bluuebunny/crossref_metadata_2025 huggingface dataset:
Novel Mechanism of Lapatinib Resistance in HER2-Positive Breast Tumor Cells: Activation of AXL
Authors: Liu Li , Greger James , Shi Hong , Liu Yuan , Greshock Joel , Annan Roland , Halsey Wendy , Sathe Ganesh M. , Martin Anne-Marie , Gilmer Tona M.
Published: August 2009

HER2-directed therapies, such as trastuzumab and lapatinib, are important treatments for breast cancer. However, some tumors do not respond or develop resistance to these agents. We isolated and characterized multiple lapatinib-resistant, HER2-positive, estrogen receptor (ER)–positive breast cancer clones derived from lapatinib-sensitive BT474 cells by chronic exposure to lapatinib. We show overexpression of AXL as a novel mechanism of acquired resistance to HER2-targeted agents in these models. GSK1363089 (foretinib), a multikinase inhibitor of AXL, MET, and vascular endothelial growth factor receptor currently in phase II clinical trials, restores lapatinib and trastuzumab sensitivity in these resistant cells that exhibit increased AXL expression. Furthermore, small interfering RNA to AXL, estrogen deprivation, or fulvestrant, an ER antagonist, decreases AXL expression and restores sensitivity to lapatinib in these cells. Taken together, these data provide scientific evidence to assess the expression of AXL in HER2-positive, ER-positive patients who have progressed on either lapatinib or trastuzumab and to test the combination of HER2-targeted agents and GSK1363089 in the clinic. [Cancer Res 2009;69(17):6871–8]
Kisspeptin-10, a KISS1-Derived Decapeptide, Inhibits Tumor Angiogenesis by Suppressing Sp1-Mediated VEGF Expression and FAK/Rho GTPase Activation
Authors: Cho Sung-Gook , Yi Zhengfang , Pang Xiufeng , Yi Tingfang , Wang Ying , Luo Jian , Wu Zirong , Li Dali , Liu Mingyao
Published: August 2009

Kisspeptin-10 (Kp-10), a decapeptide derived from the primary translation product of KISS1 gene, has been reported previously to be a key hormone for puberty and an inhibitor for tumor metastasis via the activation of G protein–coupled receptor 54. However, whether Kp-10 inhibits angiogenesis, which is critical for tumor growth and metastasis and other human diseases, is still unknown. Here we show that Kp-10 significantly inhibits human umbilical vein endothelial cell (HUVEC) migration, invasion, and tube formation, key processes in angiogenesis. Using chicken chorioallantoic membrane assay and vascular endothelial growth factor (VEGF)–induced mouse corneal micropocket assay, we show that Kp-10 inhibits angiogenesis in vivo. Furthermore, Kp-10 inhibits tumor growth in severe combined immunodeficient mice xenografted with human prostate cancer cells (PC-3) through inhibiting tumor angiogenesis, whereas Kp-10 has little effect on the proliferation of HUVECs and human prostate cancer cells. In deciphering the underlying molecular mechanisms, we show that Kp-10 suppresses VEGF expression by inhibiting the binding of specificity protein 1 to VEGF promoter and by blocking the activation of c-Src/focal adhesion kinase and Rac/Cdc42 signaling pathways in HUVECs, leading to the inhibition of tumor angiogenesis. [Cancer Res 2009;69(17):7062–70]
Paclitaxel Directly Binds to Bcl-2 and Functionally Mimics Activity of Nur77
Authors: Ferlini Cristiano , Cicchillitti Lucia , Raspaglio Giuseppina , Bartollino Silvia , Cimitan Samanta , Bertucci Carlo , Mozzetti Simona , Gallo Daniela , Persico Marco , Fattorusso Caterina , Campiani Giuseppe , Scambia Giovanni
Published: August 2009

We reported previously that Bcl-2 is paradoxically down-regulated in paclitaxel-resistant cancer cells. We reveal here that paclitaxel directly targets Bcl-2 in the loop domain, thereby facilitating the initiation of apoptosis. Molecular modeling revealed an extraordinary similarity between the paclitaxel binding sites in Bcl-2 and β-tubulin, leading us to speculate that paclitaxel could be mimetic of an endogenous peptide ligand, which binds both proteins. We tested the hypothesis that paclitaxel mimics Nur77, which, like paclitaxel, changes the function of Bcl-2. This premise was confirmed by Nur77 interacting with both paclitaxel targets (Bcl-2 and β-tubulin) and a peptide sequence mimicking the Nur77 structural region, thus reproducing the paclitaxel-like effects of tubulin polymerization and opening the permeability transition pore channel in mitochondria. This discovery could help in the development of novel anticancer agents with nontaxane skeleton as well as in identifying the clinical subsets responsive to paclitaxel-based therapy. [Cancer Res 2009;69(17):6906–14]
Epigenetic Deregulation of DNA Repair and Its Potential for Therapy
Authors: Hegi Monika E. , Sciuscio Davide , Murat Anastasia , Levivier Marc , Stupp Roger
Published: August 2009

Epigenetic silencing of essential components of DNA repair pathways is a common event in many tumor types, and comprise O6-methylguanine-DNA methyltransferase (MGMT), human mut L homolog 1 (hMLH1), Werner syndrome gene (WRN), breast cancer susceptibility gene 1 (BRCA1), and genes of the Fanconi anemia pathway. Most interestingly, some of these alterations become the Achilles heel of the affected tumors upon treatment with certain classes of anticancer agents. That is, patients whose tumors carry such defects can be stratified for respective therapy rendering some classic DNA damaging agents, such as alkylators or DNA crosslinking agents, into “targeted therapies.” Here we review some of the affected repair pathways that, when inactivated, sensitize the tumors to specific drugs and are thus exploitable for individualized therapy. (Clin Cancer Res 2009;15(16):5026–31)
miR-200 Expression Regulates Epithelial-to-Mesenchymal Transition in Bladder Cancer Cells and Reverses Resistance to Epidermal Growth Factor Receptor Therapy
Authors: Adam Liana , Zhong Meng , Choi Woonyoung , Qi Wei , Nicoloso Milena , Arora Ameeta , Calin George , Wang Hua , Siefker-Radtke Arlene , McConkey David , Bar-Eli Menashe , Dinney Colin
Published: August 2009

Purpose: The epithelial-to-mesenchymal transition (EMT) is a cell development-regulated process in which noncoding RNAs act as crucial modulators. Recent studies have implied that EMT may contribute to resistance to epidermal growth factor receptor (EGFR)–directed therapy. The aims of this study were to determine the potential role of microRNAs (miRNA) in controlling EMT and the role of EMT in inducing the sensitivity of human bladder cancer cells to the inhibitory effects of the anti-EGFR therapy.

