bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B215: Ascending single-dose study of safety, tolerability, and pharmacokinetics of bosutinib (SKI-606) in healthy subjects
Authors: Abbas Richat , Hug Bruce A. , Leister Cathie , Burns Jaime , Sonnichsen Daryl
Published: June 2012

Introduction: Bosutinib (SKI-606), a dual inhibitor of Src and Abl tyrosine kinases, is being developed for the treatment of patients with chronic myelogenous leukemia. An oral dose of 500 mg daily shows evidence of clinical efficacy in patients with Philadelphia chromosome-positive chronic myelogenous leukemia (Cortes et al. Blood 112:1098, 2008). To support the clinical pharmacology assessment of bosutinib, a single-ascending-dose safety and pharmacokinetic (PK) evaluation was performed in healthy subjects.

Methods: This was a randomized, double-blind, placebo-controlled, single-ascending dose, sequential-group study of oral bosutinib in healthy subjects. Subjects were originally to receive doses of 200, 400, 600, 800, 1000 and 1200 mg of bosutinib in the fasting state. However, subjects who received 400 mg bosutinib had multiple adverse events and then 200 mg, 400 mg, 600 mg, and 800 mg bosutinib was administered with food. Each cohort included 6 subjects who received one dose of bosutinib and 2 who received placebo. Plasma samples, collected through 96 h post-dose, were analyzed by using a liquid chromatography/tandem mass spectrometry assay for bosutinib concentrations. PK analyses were performed using a noncompartmental method and dose linearity was assessed using a power model.

Results: A total of 55 subjects (41 bosutinib, 14 placebo) were enrolled (47 males, 8 females, median age [range] 28 [18–50] y, 87%White). Thirty-three (81%) subjects had adverse events (AEs) after receiving bosutinib. The most common AEs for all subjects receiving bosutinib included diarrhea (39% subjects), nausea (29%), headache (22%), postural dizziness (22%) catheter site-related reaction and fatigue (10% each). The frequency and severity of gastrointestinal AEs appeared to be dose related. Doses up to 600 (fed) were well tolerated. After administration of single ascending doses of 200–800 mg bosutinib with food, absorption of bosutinib was relatively slow with a median tmax of 6 h, and both Cmax and AUC increased with increasing dose in a linear fashion. The apparent volume of distribution (Vz/F) was large (131–214 L/kg), mean clearance (CL/F) was 2.25–3.81 L/h/kg, and mean half-life was 32–39 h. Food increased bosutinib exposure (Cmax and AUC) by ∼2-fold and 1.5-fold after administration of the 200-mg and 400-mg doses, respectively.

Conclusions: Bosutinib was generally safe and well tolerated in healthy subjects after administration of single oral doses up to 600 mg (fed). The PK of bosutinib was linear when given with food, and the half-life supports a once-daily dosing regimen.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B215.
Abstract B216: Exposure-response (E-R) relationship of ABT-263-induced thrombocytopenia in cancer patients in phase 1 studies
Authors: Xiong Hao , Noertersheuser Peter A. , Pradhan Rajendra S. , Krivoshik Andrew P. , Humerickhouse Rod , Awni Walid
Published: June 2012

Background: ABT-263 is a novel small molecule Bcl-2 family protein inhibitor that binds with high affinity to multiple antiapoptotic Bcl-2 family proteins. The pharmacokinetics, safety and preliminary efficacy of ABT-263 in patients with relapsed or refractory lymphoid malignancies, CLL/SLL, SCLC and other non-hematological malignancies are being studied in three Phase 1/2a studies. Both a 14/21-day dosing schedule and a continuous dosing preceded with a 7-day lead-in period are being evaluated in these studies. Grade 4 thrombocytopenia has been the most prominent, mechanism-related, dose-limiting toxicity in patients in these studies. E-R relationship in this drug-induced thrombocytopenia may inform the selection of the dose and dosing schedule for a Phase 3 study in CLL patients.

Methods: ABT-263 plasma concentrations and platelet data from more than 100 patients in the three ongoing Phase 1/2a studies were included in the analysis. ABT-263 plasma concentration data were described using a two-compartment model with first-order absorption and elimination. The relationship between the circulating platelet count and ABT-263 plasma concentration was described with a semi-physiological model that had been developed using data in dogs. Simulations were preformed to compare the 14/21-day dosing schedule and the continuous dosing schedule with a low-dose lead-in period and project the incidence rates of Grades 3 and 4 thrombocytopenia in the dose range of 200 and 350 mg.

Results: The E-R relationship was generally well described by the semi-physiology model. Simulations indicated that, compared to the 14/21-day dosing schedule, continuous dosing preceded with a 7-day lead-in period minimizes platelet variability across cycles. The lead-in treatment appeared to blunt the initial sharp drop in platelet count according to the simulations with 250 mg dose. Although there seemed to be little difference in the incidence rate of Grade 4 thrombocytopenia, continuous dosing showed a trend of lower rate of Grade 3 thrombocytopenia than the 14/21-day dosing schedule. In terms of the rates of Grade 3 and 4 thrombocytopenia, the dose-response curve between 200 and 350 mg was quite shallow.

