bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1841: Selective inhibitors of nuclear export (SINE) activate multiple tumor suppressor pathways and kill prostate cancer cells across multiple genotypes in vitro and in vivo
Authors: Gravina Giovanni Luca , Ricevuto Enrico , Mancini Andrea , Shechter Sharon , Sandanayaka Vincent , Kauffman Michael G. , Shacham Sharon , Festuccia Claudio
Published: August 2012

CRM1 (Xpo1) is a key nuclear export protein which controls the nuclear export of multiple tumor suppressor proteins (TSP) and cell proliferation pathways including p53, p21, FOXO, PI3K/AKT, Wnt/ß-catenin and NF-κB. Mislocalization of TSP can abrogate their functions as well as render chemotherapies ineffective. Induction of nuclear expression of TSP and chemotherapy targets, by CRM1 inhibition can restore their tumor suppressor functions and increase drug sensitivity. We have developed orally active, small molecule SINE that irreversibly and potently inhibit CRM1 mediated nuclear export of multiple TSP and other cargoes (IC50 < 100 nM). Here, we describe in vitro results using SINE compounds, KPT-185 in vitro and KPT-251 in vitro and in vivo on seven prostate cancer (PrCa) cell lines representing distinct differentiation/progression states of disease and genotypes: LAPC-4 (Androgen receptor [AR] positive, androgen dependent with low Akt/mTOR activities, p53 wt); LnCaP (AR positive, androgen dependent with high Akt/mTOR activities, p53 wt); LnCaP-C81 and LnCaP-C4-2B (AR positive, androgen independent with high Akt/mTOR activities, p53 wt); 22rv1 (AR positive, androgen independent with low Akt/mTOR activities, p53 wt); PC3 (AR negative, with high Akt/mTOR activites and no p53 function (p53 del) and DU145 (AR negative, with low Akt/mTOR activites and mutant p53). Benign prostatic hyperplasia line (BPH1) and Prostatic epithelial line (EPN) were used as non-neoplastic controls. We show that SINE block CRM1 mediated nuclear export of FOXO and p53 with a IC50 values <100 nM in both PrCa and non-neoplastic cell lines. KPT compounds are selectively cytotoxic to PrCa lines with EC50s between 10 and 1000 nM, and show limited cytotoxicity on the non-neoplastic EPN and BPH1 lines (EC50 >5-20 μM). SINE cytotoxic effects are independent of p53 status, and induce caspase-3 activation. SINE display synergistic effects in combination with cisplatin and docetaxel in vitro with combination indices between 0.3 and 0.8. In vivo results indicate that KPT-251 show a dose-dependent inhibition of tumor growth. KPT251 was also synergistic when administered at 30mg/kg/5 days/week per os with docetaxel and cisplatin by using the aggressive p53 wt 22rv1 xenografts. Taken together, CRM1 inhibition represents a completely novel, neoplasia-selective and well- tolerated target for use as single agent or in combination with chemotherapy for PrCa.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1841. doi:1538-7445.AM2012-1841
Abstract 1849: Identification and functional analysis of SHISA2 overexpressed in prostate cancer
Authors: Tamura Kenji , Furihata Mutsuo , Satake Hirofumi , Anchi Takashi , Kamei Maiko , Fukuhara Hideo , Shimamoto Tsutomu , Ashida Shingo , Karashima Takashi , Yamasaki Ichiro , Nishikawa Hiroshi , Kamada Masayuki , Yasuda Masaharu , Inoue Keiji , Shuin Taro
Published: August 2012

Prostate cancer (PC) is usually androgen-dependent and responds well to androgen ablation therapy. However, at a certain stage some prostate cancers eventually acquire a castration-resistant phenotype. These cancer cells are originally high-grade and show very poor response to any anticancer therapies. To identify novel molecular cancer drug targets, we previously analyzed the gene expression profiles of high-grade PCs using a cDNA microarray combined with laser microbeam microdissection and found a number of genes that are transactivated in high-grade PC. Among them, we report the identification of a novel molecular target, SHISA2. We confirmed overexpression of SHISA2 in high-grade prostate cancer cells by semi-quantitative RT-PCR and immunohistochemical analysis. Knockdown of SHISA2 expression by siRNA in a prostate cancer cell line resulted in a drastic decrease of cell numbers. In contrast, SHISA2 overexpression in a prostate cancer cell line promoted cell proliferation. These findings suggest that SHISA2 could be involved in aggressive phenotype of prostate cancers, including castration-resistant prostate cancers, and that it should be a potential molecular target for development of new therapeutics and a diagnostic biomarker for aggressive prostate cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1849. doi:1538-7445.AM2012-1849
Abstract 1862: NY-ESO-1 and clinicopathological features in ovarian cancer
Authors: Topka Charlotte R. , ElMasri Wafic M. , Magyar Clara E. , Song Min , Dorigo Oliver
Published: August 2012