Experimental Design: miRNA array screening and real-time reverse transcription-PCR were used to identify and validate the differential expression of miRNAs involved in EMT in nine bladder cancer cell lines. A list of potential miR-200 direct targets was identified through the TargetScan database. The precursor of miR-200b and miR-200c was expressed in UMUC3 and T24 cells using a retrovirus or a lentivirus construct, respectively. Protein expression and signaling pathway modulation, as well as intracellular distribution of EGFR and ERRFI-1, were validated through Western blot analysis and confocal microscopy, whereas ERRFI-1 direct target of miR-200 members was validated by using the wild-type and mutant 3′-untranslated region/ERRFI-1/luciferse reporters.

Results: We identified a tight association between the expression of miRNAs of the miR-200 family, epithelial phenotype, and sensitivity to EGFR inhibitors–induced growth inhibition in bladder carcinoma cell lines. Stable expression of miR-200 in mesenchymal UMUC3 cells increased E-cadherin levels, decreased expression of ZEB1, ZEB2, ERRFI-1, and cell migration, and increased sensitivity to EGFR-blocking agents. The changes in EGFR sensitivity by silencing or forced expression of ERRFI-1 or by miR-200 expression have also been validated in additional cell lines, UMUC5 and T24. Finally, luciferase assays using 3′-untranslated region/ERRFI-1/luciferase and miR-200 cotransfections showed that the direct down-regulation of ERRFI-1 was miR-200-dependent because mutations in the two putative miR-200-binding sites have rescued the inhibitory effect.

Conclusions: Members of the miR-200 family appear to control the EMT process and sensitivity to EGFR therapy in bladder cancer cells and the expression of miR-200 is sufficient to restore EGFR dependency at least in some of the mesenchymal bladder cancer cells. The targets of miR-200 include ERRFI-1, which is a novel regulator of EGFR-independent growth. (Clin Cancer Res 2009;15(16):5060–72)
A Three-Gene Signature for Outcome in Soft Tissue Sarcoma
Authors: Hoffmann Andreas-Claudius , Danenberg Kathleen D. , Taubert Helge , Danenberg Peter V. , Wuerl Peter
Published: August 2009

Purpose: Finding markers or gene sets that would further classify patients into different risk categories and thus allow more individually adapted multimodality treatment regimens in soft tissue sarcomas is necessary. In this study, we investigated the prognostic values of hypoxia-inducible factor 1a (HIF1a), heparin-binding epidermal growth factor–like growth factor (HB-EGF), vascular endothelial growth factor (VEGF), and other angiogenesis-related gene expressions, as well as their interrelationships.

Experimental Design: Formalin-fixed paraffin-embedded tissue samples were obtained from 45 patients with soft tissue sarcoma (median age 57 years, range 16–85 years). After laser capture microdissection direct quantitative real-time reverse transcription-PCR (TaqMan) assays were done in triplicates to determine HIF1a, HB-EGF, VEGF, and other gene expression levels.

Results: Multivariate Cox regression analysis revealed significant independent associations of HB-EGF, HIF1a, and VEGF-C gene expression to the overall survival (P < 0.0001). A combined factor of these three genes showed a relative risk for shorter survival of 5.5, more than twice higher as in an increasing International Union against Cancer Stage. Receiver operating characteristic curve analysis showed a significant sensitivity of 73% and specificity of 82% of this factor for the diagnosis of short (<3 years) versus long (3-9 years) survival (P = 0.0002). VEGF-A showed significant gender differences in the association to survival.

Conclusions: Measuring HIF1a, HB-EGF, and VEGF-C expression may contribute to a better understanding of the prognosis of patients with soft tissue sarcoma and may even play a crucial role for the distribution of patients to multimodal therapeutic regimens. Prospective studies investigating the response to different adjuvant or palliative therapies seem to be warranted. (Clin Cancer Res 2009;15(16):5191–8)
Phase I Study of Vorinostat in Combination with Bortezomib for Relapsed and Refractory Multiple Myeloma
Authors: Badros Ashraf , Burger Angelika M. , Philip Sunita , Niesvizky Ruben , Kolla Sarah S. , Goloubeva Olga , Harris Carolynn , Zwiebel James , Wright John J. , Espinoza-Delgado Igor , Baer Maria R. , Holleran Julianne L. , Egorin Merrill J. , Grant Steven
Published: August 2009

Purpose: Vorinostat, a histone deacetylase inhibitor, enhances cell death by the proteasome inhibitor bortezomib in vitro. We sought to test the combination clinically.

Experimental Design: A phase I trial evaluated sequential dose escalation of bortezomib at 1 to 1.3 mg/m2 i.v. on days 1, 4, 8, and 11 and vorinostat at 100 to 500 mg orally daily for 8 days of each 21-day cycle in relapsed/refractory multiple myeloma patients. Vorinostat pharmacokinetics and dynamics were assessed.