Conclusions: A continuous dosing preceded with a 7-day lead-in period is superior to the 14/21-day dosing schedule because it minimizes platelet variability across cycles and blunts the initial sharp drop in platelet count. Drug-induced thrombocytopenia is not sensitive to the change in dose does not provide significant differentiation in the dose range of 200 mg and to 350 mg.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B216.
Abstract B218: Lentiviral shRNA screen of human kinases identifies PLK1 as a potential therapeutic target for osteosarcoma
Authors: Duan Zhenfeng , Ji Diana , Weinstein Edward , Choy Edwin , Yang Cao , Mankin Henry , Hornicek Francis
Published: June 2012

Induction of RNA interference in cancer cells has made possible high-throughput genome-wide loss-of-function studies using reverse genetic screening techniques. Kinases play an important role in the growth and survival of tumor cells. We aim to identify kinases that are vital to the survival of osteosarcoma cells and may be a key target in creating novel anticancer therapies. We describe an optimized systematic screen of known kinases using osteosarcoma cell lines (KHOS and U-2OS) and a lentiviral-based short hairpin RNA (shRNA) human kinase library. CellTiter 96® AQueous One Solution Cell Proliferation Assay was used to measure cell growth and survival. We identified several kinases, including human polo-like kinase (PLK1), which inhibit cell growth and induce apoptosis in osteosarcoma cells when knocked down. cDNA rescue and synthetic siRNA assays confirm that the observed phenotypic changes result from the loss of PLK1 gene expression. Furthermore, a small molecule inhibitor to PLK1 inhibited osteosarcoma cell growth and induced apoptosis. Western blot analysis confirmed that PLK1 is highly expressed and activated in several osteosarcoma cell lines as well as in resected tumor samples.

Immunohistochemistry analysis showed that patients with high PLK1 tumor expression levels correlated with significantly shorter survival than patients with lower levels of tumor PLK1 expression. These results demonstrate the capability and feasibility of a high-throughput screen with a large collection of lentiviral kinases and its effectiveness in identifying potential drug targets. The development of more potent inhibitors that target PLK1 may open doors to a new range of anti-cancer strategies in osteosarcoma.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B218.
Abstract B22: Dynamics of cancer cell survival dependency on Bcl-2 family members
Authors: Mc Henry Kevin T. , Donovan Jerry , Loo Alice , Fisher David E. , Zawel Leigh , Porter Dale
Published: June 2012

Apoptosis is regulated through the interactions of the Bcl-2 family of proteins. Overexpression of anti-apoptotic Bcl-2 family members plays a key role in promoting survival for cells undergoing various insults that would normally trigger cell death. Consequently, these proteins are believed to serve as contributing factors to the processes of tumor initiation, tumor progression and the development of resistance to therapeutics. ABT-263, an antagonist of Bcl-2, Bcl-xL, and Bcl-w, is currently being tested in clinical trials. Some cancer cells express high amounts of Mcl-1, another Bcl-2 family anti-apoptotic protein that shares pro-survival function with Bcl-2 but is not targeted by ABT-263. Preclinical data indicates Mcl-1 overexpressing cells are not sensitive to ABT-263. To understand this mechanism of Mcl-1 dependence, A2058 melanoma cells were infected with a lentivirus encoding inducible shRNA against Mcl-1. Upon induction of MCL1 knockdown, massive cell death occurred. Interestingly, a small subset of the cell population managed to survive. Subsequent western blot analysis confirmed a sustained depletion of Mcl-1. Further study indicated A2058 cells deficient in Mcl-1 survived by upregulating expression of Bcl-2, a process we term “switching dependency”. In addition, these cells became sensitized to ABT-263 treatment. An A2058 xenograft model indicates that combined knockdown of both Mcl-1 and Bcl-2 significantly inhibited tumor growth when compared to tumors treated with ABT-263 lacking either protein individually. Real-time kinetic studies reveal a rapid sensitization of A2058 cells to ABT-263 upon sh-RNA induction. To understand whether de novo protein synthesis of Bcl-2 plays a mechanistic role in dependency switching, A2058 sh-Mcl-1 cells were treated with the protein synthesis inhibitor cycloheximide. Kinetic analysis indicates that de novo protein synthesis is not required for dependency switching. Real-time quantitative PCR was then employed to determine if a relationship exists between the decrease in Mcl-1 mRNA, and subsequent sensitivity to ABT-263, is a consequence of a change in Bcl-2 mRNA levels. Surprisingly, the results indicate as the loss of Mcl-1 message occurs, there is no concurrent change in Bcl-2 mRNA. Taken together these data indicate that dependency switching, and therefore survival, occurs without de novo Bcl-2 protein synthesis or alterations in protein turnover. Furthermore, dependency switching is a possible mechanism cancer cells employ to develop resistance toward various anti-cancer therapies with novel implications for both the concept of oncogene addiction and development of targeted cancer therapeutics. Therefore there is an immediate need for elucidation of this novel cancer resistance mechanism.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B22.
Abstract B220: Identification and characterization of kinase targets to treat melanoma
Authors: Madhunapantula SubbaRao V. , Sharma Arati , Chandagalu Raghavendra Gowda D. , Robertson Gavin P.
Published: June 2012