Ovarian cancer ranks fifth in cancer deaths among women and the American Cancer Society estimates that in the US about 15,460 women will die from this disease in 2011. NY-ESO-1 is an 18 kDa member of the cancer/testis 6 antigen family which is thought to play a role in germ cell differentiation. In healthy adults, NY-ESO-1 is restricted to the testis. NY-ESO-1 expression has been found in various malignant neoplasms, including melanomas, neuroblastomas, soft tissue sarcomas, and carcinomas from the breast and endometrium. To further delineate the role of NY-ESO-1 in epithelial ovarian cancer, we constructed a tissue microarray of archived formalin-fixed paraffin-embedded paired tumor and normal control specimens from ovarian cancer patients. NY-ESO-1 expression was correlated with clinicopathologic features and overall survival. Specimens were collected from 242 patients who underwent cytoreductive surgery for ovarian cancer between 1989 and 2009. Immunohistochemical staining was performed with mouse monoclonal antibody clone E978 specific for NY-ESO-1. Stained slides were digitized with the Aperio ScanScope XT and images were analyzed using DEFINENS Tissue Studio software. Histological scores (H-scores) based on intensity of NY-ESO-1 staining and percent positive cells were calculated for all specimens. Positive NY-ESO-1 expression was found in 21% of papillary serous carcinomas, 18% of endometrioid carcinomas, and 20% of clear cell carcinomas. Ovarian cancer tissue had significantly higher H-scores compared to normal tissue controls from the same patients. H-scores increased with increasing FIGO stage and grade of the tumors. Caucasians had higher H-scores compared to African Americans and Asians. H-scores did not correlate with overall survival. Similarly, no differences in overall survival based on NY-ESO-1 expression levels were detected when patients were stratified by their histologic subtype. In conclusion, NY-ESO-1 is expressed in all histologic subtypes of epithelial ovarian cancer but not in normal tissues, therefore making it an ideal candidate for the development of targeted therapeutic approaches in ovarian cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1862. doi:1538-7445.AM2012-1862
Abstract 1864: Role of store-operated Ca2+ entry in proliferation and cell cycle in melanoma
Authors: Umemura Masanari , Baljinnyam Erdene , De Lorenzo Mariana S. , Feske Stefan , Xie Lai-Hua , Iwatsubo Kousaku
Published: August 2012

Background: Melanoma has a poor prognosis due to its rapid progression and a high metastatic ability. Ca2+ homeostasis plays a pivotal role in proliferation in cancer cells, but little is known about the role of Ca2+ in melanoma. Store-operated calcium entry (SOCE) is defined as Ca2+ entry from extracellular space triggered when Ca2+ store in the endoplasmic reticulum (ER) is depleted. SOCE is regulated by coordination between Orai (ORAI calcium release-activated calcium modulator) in the plasma membrane, and STIM1 (stromal interaction molecule 1) in the ER. We examined the role of Ca2+ in melanoma cells especially focusing on SOCE. Method: Intracellular Ca2+ level was measured by using Fluo4-AM, a Ca2+-sensing fluorescent dye. Cell proliferation was measured by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Cells were stained with propidium iodide and cell cycle was assessed using flow cytometer. Protein phosphorylation microarray was performed by using a commercially available kit (Cancer Signaling Phospho-Antibody Array). Results: SOCE, as demonstrated by enhancement of Ca2+ entry from extracellular space after Ca2+ depletion in the ER, was observed in SK-MEL-2, C8161, SK-MEL-24, UACC257 (human) and B16 (mouse) melanoma cell lines, suggesting that the existence of SOCE is universal among melanoma cells. SOCE was attenuated by Pyr3, a pyrazole compound which is known to inhibit SOCE, as well as by EGTA that chelates extracellular Ca2+. Pyr3 inhibits cellular proliferation in SK-MEL-2 cells (IC50: 5.1uM) and C8161 cells (IC50: 2.1uM). Cell cycle was analyzed in C8161 cells, and the cell number in the G2 phase was increased by both Pyr3 and EGTA, indicating that SOCE plays a role in the transition of G2/S phase. Protein microarray data showed that Pyr3 reduces phosphorylation of signal transducers and activators of transcription 5 (STAT5) by 93%, suggesting that the JAK/STAT pathway may be involved in SOCE-induced alteration of cell cycle. Conclusion: Our results showed that inhibition of SOCE suppresses proliferation in melanoma, thus it could be a new strategy for melanoma therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1864. doi:1538-7445.AM2012-1864
Abstract 1877: Predictive marker discovery for Sorafenib using patient derived xenograft models of hepatocellular carcinoma
Authors: Zhang Juan , Sun Yanmei , Song Xiaoming , Yang Jie , Cai Jie , Shi Qian , Chen Taiping
Published: August 2012

Sorafenib, a multikinase inhibitor targeting vascular endothelial growth factor (VEGF)- mediated angiogenesis and blocking the RAF/extracellular signal regulated kinase (ERK) kinase (MEK)/ERK cascade, is the first drug found to prolong survival of patients with advanced hepatocellular carcinoma (HCC). Although it is only used in treating advanced renal cell carcinoma and unresectable HCC, many patients with other malignant diseases as well as patient with resectable HCC may also benefit from this treatment. There is a clear need for predictive markers of Sorafenib to help selecting the patient population that may benefit the most. We set out to discover the predictive markers using patient derived xenograft (PDX) models of HCC. The response to Sorafenib was investigated in 20 PDX subcutaneous models and 6 orthotopic cell line (MHCC97H, HUH-1, HepG2, Hep3B, SK-Hep-1 and PLC/PRF/5) derived xenogrft (CDX) models. 11 out of 20 PDX models (55%), and 4 out of 6 CDX models (67%) achieved significant tumor growth inhibition by the treatment of Sorafenib at the dose range from 25-50 mg/kg in nude mice. Gene expression profile, hotspot mutation detection and copy number variations were analyzed to sift out the molecular signatures that may be predictive of the response to Sorafenib. Genes and pathways such as VEGF, PDGF, FGF, Raf/MAPK/ERK, PTEN/ PI3K/AKT/mTOR, TGF-beta/Wnt, EGFR, IGF and HGF/c-MET, were investigated using methods such as IHC analysis of pretreatment xenograft tissues. Our results indicate that more than half of the resectable HCC may be responsive to Sorafenib treatment, and markers other than the usual targets of Sorafenib may be predictive of this response. This study may shed light on the predictive markers of Sorafenib and help patients that may be mostly benefited from this drug.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1877. doi:1538-7445.AM2012-1877
Abstract 1878: Upregulation of UGT1A4 by fulvestrant: A potential elimination mechanism of anastrazole in breast cancer treatment
Authors: Edavana Vineetha Koroth , Dhakal Ishwori B. , Williams Suzanne , Kadlubar Susan A.
Published: August 2012

