bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3526: Inhibition of ABCB1 (MDR1) expression by an siRNA nanoparticulate delivery system to overcome drug resistance in osteosarcoma
Authors: Susa Michiro , Iyer Arun K. , Liu Xianzhe , Choy Edwin , Hornicek Francis J. , Mankin Henry , Milane Lara , Amiji Mansoor M. , Duan Zhenfeng
Published: February 2011

Background: Osteosarcoma accounts for approximately 60% of primary malignant bone tumors diagnosed in the first two decades of life. The use of neo-adjuvant chemotherapy in treating osteosarcoma has improved patients’ average 5 year survival rate from 20% to 70% in the past 30 years. However, for patients who progress after chemotherapy, its effectiveness diminishes due to the emergence of multi-drug resistance (MDR) after prolonged therapy. One of the innovative approaches to addressing MDR is to inhibit MDR1 mRNA expression by RNA interference (RNAi). In order to overcome both the dose-limiting side effects of conventional chemotherapeutic agents and the therapeutic failure resulting from MDR, we designed and evaluated a novel drug delivery system for MDR1 siRNA delivery.

Materials and Methods: Novel biocompatible, lipid-modified dextran-based polymeric nanoparticles were used as the platform for MDR1 siRNA delivery; and the efficacy of combination therapy with this system was evaluated. The mean particle size of the MDR1 siRNA loaded nanoparticles as determined by dynamic light scattering (DLS) measurement was 104.4+3.7 nm and the zeta potential was almost neutral (−0.19+1.13 mV). Multi-drug resistant osteosarcoma cell lines (KHOSR2 and U-2OSR2) were treated with the MDR1 siRNA nanocarriers and MDR1 protein (P-gp) expression, drug retention, and immunofluorescence were analyzed. Combination therapy of the MDR1 siRNA loaded nanocarriers with increasing concentrations of doxorubicin was also analyzed.

Results: We observed that MDR1 siRNA loaded dextran nanoparticles efficiently suppresses P-gp expression in the drug resistant osteosarcoma cell lines. With the delivery of nanoparticles loaded with MDR1 siRNA, growth inhibition with doxorubicin was substantially more marked than with the administration of 100 fold higher amounts of free drugs. The results also demonstrated that this approach may be capable of reversing drug resistance by increasing the amount of drug accumulation in MDR cell lines. Furthermore, subcellular distribution of doxorubicin in MDR cells mimicked that of the drug sensitive variant when nanoparticles were utilized.

Conclusions: Lipid-modified dextran-based polymeric nanoparticles are a promising platform for siRNA delivery. Nanocarriers loaded with MDR1 siRNA are a potential treatment strategy for reversing MDR in osteosarcoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3526.
Abstract 3561: The radiosensitizing and cytotoxic effects of PBS-1086, a Rel inhibitor of NF-KB, in breast cancer cells
Authors: Hunter Jill E. , Willmore Elaine , Irving Julie A E , Sliskovic D. Robert , Monie Dileep , Bobb Kathryn , Zhang Jie , Durkacz Barbara W.
Published: February 2011

Recent RNA interference screening has identified p53 and ras mutations to be synthetic lethal with the canonical and non-canonical pathways of NF-κB. We have synthesized inhibitors that antagonize bindings of RelA (p65), RelB and c-rel to the NF-κB sites and thus inhibit both the canonical and non-canonical paths. One such compound, PBS-1086, is evaluated here as a radiosensitizer in wild type and p65−/− mouse embryonic fibroblasts (MEFs), and as a radiosensitizing and chemotherapeutic agent in the p53 deficient and KRAS (G13D) mutant breast cancer cell line, MDA-MB-231.

We found that p65−/− MEFs were ≥ 2-fold more sensitive than p65+/+ MEFs to irradiation (IR) in clonogenic survival assays, thus demonstrating that NF-κB activation conferred radio-resistance. Incubation with PBS-1086 at 1.4 µM, a non-cytotoxic concentration, radiosensitized p65+/+ MEFs (LD50 value of 1.5 Gy for IR + PBS-1086 compared with 2.3 Gy for IR alone). PBS-1086 had no effect on radio-sensitization of p65−/− MEFs, which indicated the radiosensitizing effect of PBS-1086 was mediated through p65. Similar radiosensitizing effects were observed by using RNA interference technique to down-regulate the expression level of p65 by > 95% in p65+/+ MEFs but not in p65−/− MEFs. We also confirmed the inhibition of the NF-κB by PBS-1086 or p65 siRNA in MEFs. NF-κB-dependent luciferase expression increased by 2.5 fold after 10 Gy IR in p65+/+ MEFs. Treatment with PBS-1086 or p65 siRNA reduced luciferase expression significantly. Similarly, IR increased p65+/+ MEFs nuclear extract NF-κB DNA binding activities by 2 fold, which was significantly decreased by either PBS-1086 incubation or p65 siRNA treatment in these cells.

Next, we studied PBS-1086 in MDA-MB-231 breast cancer cells with constitutive NF-κB activation. First, we tested PBS-1086 as a single agent on the growth of the breast cancer cells. Significant cytotoxic effect to PBS-1086 was observed with an LD50 value of 0.4 μM to the MDA-MB-231 cells. Then we tested synergistic effect of PBS-1086 with irradiation in these cells. Incubation with 0.2 µM PBS-1086 significantly enhanced radiation effect by 1.5 fold (LD50 value of 1.66 Gy IR for IR + PBS-1086 compare with 2.55 Gy for IR alone).