Incidence and mortality rates for malignant melanoma continue to rise every year. Lack of functional knowledge regarding key proteins regulating melanoma development is one of the major drawbacks hindering development of targeted therapeutics. Additionally, it has been found that monotherapies targeting proteins regulating melanoma development are not as effective as combination therapies warranting research to identify new therapeutic targets that can be inhibited in combination to better treat melanoma. Therefore, we conducted a siRNA-based screen to functionally identify key kinases regulating melanoma development using the Amaxa 96-well shuttle system and a Stealth siRNA library consisting of siRNAs targeting 636 human kinases. Involvement of the identified kinases is validated in vitro using multiple metastatic melanoma cell lines and in vivo in xenografted tumor kinetics studies. The screen identified a number of known and novel kinase targets. Results of primary and secondary screens, validation of the identified targets in vitro and in vivo are presented. Since the data demonstrated in this screen encompasses all the known kinases, it helps to functionally identify key kinases involved in melanoma as well as, design and select appropriate chemotherapeutic agents or agent combinations to effectively inhibit melanoma tumor development. In conclusion the data obtained in this siRNA screen contributes significantly to the currently available knowledge by demonstrating key kinases to inhibit melanoma tumor development either singly or in combination.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B220.
Abstract B224: Multi-tyrosine kinase targeting resulting in concomitant downregulation of proliferation/survival pathways dependent on FGF-R3, Jak2 and BCMA in human multiple myeloma cells
Authors: Cassinelli Giuliana , Ronchetti Domenica , Laccabue Diletta , Mattioli Michela , Cuccuru Giuditta , Favini Enrica , Nicolini Valentina , Greco Angela , Neri Antonino , Zunino Franco , Lanzi Cinzia
Published: June 2012

Proliferation/survival pathways constitutively activated by genetic alterations in multiple myeloma (MM), or sustained by the bone marrow (BM) microenvironment, provide opportunities for the development of novel targeted therapies. Deregulated protein tyrosine kinases (TKs) involved in these pathways may play a critical role in driving the MM malignant phenotype thus representing attractive therapeutic targets. We have investigated the effects of a multi-target tyrosine kinase inhibitor (1,3-dihydro-5,6-dimethoxy-3-[(4-hydrophenyl)methylene]1-indol-2-one) (named RPI-1 in our laboratory) in a panel of human MM cell lines, including t(4;14) positive cell lines expressing the TK receptor FGF-R3. Cells harboring FGF-R3 activating mutations (KMS11 and OPM2) displayed the highest sensitivity to the drug antiproliferative effect. However, proliferation induced by the FGF-R3 ligand, aFGF, was abrogated even in cells harboring a non-constitutively active receptor. Drug treatment inhibited activation and expression of the FGF-R3Y373C mutant as well as aFGF-dependent signaling involving AKT and ERKs. Jak2 has been identified as an additional target of RPI-1 in MM cells. Inhibition of Jak2 TK resulted in STAT3 inactivation. Blockade of these proliferation/survival pathways was associated with activation of caspase-dependent apoptosis. Moreover, the TK inhibitor abrogated proliferative and pro-invasive stimuli provided by conditioned medium from mesenchymal stromal cells. Gene expression analysis of KMS11 cells showed 22 upregulated and 52 downregulated genes upon RPI-1 treatment, with an early modulation of genes implicated in MM pathobiology such as SAT-1, MYC, MIP-1α/β, FGF-R3, and the growth factor receptor B-cell maturation antigen (BCMA). Thus, concomitant blockade of FGF-R3 and Jak2 results in inhibition of several MM-promoting pathways, including BCMA-regulated signaling, and downregulation of disease-associated proteins. Overall, these data support the therapeutic interest for FGF-R3/JAK2 multi-target approaches in the treatment of t(4;14) MM.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B224.
Abstract B231: Targeting Her2+ ductal breast cancer with PDL192, a novel humanized anti-TweakR monoclonal antibody
Authors: Chao Debra T. , Su Mien , Sho Mien , Choi Doghee , Ye Shiming , Tomlinco Sonia , Kim Han , Zhang Yongke , Yin Johnny , Hsi Eric D. , Durkan Lisa , Lambert Peter , Starling Gary C. , Afar Daniel E.H. , Culp Patricia A.
Published: June 2012

TWEAK receptor (TweakR), also known as Fn14, is a cell surface protein and member of the tumor necrosis factor receptor superfamily. TNFRSF family members are appealing therapeutic targets due to their expression and function in tumor cells. The goal of the current study was to examine the potential of TweakR as a therapeutic target in various subtypes of breast cancer by 1) determining expression of TweakR using immunohistochemistry (IHC) and 2) treating cells derived from breast cancer in vitro and in vivo with PDL192, a novel, humanized anti-TweakR mAb. Overexpression of TweakR protein in breast cancer was confirmed by IHC on formalin fixed paraffin embedded (FFPE) tissues. TweakR was elevated in infiltrating tumors when compared with normal adjacent tissues. 30 to 40% of various subtypes of invasive ductal breast cancer samples stained highly for TweakR; however, fewer than 5% of lobular breast cancer samples had high levels of TweakR. In addition, a strong positive correlation of TweakR and Her2 expression was observed, irrespective of ER status. More than 60% of Her2+ invasive ductal cancer exhibited elevated levels of TweakR. Strong TweakR expression was also observed in bone metastasis samples from primary breast cancer but rarely in benign tumors, such as DCIS.

PDL192 inhibited the in vitro growth of various subtypes of TweakR-expressing breast cancer cell lines and this activity was augmented by crosslinking the mAb. In addition, PDL192 significantly inhibited the in vivo growth of a variant of the breast cancer xenograft model MDA-MB231 that exhibits metastatic potential. PDL192 administration exhibited a dose-dependent inhibition of primary tumor growth and lung metastasis and enhanced the anti-tumor activity of several chemotherapy agents currently used to treat breast cancer. These data provide the rationale to evaluate PDL192 as a potential therapy for the treatment of breast cancer. Currently, PDL192 is being tested in a phase I safety study for the treatment of patients with solid tumors.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B231.
Abstract B239: Profiling the type I insulin-like growth factor receptor (IGF1R) pathway regulated metastasis signature
Authors: Sarver Aaron , Sachdev Deepali
Published: June 2012