The pure antiestrogen, fulvestrant (Faslodex®) has been approved for use in the treatment of postmenopausal patients with hormone-receptor positive metastatic disease. It has been postulated that the combination of an aromatase inhibitor with a pure antiestrogen could provide therapeutic advantages over the administration of an aromatase inhibitor alone. The effect of co-administration of these drugs on Phase I and Phase II metabolism (i.e. glucuronidation) have not been explored. The role of pharmacogenetics in clinical benefit and overall survival in patients receiving anastrozole and fulvestrant is unexplored. In this study, we demonstrated that fulvestrant up-regulates expression of UDP glucuronosyltransferase (UGT) 1A, the major enzyme responsible for the direct conjugation and subsequent elimination anastrozole. Endogenous mRNA expression levels of UGT1A4 in HepG2 cells were induced 4.2-fold by fulvestrant treatment in the ERα-expressing cell line. In vitro assays using liver microsomal specimens from individual subjects demonstrated 163 fold variability in anastrazole glucuronidation. Anastrazole glucuronodation was correlated (r = 0.977, p <0.0001) with lamotrigine glucuronidation, a diagnostic substrate for UGT1A4. We also found a significant correlation between anastrazole glucuronidation and UGT1A4 mRNA expression levels in these HLMs (r = 0.978, p <0.0001). We also examined whether UGT1A4 promoter SNPs were associated with the observed interindividual variability in anastrozole glucuronidation. Microsomes from the subjects with mutated heterozygous UGT1A4 promoter SNPs 36G<A and SNPs 217 T<G exhibited a significantly higher activity than the microsomes from the subjects with homozygous wild type UGT1A4 promoter SNP 36G<A and SNP 217 T<G. On the other hand, microsomes from the subjects with homozygous wild typeUGT1A4 promoter SNPs 219C<T and SNPs 163G<A exhibited significantly higher activity when compared with microsomes from the subjects with mutated heterozygous UGT1A4 promoter SNPs 219C<T and SNPs 163G<A. These in vitro studies indicate that fulvestrant administration could influence the disposition of anastrazole in combination therapy of aromatase inhibitor with a pure antiestrogen by up regulating the phase II metabolizing enzyme UGT1A4. Our data also suggest that the UGT1A4 promoter polymorphisms may be associated with altered rates glucuronidation and detoxification of anastrazole in vivo. Further studies are needed to assess the complete effects of fulvestrant administration on the disposition of drugs that are substrates of UGT1A4.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1878. doi:1538-7445.AM2012-1878
Abstract 1880: Incorporating protein levels with genetic variation and gene expression in sensitivity to chemotherapy
Authors: Stark Amy L. , Hause Ronald J. , Gorsic Lidija K. , Antao Nirav N. , Chung Sophie H. , Gill Daniel F. , White Kevin P. , Jones Richard B. , Dolan M. Eileen
Published: August 2012

Genome-wide studies of genes important in chemotherapeutic sensitivity or disease susceptibility and progression have primarily relied on mRNA expression levels; however, mRNA expression levels do not always accurately reflect downstream protein abundances. We have applied a novel technology, microwestern arrays, along with reverse phase lysate arrays, to evaluate the role that over 400 transcription factor and signaling proteins play in mediating cellular sensitivity to ten chemotherapeutic agents. Utilizing unrelated International HapMap lymphoblastoid cell lines derived from the Yoruba population, we evaluated the relationship between protein abundances, genetic variation, microRNA and mRNA expression, and chemotherapeutic-induced cytotoxicity and apoptosis. Using ∼12 million single nucleotide polymorphism (SNPs) imputed from the 1000 Genomes Project, we identified SNPs associated with protein abundance (protein quantitative trait loci, pQTLs) and SNPs associated with chemotherapeutic sensitivity (drug quantitative trait loci, dQTLs). Chemotherapeutic agents included platinating agents, antimetabolites and microtubule inhibitors. For cytarabine there was a significant (p<0.0001) enrichment of pQTLs in the SNPs associated with cytotoxicity and apoptosis. We also identified 71 protein levels associated with cytarabine-induced cytotoxicity and apoptosis at p<0.05. One protein, ATF2, was associated (p<0.006) with both high (40 µM) and low (5 µM) dose cytarabine-induced apoptosis and there were six SNPs associated with apoptosis phenotypes (p<0.00007) that also associated with the protein level (p<0.00009). Knockdown of ATF2 protein with siRNA resulted in 40-51% of endogenous levels concomitant with a significant increase in apoptosis induced by cytarabine at both doses (p=0.001). This relationship was not identified in analyses correlating mRNA with phenotypes thus demonstrating the importance of incorporating protein levels into studies that aim to identify potential biomarkers. By incorporating genetic variation, mRNA, protein, and microRNA levels with chemotherapeutic sensitivity, the complex role of the genetic architecture in cellular chemotherapy response through its effects on expression and protein can be better understood. This study has broad potential for application to other chemotherapeutic drugs and complex traits.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1880. doi:1538-7445.AM2012-1880
Abstract 1882: Gemcitabine pathway SNPs are associated with its PK parameters in patients with solid tumors
Authors: Mitra Amit K. , Khatri Amit , Kirstein Mark N. , Kratzke Robert A. , Lamba Jatinder K.
Published: August 2012