Results: Twenty-three patients were treated. Patients had received a median of 7 prior regimens (range, 3-13), including autologous transplantation in 20, thalidomide in all 23, lenalidomide in 17, and bortezomib in 19, 9 of whom were bortezomib-refractory. Two patients receiving 500 mg vorinostat had prolonged QT interval and fatigue as dose-limiting toxicities. The most common grade >3 toxicities were myelo-suppression (n = 13), fatigue (n = 11), and diarrhea (n = 5). There were no drug-related deaths. Overall response rate was 42%, including three partial responses among nine bortezomib refractory patients. Vorinostat pharmacokinetics were nonlinear. Serum Cmax reached a plateau above 400 mg. Pharmacodynamic changes in CD-138+ bone marrow cells before and on day 11 showed no correlation between protein levels of NF-κB, IκB, acetylated tubulin, and p21CIP1 and clinical response.

Conclusions: The maximum tolerated dose of vorinostat in our study was 400 mg daily for 8 days every 21 days, with bortezomib administered at a dose of 1.3 mg/m2 on days 1, 4, 8, and 11. The promising antimyeloma activity of the regimen in refractory patients merits further evaluation. (Clin Cancer Res 2009;15(16):5250–7)
Prognostic Significance of Interleukin-6 Single Nucleotide Polymorphism Genotypes in Neuroblastoma:rs1800795(Promoter) andrs8192284(Receptor)
Authors: Lagmay Joanne P. , London Wendy B. , Gross Thomas G. , Termuhlen Amanda , Sullivan Nicholas , Axel Amy , Mundy Bethany , Ranalli Mark , Canner Jason , McGrady Patrick , Hall Brett
Published: August 2009

Purpose: Neuroblastoma is a childhood cancer of the sympathetic nervous system and many patients present with high-risk disease. Risk stratification, based on pathology and tumor-derived biomarkers, has improved prediction of clinical outcomes, but overall survival (OS) rates remain unfavorable and new therapeutic targets are needed. Some studies suggest a link between interleukin (IL)-6 and more aggressive behavior in neuroblastoma tumor cells. Therefore, we examined the impact of two IL-6 single nucleotide polymorphisms (SNP) on neuroblastoma disease progression.

Experimental Design: DNA samples from 96 high-risk neuroblastoma patients were screened for two SNP that are known to regulate the serum levels of IL-6 and the soluble IL-6 receptor, rs1800795 and rs8192284, respectively. The genotype for each SNP was determined in a blinded fashion and independent statistical analysis was done to determine SNP-related event-free survival (EFS) and OS rates.

Results: The rs1800795 IL-6 promoter SNP is an independent prognostic factor for EFS and OS in high-risk neuroblastoma patients. In contrast, the rs8192284 IL-6 receptor SNP revealed no prognostic value.

Conclusions: The rs1800795 SNP [−174 IL-6 (G > C)] represents a novel and independent prognostic marker for both EFS and OS in high-risk neuroblastoma. Because the rs1800795 SNP [−174 IL-6 (G > C)] has been shown to correlate with production of IL-6, this cytokine may represent a target for development of new therapies in neuroblastoma. (Clin Cancer Res 2009;15(16):5234–9)
Imaging of αvβ3 Expression by a Bifunctional Chimeric RGD Peptide not Cross-Reacting with αvβ5
Authors: Zannetti Antonella , Del Vecchio Silvana , Iommelli Francesca , Del Gatto Annarita , De Luca Stefania , Zaccaro Laura , Papaccioli Angela , Sommella Jvana , Panico Mariarosaria , Speranza Antonio , Grieco Paolo , Novellino Ettore , Saviano Michele , Pedone Carlo , Salvatore Marco
Published: August 2009

Purpose: To test whether a novel bifunctional chimeric peptide comprising a cyclic Arg-Gly-Asp pentapeptide covalently bound to an echistatin domain can discriminate αvβ3 from αvβ5 integrin, thus allowing the in vivo selective visualization of αvβ3 expression by single-photon and positron emission tomography (PET) imaging.

Experimental Design: The chimeric peptide was preliminarily tested for inhibition of αvβ3-dependent cell adhesion and competition of 125I-echistatin binding to membrane of stably transfected K562 cells expressing αvβ3 (Kαvβ3) or αvβ5 (Kαvβ5) integrin. The chimeric peptide was then conjugated with diethylenetriaminepentaacetic acid and labeled with 111In for single-photon imaging, whereas a one-step procedure was used for labeling the full-length peptide and a truncated derivative, lacking the last five C-terminal amino acids, with 18F for PET imaging. Nude mice bearing tumors from Kαvβ3, Kαvβ5, U87MG human glioblastoma, and A431 human epidermoid cells were subjected to single-photon and PET imaging.

Results: Adhesion and competitive binding assays showed that the novel chimeric peptide selectively binds to αvβ3 integrin and does not cross-react with αvβ5. In agreement with in vitro findings, single-photon and PET imaging studies showed that the radiolabeled chimeric peptide selectively localizes in tumor xenografts expressing αvβ3 and fails to accumulate in those expressing αvβ5 integrin. When 18F-labeled truncated derivative was used for PET imaging, αvβ3- and αvβ5-expressing tumors were visualized, indicating that the five C-terminal amino acids are required to differentially bind the two integrins.

Conclusion: Our findings indicate that the novel chimeric Arg-Gly-Asp peptide, having no cross-reaction with αvβ5 integrin, allows highly selective αvβ3 expression imaging and monitoring. (Clin Cancer Res 2009;15(16):5224–33)
CIP2A Is Associated with Human Breast Cancer Aggressivity
Authors: Côme Christophe , Laine Anni , Chanrion Maïa , Edgren Henrik , Mattila Elina , Liu Xiaoling , Jonkers Jos , Ivaska Johanna , Isola Jorma , Darbon Jean-Marie , Kallioniemi Olli , Thézenas Simon , Westermarck Jukka
Published: August 2009

Purpose: To investigate the clinical relevance of the recently characterized human oncoprotein cancerous inhibitor of protein phosphatase 2A (CIP2A) in human breast cancer.

Experimental Design: CIP2A expression (mRNA and protein) was measured in three different sets of human mammary tumors and compared with clinicopathologic variables. The functional role of CIP2A in breast cancer cells was evaluated by small interfering RNA–mediated depletion of the protein followed by an analysis of cell proliferation, migration, anchorage-independent growth, and xenograft growth.

