bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 646: Structurally guided development of meditope-based antibody-drug conjugates
Authors: Matho Michael M. , Gardiner Elisabeth M. , Dumitru Calin D.
Published: August 2015

Meditopes are cyclic peptides that bind the Fab fragment noncovalently, in a novel and unique site, outside the paratope. This property can be utilized to harness cytotoxic payloads to an antibody in a site-specific and consistent manner. One of the originally described peptides, named cQFD, presents a moderate affinity for the humanized antibody cetuximab (KD = 0.43 uM). The binding site involves residues from the heavy as well as the light chain of the antibody, and from both sides of the variable-to-constant hinge of the Fab domain, with a predominance for residues of the variable region. A similar affinity of cQFD to trastuzumab was achieved by mutating thirteen specific residues of the antibody, all located in the variable (Fv) region of the Fab domain; Nine of which are located in the light chain (LC: V9, I10, R39, T40, N41, G42, S43, R45, I83, D85, A100), two in the heavy chain (HC: S40 and I89). We refer to those mutations as to the “meditope enabling”. Via a combined approach, we (i) looked into peptide modifications to increase meditope's affinity for the antibody, and (ii) evaluated the transferability of this approach onto a panel of human antibodies of therapeutic relevance. Based on X-ray crystallography analysis, antibody modeling—in the case of antibodies for which no structure is available—and guided docking, we suggested a number of modifications (point mutations and/or peptide extension) that were subsequently tested via surface plasmon resonance. The top meditope leads were conjugated to MMAE and DM1 and their binding was reassessed to a number of humanized/IgG1 antibodies-confirming that payload conjugation does not interfere with meditope binding and also that meditope enabling and binding does not interfere with antigen binding. Finally, the complexes between antibodies and meditope-drug conjugates were tested in vitro in a panel of tumor cell lines, to measure and rank the efficacy of those leads in our ADC pipeline for further development as novel oncology therapeutics.

Citation Format: Michael M. Matho, Elisabeth M. Gardiner, Calin D. Dumitru. Structurally guided development of meditope-based antibody-drug conjugates. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 646. doi:10.1158/1538-7445.AM2015-646
Abstract 648: Optimization of a PEGylated glucuronide-auristatin linker for antibody-drug conjugates
Authors: Burke Patrick J. , Hamilton Joseph Z. , Jeffrey Scott C. , Hunter Joshua H. , Doronina Svetlana O. , Okeley Nicole M. , Anderson Martha E. , Senter Peter D. , Lyon Robert P.
Published: August 2015

The clinical success of antibody-drug conjugates (ADCs) brentuximab vedotin and ado-trastuzumab emtansine has led to an increased effort to identify new antibody payloads and develop improved linker technologies. Most antibody payloads impart significant hydrophobicity to the ADC resulting in accelerated plasma clearance and diminished in vivo activity, particularly for conjugates with high drug to antibody ratios (DAR). Consequently, drug-linkers have been designed to minimize or mask hydrophobicity, enabling a higher DAR and improved pharmacokinetic properties, translating to enhanced in vivo potency. We recently reported on the incorporation of a discrete PEG polymer as a side chain in the β-glucuronide-monomethylauristatin E (MMAE) linker to provide homogeneous DAR 8 conjugates with decreased plasma clearance and increased antitumor activity in xenograft models relative to a non-PEGylated control. The PEG-glucuronide-MMAE lead has now been optimized by minimizing the size of the PEG side chain and incorporating a self-stabilizing maleimide to prevent payload de-conjugation in vivo. Multiple PEG-glucuronide-MMAE linker constructs were prepared with PEG size varying from 0 to 24 ethylene oxide units, and uniform DAR 8 ADCs were evaluated. PEG size had small but variable effects in vitro, with diminished activity observed in some cell lines. In contrast, a clear relationship was observed between PEG length and plasma pharmacokinetics. Longer PEG chains resulted in slower clearance, with a ‘threshold’ length beyond which clearance was not impacted. Conjugates bearing PEG of sufficient length to minimize ADC plasma clearance provided a wider therapeutic window relative to conjugates bearing shorter PEGs. Thus, sufficient masking of drug hydrophobicity is a viable strategy to provide uniform DAR 8 ADCs that preserve ADC pharmacokinetics and increase potency through higher drug loading.

Citation Format: Patrick J. Burke, Joseph Z. Hamilton, Scott C. Jeffrey, Joshua H. Hunter, Svetlana O. Doronina, Nicole M. Okeley, Martha E. Anderson, Peter D. Senter, Robert P. Lyon. Optimization of a PEGylated glucuronide-auristatin linker for antibody-drug conjugates. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 648. doi:10.1158/1538-7445.AM2015-648
Abstract 652: Antibody-drug conjugates (ADCs) of indolino-benzodiazepine DNA-alkylating agents
Authors: Miller Michael L. , Shizuka Manami , Fishkin Nathan , Reid Emily , Archer Katie , Maloney Erin , Bai Chen , Ab Olga , Yoder Nick C. , Wu Rui , Hong Erica , Bogalhas Megan , Wilhelm Alan , Whiteman Kathleen , Chari Ravi
Published: August 2015

There are over thirty ADCs in the clinic today, reflecting the interest in ADCs for the treatment of cancer. Most of these ADCs utilize a tubulin-interacting small molecule as their cytotoxic payload. In order to continue to extend the utility of ADCs to more types of cancers, there is a strong interest in developing cytotoxic payloads with other mechanisms of action. Previously we have described the development of a class of indolino-benzodiazepine dimers (IGNs) with high potency and specificity. ADCs made with a mono-imine containing IGN, which only alkylate DNA, were found to display improved tolerability, along with the absence of delayed toxicity, compared to their DNA-cross-linking di-imine counterparts.