In conclusion, we have demonstrated that a Rel inhibitor of NF-KB can potentiate radioactivity through blocking the canonical pathway. It can also selectively kill tumor cells with p53 and KRAS mutations, perhaps through blocking the canonical and non-canonical pathways.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3561.
Abstract 3568: Treatment of Organ-Specific Chloroleukemia by Cytotech Labs API 31510 and Chemotherapy
Authors: Narain Niven R. , Mauro Lucia M. , McCook John P. , Jimenez Joaquin J.
Published: February 2011

The development of chloroleukemia in the rat after transplantation leads to 100% mortality. Recently, two novel clones have been developed. Following intraperitoneal injection, one clone preferentially migrates and forms tumors in the lungs (CHL-L), while another clone forms tumors in the liver (CHL-Li). In vitro, proliferation of both clones were inhibited in a dose-dependent manner by a novel IV solution of API 31510. Accordingly, in the present study we examined the hypothesis that Cytotech Labs API 31510 alone or in combination with cyclophosphamide (CTX) could be effective in the treatment of both clones in vivo. In the first experiments, 24 rats were randomized into 4 groups of six rats each and injected intraperitoneally with CHL-L. In the control group, 100% developed CHL-L. In the group receiving CTX, 5 out of 6 developed CHL-L, while 1 animal remained disease-free. In the group receiving API 31510, 3 out of 6 rats remained disease-free and 3 out of 6 had a partial therapeutic response. In the group which received API 31510 and CTX, 100% of the animals remained disease-free. In a separate experiment, the same protocol was followed using CHL-Li. As observed with the other clone, animals injected with all control animals developed CHL-Li tumors. Similarly, in the group receiving CTX alone, 5 out of 6 developed CHL-Li and 1 remained disease-free. In the group receiving API 31510, 3 out of 6 animals remained disease-free and 3 out of 6 had a partial response. When combining 31510 and CTX, 5 out of 6 animals remained disease-free and 1 had a partial response. No side effects were observed in the animals receiving API 31510 as evidenced by weight gain and behavior. Taken together, these results indicate that API 31510 as a single agent was effective against both chloroma clones used and as an adjuvant to chemotherapy, API 31510 exhibited treatment efficacy in over 90% of animals treated. These data provide a strong foundation for further development of Cytotech Labs API 31510 intravenous formulation as a viable agent for cancer.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3568.
Abstract 3570: Maitake mushroom extract induces apoptosis in breast cancer MCF7 cells through BAK1 activation
Authors: Soares Raquel , Meireles Manuela , Obiol Diego , Alonso Eliana N. , Rocha Ana , Pirraco Ana , Balogh Gabriela Andrea
Published: February 2011

Background: Maitake mushroom has various bioactive compounds. D-fraction, a protein-bound polysaccharide compound, seems to be the most potent for enhancing the immune system via oral administration or injection. In previous studies, the combination of this extract with chemotherapy suggested that the D-Fraction may have the potential to decrease the size of lung, liver, and breast tumors in cancer patients.

Objective: The aim of this study was to examine the in vitro effect of maitake D- Fraction on breast cancer MCF7 cells.

Methods: Cultures of MCF7 cells, incubated with increased concentrations of liquid maitake D-fraction, 0mg/ml; 18,3pg/ml; 36pg/ml; 91pg/ml; 183pg/ml; 367pg/ml, were evaluated for viability, by MTS assay and apoptosis, by TUNEL assay. All experiments were performed in triplicate.

Results: Results seem to indicate that maitake-treated cells have higher apoptosis levels and poorer viability when compared to non-treated cell cultures.

Conclusion: The results obtained in this study, focusing the effect of maitake D- fraction in breast cancer cell proliferation, can drive us to new therapeutic strategies against breast cancer and show that we can find a lot of bioactive compounds in traditional food.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3570.
Abstract 3571: The Thioredoxin Reductase Inhibitor Chaetocin has Potent Antineoplastic Effects in Solid Tumors
Authors: Isham Crescent R. , Tibodeau Jennifer D. , Merchan Jaime R. , Bible Keith Christopher
Published: February 2011