Results: CIP2A mRNA is overexpressed (n = 159) and correlates with higher Scarff-Bloom-Richardson grades (n = 251) in samples from two independent human breast cancer patients. CIP2A protein was found to be overexpressed in 39% of 33 human breast cancer samples. Furthermore, CIP2A mRNA expression positively correlated with lymph node positivity of the patients and with the expression of proliferation markers and p53 mutations in the tumor samples. Moreover, CIP2A protein expression was induced in breast cancer mouse models presenting mammary gland–specific depletion of p53 and either BRCA1 or BRCA2. Functionally, CIP2A depletion was shown to inhibit the expression of its target protein c-Myc. Loss of CIP2A also inhibited anchorage-independent growth in breast cancer cells. Finally, CIP2A was shown to support MDA-MB-231 xenograft growth in nude mice.

Conclusions: Our data show that CIP2A is associated with clinical aggressivity in human breast cancer and promotes the malignant growth of breast cancer cells. Thus, these results validate the role of CIP2A as a clinically relevant human oncoprotein and warrant further investigation of CIP2A as a therapeutic target in breast cancer treatment. (Clin Cancer Res 2009;15(16):5092–100)
MicroRNA Regulation of Oncolytic Herpes Simplex Virus-1 for Selective Killing of Prostate Cancer Cells
Authors: Lee Cleo Y. F. , Rennie Paul S. , Jia William W.G.
Published: August 2009

Purpose: Advanced castration-resistant prostate cancer, for which there are few treatment options, remains one of the leading causes of cancer death. MicroRNAs (miRNA) have provided a new opportunity for more stringent regulation of tumor-specific viral replication. The purpose of this study was to provide a proof-of-principle that miRNA-regulated oncolytic herpes simplex virus-1 (HSV-1) virus can selectively target cancer cells with reduced toxicity to normal tissues.

Experimental Design: We incorporated multiple copies of miRNA complementary target sequences (for miR-143 or miR-145) into the 3′-untranslated region (3′-UTR) of an HSV-1 essential viral gene, ICP4, to create CMV-ICP4-143T and CMV-ICP4-145T amplicon viruses and tested their targeting specificity and efficacy both in vitro and in vivo.

Results: Although miR-143 and miR-145 are highly expressed in normal tissues, they are significantly down-regulated in prostate cancer cells. We further showed that miR-143 and miR-145 inhibited the expression of the ICP4 gene at the translational level by targeting the corresponding 3′-UTR in a dose-dependent manner. This enabled selective viral replication in prostate cancer cells. When mice bearing LNCaP human prostate tumors were treated with these miRNA-regulated oncolytic viruses, a >80% reduction in tumor volume was observed, with significantly attenuated virulence to normal tissues in comparison with control amplicon viruses not carrying these 3′-UTR sequences.

Conclusion: Our study is the first to show that inclusion of specific miRNA target sequences into the 3′-UTR of an essential HSV-1 gene is a viable strategy for restricting viral replication and oncolysis to cancer cells while sparing normal tissues. (Clin Cancer Res 2009;15(16):5126–35)
Tumor and Host-Mediated Pathways of Resistance and Disease Progression in Response to Antiangiogenic Therapy
Authors: Ebos John M. L. , Lee Christina R. , Kerbel Robert S.
Published: August 2009

Despite early benefits seen in cancer patients treated with antivascular endothelial growth factor (VEGF) pathway-targeted drugs, the clinical benefits obtained in terms of progression-free or overall survival have been more modest than expected. This outcome is, at least in part, due to antiangiogenic drug resistance mechanisms that involve pathways mediated largely by the tumor, whether intrinsic or acquired in response to therapy, or by the host, which is either responding directly to therapy or indirectly to tumoral cues. The focus of this review is to distinguish, where possible, between such host and tumor-mediated pathways of resistance and discuss key challenges facing the preclinical and clinical development of antiangiogenic agents, including potential differences in drug efficacies when treating primary tumors or various stages of metastatic disease. (Clin Cancer Res 2009;15(16):5020–5)
A Limited Sampling Schedule to Estimate Individual Pharmacokinetic Parameters of Fludarabine in Hematopoietic Cell Transplant Patients
Authors: Salinger David H. , Blough David K. , Vicini Paolo , Anasetti Claudio , O'Donnell Paul V. , Sandmaier Brenda M. , McCune Jeannine S.
Published: August 2009

Purpose: Fludarabine monophosphate (fludarabine) is frequently administered to patients receiving a reduced-intensity conditioning regimen for allogeneic hematopoietic cell transplant (HCT) in an ambulatory care setting. These patients experience significant interpatient variability in clinical outcomes, potentially due to pharmacokinetic variability in 2-fluoroadenine (F-ara-A) plasma concentrations. To test such hypotheses, patient compliance with the blood sampling should be optimized by the development of a minimally intrusive limited sampling schedule (LSS) to characterize F-ara-A pharmacokinetics. To this end, we sought to create the first F-ara-A population pharmacokinetic model and subsequently a LSS.

Experimental Design: A retrospective evaluation of F-ara-A pharmacokinetics was conducted after one or more doses of daily i.v. fludarabine in 42 adult HCT recipients. NONMEM software was used to estimate the population pharmacokinetic parameters and compute the area under the concentration-time curve.

Results: A two-compartment model best fits the data. A LSS was constructed using a simulation approach, seeking to minimize the scaled mean squared error for the area under the concentration-time curve for each simulated individual. The LSS times chosen were 0.583, 1.5, 6.5, and 24 hours after the start of the 30-minute fludarabine infusion.

Discussion: The pharmacokinetics of F-ara-A in an individual HCT patient can be accurately estimated by obtaining four blood samples (using the LSS) and maximum a posteriori Bayesian estimation.