The insulin-like growth factors (IGFs) acting via the type I IGF receptor (IGF1R) regulate growth and metastasis of multiple types of cancers. Agents targeting IGF1R are currently being tested in phase II/III clinical trials. We have previously shown that IGF1R can regulate metastasis independently of primary tumor growth in a model of high risk metastatic cancer using the MDA435/LCC6 metastatic cancer cells. Here we sought to develop an IGF1R driven metastasis signature that may be useful in selecting patients that could benefit from IGF1R targeted therapy and determine if changes in expression of the genes in the signature can be useful in predicting response. Serum starved LCC6-WT cells (with wild-type functional IGF1R) and LCC6-DN (cells with dominant negative IGF1R) were treated with IGF-I, AVE1642 (antibody against IGF1R that inhibits metastasis but not tumor growth in the mammary fat pad), or AVE1642+ IGF-I for 4 and 24 hours and microarray analysis was performed using the U133 Plus 2.0 human arrays with 47,000 transcripts. Array data were analyzed to determine genes regulated by IGF1R signaling in these cells and if disruption of signaling with AVE1642 affected genes whose expression was regulated by IGF-I signaling. A total of 91 transcripts were regulated by IGF signaling in LCC6-WT cells and AVE1642 treatment reversed the effects of IGF1R signaling. We are currently querying human tumor databases to determine if genes regulated by activation of IGF1R are associated with lymph node metastasis and poor prognosis in human tumors. Our results suggest that identifying IGF1R driven metastasis signatures may be useful to identify and monitor patients who could benefit from IGF1R targeted therapy for inhibition of metastatic disease and that patients with this IG

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B239.
Abstract B243: Kras mutation status as biomarker for response to TRAIL treatment in colorectal cancer
Authors: Van Schaeybroeck Sandra , Kyula Joan , Longley Dan , Johnston Patrick
Published: June 2012

Background: Kras is a member of the ras family of GTPase proteins that regulate cellular growth and survival in response to stimulation of cell surface receptors such as the Epidermal Growth Factor Receptor (EGFR). Activating mutations in the Kras gene are found in 30–50% of colorectal tumors (CRC). The presence of mutant Kras in tumors has been shown to correlate with lack of response to EGFR inhibitors. Recently, we found that CRC cells respond to TRAIL with an EGFR/HER2-mediated survival response and that by blocking this response, apoptosis is enhanced. A phase I/IIa study is currently investigating effect of AMG655 (DR5 agonist) with panitumumab (EGFR monoclonal antibody) in patients with chemorefractory CRC. The aim of this study was to evaluate the role of Kras mutational status in determining response to TRAIL alone and in combination with EGFR- or MEK-targeted therapies in a panel isogenic paired Kras wild type (WT) and mutant (MT) CRC cells.

Methods: Erk1/2 phosphorylation and expression were measured by Western blotting. Apoptosis was measured by flow cytometry andWestern blotting for PARP, caspase-8 and caspase 3 cleavage.

Results: Using paired isogenic KrasMT HCT116 and KrasWT HKH-2 and HKe-3 cell lines, we found higher basal levels of pErk1/2 in the KrasMT HCT116 cell line compared to the KrasWT HKH-2 and HKe-3 cell lines. Furthermore, we found that the HKH-2 and HKe-3 KrasWT cells were more sensitive to rhTRAIL compared to the isogenic parental and KrasMT HCT116 cell line. Moreover, we found a statistical significant increase in apoptosis when rhTRAIL was combined with the EGFR monoclonal antibodies panitumumab or cetuximab in KrasWT HKH-2 and HKe-3 cell lines but not in the parental KrasMT HCT116 cell line. In contrast, KrasMT HCT116 cell line was more sensitive to combined TRAIL/MEK-targeted therapy compared to KrasWT HKH-2 clone.

Conclusions: Our findings indicate that Kras mutations can predict response to TRAIL alone and in combination with EGFR-targeted agents in CRC cells. Inhibition of MEK1/2 in combination with TRAIL-targeted therapies may provide a strong treatment strategy in Kras mutant CRC tumors. We are currently investigating mechanism of KrasMT induced resistance to TRAIL treatment and these data will be presented during the meeting.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B243.
Abstract B253: The IGF1R inhibitor NVP-AEW541 disrupts a pro-survival and pro-angiogenic IGF-Stat3-HIF1 pathway in human glioblastoma cells
Authors: Monti Elena , Ravizza Raffaella , Molteni Roberta , Gariboldi Marzia B.
Published: June 2012