Background: We had previously reported that the natural product chaetocin has potent and selective in vitro, ex vivo and in vivo anti-myeloma activity attributable to the imposition of cellular oxidative stress (ROS) in part mediated via competitive inhibition of the redox enzyme thioredoxin reductase. Having also observed chaetocin-induced cytotoxicity in solid tumor cell lines, we now extend prior work to characterize the effects of chaetocin in solid tumor cell lines and in human umbilical vein endothelial cells (HUVECs). Methods: The effects of chaetocin in solid tumor cell lines were assessed using colony forming assays, trypan blue exclusion assays, apoptosis and autophagy assays, electron microscopy, transcriptional profiling, and the National Cancer Institute 60 cell line screen. Results: Chaetocin demonstrated potent anti-cancer activity in all assessed solid tumor cell lines with IC50 values between 2-10 nM (24 h exposures, colony forming assays). While apoptosis was induced in a cell line-dependant fashion, it was not required for chaetocin-induced cytotoxicity, as ZVAD-fmk prevented apoptosis but not cell death. Markers of autophagy were not altered by chaetocin treatment. Interestingly, results form the NCI 60 cell line screen showed that hematological cell lines were generally more resistant to chaetocin than solid tumor lines despite our prior report indicating the activity of chaetocin in myeloma. Transcriptional profiling results were consistent with those anticipated from an agent producing cell death via imposition of cellular ROS, with heme oxidase-1 prominently induced along with other transcripts in pathways related to inflammatory response and cell death. Results from OxyBlot protein oxidation kit analyses (Millipore, Billerica, MA) confirmed a generalized increase in the carbonyl modification of proteins, a hallmark of cellular oxidative damage, in response to chaetocin treatment. Experiments using Rho0 mitochondrial inactive cells indicated that cellular ROS is induced by chaetocin independent of respiratory functional mitochondria. Chaetocin was also shown to block the interleukin-, fibroblast growth factor- or EGM2 media-induced proliferation of HUVEC cells at low-nanomolar concentrations. Conclusions: Chaetocin has wide-ranging antineoplastic activity across not only hematological, but also solid tumor, cell lines and displays evidence of antiangiogentic activity in HUVEC proliferation assays. Overall, chaetocin appears to be an attractive agent for further development as a candidate anti-cancer therapeutic in a variety of neoplasms. Supported in part by CA125750.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3571.
Abstract 3585: Taqman genotyping assays can be used on decalcified and paraffin embedded tissue from patients with osteosarcoma
Authors: Hagleitner Melanie M. , Coenen Marieke J.H. , Jeuken Judith W.M. , Flucke Uta , Schreuder Bart H.W. , Loo Maroeska D.W.M. te , Hoogerbrugge Peter M.
Published: February 2011

Insight in genetic variants predicting treatment outcome and/or side effects related to cancer medication will contribute to optimize treatment for the individual patients. However, most clinical trials performed so far in cancer treatment did not routinely collect and store patient germ line DNA. Formalin-fixed paraffin-embedded tissues (FFPE) from archived clinical samples could provide a source of germ line DNA for genetic research. Specially, in cancers like osteosarcoma with a 5-year overall survival of approximately 50-60%, archived histological specimens are often the only accessible source of biological material. This material, however, generally has been decalcified and paraffin-embedded for many years. DNA from formalin-fixed and paraffin embedded (FFPE) tissues, specially, in decalcified FFPE tissues has a higher frequency of non-reproducible sequence alterations compared with germ line DNA. In our study, we investigated whether DNA obtained from these tissues can be used for reliable single nucleotide polymorphism (SNP) genotyping. Therefore, we analyzed whether the genotypes obtained from DNA of these preserved tissues, were similar as those found in freshly DNA isolated from peripheral blood cells from the same patients. We studied two SNPs in the drug transporter MDR1 (ABCB1) using Taqman® SNP genotyping assays. Genotypes of the germ line DNA derived from freshly isolated DNA of 20 surviving patients with osteosarcoma were compared with genotypes obtained from archived material from decalcified FFPE blocks of the same patients. Decalcified FFPE-derived DNA yielded smaller PCR fragments compared to DNA extracted from peripheral blood cells, with a maximum reliable size of ∼300bp. We were able to evaluate each SNP in 19 of the 20 cases included in this study. All successfully genotyped samples showed 100%-concordance between genotypes obtained from DNA of FFPE tissue and the genotypes obtained from DNA of blood. In conclusion, we have demonstrated that decalcified FFPE tissue can be used for genetic polymorphism analysis using Taqman® allelic discrimination assays. This forms an unique opportunity to combine new insights in genetic research with historical patient cohorts, for instance of patients with osteosarcoma.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3585.
Abstract 3589: Characterization of 17-dihydroexemestane glucuronidation. A role for the UGT2B17 deletion in exemestane pharmacogenetics
Authors: Sun Dongxiao , Chen Gang , Dellinger Ryan W. , Sharma Arun K. , Lazarus Philip
Published: February 2011

Exemestane is a 3rd-generation aromatase inhibitor (AI) used in the treatment of breast cancer in postmenopausal women. Reduction to form 17-dihydroexemestane and subsequent glucuronidation to hydroexemestane-17-O-glucuronide is a major pathway for exemestane metabolism. In the present study, homogenates from a newly-developed HEK293 aromatase-overexpressing cell line (HEK293-aro) were used to examine the activity of exemestane versus 17-dihydroexemestane in vitro. 17-dihydroexemestane exhibited an inhibition of estrone formation in HEK293-aro cell homogenates (IC50=2.3±0.83 μM) that was similar to that observed for exemestane (IC50=1.4±0.42 μM). UGT-overexpressing cell lines were screened for activity against 17-dihydroexemestane and UGTs 2B17>1A10>1A8>1A4 were the only UGTs that exhibited activity. Of the hepatic UGTs, UGT2B17 exhibited a 17-fold higher Vmax/KM than UGT1A4. In human liver microsome (HLM) specimens, the rate of hydroexemestane-17-O-glucuronide formation using 17-dihydroexemestane as substrate was shown to be significantly (p<0.001) decreased (14-fold) in HLMs from subjects with the UGT2B17 (0/0) deletion genotype as compared with HLMs from subjects wild type (+/+) for UGT2B17. A significant (r2=0.72) correlation was observed between HLM hydroexemestane-17-O-glucuronide formation and liver UGT2B17 expression in the same subjects, and the KM for UGT2B17(+/+) HLMs against 17-dihydroexemestane was 7.3 μM, ∼ 2-fold lower than that observed for UGT2B17 against 17-dihydroexemestane in vitro (14.5 μM). UGT2B17 (0/0) HLMs exhibited a 1.7-fold higher KM and a 36-fold lower Vmax/KM than UGT2B17