Conclusion: These are essential tools for prospective pharmacodynamic studies seeking to determine if clinical outcomes are related to F-ara-A pharmacokinetics in patients receiving i.v. fludarabine in the ambulatory clinic. (Clin Cancer Res 2009;15(16):5280–7)
Inhibition of Insulin-Like Growth Factor 1 Receptor by CP-751,871 Radiosensitizes Non–Small Cell Lung Cancer Cells
Authors: Iwasa Tsutomu , Okamoto Isamu , Suzuki Minoru , Hatashita Erina , Yamada Yuki , Fukuoka Masahiro , Ono Koji , Nakagawa Kazuhiko
Published: August 2009

Purpose: Therapeutic strategies that target the insulin-like growth factor I receptor (IGF-1R) hold promise for a wide variety of cancers. We have now investigated the effect of CP-751,871, a fully human monoclonal antibody specific for IGF-IR, on the sensitivity of human non–small cell lung cancer (NSCLC) cell lines to radiation.

Experimental Design: The radiosensitizing effect of CP-751,871 was evaluated on the basis of cell death, clonogenic survival, and progression of tumor xenografts. Radiation-induced damage was evaluated by immunofluorescence analysis of the histone γ-H2AX and Rad51.

Results: A clonogenic survival assay revealed that CP-751,871 increased the sensitivity of NSCLC cells to radiation in vitro. CP-751,871 inhibited radiation-induced IGF-IR signaling, and potentiated the radiation-induced increases both in the number of apoptotic cells and in the activity of caspase-3. Immunofluorescence analysis of the histone γ-H2AX and Rad51 also showed that CP-751,871 inhibited the repair of radiation-induced DNA double-strand breaks. Finally, combination therapy with CP-751,871 and radiation delayed the growth of NSCLC tumor xenografts in nude mice to a greater extent than did either treatment modality alone.

Conclusions: These results show that CP-751,871 sensitizes NSCLC cells to radiation both in vitro and in vivo, and that this effect of CP-751,871 is likely attributable to the inhibition of DNA repair and enhancement of apoptosis that result from attenuation of IGF-IR signaling. Combined treatment with CP-751,871 and radiation thus warrants further investigation in clinical trials as a potential anticancer strategy. (Clin Cancer Res 2009;15(16):5117–25)
Unique Clinicopathologic Features Characterize ALK-Rearranged Lung Adenocarcinoma in the Western Population
Authors: Rodig Scott J. , Mino-Kenudson Mari , Dacic Sanja , Yeap Beow Y. , Shaw Alice , Barletta Justine A. , Stubbs Hannah , Law Kenny , Lindeman Neal , Mark Eugene , Janne Pasi A. , Lynch Thomas , Johnson Bruce E. , Iafrate A. John , Chirieac Lucian R.
Published: August 2009

Purpose: The anaplastic large cell kinase gene (ALK) is rearranged in ∼5% of lung adenocarcinomas within the Asian population. We evaluated the incidence and the characteristics of ALK-rearranged lung adenocarcinomas within the western population and the optimal diagnostic modality to detect ALK rearrangements in routine clinical practice.

Experimental Design: We tested 358 lung adenocarcinomas from three institutions for ALK rearrangements by fluorescent in situ hybridization (FISH) and immunohistochemistry with and without tyramide amplification. The clinicopathologic characteristics of tumors with and without ALK rearrangements were compared.

Results: We identified 20 (5.6%) lung adenocarcinomas with ALK rearrangements within our cohort of western patients. ALK rearrangement was associated with younger age (P = 0.0002), never smoking (P < 0.0001), advanced clinical stage (P = 0.0001), and a solid histology with signet-ring cells (P < 0.0001). ALK rearrangement was identified by FISH in 95% of cases and immunohistochemistry with and without tyramide amplification in 80% and 40% of cases, respectively, but neither FISH nor immunohistochemistry alone detected all cases with ALK rearrangement on initial screening. None of the ALK-rearranged tumors harbored coexisting EGFR mutations.

Conclusions: Lung adenocarcinomas with ALK rearrangements are uncommon in the western population and represent a distinct entity of carcinomas with unique characteristics. For suspected cases, dual diagnostic testing, with FISH and immunohistochemistry, should be considered to accurately identify lung adenocarcinomas with ALK rearrangement. (Clin Cancer Res 2009;15(16):5216–23)
Differential Clinical Significance of Individual NKG2D Ligands in Melanoma: Soluble ULBP2 as an Indicator of Poor Prognosis Superior to S100B
Authors: Paschen Annette , Sucker Antje , Hill Bettina , Moll Iris , Zapatka Marc , Nguyen Xuan Duc , Sim Geok Choo , Gutmann Isabelle , Hassel Jessica , Becker Jürgen C. , Steinle Alexander , Schadendorf Dirk , Ugurel Selma
Published: August 2009

Purpose: Cytotoxic lymphocytes interact with human tumor cells via the activating immunoreceptor NKG2D, recognizing a variety of stress-associated MIC and ULBP surface molecules. However, tumors can escape from this immunosurveillance by shedding NKG2D ligands (NKG2DL), rendering the soluble products detectable in patients' sera.

Experimental Design: To elucidate the clinical significance of NKG2DL diversity, we studied their expression on melanoma tissues and their presence as soluble molecules in sera from >200 melanoma patients and compared the latter with the well-established serum marker S100B.

Results: Immunohistochemistry revealed a heterogeneous expression of MIC and ULBP2 molecules between and within melanoma metastases. Compared with MIC, ULBP2 was less frequently expressed. Accordingly, elevated levels of soluble ULBP2 (sULBP2) were detected in sera of melanoma patients less frequently than elevated levels of soluble MICA (sMICA), although both soluble NKG2DL (sNKG2DL) were significantly increased compared with sera of healthy controls (P < 0.0001). Strikingly, elevated concentrations of sULBP2, but not of sMICA, were strongly associated with disease progression (P < 0.0001) and tumor load (P = 0.0003). Elevated serum levels of either sNKG2DL correlated with reduced overall survival, albeit considerably stronger for sULBP2 (P < 0.0001) than for sMICA (P = 0.011). In early-stage (I-III) melanoma patients, only sULBP2 (P < 0.0001) but neither sMICA nor S100B revealed prognostic significance. Multivariate analysis identified sULBP2 (P = 0.0015) and S100B (P = 0.013) but not sMICA as independent predictors of prognosis.