Additional structure-activity relationship studies with mono-imine containing IGNs have resulted in two promising new leads, IGN-D1 and IGN-P1, which are conjugated to an antibody via either a disulfide or peptide linker, respectively. ADCs of IGN-D1 and IGN-P1 were prepared for evaluation using a folate receptor α (FRα)-binding antibody, anti-FRα, and EGFR-binding antibody, anti-EGFR.

Anti-FRα-IGN-D1 and anti-FRα-IGN-P1 were found to be highly potent against the FRα-expressing KB cell line (IC50 values of 8 pM and 4 pM, respectively) and also demonstrated strong activity against nearby target-negative cancer cells. The addition of excess unconjugated antibody blocked the cytotoxic effect of these conjugates, thus demonstrating that their activity is antigen specific. In addition, both anti-FRα-IGN conjugates were found to be highly potent (IC50 values of 100 pM and 30 pM, respectively) against the breast ductal carcinoma cell line, T47D, despite only a moderate level of antigen expression on the target cells. Similarly, anti-EGFR-IGN-D1 and anti-EGFR-IGN-P1 were found to have IC50 values in the picomolar range against a panel of EGFR-expressing cell lines. In vivo, anti-FRα-IGN-D1 and anti-FRα-IGN-P1 were found to be effective against NCI-H2110 non-small cell lung cancer xenografts, with anti-tumor activity observed at single doses as low as 3 μg/kg (payload dose, equivalent to 0.15 mg/kg Ab dose).

These data support evaluation of new ADC therapies with these two new IGNs for solid tumor indications as well as tumors with low antigen expression.

Citation Format: Michael L. Miller, Manami Shizuka, Nathan Fishkin, Emily Reid, Katie Archer, Erin Maloney, Chen Bai, Olga Ab, Nick C. Yoder, Rui Wu, Erica Hong, Megan Bogalhas, Alan Wilhelm, Kathleen Whiteman, Ravi Chari. Antibody-drug conjugates (ADCs) of indolino-benzodiazepine DNA-alkylating agents. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 652. doi:10.1158/1538-7445.AM2015-652
Abstract 68: Exomic landscape of uterine leiomyosarcomas
Authors: Mäkinen Netta , Heikkinen Tuomas , Aavikko Mervi , Bützow Ralf , Vahteristo Pia
Published: August 2015

Uterine leiomyosarcomas are rare malignant tumors originating from the smooth muscle lining of the uterus, the myometrium. They account for 1-2% of all uterine malignancies with an estimated incidence of 0.3-0.4/100,000 women per year. The symptoms, signs, and histopathological features of uterine leiomyosarcomas widely overlap with those of uterine leiomyomas, their benign counterparts, which can cause diagnostic challenges in daily pathological practice. Because leiomyosarcomas are clinically aggressive tumors with poor 5-year survival and a high recurrence rate, early diagnosis is important for prolonged survival. Currently, the diagnosis of these tumors is often incidental and postoperative. Most leiomyosarcomas are aneuploid with complex numerical and structural chromosomal aberrations and show substantial cytogenetic heterogeneity. Although a few cancer genes have been implicated in uterine leiomyosarcomas, such as TP53, RB1, and CDKN2A, these genes represent, however, common cancer genes not specific for smooth muscle malignancies. The aim of this study is to examine somatic variation in the coding regions of uterine leiomyosarcomas by exome sequencing to identify potential new molecular biomarkers. Altogether 18 formalin-fixed paraffin-embedded tumor samples entered whole-exome sequencing. After the filtering of exome sequencing data, a list of genes which were mutated in at least two tumors was created. The list included over 50 genes TP53 being the most frequently mutated gene. The most prominent candidate variants are validated with Sanger sequencing and the validations are ongoing. Also a series of 35 additional tumors is available on tissue microarray for possible immunohistochemical validations. Identification of novel molecular genetic characteristics in uterine leiomyosarcomas is clinically relevant to further improve the diagnosis and prognosis of the patients. New knowledge on the mechanisms underlying the progression to malignancy is also essential for developing new treatments.