Glioblastomas (GBMs) typically develop hypoxic areas within the tumor mass, which for some cells results in necrosis, but for others leads to pleiotropic adaptive changes promoting tumor progression through activation of survival pathways, disruption of apoptosis, increase in invasive potential and neovascularization. The transcription factor HIF1 (hypoxia-inducible factor 1) is known to play a major role in directing this adaptive program, following stabilization of its inducible -subunit under hypoxic conditions; however, HIF1 activity can also be enhanced by oxygen-independent mechanisms upon activation of growth factor receptors and their downstream signal transduction pathways. The present study investigates a) the possible presence of a regulatory loop linking IGF1R and HIF1 in human glioblastoma cells; b) the role played by the signal transducer and activator of transcription 3 (Stat3) in this pathway; and c) the ability of the IGF1R inhibitor NVP-AEW541 (Novartis Pharma, Basel, CH) to disrupt this pathway. We show that activation of IGF1R by its ligand IGF1 causes Stat3 activation, increased levels of HIF1α and increased release of VEGF, the product of one of the major target genes for HIF1 transcriptional activity. Treatment with NVP-AEW541 reverses all these changes, and so does transfection of the cells with a dominant negative form of Stat3. Interestingly, we also observe that IGF-independent activation of HIF1, obtained in the presence of the hypoxia mimic CoCl2, results in IGF1R activation, thereby indicating that a two-way relationship exists between growth factor receptor activation and increase in HIF1 transcriptional activity; NVP-AEW541 treatment significantly reduces both HIF1α protein levels and IGF1R tyrosine-specific phosphorylation in CoCl2-treated cells. Because cells surviving under hypoxic condition often become resistant to radiotherapy and chemotherapy, modulation of the IGF1R-HIF1 pathway could be a viable approach to increase the effectiveness of conventional cytotoxic agents; as GBM typically presents hypoxic areas, this strategy might prove particularly valuable in improving the poor prognosis of patients affected by this tumor type.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B253.
Abstract B254: Extended target-coverage by selective Chk1 inhibitors enhances pharmacodynamic inhibition of Chk1 signaling and antitumor activity in vivo
Authors: Humphries Michael J. , von Carlowitz Ira , Le Huerou Yvan , Randolph Nikole , Mohr Peter J. , Wang Bin , Lyon Michael A. , Gunawardana Indrani , Chicarelli Mark , Blake Jim F. , Schrag Michael L. , Winski Shannon L. , Wallace Eli
Published: June 2012

Loss of coordination between cell cycle checkpoints and DNA damage repair is a fundamental feature tumor cells rely on for unregulated growth and developing chemotherapeutic resistance. The protein kinase Checkpoint kinase 1 (Chk1) is a sentinel molecule essential for cell cycle arrest at the S and G2M checkpoints, as well as regulating homologous recombination DNA repair. In tumor cells exposed to chemotherapy, Chk1 inhibition overrides cell cycle arrest and DNA repair functions, effectively driving tumor cells into a state of mitotic catastrophe and, ultimately, cell death. We have previously reported in schedule-dependence studies, using Chk1 inhibitors and irinotecan (CPT-11), that oral administration of Chk1 inhibitors allows for multi-day target-coverage, and thus continuous inhibition of Chk1 for a finite period of time, which maximizes anti-tumor efficacy. Here, we extend these studies and investigate the pharmacodynamic relationship to efficacy, as well as the specific biomarkers that are predictive of an anti-tumor effect, when Chk1 inhibitors are administered on a multi-day dosing schedule. Utilizing potent (IC50=24–27nM), selective, and orally bio-available small molecule Chk1 inhibitors of which Chk1-A and Chk1-C are representative, we find only modest inhibition of the functional biomarker phospho-cdc2, following a single dose of a Chk1 inhibitor. Alternatively, on multi-day Chk1 inhibitor dose schedules, we find dose-related pharmacodynamic inhibition of Chk1 signaling that is maximized at doses where we see significant tumor growth inhibition in efficacy experiments. Furthermore, multi-day dosing of Chk1 inhibitors induces marked inhibition of phospho-cdc2 and Rad51 protein levels, suggesting tumoricidal activity related to Chk1 inhibition is due to both checkpoint override and impairment of DNA damage repair. In human tumor xenografts administered combination therapy with gemcitabine, an orally-delivered Chk1 inhibitor dosed on a multi-day schedule shows superior efficacy over an IV administered compound. Taken together, our findings show a clear correlative relationship between pharmacodynamic target inhibition and anti-tumor activity that is exclusively achieved on multi-day dose schedules. These results demonstrate the need for prolonged Chk1 inhibition to provide robust pharmacodynamic inhibition and maximal anti-tumor efficacy.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B254.
Abstract B258: Investigating PERK biological pathway using protein/peptide microarrays and SAR with small molecule inhibitors
Authors: Otlik Hubert , Timofeevski Sergei , Quenzer Terri , Briere David , Fanjul Andrea , Alton Gordon , Ermolieff Jacques
Published: June 2012

Accumulation of unfolded proteins in the endoplasmic occurs when the cell is subject to stress caused by various pathological conditions such as hypoxia, viral infection, and glucose depravation. Under such stress the cell will initiate an unfolded protein response (UPR), a protective mechanism that is specifically designed to re-establish homeostasis and normal endoplasmic reticulum (ER) function. This adaptive mechanism inhibits overall protein translation, but enhances the translation of a small number of key stress response proteins that will clear the ER of unfolded proteins and send them to the cytoplasm for degradation. The UPR is initiated by several proteins such as IRE1a, ATF6a and PERK, the later being a key Ser/Thr protein kinase in UPR signaling. In response to inducers of ER stress, BiP (GRP78) dissociates from the luminal ER domain of PERK, resulting in the oligomerization, autophosphorylation, and activation of PERK which in turn phosphorylates eIF-2a on Ser51 and Nrf2 (unknown site of phosphorylation). In order to further identify potential substrates for PERK, this enzyme was tested against large protein and peptide microarrays. A significant number of proteins and peptides in the microarrays were found to be phosphorylated by PERK. After additional selection, several of these substrates were further characterized using a microfluidic mobility-shift assay and submitted to LC/MS analysis to identify the site of phosphorylation by PERK. These newly identified peptide substrates were used to develop a robust biochemical assay for the testing of small molecule inhibitors of PERK activity. A limited SAR was established for a set of compounds against PERK and two other related protein kinases, GCN2 and PKR. One of the most potent and selective PERK inhibitors was found to modulate PERK cellular signaling as evidenced by blocking the phosphorylation of (Ser51)eIF-2a as judged by Elisa and Western blot analysis.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B258.
Abstract B263: Mechanistic, functional, and in vivo efficacy of inhibiting ETK/BMX in cancer models
Authors: Gourley Eric S. , Liu Xiao-Hui , Hewitt William , Walker Rusty , Warner Steven L. , Vankayalapati Hariprasad , Bearss David J.
Published: June 2012