Conclusion: Our data reveal marked differences in the clinical significance of individual sNKG2DL. Only sULBP2 is an independent predictor of prognosis, the significance of which is superior to the well-established and widely used melanoma serum marker S100B. (Clin Cancer Res 2009;15(16):5208–15)
Impact of Epidermal Growth Factor Receptor and KRAS Mutations on Clinical Outcomes in Previously Untreated Non–Small Cell Lung Cancer Patients: Results of an Online Tumor Registry of Clinical Trials
Authors: Jackman David M. , Miller Vincent A. , Cioffredi Leigh-Anne , Yeap Beow Y. , Jänne Pasi A. , Riely Gregory J. , Ruiz Marielle Gallegos , Giaccone Giuseppe , Sequist Lecia V. , Johnson Bruce E.
Published: August 2009

Purpose: The impact of epidermal growth factor receptor (EGFR) and KRAS genotypes on outcomes with erlotinib or gefitinib therapy continues to be debated. This study combines patient data from five trials in predominantly Western populations to assess the impact of EGFR and KRAS mutations on first-line therapy with an EGFR–tyrosine kinase inhibitor (TKI) and compare clinical versus molecular predictors of sensitivity.

Experimental Design: Chemotherapy-naïve patients with advanced non–small cell lung cancer and known EGFR mutation status treated with erlotinib or gefitinib monotherapy as part of a clinical trial were eligible for inclusion. Patients received daily erlotinib (150 mg) or gefitinib (250 mg) until disease progression or unacceptable toxicity. Data were collected in a password-protected web database. Clinical outcomes were analyzed to look for differences based on EGFR and KRAS genotypes, as well as clinical characteristics.

Results: Patients (223) from five clinical trials were included. Sensitizing EGFR mutations were associated with a 67% response rate, time to progression (TTP) of 11.8 months, and overall survival of 23.9 months. Exon 19 deletions were associated with longer median TTP and overall survival compared with L858R mutations. Wild-type EGFR was associated with poorer outcomes (response rate, 3%; TTP, 3.2 months) irrespective of KRAS status. No difference in outcome was seen between patients harboring KRAS transition versus transversion mutations. EGFR genotype was more effective than clinical characteristics at selecting appropriate patients for consideration of first-line therapy with an EGFR-TKI.

Conclusion: EGFR mutation status is associated with sensitivity to treatment with an EGFR-TKI in patients with advanced non–small cell lung cancer. Patients harboring sensitizing EGFR mutations should be considered for first-line erlotinib or gefitinib. (Clin Cancer Res 2009;15(16):5267–73)
A Randomized, Double-Blind, Placebo-Controlled, Phase II Study Comparing the Tolerability and Efficacy of Ipilimumab Administered with or without Prophylactic Budesonide in Patients with Unresectable Stage III or IV Melanoma
Authors: Weber Jeffrey , Thompson John A. , Hamid Omid , Minor David , Amin Asim , Ron Ilan , Ridolfi Ruggero , Assi Hazem , Maraveyas Anthony , Berman David , Siegel Jonathan , O'Day Steven J.
Published: August 2009

Purpose: Diarrhea (with or without colitis) is an immune-related adverse event (irAE) associated with ipilimumab. A randomized, double-blind, placebo-controlled, multicenter, multinational phase II trial was conducted to determine whether prophylactic budesonide (Entocort EC), a nonabsorbed oral steroid, reduced the rate of grade ≥2 diarrhea in ipilimumab-treated patients with advanced melanoma.

Experimental Design: Previously treated and treatment-naïve patients (N = 115) with unresectable stage III or IV melanoma received open-label ipilimumab (10 mg/kg every 3 weeks for four doses) with daily blinded budesonide (group A) or placebo (group B) through week 16. The first scheduled tumor evaluation was at week 12; eligible patients received maintenance treatment starting at week 24. Diarrhea was assessed using Common Terminology Criteria for Adverse Events (CTCAE) 3.0. Patients kept a diary describing their bowel habits.

Results: Budesonide did not affect the rate of grade ≥2 diarrhea, which occurred in 32.7% and 35.0% of patients in groups A and B, respectively. There were no bowel perforations or treatment-related deaths. Best overall response rates were 12.1% in group A and 15.8% in group B, with a median overall survival of 17.7 and 19.3 months, respectively. Within each group, the disease control rate was higher in patients with grade 3 to 4 irAEs than in patients with grade 0 to 2 irAEs, although many patients with grade 1 to 2 irAEs experienced clinical benefit. Novel patterns of response to ipilimumab were observed.

Conclusions: Ipilimumab shows activity in advanced melanoma, with encouraging survival and manageable adverse events. Budesonide should not be used prophylactically for grade ≥2 diarrhea associated with ipilimumab therapy. (Clin Cancer Res 2009;15(17):5591–8)
Phosphoinositide 3-Kinase Mutations in Breast Cancer: A “Good” Activating Mutation?
Authors: Di Cosimo Serena , Baselga José
Published: August 2009

Gain-of-function mutations in the catalytic subunit of phosphoinositide-3-kinase (PI3KCA) occur frequently in breast cancer. Kalinsky and colleagues show that PI3KCA mutations are associated with favorable clinicopathologic features and better clinical outcome, including survival. These findings will have to be considered in the design and interpretation of clinical trials with inhibitors of the PI3K pathway. (Clin Cancer Res 2009;15(16):5017–9)
Antitumor efficacy of IPI-504, a selective heat shock protein 90 inhibitor against human epidermal growth factor receptor 2–positive human xenograft models as a single agent and in combination with trastuzumab or lapatinib
Authors: Leow Ching Ching , Chesebrough Jon , Coffman Karen T. , Fazenbaker Christine A. , Gooya John , Weng David , Coats Steve , Jackson Dowdy , Jallal Bahija , Chang Yong
Published: August 2009