Gemcitabine (2′, 2′-difluoro 2′-deoxycytidine, dFdC) is a cytotoxic pyrimidine antimetabolite used as first-line chemotherapeutic agent for the treatment of a wide range of solid tumors and hematologic malignancies. It is as a pro-drug that undergoes cellular uptake through transporters like human equilibrative nucleoside transporter (hENT1/SLC29A1) and human concentrative nucleoside transporters (hCNT1/SLC28A1 and hCNT3/SLC28A3) followed by extensive intracellular phosphorylation by kinases like deoxycytidine kinase (DCK) and cytidine monophosphate kinase (CMPK) to its active diphosphorylated (dFdCDP) and triphosphorylated (dFdCTP) forms. Gemcitabine is primarily inactivated via deamination to its less potent metabolite 2′, 2′-difluorodeoxyuridine (dFdU) by cytidine deaminase (CDA). High inter-patient variability in response to gemcitabine treatment has been a major concern in gemcitabine chemotherapy. However, association of pharmacogenomics with gemcitabine pharmacokinetics (PK) has not been extensively studied so far. In the present study, therefore, we investigated association between single nucleotide polymorphisms (SNPs) within genes involved in gemcitabine transport/metabolic pathway and pharmacokinetic parameter estimates for gemcitabine and metabolites in patients diagnosed with solid tumors. Pharmacokinetic blood sampling (n=40) was as follows: pre-infusion, 5, 15, 30, 45 min, and 1, 1.25, 1.5, 2, 6, 24, 48 and 72 hours after the end of gemcitabine infusion. Gemcitabine and dFdU concentrations in plasma and intracellular dFdCTP levels in PBMCs were measured using HPLC-UV and LC MS/MS. A total of 86 SNPs in 14 genes viz. CDA, CMPK, CTPS, DCK, DCTD, NME1, NME2, NME3, NME4, NT5C2, NT5C3, SLC28A1, SLC28A3, and SLC29A1 were genotyped using Sequenom-based assays. Pharmacogenomic association was evaluated using Wilcoxon rank-sum test and Kruskal-Wallis one-way analysis of variance by ranks (p≤0.05). SNPs in 5′ nucleotidase genes- rs1163075, rs1926029, rs11598702 and rs1712517 in NT5C2 and rs3750117 in NT5C3 were associated with gemcitabine elimination clearance. SNPs in CDA- rs1048977 and rs4655226 were associated with dFdU clearance. SNPs in CTPS- rs4430318, rs61780437 and rs1044457 were strongly associated with higher dFdCTP formation clearance. Additionally, CDA, DCTD, SLC28A1, SLC28A3 and SLC29A1 SNPs were also associated with dFdCTP formation clearance. The results from our study indicate pharmacogenomic association between gemcitabine pharmacokinetic pathway SNPs and PK parameters. Presence of these SNPs could impact outcomes for patients receiving gemcitabine therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1882. doi:1538-7445.AM2012-1882
Abstract 189: miRNAs regulating members of SLC transporter family genes in penile carcinomas
Authors: Muñoz Juan J. , Linde Sandra A. , Barros Mateus C. , Busso Ariane F. , Marchi Fábio , Soares Fernando A. , Rogatto Silvia R.
Published: August 2012