(+/+) HLMs. These data suggest that UGT2B17 is important in the metabolism of 17-dihydroexemestane, a major active metabolite of exemestane, and that the UGT2B17 deletion could have an important role in overall patient response to exemestane.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3589.
Abstract 3591: Efficacy and distribution of intratumoral paclitaxel given alone or in combination with doxorubicin using hydrogel in nude mice
Authors: Lee Joo-Ho , Kim Kwang-Suck , Kim Soo-Yeon , Kim Yu Jin , Al-abd Ahmed M. , Hong Ki-Yun , Song Soo-Chang , Kuh Hyo-Jeong
Published: February 2011

Intralesional chemotherapy is suggested to improve local tumor control as well as systemic toxicity profile of antitumor agents against solid tumors. Thermosensitive poly-(organophosphazene) hydrogel is a novel injectable polymer that transforms from sol to gel at body temperature. In this study, we evaluated distribution and efficacy of paclitaxel (PTX) when given as intratumoral injection using the hydrogel or solution formulation in human SNU-601 tumor xenograft-bearing nude mice. Following intratumoral injection of 60 mg/kg of PTX, plasma drug concentrations were lower than 0.5 μg/ml(LOQ) for both hydrogel and solution. For PTX tumor concentration, Cmax was 1.2 folds higher and T1/2 3.7 folds longer with hydrogel compared to solution. Over 21d, AUCtumor was 1.5 folds greater in hydrogel compared to solution, indicating greater drug exposure and retention at target site. The antitumor activity of PTX (30 mg/kg) when given alone or in combination with doxorubicin (DOX, 15 mg/kg) was evaluated after intratumoral hydrogel injection in SNU-601 (Td= 21 d) and SNU-398 (Td= 5 d) xenograft models. The synergism between DOX and PTX, independent of dosage form, was observed in SNU-398, but not in SNU-601. No toxicity was observed in hydrogel group in neither single nor combination treatment. For slow-growing SNU-601 tumor, combination of PTX and DOX given in hyrogel mixture showed greater activity than that of solution. In conclusion, poly-(organophosphazene) polymer may be useful in intralesional administration of PTX to achieve greater drug exposure at target site. Also, combination of DOX and PTX, showed a potential for greater antitumor efficacy, which warrants further evaluation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3591.
Abstract 3597: Association between the ABCG2 polymorphisms and the gefitinib-induced side effects in Japanese patients with non-small cell lung cancer
Authors: Imai Yasuo , Akasaka Keiichi , Kaburagi Takayuki , Imura Johji , Ueda Yoshihiko
Published: February 2011

ABCG2 is a half-size ATP-binding cassette transporter implicated in cellular gefitinib transport. Reportedly, the c.421C>A ABCG2 gene variant was associated with gefitinib-induced diarrhea in Caucasian patients with non-small cell lung cancer. c.421C>A ABCG2, resulting in p.Q141K substitution, is more prevalent in Asian populations. Therefore, the putative relationship between gefitinib-induced adverse effects and this functional polymorphism was investigated in 75 Japanese patients with non-small cell lung cancer treated with gefitinib 250 mg/d orally, and the results were correlated with treatment-related adverse effects. c.376 C>T, resulting in truncated, non-functional ABCG2, was also investigated. Forty one (54.7%) patients harbored 376T or 421A ABCG2 on at least one allele, while the remaining 34 (45.3%) were wild type for ABCG2. No significant group differences were observed in frequency of gefitinib-related diarrhea or other adverse effects. Next, DLD-1 colon cancer cells expressing wild-type (DLD-1/WT) or 141K mutant ABCG2 (DLD-1/Q141K) were established for investigation of in-vitro cell sensitivity to the ABCG2-substrate drugs, gefitinib and SN-38. ABCG2 expression was much lower in DLD-1/Q141K cells than in DLD-1/WT cells, despite similar ABCG2 mRNA levels. DLD-1/WT cells acquired more resistance to SN-38 than did DLD-1/Q141K cells, but neither cell line acquired gefitinib resistance compared with parental cells. In-vitro data also suggested that ABCG2 has only a limited role in toxicity of gefitinib, but not SN-38, in colon-derived cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3597.
Abstract 3630: In vitro and in vivo antitumor activity of multi-tyrosine Kinase inhibitors in neuroblastoma
Authors: Calero Raul , Serrano Rosario , Avendaño Laura , Poblet Enrique , Ocana Alberto
Published: February 2011

Background: Despite Neuroblastoma (NB) is a curable tumor in its early stages, high risk patients continue to have a bad prognosis and a poor survival. Tyrosine kinase receptors mediate proliferation and survival and have been associated with oncogenic transformation in many human tumour types including NB. Sorafenib and sunitinib are two multityrosine kinase inhibitors that have shown antitumor activity in some solid tumours, like renal cell carcinoma. Evaluation of approved multi TK inhibitors in NB is a main goal in order to identify novel therapeutic options of treatment.