Epithelial and endothelial tyrosine kinase (ETK), also known as bone marrow tyrosine kinase gene in chromosome X (BMX), belongs to the Burton's Tyrosine Kinase (BTK) family. ETK is a nonreceptor tyrosine kinases involved in tumorigenicity, adhesion, motility, angiogenesis, proliferation, and differentiation. It is overexpressed in metastatic breast and prostate cancers and is implicated in the neuroendocrine transformation of prostate cells. Significant oncogenes such as Src, focal adhesion kinase (FAK), and phosphatidyl-inositol (PI)-3 kinase are upstream regulators of ETK while several hub proteins, also critical for cancer development and progression, such as AKT, STAT3, and p21 activated kinase 1 (PAK1), are downstream of ETK. Cell-based validation studies also demonstrate that inhibition of ETK diminishes cellular transformation, proliferation, and migration. Such evidence provides substantial reasoning for developing new small molecule inhibitors against ETK. Applying SuperGen's proprietary small molecule drug discovery process, CLIMB™ a series of lead compounds were effectively developed to potently and selectively inhibit ETK. Biochemical data from kinase screens against ETK demonstrated nanomolar potency of SuperGen's inhibitors and good selectivity toward ETK from a panel of 75 kinases. To mechanistically demonstrate ETK inhibition in cell-based models, immunoprecipitations were carried out and phospho-ETK levels were detected. These compounds exhibited a dose-dependent response against phospho-ETK. Functionally, the inhibition of ETK will produce changes in phospho-AKT (S473) levels as well as phospho-STAT3 (Y705) levels downstream of EGF stimulation. Using Luminex bead-based technology and western blot techniques respectively, SuperGen's lead ETK compounds also yielded concentration-dependent responses with EC50 values in the nanomolar region. Given ETK's reported angiogenic role in endothelial cell migration and proliferation, these compounds were evaluated in an in vitro tube formation assay. Likewise, these ETK inhibitors demonstrated potent nanomolar inhibition of endothelial tube formation. Finally, having shown mechanistically and functionally the efficacy of ETK inhibitors through in vitro assays, the compounds were tested with in vivo solid tumor efficacy models in nude mice. All five lead inhibitors effectively decreased tumor burden volume relative to the vehicle treatment group in an AN3CA xenograft model. Taken together, these data suggest SuperGen's small molecule ETK inhibitors represent a potentially new therapeutic avenue for treating patients with solid tumor malignancies in the future.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B263.
Abstract B265: A Phase 1 safety and pharmacokinetic study of XL765 (SAR245409), a novel PI3K/TORC1/TORC2 inhibitor, in combination with temozolomide (TMZ) in patients (pts) with malignant glioma
Authors: Wen Patrick Y. , Omuro Antonio M. , Batchelor Tracy T. , Lai Albert , Mellinghoff Ingo K. , Nghiemphu Leia , Norden Andrew , Gendreau Steven B. , Laird A. Douglas , Nguyen Linh , Cloughesy Timothy
Published: June 2012

Methods: Pts with glioblastoma or anaplastic/mixed gliomas receive TMZ 200 mg/m2/day on Days 1–5, and XL765 qd in 28-day cycles. Pts must be on a maintenance TMZ dose of 200 mg/m2/day for at least one 28-day cycle prior to enrollment. Cycle 1 safety data determine dose-limiting toxicities (DLTs). Tumor response is assessed by the modified Macdonald criteria every 8 weeks.

Results: As of 24Aug2009, 15 pts (12 with glioblastoma, 2 with mixed glioma and 1 diagnosed with anaplastic oligodendroglioma) have been treated with XL765 in combination with TMZ across 3 dose levels of XL765 (30, 40, and 60 mg qd). The MAD has been identified as 60 mg for the once-daily schedule. Two of 6 pts at the 60 mg XL765 dose level experienced DLTs: one pt experienced DLTs of Grade 3 brain edema and Grade 4 thrombocytopenia (these events were also reported as related SAEs) and the other pt experienced a DLT of Grade 3 diffuse maculopapular rash. Three additional pts experienced SAEs that were not attributed to study treatment. With the exception of the DLTs described above, all AEs reported for 12 pts (as of 03Aug2009) were Grade 1 or Grade 2. Preliminary data suggest no PK interaction between XL765 and TMZ. XL765 PK when given in combination with TMZ are consistent with the PK of single-agent XL765 (Exelixis study XL765-001), and TMZ exposure in this study is similar to that published for single-agent TMZ. At the 30 and 40 mg qd XL765 dose levels, inhibition of PI3K pathway signaling has been demonstrated in paired skin biopsies using immunofluorescent analysis of pAKT-T308, pAKT-S473, and p4EBP1. Based on an initial molecular assessment of archival tumor material the most common molecular alterations detected included MGMT promoter methylation (5 of 12 evaluated pts), PTEN point mutations (8 of 14 evaluated pts), and EGFR gene amplification (7 of 11 evaluated pts). Of the first 12 pts that were enrolled, six pts have remained on study for at least 16 weeks, one of whom continues on study for 50+ weeks (a total of 78+ weeks following chemoradiation). Archival tumor tissue from this pt exhibited MGMT promoter methylation, PTEN mutation, and EGFR amplification.