IPI-504 is a novel, highly soluble small-molecule inhibitor of heat shock protein 90 (Hsp90), a protein chaperone essential for regulating homeostasis of oncoproteins and cell signaling proteins. Human epidermal growth factor receptor 2 (HER2; ErbB2) oncoprotein, expressed in a subset of metastatic breast cancers, is a Hsp90 client protein. In this study, we investigated the antitumor activity and the mechanism of action of IPI-504 in HER2+, trastuzumab-sensitive and trastuzumab-refractory cell lines in vitro and in vivo. IPI-504 exhibited potent antiproliferative activities (range of IC50, 10-40 nmol/L) against several tumor cell lines examined, whereby mechanism of action was mediated through HER2 and Akt degradation. Both intravenous and oral administration of IPI-504 assessed in multiple schedules showed potent tumor growth inhibition in vivo with corresponding degradation of HER2. The tolerability and efficacy of IPI-504 combined with either trastuzumab or lapatinib were also investigated in HER2+ tumor xenograft models. Combination of IPI-504 with trastuzumab significantly enhanced tumor growth delay and induced greater responses when compared with either agent alone. Although, as expected, trastuzumab alone did not exhibit any significant antitumor activity in the trastuzumab-resistant JIMT-1 model, IPI-504 administered in combination with trastuzumab yielded greater antitumor efficacy than either agent alone. Finally, combination of IPI-504 and lapatinib was well tolerated up to 50 mg/kg IPI-504 and 100 mg/kg lapatinib and resulted in significant delay in tumor growth, including partial and complete tumor responses. These lines of evidence support the development of IPI-504 in HER2-positive breast cancers as a single agent and in combination with either trastuzumab or lapatinib.[Mol Cancer Ther 2009;8(8):2131–41]
Enhancement of vascular endothelial growth factor–mediated angiogenesis in tamoxifen-resistant breast cancer cells: role of Pin1 overexpression
Authors: Kim Mi Ra , Choi Hong Seok , Yang Jin Won , Park Byung Chul , Kim Jung-Ae , Kang Keon Wook
Published: August 2009

Acquired resistance to tamoxifen (TAM) is a serious therapeutic problem in breast cancer patients. Here, we found that TAM-resistant MCF-7 cells (TAMR-MCF-7 cells) produced higher levels of vascular endothelial growth factor (VEGF) than control MCF-7 cells. Molecular analyses using reporter genes and Western blots supported the involvement of c-Jun/activator protein-1 and hypoxia-inducible factor 1α in enhanced VEGF transcription in TAMR-MCF-7 cells. Pin1, a peptidyl prolyl isomerase, was consistently overexpressed in TAMR-MCF-7 cells, and c-Jun/activator protein-1–dependent VEGF transcription in TAMR-MCF-7 cells was almost completely inhibited by Pin1 siRNA and by the Pin1 inhibitor juglone. Chick chorioallantoic membrane assays confirmed that the increased angiogenic intensity of TAMR-MCF-7 cells was significantly suppressed by Pin1 inhibition. These results show that Pin1 overexpression is closely associated with VEGF-mediated angiogenesis and suggest that Pin1 is a potential therapeutic target of excessive angiogenesis in TAM-resistant breast cancer cases.[Mol Cancer Ther 2009;8(8):2163–71]
Oleandrin-mediated inhibition of human tumor cell proliferation: Importance of Na,K-ATPase α subunits as drug targets
Authors: Yang Peiying , Menter David G. , Cartwright Carrie , Chan Diana , Dixon Susan , Suraokar Milind , Mendoza Gabriela , Llansa Norma , Newman Robert A.
Published: August 2009

Cardiac glycosides such as oleandrin are known to inhibit the Na,K-ATPase pump, resulting in a consequent increase in calcium influx in heart muscle. Here, we investigated the effect of oleandrin on the growth of human and mouse cancer cells in relation to Na,K-ATPase subunits. Oleandrin treatment resulted in selective inhibition of human cancer cell growth but not rodent cell proliferation, which corresponded to the relative level of Na,K-ATPase α3 subunit protein expression. Human pancreatic cancer cell lines were found to differentially express varying levels of α3 protein, but rodent cancer cells lacked discernable expression of this Na,K-ATPase isoform. A correlation was observed between the ratio of α3 to α1 isoforms and the level of oleandrin uptake during inhibition of cell growth and initiation of cell death; the higher the α3 expression relative to α1 expression, the more sensitive the cell was to treatment with oleandrin. Inhibition of proliferation of Panc-1 cells by oleandrin was significantly reduced when the relative expression of α3 was decreased by knocking down the expression of α3 isoform with α3 siRNA or increasing expression of the α1 isoform through transient transfection of α1 cDNA to the cells. Our data suggest that the relative lack of α3 (relative to α1) in rodent and some human tumor cells may explain their unresponsiveness to cardiac glycosides. In conclusion, the relatively higher expression of α3 with the limited expression of α1 may help predict which human tumors are likely to be responsive to treatment with potent lipid-soluble cardiac glycosides such as oleandrin. [Mol Cancer Ther 2009;8(8):2319–28]
Three-kinase inhibitor combination recreates multipathway effects of a geldanamycin analogue on hepatocellular carcinoma cell death
Authors: Pritchard Justin R. , Cosgrove Benjamin D. , Hemann Michael T. , Griffith Linda G. , Wands Jack R. , Lauffenburger Douglas A.
Published: August 2009