Background: Penile cancer (PeCa) is a rare disease in developed countries, but with high incidence in South America and Africa. Surgery is frequently used for local control of the disease and it can lead to partial or complete amputation of the organ, with subsequent psychosexual morbidity. Integrating data previously obtained using CGH-array and large-scale gene expression in PeCa, we found deregulation of members of solute carriage (SLCs) gene family that was not associated with genomic alterations. These findings suggested that other regulatory mechanisms could be involved, such as miRNAs. Aim: The purpose of this study was to confirm the gene expression pattern of six SLC genes (SLC1A3, SLC8A1, SLC25A4, SLC25A16, SLC25A25 and SLC27A4) and their putative regulatory miRNAs (miR-22, miR223, miR-23b, miR-26b, miR-27b and, miR-133b, respectively) in PeCa and associate the findings with clinical and pathological data. Patients and Methods: RNA was obtained from fresh PeCa (27 training and 13 validation set samples) and 4 normal penile tissues. SLC genes and miRNAs expression were evaluated by RT-qPCR. HPV genotyping was assessed using Linear Array HPV Test Genotyping (Roche). Results: Positive correlation between SLC1A3, SLC8A1, SLC25A4 and SLC27A4 expression levels detected by microarray and RT-qPCR was observed in the training set samples. Downexpression of SLC8A1 (P=0.009) and SLC25A25 (P=0.002) and overexpression of SLC27A4 (P=0.006) were detected in tumors (training and validation set) compared with normal controls, thus validating the previous findings of global gene expression. Significant downexpression of miR-23b, miR-26b, miR-27b and miR-133b and overexpression of miR-223 were detected in tumors compared with normal controls. Interestingly, lower levels of miR-133b were observed in 96% of the tumors (P=0.0007). Significant negative correlation was detected between SLC8A1 transcript levels and its putative regulatory miRNA (miR-223, P=0.015, r = −0.482) and between SLC27A4 e miR-133b (P=0.023, r = –0.454), suggesting that these genes are regulated by these miRNAs. Higher levels of SLC25A16 were found in HPV positive tumors (P=0.012). A significant association was found between lower levels of miR-23b and miR-27b and perineural invasion (P=0,032 and P=0,032; respectively). Conclusions: This study showed the deregulation of members of SLC gene family in PeCa and the involvement of putative regulatory miRNAs. miR-23b and miR-27b were found as putative prognostic markers in PeCa patients and with potential as therapeutic targets.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 189. doi:1538-7445.AM2012-189
Abstract 1900: HSP 70 may cause EGFR-TKIs-resistance due to inhibit drug binding to EGFR in the cells that expressed mutant EGFR
Authors: Ohmori Tohru , Kadofuku Tsuyoki , Ichihashi Yoko , Yamaoka Toshimitsu , Hirose Takashi , Adachi Mitsuru , Saijo Nagahiro
Published: August 2012

Cancer cells that expressed mutant EGFR are more sensitive to EGFR tyrosine kinase inhibitors (EGFR-TKIs) than that expressed wild type EGFR. To elucidate difference of the characteristics between wild type and 15 bp deletion mutant EGFR, we explored the difference of EGFR-binding proteins using respective stable transfectants, 293_pEGFR and 293_p≤15. As the result, we detected heat shock protein 70 (HSP70) specifically binds to the mutant EGFR but wild type EGFR. To examine whether HSP70 influenced EGFR-TKI sensitivity, we examined the effect of HSP70 inhibition. [Methods] HSP70 siRNA was exposed to the cells for 2 days. Cytotoxicity of the drugs was measured by MTT assay. The drug binding to EGFR was measured by [14C]gefitinib. [Results] Suppression of HSP70 by siRNA increased gefitinib and erlotinib sensitivities and enhanced gefitinib binding to this receptor, in 293_p≤15 cells. Same phenomena were observed when HSP70 was inhibited by HSP70 specific inhibitor, 2-phenylethynesulfonamide (PES). PES inhibits HSP70 binding to the mutant EGFR. Interestingly, this compound enhanced the EGFR autophosphorylation and c-Cbl binding to the receptor in 293_p≤15 cells. These results suggest that HSP70 may modulate EGFR activity and may cause EGFR-TKIs-resistance due to inhibit drug binding to the mutant EGFR. HSP70 inhibitor may possibly enhance the efficacy of EGFR-TKIs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1900. doi:1538-7445.AM2012-1900
Abstract 1907: Heat shock protein 90 inhibition overcomes hepatocyte growth factor-triggering resistance to EGFR tyrosine kinase inhibitors in EGFR mutant lung cancer by decreasing client protein expression and angiogenesis
Authors: Yamada Tadaaki , Koizumi Hitomi , Takeuchi Shinji , Nakagawa Takayuki , Yano Seiji
Published: August 2012

The three major, clinically relevant mechanisms of acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR mutant lung cancer are a second mutation in the EGFR gene (T790M), Met amplification, and increased expression of hepatocyte growth factor (HGF). Heat shock protein90 (Hsp90) is a 90kDa molecular chaperone for proteins that include EGFR, Met and EML4-ALK. Hsp90 inhibitors have been reported to reverse resistance to EGFR-TKIs caused by EGFR T790M mutation and Met amplification. Here, we determined whether inhibition of Hsp90 could overcome HGF-triggered EGFR-TKI resistance in EGFR mutant lung cancer cells. EGFR-TKI erlotinib inhibited the growth of PC-9, HCC827, and Ma-1 cells, with deletions in exon19 of EGFR, but not HGF-gene transfected Ma-1 (Ma-1/HGF) cells and H1975 cells, which contain the EGFR L858R and T790M mutations, respectively. Erlotinib also did not inhibit the growth of PC-9 and Ma-1 cells in the presence of HGF. However, an Hsp90 inhibitor, 17-DMAG, induced apoptosis and markedly inhibited the growth of these cell lines, even in the presence of HGF. This inhibition by 17-DMAG was associated with decreased expression of EGFR and Met, thereby decreasing Akt phosphorylation in tumor cells. In an in vivo model of HGF-triggered erlotinib-resistance, using Ma-1/HGF cells, 17-DMAG markedly suppressed tumor growth by decreasing angiogenesis and increasing apoptosis. These results indicate that Hsp90 inhibitors may overcome HGF-triggered resistance to EGFR-TKIs and that the use of Hsp90 inhibitors may result in more successful treatment of EGFR mutant lung cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1907. doi:1538-7445.AM2012-1907
Abstract 1908: MDR1 gene overexpression confers resistance to C-MET inhibitor in multidrug resistant cancer cell line
Authors: Hung Tsai-Hsien , Li Yu-Hsin , Chong Kowit-Yu
Published: August 2012