Conclusions: XL765 in combination with TMZ is generally well tolerated at doses up to 40 mg qd. No apparent PK interaction between XL765 and TMZ has been observed.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B265.
Abstract B268: Tyrosine kinase profiling in head and neck cancer tumors in combination with clinical prognosis factors guides therapeutic combination strategies
Authors: Rodriguez-Barrueco Ruth , Ortiz-Ruiz Maria , Lopez-Castro Rafael , Cruz Juan Jesus , Ocana Alberto , Pandiella Atanasio
Published: June 2012

Background: Tyrosine kinases (TK) are a particular subgroup of proteins implicated in the genesis/progression of Squamous cell carcinoma of the head and neck (SCCHN). Therapeutic inhibition of several of them has shown to be clinically useful, as is the case of EGFR inhibition with the antibody cetuximab. Identification of the TKs activated in SCCHN is a must in order to adequately target these kinases with available inhibitors.

Methods: Here we have investigated activated tyrosine kinases in head and neck cancer tumors derived from patients using a human phospho protein array for 42 receptor tyrosine kinases (RTK). Clinicopathological parameters were reviewed from patient charts to associate activated kinases with prognosis. In vivo xenografted models were used to study the antiproliferative effect of the combination of specific TK inhibitors against them.

Results: TK receptors of the EGF and the VEGF family were the mostly activated in tumors derived from patients. EGFR/HER family receptors, such as HER3, were also activated (phosphorylated). These data were corroborated in SCCHN cell lines. In these cells, Akt and FAK kinases, other RTK signalling intermediates, were also activated. Combination of the anti-EGFR-HER2 TK inhibitor lapatinib with dasatinib (that targets FAK) was synergistic in vitro. Combination of lapatinib with the anti-VEGFR TK inhibitor pazopanib resulted in a better trend in response in the in vivo xenografted models.

Conclusions: Rational target drug combinations should be based on the identification of activated TK receptors or downstream signalling molecules in association with prognosis. Combination strategies using anti-EGFR/HER, anti-FAK, and anti-VEGFR compounds increases the action of individual treatments and should be considered when design rational clinical trials.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B268.
Abstract B28: Selective antagonism of IAP proteins for regulating apoptosis and TNF pathways
Authors: Ndubaku Chudi
Published: June 2012

The inhibitor of apoptosis (IAP) proteins are important regulators of cancer cell survival, making them important targets for oncological intervention. There are eight known members of the IAP family of proteins. Using a combination of structure-based design and a merger of existing inhibitor motifs, we generated selective inhibitors that will interrogate the biological relevance of antagonism of XIAP and c-IAP1 and 2. The cellular biology of selective antagonism of these proteins will be presented.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B28.
Abstract B33: Prognostic role of Wnt/beta-catenin pathway in colorectal carcinoma
Authors: Stanczak Aleksandra , Stec Rafal , Bodnar Lubomir , Olszewski Wojciech , Ciechowicz Marzena , Kozlowski Wojciech , Szczylik Cezary , Lamparska-Przybysz Monika , Wieczorek Maciej
Published: June 2012

Introduction: Beta-catenin is a member of a cadherin-catenin complex mediating cell adhesion. It is also a key protein of Wnt/Beta-catenin pathway responsible for cell cycle and proliferation. This is important for initiation and progression of colorectal cancer. The aim of this study was to determine expression and localization of E-cadherin, Beta-catenin and Wnt-1 proteins in colorectal tumors.

Description: Forty-four unrelated patients with advanced colorectal carcinoma (CRC) were enrolled consecutively from CRC patients attending palliative care. Expression of Beta-catenin, E-cadherin and Wnt-1 was determined by immunohistochemistry on formalin-fixed, paraffin-embedded primary colorectal tissue. The expression was scored by two independent observers who had no knowledge of the clinical data. Correlation was measured with Spearman test, overall survival were analyzed with Kaplan-Meier method. Log-rank test was used for univariate analysis, while Cox regression model for multivariate analysis. A value of p < 0,05 was considered significant.

Results: Decreased expression of membrane E-cadherin and Beta-catenin was observed in 34% and 27% of patients, respectively. Abnormal cytoplasmic E-cadherin was detected in 32% of cases and abnormal cytoplasmic and nuclear Beta-catenin was detected in 50% and 25% of patients, respectively. 60% of patients had decreased cytoplasmic Wnt-1 expression. Correlation was seen between cytoplasmic E-cadherin and cytoplasmic Beta-catenin (R 0,35; p=0,019). Nuclear localization of Betacatenin was correlated with localization of primary tumor (the highest expression in rectum) (R 0,48; p=0,001) and tumor size (R 0,32; p=0,035). Lower Wnt-1 expression was correlated with higher nodal stage (R -0,31; p=0,047). Nuclear Beta-catenin was an adverse prognostic factor in univariate analysis (log-rank test p=0,034) which was confirmed in multivariate analysis (HR 4,27 95% CI 1,70–10,72; p=0,002). Higher CEA serum level before treatment (HR 4,30 95% CI 1,35–13,66; p=0,013) and female sex (HR 2,75 95% CI 1,24–6,10; p=0,013) were also independent prognostic factors.

