We had earlier demonstrated that short-term treatment with pregnancy levels of estradiol was very effective in delaying tumor progression and decreasing tumor burden in activated Her2/neu transgenic mice. In this present investigation, we have attempted to understand the molecular mechanisms underlying this anti-tumor effect of short-term pregnancy levels of estradiol treatment against Her2/neu driven mammary carcinogenesis. Seven week old activated Her2/neu transgenic mice were treated for 3 weeks with 100micrograms of estradiol in silastic capsules. We have shown that this dose of estradiol treatment mimics pregnancy levels of the hormone in circulation. Control animals received empty silastic capsules for the same duration. Mice were palpated once every week for nine months to monitor the development of mammary cancer. Histopathological examination was performed to confirm the carcinomatous nature of the palpable tumors. A group (n=3) of mice were terminated from control and estradiol treated groups at 6 and 12 weeks post withdrawal of the treatment. Mammary tumors were excised, snap-frozen in liquid nitrogen and stored at -80 C. Protein and total RNA were extracted from these tissues. Pathway focused microarray analysis was done on the PI3K/AKT and P53 pathway. Results from the PI3K/AKT pathway demonstrate that, in estrogen treated mice, proteins and genes involved in tumor-suppression (Mtcp1,Rbl2,Srf and Ywhah) and immune response (Cd14,Tlr4) (which have been shown to play a role in reducing tumors by induction of apoptosis) were up-regulated. Many genes involved in promoting tumorigenesis were significantly down-regulated (Birc5,Ccnb2,Mdm2) while genes involved in cell cycle regulation and apoptosis were upregulated (Dapk1, Cul9, Sesn2).This trend in gene expression demonstrates that short-term pregnancy levels estrogen is altering key pathways leading to the delay and inhibition of Her-2 positive mammary cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5567. doi:10.1158/1538-7445.AM2011-5567
Abstract 5569: The association between statin use and risk of biochemical recurrence in men treated with radical prostatectomy in a cohort of men with inherited forms of prostate cancer
Authors: Ishak Miriam , Cooney Kathleen
Published: June 2012

Background: Evidence exists that statin medications are associated with decreased risk of advanced prostate cancer. The risk factors for prostate cancer are increasing age, African-American race and family history of the disease. To date, studies of the association between statin medication and risk of prostate cancer have not considered men with familial or hereditary prostate cancer.

Methods: Men enrolled in the University of Michigan Prostate Cancer Genetics Project (PCGP) who had been previously diagnosed with prostate cancer and treated with radical prostatectomy were surveyed as to their past and current statin medication use status, type of statin medication, duration of statin medication use and their incidence of biochemical recurrence. PCGP is a large, family-based study of inherited forms of prostate cancer. The PCGP enrollment criteria included families with two or more living-affected family members in a first-degree or second-degree relationship. The risk of biochemical recurrence (BCR) was modeled using logistic regression models and odds ratios and 95% confidence intervals were calculated.

Results: Of 595 unrelated men with past radical prostatectomy surveyed, 91 men experienced BCR. At time of survey, 40% of men were current statin users. Median follow-up time was 67 months for statin users was 75 months for non-statin users. Current statin-use at time of recurrence was associated with a decreased risk of BCR (OR = 0.685; 95% C.I.:0.438 – 1.072; p=0.09); while ever statin-use was not associated with risk of BCR in multivariate models adjusted for age and BMI.

Discussion: This represents the only study to date to assess statin medication use and the risk of biochemical recurrence in men with familial and hereditary prostate cancer. Given the importance of family history as a risk factor for prostate cancer, these findings should be confirmed in other studies and may implicate subgroups of men at risk for prostate cancer who may benefit from statin medication use.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5569. doi:10.1158/1538-7445.AM2011-5569
Abstract 5575: Tetrandrine blocks emergence of castrate resistant phenotype in prostate cancer by promoting apoptosis and sensitizing prostate cancer cells to androgen deprivation
Authors: Koul Sweaty , Koul Hari K.
Published: June 2012