Methods: Here, we tested the antitumor activity of sorafenib and sunitinib, two multityrosine kinase inhibitors, in NB cell lines and in an animal model. To this end, we have used cancer cell lines and an in vivo xenografted mouse model. MTT proliferation, apoptosis assays, immunohistochemical analysis, and biochemical experiments were performed to investigate the molecular bases of the action of both compounds, of sunitinib alone, or of sunitinib in combination with standard chemotherapy.

Results: Using a representative panel of NB cell lines, we report that sunitinib exhibits a potent antitumor activity in vitro (IC50 0.8-2.6 µM) and a better growth inhibition profile compared with sorafenib (IC50 0.5-5.2 µM). Sunitinib also shows a high therapeutic index, when tested in non-transformed cells, and is synergic at very low doses (0.1 µM) with chemotherapies used for the treatment of NB tumors. Mechanistically sunitinib inhibits several tyrosine kinase receptor proteins and the downstream PI3-AKT pathway. Administration of sunitinib to NB cells induces cell cycle arrest and stimulates apoptosis. In vivo xenograft studies revealed a significant antitumor activity of sunitinib compared with the control group in two types of cell lines (SH-SY5Y, P=0.02 and SK-N-BE, P=0.08). Sunitinib demonstrated an anti-angiogenic effect in vivo with a reduction of markers associated with angiogenesis (e.g. factor VIII).

Conclusion: These results demonstrate the antitumor activity of sunitinib in NB tumors and support their further development in clinical studies alone or in combination with chemotherapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3630.
Abstract 3635: Novel chemotherapeutic agent AN019 for lapatinib-resistant breast cancer
Authors: Gondi Christopher S. , Gorantla Bharathi , Rao A.K.S. Bhujanga , Amala K. , Naidu M.U.R. , Jogi K.V. , Ramana G.V. , Chowdary M. Praveen , Rao Jasti S.
Published: February 2011

Breast cancer is the most frequently diagnosed cancer in women. Overexpression of human epidermal growth factor receptors (EGFRs) represents a biological subclass of breast cancer with distinct molecular alterations, clinical behavior, and response to systemic therapy. In this study, we introduce a novel compound (AN019) that has greater anti-tumor activity than Lapatinib. We demonstrate that AN019 is more effective at inhibiting the angiogenic potential and proliferation of both MDAMB231 and HTB20/BT474 cells. FACS analysis shows that AN019 treatment also caused the accumulation of MDAMB231 and HTB20/BT474 cells in the sub-G0/1 phase in a dose-dependent manner; this effect was accompanied by increased PARP cleavage indicative of apoptosis. Strong inhibition of EGFR phosphorylation was also observed in both MDAMB231 and HTB20/BT474 cells. HTB20/BT474 cells did not respond well to Lapatinib treatment when compared to AN019 treatment. From our animal studies using SCID mice implanted with HTB20/BT474 cells, we observed a dose-dependent inhibition of tumor growth when compared to controls or Lapatinib-treated mice at comparable concentrations. A dose-dependent inhibition of EGFR phosphorylation was also observed from immunohistochemical analysis of tumor sections. Our results show that AN019 is superior to Lapatinib in EGFR-overexpressing cells and has strong anti-angiogenic, anti-proliferative and pro-apoptotic properties. In vivo studies demonstrate the increased anti-tumor activity of AN019 over Lapatinib. These results suggest the therapeutic potential of AN019 as anti-angiogenic and anti-proliferative agent in Lapatinib-resistant and EGFR-overexpressing cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3635.
Abstract 3643: Manipulation of HuR expression alters the efficacy of PARP-inhibitors in pancreatic and ovarian cancer cells
Authors: Costantino Christina L. , Richards Nathan G. , Cozzitorto Joseph , Rittenhouse David , Gonye Greg , Yeo Charles J. , Witkiewicz Agnes , Brody Jonathan R.
Published: February 2011