Conclusions: This finding supports that Beta-catenin is an independent prognostic factor for colorectal carcinoma.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B33.
Abstract B44: Survivin protein detection is a promising biomarker in the serum of gastroenteropancreatic neuroendocrine tumor (GEP-NET) patients
Authors: Georgieva Inna , Hein Sibyll , Wang Yawen , Kiewe Philipp , Hopfenmüller Werner , Bläker Michael , Zeitz Martin , Grabowski Patricia
Published: June 2012

Background: The bifunctional protein survivin is a powerful prognostic marker in GEP-NET disease as measured by immunohistochemical methods in tumor tissues. The present study was focused to investigate if survivin can also be detected in the serum of GEP-NET patients and to evaluate its potential diagnostic role.

Aim: To establish a method by which the serum levels of survivin can be used as a predictive value in the treatment outcome of patients with GEP-NET disease.

Materials and Methods: In 2007, serum samples were obtained from patients with well-differentiated GEP-NET disease (n=22) either at Charité-Campus Benjamin Franklin (n=8) or at the University Hospital of Hamburg-Eppendorf (n=14), hematological malignancies (HM) and solid cancers (n=22), as well as healthy volunteers (n=25), the last all obtained at the Charité. All patients had given their informed consent to this pilot study. The patients of the GEP-NET group had either stable disease with metastasis, predominantly to the liver (n=14), or were progressive at the time of venipuncture (n=6). 9 foregut tumors, 10 midgut tumors and 3 hindgut tumors were included, a third of those functionally active. Most of the patients received any kind of bio- or chemotherapy. The mean age of GEP-NET patients was 61.1 years (range 29–81), whereas the mean age of patients with HM and solid tumors was 55.9 years (range 30–82). Enzyme-linked immunosorbent assay (ELISA) analysis was used for determination of survivin protein serum levels.

Results: There was a statistically significant increase of the survivin protein serum levels in patients with HM and solid cancers compared to the healthy controls (p=0.009). The difference in survivin serum levels between healthy controls and patients with GEP-NETs did not reach statistical significance (p=0.089). No correlation to clinical features like localization of the tumor, functional activity, Chromogranin A levels or kind of therapy was found. However, survivin serum levels tended to be higher in patients with a greater tumor burden.

Conclusion: This is the first study to quantify survivin protein levels in the serum of GEP-NET patients. The results of this small pilot study should be evaluated in further clinical trials by correlating survivin serum levels before and after therapy in order to define its potential diagnostic and prognostic use.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B44.
Abstract B49: FOLPI (picoplatin/5-fluorouracil/leucovorin) versus modified FOLFOX-6 as a neuropathy-sparing first-line therapy for colorectal cancer (CRC)
Authors: De Jager Robert , Cheporov Sergey , Gladkov Oleg , Biakhov Mikhail , Breitz Hazel , Earhart Robert
Published: June 2012

Introduction: Picoplatin (Pico) was designed to overcome platinum resistance and has the potential for improved safety compared to other platinum agents. FOLFOX (5-FU, LV, oxaliplatin [oxali]) treatment for advanced CRC has dose-limiting oxali-related neurotoxicity. The incidence of grade (G) 3–4 neurotoxicity with single-agent Pico across studies was <2%, suggesting that Pico may be a neuropathy-sparing alternative to oxali. The present randomized Phase 2 study evaluates Pico when administered Q4W with Q2W FU and LV (FOLPI) vs modified FOLFOX-6 (FOLFOX) as 1st line treatment for patients (pts) with advanced CRC.

Methods: Each pt with no prior chemotherapy for advanced CRC received LV and infusional FU per FOLFOX Q2W. Pts received Pico Q4W (150 mg/m2) or oxali Q2W (85 mg/m2). Tumor response was assessed by RECIST using CT scans. Adverse events (AEs) were assessed with CTCAE and neuropathy was evaluated using the FACT-Neurotoxicity questionnaire and by a neurologist blinded to treatment.

Results: 51 pts received FOLPI and 50 received FOLFOX. Platinum exposure was similar between treatment arms. With FOLFOX, the most frequent G 3/4 AEs were neutropenia (22%), neuropathy (16%) and thrombocytopenia (12%). With FOLPI, the most frequent G 3/4 AEs were neutropenia (57%), thrombocytopenia (43%) and anemia (22%). There was no G3/4 neuropathy on the FOLPI arm. FOLPI had significantly less neurotoxicity (G 2–4) than FOLFOX (p<0.0002). One FOLPI pt had febrile neutropenia, was retreated and achieved a CR. No pt was discontinued due to bleeding. Hematologic support (neutrophil growth factors, platelet transfusions, RBC transfusions and erythropoietin) were 4%, 2%, 12% and 12% for FOLPI, and 0%, 0%, 6% and 4% for FOLFOX, respectively. Drug-related AEs resulting in study drug discontinuation occurred in 20% of pts on both FOLPI and FOLFOX: neuropathy (0%, 10%), hematology (14%, 6%) and increased creatinine (6%, 2%), respectively. Most pts discontinued study drug for progressive disease on both FOLPI and FOLFOX. On FOLPI there were 2 CR (4%) and 13 PR (25%). On FOLFOX there were 3 CR (6%) and 16 PR (32%). Disease control (CR+PR+SD) was 75% for FOLPI and 76% for FOLFOX. Median overall survival has not yet been determined; however, 12-month survival rates (95% CI) for FOLPI and FOLFOX are 52% (37–65%) and 55% (40–68%), respectively.

Conclusions: Neurotoxicity with FOLPI was less frequent and less severe compared to FOLFOX. Hematologic toxicity with FOLPI was manageable. FOLPI treatment of 1st line CRC had similar disease control and survival rates to FOLFOX supporting picoplatin as a potential neuropathy-sparing alternative to the use of oxaliplatin.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B49.