Prostate cancer (PCa) is the second leading cause of cancer deaths in men, and the majority of prostate cancer deaths are a result of emergence of castrate resistant phenotype. One of the six hallmarks of cancer is resistance to apoptosis, and we hypothesized that resistance to apoptosis is in part responsible for emergence of castrate resistant phenotype in prostate cancer. Specifically we investigated the effects of Tetrandrine (TET), a bis-benzylisoquinoline alkaloid isolated from the root of Stephania tetrandra, on apoptosis of PCa cells in vitro. TET decreased viability of PCa cells. Long-term treatment with TET resulted in a dramatic increase in sub G0/G1 cell population suggestive of apoptosis. Treatment with TET resulted in decrease in anti-apoptotic proteins (Bcl-xl and survivin) and concomitant activation of caspase 3 and PARP cleavage. Moreover, Tet treatment induced DNA fragmentation and nuclear condensation, all hallmarks of apoptosis. We also observed that pretreatment with Tet sensitized LNCaP cells to androgen depletion and prevented emergence of castrate resistant phenotype in vitro culture. Moreover, Tet induced apoptosis in LNCaP C-4 and C4-B (AR positive castrate resistant prostate cancer) cells, indicated effectiveness against established castrate resistant phenotypes in prostate cancer. These results show that Tet by promoting apoptosis of prostate cancer cells and by sensitizing prostate cancer cells to androgen depletion may serve as a chemopreventive/ therapeutic agent in prostate cancer. If these results hold true in additional in vivo studies, Tet may serve as a mechanism based agent against emergence of castrate resistant prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5575. doi:10.1158/1538-7445.AM2011-5575
Abstract 558: Targeting tumour heparanase activity using fully human monoclonal antibodies
Authors: Rathanaswami Palaniswami , Pan Oscar , Ross Sarah , King Chadwick , Foltz Ian N. , Elvin Paul
Published: June 2012

Modification of heparan sulfate proteoglycans (HSPG) in the tumour microenvironment can have a significant influence on tumour cell behavior, tumour growth and metastasis. HSPG play an important role in shaping the integrity of the extracellular matrix, and cell surface HSPG are determinants of cell-cell interactions and receptor function. Heparanase (Hpa1), an endoglycosidase, cleaves the glycosidic bonds within heparan sulfate domains resulting in HSPG with impaired or altered functionality. Elevated expression of heparanase has been correlated with disease progression and poor survival in a number of human cancers. Using Xenomouse™ technology and immunising with recombinant human Hpa1, we have generated fully human monoclonal antibodies that bind to heparanase with high affinity (<100pM). A number of antibodies were shown to inhibit recombinant human, murine and cynomolgus heparanase activity in a HTRF (homogeneous time resolved fluorescence) heparan sulfate degradation assay with IC50s in the nM range (0.2-3nM) for human Hpa1. The antibodies also inhibited cell surface localised heparanase activity and appeared to interfere with secretion and activation of heparanase. Targeting heparanase activity with an inhibitory therapeutic antibody may be a promising alternative strategy to the use of heparan sulfate mimetics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 558. doi:10.1158/1538-7445.AM2011-558
Abstract 5582: The inhibitory effects of Magnoliae Flos extract and its active components on receptor activator of nuclear factor-κB ligand-mediated osteoclastogenesis and on expression of osteolytic factors in breast cancer cells
Authors: Jun Ah Young , Park Kwang-Kyun , Chung Won-Yoon
Published: June 2012