PARP-inhibitors (PARPi) are a novel class of agents that target cancer cells deficient in DNA repair. Early phase trials show promising success with PARP-inhibitors, yet drug resistance mechanisms and off-target pathways affected by drug exposure have not been explored. HuR (ELAV1) is a protein that binds and stabilizes mRNA transcripts upon certain stress stimuli. These HuR mRNA targets result in an increase in protein expression; and thus, protein function. Recently, we demonstrated that HuR binds and regulates deoxycytidine kinase, the gemcitabine metabolizing enzyme. Thus, HuR expression enhances gemcitabine efficacy against pancreatic cancer cells. We subsequently screened other chemotherapeutics that may stimulate the HuR stress response. In isogenic pancreatic cancer cell lines, stable HuR overexpression rendered cells up to 2-fold more sensitive to PARPi compared to control cells. Accordingly, siRNA knock down of HuR expression in both pancreatic and ovarian cancer cell lines caused PARPi resistance compared to a scramble-sequence control siRNA. In a pancreatic cancer cell line, MiaPaCa2, a 2.5-fold siRNA knock down of HuR mRNA expression detected by qPCR rendered cells approximately 4-fold more resistant to PARPi. Similarly, a modest siRNA knock down of HuR in an ovarian cancer cell line, A2780, rendered cells more resistant to PARPi. Immunofluorescence studies indicate that a 5 hour treatment with 75 µM PARPi directly caused HuR to transport from the nucleus to the cytoplasm, presumably moving ARE-rich mRNA cargoes after drug exposure. Focused gene expression analysis on HuR bound, immunoprecipitated RNA defined functional HuR downstream targets (including deoxycytidine kinase). Current studies will reveal and validate the importance of these downstream mRNA targets in PARP-inhibitor metabolism and efficacy. We hypothesize that HuR may be central to PARPi effectiveness and may be useful in understanding PARPi de novo drug resistance mechanisms. Future work will reveal whether HuR status can be utilized as a predictive marker for PARP-inhibitor-based therapy and if all clinically available PARP-inhibitors engage HuR's regulation of unique mRNA cargoes in cancer cells upon treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3643.
Abstract 3648: Proteomic profiling of vorinostat in breast cancer
Authors: Shaw Patrick G. , Chearkady Raghothama , Pandey Akhilesh , Davidson Nancy
Published: February 2011

The lysine deacetylase (KDAC) inhibitor, vorinostat or suberoylanilide hydroxamic acid (SAHA), is currently being used for treatment of cutaneous T cell lymphoma, however, its anti-breast cancer effects are under evaluation. SAHA's pluripotent effect on histone and non-histone lysine acetylation across cancer types raises the theoretical concern that it could both re-express epigenetically silenced tumor suppressors and activate oncogenic pathways as well. In order to investigate the potential role of SAHA in tumor suppressor, cell death and signaling pathways, we used quantitative mass spectrometric analysis of MCF10A normal mammary epithelial cells and MDA-MB-231 breast cancer cells treated with SAHA for 12 and 24 hours. We observed downregulation of EGFR, Ephrin type A receptors and cell cycle proteins including CDC2 and CDC42 protein kinase, which is consistent with anti-tumor activity. We found that SAHA treatment also promoted activation of proteins including Ephrin type B receptors, STAT3 and p120 Catenin. Such molecular effects must be characterized in greater detail in order to determine the potential efficacy of this drug in breast cancer. Our findings show that SAHA-mediated inhibition of KDACs results in a wide range of downstream consequences in mammary epithelial and breast cancer cells. Candidate molecules identified in this study can be used as biomarkers to assay the breast cancer response to SAHA.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3648.
Abstract 3652: HDAC inhibitors activate NF-κB in human leukemia cells through an ATM/NEMO-dependent DNA Damage-related pathway
Authors: Rosato Roberto R. , Kolla Sarah S. , Hock Stefanie K. , Almenara Jorge A. , Atadja Peter , Dent Paul , Grant Steve
Published: February 2011

The ability of HDAC inhibitors (HDACIs) to induce sustained activation of NF-κB (p65/RelA) has been attributed to reversible RelA acetylation (Chen et al., Science 293:1653-7, 2001), a phenomenon that diminishes HDACI antineoplastic activity (Dai et al., Mol Cell Biol. 25:5429-44, 2005). However, the process by which HDACIs initially activate NF-κB has yet to be elucidated. To address this question, mechanisms underlying HDAC inhibitor (HDACI)-mediated NF-κB activation were investigated in human myeloid leukemia cells. Exposure of U937 and other leukemia cells to the pan-HDACI LBH-589 induced reactive oxygen species (ROS) and p65/RelA activation, the NF-κB-dependent induction of Mn-SOD2 mRNA and protein accompanied by ROS elimination, and sequential induction of XRCC1 (single strand) followed by γ-H2A. X (double strand) DNA breaks. LBH-589 lethality was significantly attenuated by siRNA knockdown of the chromatin-linked DNA damage protein histone H1.2. U937 IκBα super-repressor cells lacking serine 32 and 36 IκBα phosphorylation sites displayed diminished HDACI-mediated NF-κB activation/Mn-SOD2 induction, accompanied by enhanced ROS accumulation, DNA damage, and apoptosis. In contrast, TRAF2 siRNA knockdown blocked TNFα- but not HDACI-mediated NF-κB activation and lethality. The Mn-SOD2 mimetic TBAP prevented HDACI-induced NF-κB activation and nuclear localization, while dramatically attenuating DNA damage and apoptosis. Notably, LBH-589 exposure activated ATM (on serine 1981) and increased its association with NEMO, events comprising a recently described “inside-out” nuclear NF-κB activation pathway (Wu et al., Science 311:1110-1, 2006). Notably, siRNA NEMO or ATM knockdown blocked HDACI-mediated NF-κB nuclear translocation/activation, diminished MnSOD2 induction, and potentiated oxidative injury, DNA damage and cell death. Similarly, SUMOylation-site mutant NEMO (K277A and/or K309A) cells exposed to LBH-589 displayed diminished association of ATM with NEMO, reduced NEMO and p65/RelA nuclear localization/activation, and MnSOD2 down-regulation. These events were accompanied by increased ROS production, γ-H2A. X formation, and enhanced lethality. Collectively, these findings indicate that in human leukemia cells, HDACIs activate the cytoprotective NF-κB pathway through an ATM/NEMO-dependent process involving the induction of ROS and DNA damage. They also raise the possibility that blocking NF-κB activation via the atypical DNA damage-related ATM/NEMO nuclear pathway has the potential to enhance HDACI antileukemic activity.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3652.
Abstract 3679: Targeting drug-resistant GIST cells with a quadruplex-binding small molecule
Authors: Gunaratnam Mekala , Collie Gavin , Reszka Anthony , Todd Alan , Parkinson Gary , Fletcher Jonathan , Neidle Stephen
Published: February 2011