c-MET signaling is implicated in a wide variety of human malignancies. Hepatocyte growth factor (HGF) binding to the proto-oncogenic c-MET receptor lead to activate multiple tyrosine kinase signaling pathway. Inappropriate c-MET signaling in cancer can enhance tumor cell proliferation, survival, motility, and invasion. Recently, several reports have indicated that inhibition of c-MET signaling induced apoptosis in a variety of cancer cells and recognized as a novel anticancer therapy approach. Previous study from our laboratory also showed that the higher mRNA expression level of HGF, and its downstream genes including HGF receptor (c-Met) and urokinase (uPA) were detected in drug resistant Dx5-C5 cell compared with parental human uterus sarcoma MES-SA cell line. Furthermore, report indicated that the constitutive expression of P-glycoprotein (P-gp) is involved in the HGF/MET related pathway of MDR-positive human hepatocellular carcinoma cell line. Moreover, MDR1 gene overexpressed leukemia cell line also shown more resistance to tyrosine kinase inhibitor imatinib mesylate. These finding suggested that chemoresistant cancer cells may also develop a similar mechanism to against c-MET inhibitor. Our results shown: when treated both cell cancer lines with c-MET inhibitor PHA665752, we found that compared to parental MES-SA cells, the Dx5-C5 cells with highly expressed P-gp was more resistant to PHA665752, the resistance was also reversed by co-treatment of MDR inhibitor verapamil and PHA665752. In addition, when transfection of c-MET shRNA into Dx5-C5 and MES-SA cell line for 48 hrs, we found that the viability was decreased in the c-MET shRNA transfected Dx5-C5 cell line in a dose dependent manner but not in the MES-SA cell line. In conclusion, our study demonstrated that existence of P-glycoprotein in MDR cell line would attenuate c-MET inhibitor induced cell death and may help us to develop a new strategy for treatment of multiple drug resistance cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1908. doi:1538-7445.AM2012-1908
Abstract 1910: Acquired resistance to HER2 targeted therapy: A common mechanism
Authors: Christenson Jessica , Kane Susan E.
Published: August 2012

There are currently two FDA approved targeted therapies being utilized in the clinic to treat Her2+ breast cancer, trastuzumab and lapatinib. Acquired resistance has become a major concern and several mechanisms for resistance have been proposed. The upregulation of a truncated transcriptional variant of Darpp-32, known as t-Darpp, is a characteristic shared by several independently developed trastuzumab-resistant cell lines. Moreover, exogenous t-Darpp overexpression is sufficient for conferring trastuzumab resistance. No consistent mechanism, however, has been identified for lapatinib resistant cells. By inhibiting signaling through Her2+ pathways such as PI3K/Akt and Erk, lapatinib induces a strong apoptotic response by down-regulating survivin and inducing BIM. Darpp-32 and t-Darpp both have been linked to apoptosis, with t-Darpp identified as having anti-apoptotic properties through its regulation of Bcl-2. The purpose of the current study was to determine if t-Darpp or other dysregulated apoptosis proteins play a role in lapatinib resistance. Lapatinib-resistant SKBR3 cells (Her2+, ER-) were selected by continuous exposure to increasing concentrations of lapatinib (0.3-2.0µM) over the course of one year. Using Sulforhodamine B and colony forming assays, we found that individual clones and pools of SK/LapR cells were, on average, 50-fold resistant to lapatinib, relative to parental SKBR3 cells. This resistance phenotype was stable even after cells were maintained for 1 month outside of drug. As observed in trastuzumab resistance and similar to previous reports of lapatinib-resistant cells, we found that SK/LapR cells had sustained PI3K/Akt signaling in the presence of lapatinib, whereas parental cells did not. Additionally, survivin inactivation and BIM induction by lapatinib seemed to be partially impaired in SK/LapR cells. Finally, SK/LapR cells appeared to down-regulate Darpp-32 and up-regulate t-Darpp, similar to what we have reported in BT474 cells selected for resistance to trastuzumab. Although t-Darpp overexpression is sufficient to confer resistance to trastuzumab, this does not appear to be the case for lapatinib resistance – SK.tDp cells transfected with a t-Darpp cDNA are not resistant to lapatinib. Experiments are currently underway to determine if the t-Darpp overexpression that we observe in SK/LapR cells is required for lapatinib resistance. Identifying a shared mechanism of resistance to lapatinib and trastuzumab would provide a novel therapeutic target that could be widely used among Her2+ breast cancer patients to ameliorate acquired drug resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1910. doi:1538-7445.AM2012-1910
Abstract 1914: Potent anticancer activity against both BRAF-mutant and BRAF wild-type melanoma cell lines using a novel CRM1 nuclear export inhibitor
Authors: Fragomeni Roberto A. Salas , Chung Hye-Won , Shacham Sharon , Kauffman Michael , Cusack James C.
Published: August 2012