Breast cancer is the first of the top ten cancer killers among the female population of OECD member countries. Breast cancer cells have spread to bone easily. The bone is the preferred site of metastasis for breast cancer and is associated with significant morbidity and poor prognosis. The interaction between breast cancer cells and the bone microenvironment results in a ‘vicious cycle’ that increases both bone destruction and tumor burden. Breast cancer induces up-regulation of receptor activator of nuclear factor-κB ligand (RANKL) in osteoblast by several osteolytic factors derived from breast cancer cells, and subsequently osteoclast formation through the interaction between RANKL in osteoblasts and RANK in osteoclast precursors. Growth factors, such as transforming growth factor-β and insulin-like growth factor-1, are released from bone matrix by osteoclast-derived bone erosion. Natural plant-derived components have received much attention as sources of potential therapeutic and preventive drugs for human diseases. Magnoliae Flos (MF), the buds of Magnolia denudata Desr., is a commonly used Asian medicinal herb for symptomatic relief of allergic rhinitis, sinusitis and headache. To evaluate the inhibitory effects of MF extract and its active components (aschatin, fargesin, lirioresinol B dimethyl ether and magnolin) on cancer-associated bone disease, we first examined their effects on RANKL-induced osteoclastogenesis and bone resorption. MF extract and active components inhibited RANKL-mediated osteoclast differentiation in mouse bone marrow macrophages in a dose dependant manner without any evidence of cytotoxicity, and bone-resorbing activity of osteoclast. In addition, MF extract and active components suppressed the mRNA expression of osteolytic factors in MDA-MB-231 human breast cancer cells, but did not affect the viability of MDA-MB-231 cells. These results demonstrate that MF extract and its active components has the potential to serve as beneficial supplements to treat and prevent cancer-induced bone diseases in patients with metastatic breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5582. doi:10.1158/1538-7445.AM2011-5582
Abstract 5585: Generation of novel cell lines for analyzing the role of TLR4 in breast cancer
Authors: Korte Mike , Rajput Sandeep , Ran Sophia
Published: June 2012

Background: Toll like receptor 4 (TLR4) is a transmembrane receptor expressed in macrophages that plays a major role in pathogen recognition and activation of innate immunity. TLR4 signaling has been linked to tumorigenesis and promotion of metastasis in ovarian, lung, prostate and head-neck cancers through up-regulation of the expression of inflammatory cytokines (e.g., IL-6, IL-8 and TNF-alpha). These factors increase cancer cell survival and resistance to chemotherapy through up-regulation of anti-apoptotic and down-regulation of apoptotic proteins, respectively. The goal of this study was to generate new isogenic cell lines that would allow analysis of the TLR4 role in paclitaxel resistance in breast cancer. To this end, we used TLR4-positive and negative lines (MDA-MB-231 and HCC1806, respectively) to generate stable TLR4 knockdown and over-expressing line derivatives.

Methods: To generate a TLR4 knockdown line, MDA-MB-231 cells were transfected with pisiRNA-TLR4 plasmid and selected with zeocin (50μg/ml). To generate a TLR4 over-expressing line, HCC1806 cells were transfected with pKT2-TLR4-PURO plasmid and selected with puromycin (1μg/ml). Ten to 20 monoclonal colonies were selected from each line and characterized for TLR4 expression and TLR4-dependent cell activities by qRT-PCR and western blot.

Results: TLR4 mRNA expression was reduced by >90% as compared with parental line in five out of 8 MDA-MB-231TLR4− clones examined. One clone had 60% reduction and two clones had no effect on TLR4 mRNA determined by qRT-PCR. TLR4 mRNA was up-regulated 1,000-12,000 fold in five (out of 11) clones derived from HCC1806 line. Two clones overexpressed TLR4 mRNA by 300-500 fold and four clones had increased TLR4 expression by 1-6 fold. All selected clones were analyzed for TLR4 functional signaling by exposure to LPS (1μg/ml), which was reduced by 50-70% in MDA-MB-231TLR4− clones and increased by 15-25 folds in HCC1806TLR4+ sub-lines, as compared with cells transfected with empty vectors.

Conclusions: These data show that we successfully created sub-lines of MDA-MB-231 with stable knockdown of TLR4 and sub-lines of breast carcinoma HCC1806 line with stable over-expression of TLR4. Ectopically expressed human TLR4 as well as TLR4 downregulating sequence had significant functional effects on the TLR4 signaling as demonstrated by increase and decrease of LPS-activated targets (IL-6, IL-8 and TNF-alpha) in MDA-MB-231TLR4− and HCC1806TLR4+ sub-lines, respectively. These new sub-lines will serve as excellent tools for analyzing the role of the TLR4 pathway signaling in breast cancer progression, metastasis and resistance to therapy.