Gastrointestinal tumours (GIST) originate, at least in part, from disregulation and consequent aberrant expression of the c-kit oncogene. Imatinib is an effective inhibitor of the ATP-binding activity of the kinase domain in the c-kit protein, and is used clinically to treat GIST patients. However acquired resistance to Imatinib is a severe problem hindering long-term patient survival. We have previously reported on an alternative approach to c-kit therapy that involves targeting and stabilising quadruplex DNA sequences with a small molecule, (i) in two sequences within the c-kit promoter so that c-kit expression is down-regulated, and (ii) in the single-stranded overhang of telomeric DNA, inhibiting telomerase activity in GIST cells. We have previously reported that both of these are targeted by a potent (IC50 ≪ 1μM) tetra-substituted naphthalene diimide compound in the Imatinib-sensitive GIST882 patient-derived cell line.

This compound is equally potent in the patient-derived GIST48 Imatinib-resistant line; we report here that it has no effect on c-kit expression, telomerase activity, or the expression of other oncogenes in which promoter quadruplexes have been identified. Instead we find that the translation of the apoptosis-related bcl-2 gene is profoundly affected, but not its transcription. The bcl-2 gene has a potential quadruplex-forming region in the 5′-UTR sequence, and we report here that this forms a stable RNA quadruplex structure, which is further stabilised by the binding of the naphthalene diimide ligand. A reporter assay approach using a dual luciferase psiCHECK-2 vector, containing the bcl-2 5′-UTR sequence, is being used to validate the key concept that this G-rich sequence is involved in modulating translation in vitro, and in vivo through transfection into mammalian cells. Our current experiments in an Imatinib-resistant GIST cell line in the presence of the compound are enabling us to substantiate this cellular target and characterize the mode of action.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3679.
Abstract 3690: Development of a cell based high-throughput assay for drug discovery to treat ovarian cancer
Authors: Arslan Ahmet D. , He Xiaolong , Beck William T.
Published: February 2011

PTBP1 (polypyrimidine tract-binding protein, PTB) is a 59.6-kD nuclear protein involved in pre-mRNA splicing. Its molecular functions include regulation of alternative splicing (AS) and internal ribosomal entry site (IRES)-mediated translation. Our previous work revealed that PTB was overexpressed in human ovarian tumors and that knockdown of PTB expression by shRNA impaired ovarian tumor cell growth and malignant properties in vitro (Oncogene 26:4961-68, 2007). We have also observed slower tumor growth in a xenograft mouse model of ovarian cancer when PTB was knocked down (unpublished). These data suggest that PTB is a novel therapeutic target. To attempt to identify small molecules that have potential to act as shRNA and inhibit PTB, we have designed a cell-based high-throughput screening (HTS) system that allows fast and reliable detection of small molecule inhibitors of PTB protein activity that have potential to alter mRNA splicing. Our approach is based on differential splicing of a PTB target gene (identified by microarray analysis) at different levels of PTB activity in the cell. We have subcloned exons 14 - 16 of one of these PTB targets, Gamma-aminobutyric acid type B receptor 1 (GABBR1, upstream of two different fluorescent markers; the resulting two minigenes were designated “Gred” [short form], and “Ggreen” [long form]). We further verified that the splicing of the minigenes is dependent on the PTB level in the cells. We are presently establishing stable reporter cell lines expressing both minigenes, and validating their compatibility for HTS assay by measuring the stability of fluorescence protein expression over time and response to known PTB inhibitory stimuli (e.g. PTBshRNA). Optimization for HTS includes testing cell seeding density, DMSO tolerance, pilot experiments with specific chemical libraries, data analyses, and verification of “hits” in vitro, followed by pilot HTS with small libraries of chemical diversity to determine if any of these agents will block the activity of PTB. Since overexpressed PTB plays an integral part in maintaining ovarian tumor cell growth and malignant properties, identification of small molecule PTB inhibitors by this approach may uncover novel drugs for the treatment of ovarian and possibly other cancers.