INTRODUCTION: BRAF and NRAS oncogenic mutations occur in over half of all human melanomas. BRAF-mutant tumors are highly sensitive to small molecule inhibitors directed against the oncogenic BRAF protein. However, in tumors that lack the specific mutation, patients have few effective therapeutic options. To address the need for new therapies for patients with BRAF-wild-type (wt) melanoma, we hypothesized that the inhibition of the major nuclear export receptor CRM1 would result in nuclear accumulation of nuclear proteins, increase apoptosis and induce cell cycle arrest independent of BRAF mutational status. To test our hypothesis, we examined the effects of a novel CRM1 inhibitor on melanoma cell lines with various BRAF and NRAS mutational backgrounds. METHODS: We used human malignant melanoma cell lines with BRAF mutation (Sk-Mel-28, A375, Mel-624, UACC903, MalMe-3M), BRAF-wt (MeWo) and BRAF-wt and NRAS mutation (Hs940T) to assess the effects of CRM1 inhibition, BRAF inhibition (PLX4032) or MEK inhibition (U0126) using both mono- or combination therapy. MTT assays were used to evaluate the effects of treatment on cell proliferation. FACS analysis with annexin/PI staining was performed to assess effects on cell cycle and cell death. The Chou-Talalay method was used to assess synergistic drug combination effects. Treatment effects on cleaved caspase-3, PARP cleavage, P53, and FOXO3 were analyzed by western blot analysis. RESULTS: As mono therapy, CRM1 inhibition led to a significant decrease in cell proliferation, cell cycle arrest and an increase in apoptosis in both BRAF wt and mutant cell lines. The degree of inhibition and cell death achieved by CRM1 inhibition across all melanoma cell lines is comparable to the results achieved by BRAF inhibitor PLX4032 in BRAF mutant cell lines. Furthermore, the combination of CRM1 inhibition and BRAF inhibition results in a strongly synergistic combination against BRAF mutant cell lines with a statistically significant decrease in cell proliferation and increased apoptotic cell death. CONCLUSION: We found that relative to other cancer cell types, both BRAF-mutant and BRAF-wt melanoma cells are exquisitely sensitive to CRM1 inhibition. In addition, CRM1 inhibition significantly enhanced the effect of PLX4032 in BRAF-mutant cell lines, suggesting that the combination of these two drugs may address the emergence of acquired resistance to BRAF monotherapy. Based on these results, CRM1 inhibition warrants further investigation to determine its potential benefit in treating melanoma patients independent of BRAF mutation status.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1914. doi:1538-7445.AM2012-1914
Abstract 1915: Novel sorafenib derivatives (SC-40, SC-43 and SC-60) show better anti-tumor effects than sorafenib in hepatocellular carcinoma through SHP-1/STAT3-mediated signaling pathway
Authors: Tai Wei-Tien , Shiau Chung-Wai , Cheng Ann-Lii , Chen Pei-Jer , Chen Kuen-Feng
Published: August 2012

Recently, we have reported that sorafenib sensitized hepatocellular carcinoma (HCC) cells to TRAIL via SHP-1 dependent inhibition of signaling transducer and activator of transcription 3 (STAT3) (Clinical Cancer Res, 2010). In addition, we identified the effect of sorafenib on STAT3 was independent to kinases and discovered a series of sorafenib derivatives as SHP-1 enhancers (J Hepatol, 2011; Euro J Med Chem, 2011). Here, we report effects of novel sorafenib derivatives (SC-40, SC-43 and SC-60) in HCC. SC-43 induced apoptotic cell death significantly and inhibited the phosphorylation of STAT3 (p-STAT3) at the concentration of 0.5 μM, which was more potent than sorafenib (5 μM). In addition, SC-60 also exhibited similar effects on down-regulating STAT3 transcription activity and subsequently inhibited the expression of STAT3-related proteins, including cyclin D1, survivin, and Mcl-1 in HCC cells. Furthermore, these analogues showed strong induction of SHP-1 activity while incubated with pure recombinant SHP-1 protein or SHP-1-containing cell extract at the concentrations of low nanomolar ranges. In addition, we found that SHP-2, sharing high homology in critical domains with SHP-1, was not affected neither in cultured HCC cells nor recombinant SHP-2 protein. Knockdown of SHP-1, but not SHP-2, by small interference RNA reduced apoptosis and reversed blockage of p-STAT3 induced by SC-43, 40, 60. Finally, three novel compounds, including SC-43, SC-40, and SC-60 all showed significant and better effects than sorafenib on growth inhibition of orthotopic and subcutaneous xenograft HCC tumors. In conclusion, targeting SHP-1 may be feasible for the treatment of HCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1915. doi:1538-7445.AM2012-1915
Abstract 1916: Blockade of the fibroblast growth factor receptor (FGFR) system by the novel FGFR inhibitor BGJ398 modulates oncogenic signaling in human gastric cancer cells, endothelial cells and pericytes
Authors: Moser Christian , Hackl Christina , Schlitt Hans J. , Geissler Edward K. , Lang Sven A.
Published: August 2012