* These authors contributed equally in this work.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5585. doi:10.1158/1538-7445.AM2011-5585
Abstract 5591: A homologous recombination assay for classifying BRCA2 missense variants
Authors: Guidugli Lucia , Goldgar David , (Shane) Pankratz Vernon S. , Erding Catherine A. , Couch Fergus J.
Published: June 2012

Germline mutations in BRCA2 predispose to breast, ovarian and other cancers. Although most mutations in the gene are known to be pathogenic, there is an increasing number of missense variants for which the contribution to cancer risk is unclear. These missense substitutions are referred to as VUS (variants of uncertain significance). Because loss of BRCA2 HR function leads to defective DNA double-strand break repair and an increased risk of cancer, we evaluated the utility of a cell-based Homologous Recombination (HR) assay for determination of the cancer relevance of BRCA2 VUS.

Specifically, we studied VUS localized in the BRCA2 DNA binding domain (DBD) for effects on HR. We transfected full length wild-type and mutant BRCA2 constructs into V-C8 BRCA2 deficient cells along with an I-Sce1-expressing plasmid which induces an unique double strand break in an integrated DR-GFP reporter plasmid. We evaluated the ability of each BRCA2 mutant to repair the induced break by quantifying the GFP positive cells, generated by HR repair of the DNA break, with fluorescent activated cell sorting (FACS).

To establish the sensitivity and specificity of the HR assay we evaluated a series of known deleterious and known neutral with the HR assay. These mutations of known effect were identified using a multifactorial likelihood approach, based on available family data, with a prior probability approach based on sequence conservation (A-GVGD) algorithm (http://agvgd.iarc.fr/agvgd_input.php). We calculated the total odds of pathogenicity for co-segregation of variants with the disease in affected carrier families, co-occurrence of VUS (in trans) with deleterious mutations and for family history of cancer of VUS and combined this odds with the A-GVGD based prior probabilities to generate the posterior probability of pathogenicity for each variant. Using this approach we successfully classified 31 VUS as either neutral or deleterious. By comparing these data with results from the HR assay we established that the assay is 100% sensitive (95% CI: 75.3% – 100%) and 100% specific (95% CI: 82.4% – 100%) for deleterious mutations in the BRCA2 DBD domain.

In summary we evaluated the sensitivity and specificity of the HR assay for VUS in the DBD relative to a probability model based on family data and A-GVGD algorithm and we showed that the assay was highly sensitive and specific. Based on these results we propose that the HR assay can be used as a classifier of the clinical relevance of the VUS in the absence of direct genetic and family data.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5591. doi:10.1158/1538-7445.AM2011-5591
Abstract 560: The use of molecular biomarkers to quantify drug distribution in solid tumors
Authors: Saggar Jasdeep K. , Tannock Ian F.
Published: June 2012

The tumor microenvironment plays a critical role in the efficacy of chemotherapy. The disorganized vasculature within solid tumors leads to decreased drug delivery to the tumor and poor penetration of drugs from blood vessels to the tumor cells. Our laboratory has shown limited distribution of the auto-fluorescent drugs doxorubicin and mitoxantrone using quantitative immunohistochemistry. However, most anti-cancer drugs are not fluorescent and do not have antibodies that recognize them. The aim of the present study is to characterize the spatial distribution within solid tumors of biomarkers of drug effect: cleaved caspase 3 (a marker of apoptosis), γH2aX (a marker of DNA damage) and Ki67 (a marker of cell proliferation) in relation to blood vessels (recognized by an antibody to CD31). We have quantified the time-dependent distribution of these biomarkers in two models: human breast cancer MDA-MB-231 xenografts and murine mammary EMT6 syngeneic tumors following treatment with doxorubicin and melphalan respectively. Biomarker distributions have been compared to distributions of doxorubicin fluorescence. Melphalan's distribution within a solid tumor has been assessed for the first time with the use of a melphalan-DNA adduct recognizing monoclonal antibody. These studies are designed to provide a basis for selecting biomarkers to reflect the distribution of other non-fluorescent agents. The distribution of doxorubicin decreases exponentially with increasing distance from blood vessels, and is similar to the distribution of γH2aX at 10 minutes after drug injection. The activation of cleaved caspase 3 (indicating apoptosis) at 24 and 48 hours following treatment also mirrors the distribution of doxorubicin. The analysis of biomarker distributions in EMT6 tumors following treatment with melphalan in relation to tumor blood vessels is in progress. The results of these studies will be applied to quantify the intratumoral distribution of other non-fluorescent anticancer drugs (hypoxia-activated pro-drugs).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 560. doi:10.1158/1538-7445.AM2011-560
Abstract 5600: Genetic variants associated with reproductive aging and the risk of breast cancer
Authors: He Chunyan , Chasman Daniel , Ruiter Rikje , Dreyfus Jill , Porcu Eleonora , Hwang Shih-Jen , Hankinson Susan , Buring Julie , Stolk Lisette , Franceschini Nora , Sannna Serena , Lunetta Kathryn , Ridker Paul , Stricker B , Demerath Ellen , Crisponi Laura , Murabito Joanne , Hunter David
Published: June 2012