(Support in part by grants RO1 CA40570 and RO1 CA138762 (to WTB), by OCRF (to XH), and by UIC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3690.
Abstract 3696: Pharmacokinetics (PK) of XMT-1001, a novel polymeric prodrug of camptothecin, in mice bearing HT-29 human colon carcinoma xenografts
Authors: Walsh Mark D. , Hanna Suzan K. , Sen Jeremy M. , Benson John D. , Cabral Carolina B. , Yurkovetskiy Alex , Lowinger Timothy B. , Zamboni Beth A. , Zamboni William C.
Published: February 2011

Introduction: XMT-1001 is a water soluble macromolecular conjugate of camptothecin (CPT). In this novel CPT prodrug, CPT is conjugated with a ∼60 kDa biodegradable hydrophilic polyacetal, poly (1-hydroxymethylethylene hydroxymethylformal) (PHF). Dual-phase release of CPT from XMT-1001 occurs via lipophilic intermediates, CPT-20-O-(N-succinimido-glycinate) (CPT-SI) and CPT-20-O-(N-succinamidoyl-glycinate) (CPT-SA). Hydrolysis of intermediates results in formation of an active-lactone form of CPT. XMT-1001 is designed to produce a prolonged exposure in plasma and tumor and to improve CPT's therapeutic index. We evaluated plasma, tumor and tissue PK of XMT-1001, CPT-SI, CPT-SA and CPT in mice bearing HT-29 human colon cancer xenografts.

Methods: XMT-1001 was administered at 60 mg CPT equivalents/kg IV x 1 via a tail vein. Mice (n = 3) were sacrificed at 0.083 to 24, 48, 72, 168 and 336 h after administration. Plasma, tumor and tissue samples were processed to measure the total (lactone + hydroxy acid) forms of XMT-1001, CPT-SI, CPT-SA and CPT via LC-MS/MS. Area under concentration versus time curves (AUC0-t) and time of last detectable concentration (TLast) were calculated.

Results: PlasmaTumorLiver AUC0-t (ng/mL·h)TLast (h)AUC0-t (ng/mL·h)TLast (h)AUC0-t (ng/mL·h)TLast (h)XMT-10013,639,034168138,156336663,493336CPT-SI53,653248,85033613,239336CPT-SA46,010241,59516814,063336CPT8,86024853481,11616

Conclusions: CPT conjugation to the polymer carrier, PHF, provides a water soluble formulation with potential advantages of extended systemic exposure to the conjugated drug XMT-1001, conjugated release products CPT-SI and CPT-SA, and active drug CPT. After XMT-1001 administration, the majority of plasma exposure is accounted for by conjugated CPT. The analysis of XMT-1001 revealed substantial tumor and tissue accumulation and extended exposure of conjugated drug and conjugated release products CPT-SI, CPT-SA and CPT. The improved tumor distribution and extended localized release of small molecule prodrugs and active CPT by XMT-1001 are hypothesized to improve safety and efficacy over existing drugs of the same class.

Supported by Mersana Therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3696.
Abstract 3717: Randomized phase II trial of cladribine with or without immediate rituximab for newly diagnosed or previously treated hairy cell leukemia to eliminate minimal residual disease: Interim report
Authors: Kreitman Robert J. , Noel Pierre , Yuan Constance M. , Raffeld Mark , Stetler-Stevenson Maryalice
Published: February 2011

Purine analogs including cladribine can induce complete remission (CR) in a high percentage of patients with previously untreated hairy cell leukemia (HCL), but late relapses are common and disease-free survival curves show no convincing evidence of cure. Many patients in CR after cladribine have minimal residual disease (MRD) which strongly expresses CD20, and the anti-CD20 Mab rituximab, particularly with a purine analog, can eliminate MRD to undetectable levels. To determine the potential benefit of adding rituximab to cladribine for initial or 2nd-line treatment of HCL, a randomized trial was begun in 150 HCL patients with 0-1 prior courses of cladribine requiring therapy. Patients receive cladribine 0.15 mg/Kg by 5 daily 2-hr i.v. infusions, and 8 weekly i.v. infusions of rituximab 375 mg/m2. Rituximab in half the patients is begun with the 1st dose of cladribine, and in the other half is delayed at least 6 months when MRD is detectable in blood. Endpoints include determining whether MRD rates are lower at 6 months with immediate rituximab, and whether blood MRD-free survival is longer when rituximab is delayed at least 6 months or begun with cladribine. HCL variant (HCLv) patients, known to respond poorly to initial cladribine alone, are not randomized and receive rituximab with cladribine. Bone marrow and blood are assessed for MRD by immunohistochemistry, PCR and flow cytometry at 1 and 6 months after cladribine. At this time 14 patients are enrolled, including 6 untreated and 8 with 1 prior course of cladribine, the latter group including 1 with HCLv. So far, the overall response rate is 100% with 10 (83%) CRs in 12 evaluable patients at 61-243 (median 202) days of follow-up. By 1 month, 0 of 5 patients after cladribine alone vs 5 (71%) of 7 after cladribine with immediate rituximab were MRD-free (p=0.028), including the patient with HCLv. Neutropenic fever was more frequent in patients receiving immediate rituximab, and was associated with both infusional reactions and reversible infections. We conclude that rituximab with cladribine can result in early clearing of MRD which is frequently present after cladribine alone. Additional accrual and follow-up is needed to determine whether simultaneous cladribine and rituximab can reduce MRD rates at 6 months, whether MRD-free survival might be longer or shorter with delayed rituximab, and whether rituximab added to cladribine for 1st or 2nd line treatment of HCL can prevent or delay clinical relapse.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3717.