Multitarget compounds that act on a diverse set of regulatory pathways are emerging as a therapeutic approach for a variety of cancers. Toward a more specified use of this approach, we hypothesize that the desired efficacy can be recreated in terms of a particular combination of relatively more specific (i.e., ostensibly single target) compounds. We test this hypothesis for the geldanamycin analogue 17-Allylamino-17-demethoxygeldanamycin (17AAG) in hepatocellular carcinoma cells, measuring critical phosphorylation levels that indicate the kinase pathway effects correlating with apoptotic responsiveness of the Hep3B cell line in contrast to the apoptotic resistance of the Huh7 cell line. A principal components analysis (PCA) constructed from time course measurements of seven phosphoprotein signaling levels identified modulation of the AKT, IκB kinase, and signal transducer and activator of transcription 3 pathways by 17AAG treatment as most important for distinguishing these cell-specific death responses. The analysis correctly suggested from 17AAG-induced effects on these phosphoprotein levels that the FOCUS cell line would show apoptotic responsiveness similarly to Hep3B. The PCA also guided the inhibition of three critical pathways and rendered Huh7 cells responsive to 17AAG. Strikingly, in all three hepatocellular carcinoma lines, the three-inhibitor combination alone exhibited similar or greater efficacy to 17AAG. We conclude that (a) the PCA captures and clusters the multipathway phosphoprotein time courses with respect to their 17AAG-induced apoptotic responsiveness and (b) we can recreate, in a more specified manner, the cellular responses of a prospective multitarget cancer therapeutic. [Mol Cancer Ther 2009;8(8):2183–92]
Double EGFR mutants containing rare EGFR mutant types show reduced in vitro response to gefitinib compared with common activating missense mutations
Authors: Tam Issan Yee-San , Leung Elaine Lai-Han , Tin Vicky Pui-Chi , Chua Daniel Tsin-Tien , Sihoe Alan Dart-Loon , Cheng Lik-Cheung , Chung Lap-Ping , Wong Maria Pik
Published: August 2009

Epidermal growth factor receptor (EGFR) mutations are common in lung adenocarcinomas, especially from nonsmoking women of Asian descent. We have previously shown EGFR mutations occur in >70% of lung adenocarcinoma from nonsmokers in our population with a complex mutational profile, including 13% of EGFR double mutations. In this study, we investigated the in vitro gefitinib response of four EGFR double mutants identified in untreated patients, including Q787R+L858R, E709A+G719C, T790M+L858R, and H870R+L858R. The phosphorylation profiles of EGFR and downstream effectors AKT, STAT3/5, and ERK1/2 were compared by immunoblot analyses among the single and double mutants transfected into H358 cells. Results showed that mutants responded to in vitro gefitinib treatment with different sensitivities. The G719C and L858R single mutants showed the highest gefitinib sensitivity compared with the corresponding coexisting single mutants E709A, Q787R, H870R, and T790M. The double mutants E709A+G719C, Q787R+L858R, and H870R+L858R showed attenuated responses to gefitinib in the EGFR and downstream effector phosphorylation profiles compared with G719C or L858R alone. T790M+L858R showed strong resistance to gefitinib. Clinically, the patient whose tumor contained H870R+L858R showed tumor stabilization by 250 mg oral gefitinib daily but cerebral metastasis developed 6 months later. Correlation with the in vitro phosphorylation profile of H870R+L858R suggested that treatment failure was probably due to inadequate suppression of EGFR signaling by the drug level attainable in the cerebrospinal fluid at the given oral dosage. Overall, the findings suggested that rare types of EGFR substitution mutations could confer relative gefitinib resistance when combined with the common activating mutants. [Mol Cancer Ther 2009;8(8):2142–51]
Stromal endothelin B receptor–deficiency inhibits breast cancer growth and metastasis
Authors: Binder Claudia , Hagemann Thorsten , Sperling Swetlana , Schulz Matthias , Pukrop Tobias , Grimshaw Matthew J. , Ehrenreich Hannelore
Published: August 2009

The endothelin (ET) axis, often deregulated in cancers, is a promising target for anticancer strategies. Whereas previous investigations have focused mostly on ET action in malignant cells, we chose a model allowing separate assessment of the effects of ETs and their receptors ETAR and ETBR in the tumor cells and the stromal compartment, which is increasingly recognized as a key player in cancer progression. In homozygous spotting lethal rats (sl/sl), a model of constitutive ETBR deficiency, we showed significant reduction of growth and metastasis of MAT B III rat mammary adenocarcinoma cells overexpressing ETAR and ET-1 but negative for ETBR. Lack of stromal ETBR expression did not influence angiogenesis. However, it was correlated with diminished infiltration by tumor-associated macrophages and with reduced production of tumor necrosis factor-α, both known as powerful promoters of tumor progression. These effects were almost completely abolished in transgenic sl/sl rats, wherein ETBR function is restored by expression of an intact ETBR transgene. This shows that tumor growth and metastasis are critically dependent on ETBR function in cells of the microenvironment and suggests that successful ETR antagonist therapy should also target the stromal component of ET signaling.[Mol Cancer Ther 2009;8(8):2452–60]
Meayamycin inhibits pre–messenger RNA splicing and exhibits picomolar activity against multidrug-resistant cells
Authors: Albert Brian J. , McPherson Peter A. , O'Brien Kristine , Czaicki Nancy L. , DeStefino Vincent , Osman Sami , Li Miaosheng , Day Billy W. , Grabowski Paula J. , Moore Melissa J. , Vogt Andreas , Koide Kazunori
Published: August 2009

FR901464 is a potent antitumor natural product that binds to the splicing factor 3b complex and inhibits pre-mRNA splicing. Its analogue, meayamycin, is two orders of magnitude more potent as an antiproliferative agent against human breast cancer MCF-7 cells. Here, we report the picomolar antiproliferative activity of meayamycin against various cancer cell lines and multidrug-resistant cells. Time-dependence studies implied that meayamycin may form a covalent bond with its target protein(s). Meayamycin inhibited pre-mRNA splicing in HEK-293 cells but not alternative splicing in a neuronal system. Meayamycin exhibited specificity toward human lung cancer cells compared with nontumorigenic human lung fibroblasts and retained picomolar growth-inhibitory activity against multidrug-resistant cells. These data suggest that meayamycin is a useful chemical probe to study pre-mRNA splicing in live cells and is a promising lead as an anticancer agent. [Mol Cancer Ther 2009;8(8):2308–18]