Early menarche and later menopause are well-established risk factors for breast cancer. Recent genome-wide association studies have identified several novel loci for these two traits. However, the association between these loci and breast cancer risk remains unknown. In this study, we investigated 19 and 17 newly identified SNPs from ReproGen Consortium that were associated with age at menarche and age at natural menopause, respectively, and assessed their association with breast cancer in 6 population-based studies. We further used these SNPs to calculate genetic risk scores (GRSs) based on their association with each trait. After adjustment for age and potential population stratification, two age-at -menarche associated SNPs (rs1079866 and rs7821178) and one age-at-natural-menopause associated SNP (rs2517388) were associated with an increased risk of breast cancer (p values, 0.003, 0.009 and 0.023, respectively); although overall we did not observe significant association between the GRSs and breast cancer risk, the 4th and 5th highest quintile of the menarche GRS had relative ratios of 1.14 (CI, 1.01 to 1.28) and 1.13 (CI, 1.00 to 1.27), respectively, compared to the lowest quintile; and these associations did not appear to be attenuated by controlling for age at menarche, age at natural menopause, or other conventional risk factors. Our study suggested that three genetic variants, independent of their association with onset of menarche or natural menopause, were associated with breast cancer risk, and a GRS that combines multiple menarche-associated SNPs might be useful for identifying a high risk subgroup for breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5600. doi:10.1158/1538-7445.AM2011-5600
Abstract 5601: The role of UDP-Glucuronosyltransferases in human breast cancers
Authors: Starlard-Davenport Athena , Lyn-Cook Beverly , Word Beverly , Pogribny Igor
Published: June 2012

Reports of an unanticipated increase in cancer risk following exposure to tamoxifen and estrogen-containing drugs suggest that long-term consequences of genetic variations in drug metabolism may need to be elucidated more thoroughly during product development. Variations in UGTs that metabolize these drugs may result in reduced drug efficacy in estrogen target tissues, such as the breast in women. In this study, we characterized the expression of UGTs in normal (n = 31) and malignant (n = 17) human breast tissues from African American and Caucasian women. Several UGT isoforms, including UGT1A1, -1A4, -1A10, and -2B7 mRNA were down-regulated in malignant human breast specimens compared to normal breast specimens as determined by real-time PCR. In addition, UGT1A4, which is the major UGT responsible for conjugating tamoxifen, was genotyped for the UGT1A424Pro/48Val and UGT1A424Thr/48Leu variant. However, the UGT1A448Val variant was not significant prevalent within these population of samples. It is anticipated that this study will provide novel information on the role of UGTs in the breast and provide insight as to how alterations in UGT expression will impact the clearance of estrogens and tamoxifen in women. It is anticipated that this study will be instrumental in predicting tamoxifen toxicity and estrogen carcinogenicity in women with a non-functional UGT variant.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5601. doi:10.1158/1538-7445.AM2011-5601