Expression of the FGFR system has been associated with poor prognosis in patients suffering from gastric cancer. Moreover, activation of FGFRs has been demonstrated to mediate tumor angiogenesis through regulation of endothelial cell (EC) and pericyte motility and survival. Therefore, in this study we sought to investigate the effects of targeting FGFR on gastric cancer cells as well as on important components of the tumor microenvironment by the novel FGFR inhibitor BGJ398. For the experiments, human gastric cancer cell lines (TMK-1 and KKLS) as well as ECs and pericytes (vascular smooth muscle cells, VSMCs) were used. The FGFR inhibitor BGJ398 was provided by Novartis Oncology (Basel, Switzerland). Effects of BGJ398 on tumor cell growth and chemoresistance were determined by MTT assays. Impact of BGJ398 on motility of cancer cells was evaluated by modified Boyden chambers. Signaling pathways affected by BGJ398 treatment were assessed by Western blot analyses. To determine effects of FGFR blockade on ECs and pericytes conditioned media from gastric cancer cells was used. Results show that targeting FGFR by BGJ398 impaired growth of gastric cancer cells in a dose-dependent manner in vitro. Furthermore, activation of Akt and Erk in cancer cells was effectively inhibited while expression of c-Myc and HIF-1α was substantially reduced. Additionally, therapy with BGJ398 led to significant inhibition of constitutive migratory and invasive properties of gastric cancer cells (P<0.05). Interestingly, interference with FGFR by BGJ398 increased sensitivity for chemotherapy with oxaliplatin and 5-FU in tumor cells. In ECs as well as pericytes blockade of FGFR by BGJ398 significantly inhibited cell growth and motility even upon stimulation with conditioned media (P<0.05). In conclusion, inhibition of the FGFR system by BGJ398 disrupts oncogenic signaling, impairs tumor cell motility and improves chemosensitivity in gastric cancer. Furthermore, antiangiogenic effects might be mediated via direct effects on ECs and pericytes. Hence, targeting the FGFR system may represent a promising approach to improve current therapy strategies for gastric cancer and warrants further preclinical evaluation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1916. doi:1538-7445.AM2012-1916
Abstract 1920: Small molecule inhibitor of polo-like kinase 1 (Plk1) causes significant melanoma growth delay and regression in vivo
Authors: Cholewa Brian D. , Desotelle Joshua A. , Ahmad Nihal
Published: August 2012

Melanoma is an aggressive skin cancer that rapidly metastasizes to become lethal, if not diagnosed early. An analysis of the Surveillance, Epidemiology, and End Results (SEER) program has suggested that the age-adjusted annual incidence of melanoma, both for young men and women, has increased over the past 20 years. In 2011, it is projected that over 70,000 new cases of melanoma will be diagnosed with nearly 9,000 cases resulting in death. Current preventive and treatment approaches have not been able to effectively manage this neoplasm. Therefore, new mechanism- and target- based approaches are urgently needed. Better understanding of the genetic control of cellular proliferation and cell division may provide the basis for the rational design of specific therapeutics for the management of cancers, including melanoma. We have previously demonstrated that the mitotic regulator Polo-like kinase 1 (Plk1) is over-expressed in clinical melanoma and a targeted depletion of Plk1 through lentiviral shRNA or a small-molecule inhibitor causes mitotic catastrophe and induction of apoptosis in human melanoma cells (J Invest Dermatol 129: 2843-53, 2009). In this study, we assessed the effect of BI6727, a second generation Plk1 specific small molecule inhibitor, in vivo in athymic nu/nu nude mice implanted with human melanoma cells. A375 cells (1x106 cells) were implanted in athymic nude by subcutaneous injection on the right flanks. The tumors were allowed to grow and the mice bearing established tumors (>100 mm3) were randomized into 3 groups (containing 12 mice each). The mice were treated with BI6727 at two doses (10 mg/kg or 25 mg/kg) or a vehicle control via tail vein injections twice weekly. Our data demonstrates that in BI6727 treated animals, the rate of tumor growth (to reach to a 1000 mm3 target volume) was significantly delayed. We also found that BI6727 at 25 mg/kg resulted in a near complete tumor regression. The observed tumor regression was stable for several days following discontinuation of treatment, after which, the tumors started growing again. Additionally, we found that BI6727 caused i) a significant inhibition of cell growth and viability, and ii) induction of apoptosis in multiple melanoma cell lines. Based on this study, we suggest that Plk1 is a druggable target for melanoma and pharmacological inhibitors of Plk1 could be clinically useful in melanoma therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1920. doi:1538-7445.AM2012-1920
Abstract 1932: Phenylmethimazole decreases constitutively high basal levels of Interleukin 6 (IL-6) in a triple negative breast cancer cell line
Authors: O'Brien John D. , Goetz Douglas J. , Alapati Anuja , Deosarkar Sudhir P. , Kohn Leonard D. , McCall Kelly D.
Published: August 2012

Interleukin 6 (IL-6) is a pro-inflammatory cytokine that plays an important role in the progression of many types of cancers by regulating macrophage recruitment and differentiation, as well as modulating the activation of the JAK/Stat signal cascade, and inducing pro-angiogenic factors. Several studies have correlated a negative prognosis in cancer with a high level of IL-6 in patient serum. Triple (i.e. progesterone, human epidermal growth factor-2, and estrogen receptor) negative breast cancer has been reported to be highly aggressive and invasive. MDA-MB-231, a triple negative breast cancer cell line, expresses high basal levels of IL-6. Recently our laboratory demonstrated that phenylmethimazole (C10), a novel toll-like receptor inhibitor, diminishes basal expression of IL-6 in pancreatic cancer and melanoma cells as well as their growth and migration. Based on these observations, we sought to test the hypothesis that C10 inhibits IL-6 expression in MDA-MB-231 cells. Here we report that triple negative breast cancer cells express considerably higher levels of IL-6 than estrogen receptor positive breast cancer cells and that C10 significantly reduces IL-6 expression in the triple negative breast cancer cells. This inhibition was observed at both the RNA and protein level. Given the importance of IL-6 in cancer progression, and our finding that C10 attenuates IL-6 expression in the MDA-MB-231 cell line, this study suggests that C10 may have therapeutic potential for triple negative breast cancers. Additional studies are currently underway to further explore these hypotheses.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1932. doi:1538-7445.AM2012-1932