Citation Format: Netta Mäkinen, Tuomas Heikkinen, Mervi Aavikko, Ralf Bützow, Pia Vahteristo. Exomic landscape of uterine leiomyosarcomas. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 68. doi:10.1158/1538-7445.AM2015-68
Abstract 685: Mechanisms of intrinsic and acquired resistance to third generation inhibitors in EGFR mutant NSCLC
Authors: Niederst Matt J.
Published: August 2015

Acquired resistance is a fundamental problem limiting the efficacy of tyrosine kinase inhibitors (TKIs) in EGFR mutant NSCLC. In over half of the cases of acquired resistance to the first generation inhibitors gefitinib and erlotinib, a secondary mutation to EGFR, T790M, promotes resistance by preventing these drugs from fully suppressing the receptor. The recent development of next generation EGFR TKIs has proven to be a major breakthrough in the treatment of these patients. In clinical trials testing the third generation EGFR inhibitors CO-1686 and AZD9291, response rates of greater than 50% in T790M+ patients have been reported. While these drugs hold a great deal of promise, they too will be limited, initially by intrinsic resistance, and eventually by acquired resistance. To study intrinsic resistance in this setting, we have developed a panel of cell lines derived from biopsies of T790M+ erlotinib-resistant patients, tested their sensitivity to third generation inhibitors and identified biomarkers that predict a lack of response. To study acquired resistance, we have generated resistance in these cells lines in vitro and derived additional models directly from patient biopsies following progression on the clinical trial. Using a combination of functional, sequencing, and drug screening approaches we have begun to identify resistance mechanisms to third generation inhibitors.

Citation Format: Matt J. Niederst. Mechanisms of intrinsic and acquired resistance to third generation inhibitors in EGFR mutant NSCLC. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 685. doi:10.1158/1538-7445.AM2015-685
Abstract 690: Characterization of serine/threonine phosphatase LAPP1 as a diagnostic and therapeutic target for lung cancer
Authors: Daigo Yataro , Takano Atsushi , Nakamura Yusuke
Published: August 2015

Lung cancer is the leading cause of cancer deaths worldwide. Since the number of patients responding well to current standard therapies is still small, further development of new anti-cancer drugs with minimum risk of adverse event is urgently awaited. To screen oncoantigens which could be used for the development of new diagnostic biomarkers and molecular targeted drugs, we have established a screening system as follows; i) To identify overexpressed genes in 120 lung cancers by genome-wide screening using the microarray representing 27,648 genes and pure populations of tumor cells taken from cancer tissues by laser microdissection, ii) To verify the candidate genes for their very low or absent expression in normal tissues by northern-blotting, iii) To validate the clinicopathological significance of their expression with tissue microarray covering hundreds of archived lung cancers, iv) To verify whether the target genes are essential for the growth or survival of cancer cells by RNAi and cell growth assays. During this process, we selected dozens of druggable oncoproteins with various enzymatic activities, and identified overexpression of Serine/Threonine phosphatase LAPP1 (lung cancer-associated protein phosphatase 1) in the majority of lung cancers and its scarce expression in normal tissues except testis. Strong LAPP1 expression was an independent prognostic factor for lung cancer patients who underwent curative surgery (P<0.0001). We identified EF-1δ (a cadmium-responsive proto-oncogene) as a LAPP1-interacting protein that was also over-expressed in cancer cells. Knock-down of LAPP1 or EF-1δ expression by siRNAs inhibited cell growth, whereas exogenous expression of LAPP1 in EF-1δ(+) cells enhanced the dephosphorylation of EF-1δ(+), and subsequent increase in the levels of active phospho-AKT. Inhibition of interaction between LAPP1 and EF-1δ by a cell-permeable peptide corresponding to a leucine zipper motif in EF-1δ, resulted in the destabilization of EF-1δ and inhibited cancer cell growth and invasion. LAPP1 is likely to be a prognostic biomarker and a potential therapeutic target for lung cancer.

Citation Format: Yataro Daigo, Atsushi Takano, Yusuke Nakamura. Characterization of serine/threonine phosphatase LAPP1 as a diagnostic and therapeutic target for lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 690. doi:10.1158/1538-7445.AM2015-690
Abstract 693: A yeast synthetic lethal screen identifies a conserved interaction between PLK1 and CKS1b affecting cancer cell viability
Authors: Reid Robert J. , Du Xing , Sunjevaric Ivana , Rayannavar Vinayak , Dittmar John , Maurer Matthew , Rothstein Rodney
Published: August 2015

The CKS1b gene is located on chromosome 1q21 and is frequently amplified in breast, lung and liver cancers. CKS1b codes for a conserved regulatory subunit of cyclin-CDK complexes that functions at multiple stages of cell cycle progression. We used a high throughput screening protocol to mimic cancer-related over-expression in Saccharomyces cerevisiae mutants to identify genes whose functions become essential only when CKS1 is over- expressed, a genetic interaction termed synthetic dosage lethality (SDL). Mutations in genes affecting the mitotic entry checkpoint and genes affecting mitotic exit are evident among the SDL interactions identified. The mitotic entry checkpoint controls the stability of the mitotic inhibitor Swe1 (Human WEE1), resulting in delayed mitosis when cell polarity is disrupted. Swe1 is a tyrosine kinase that inhibits the cyclin-dependent kinase (CDK) complex by phosphorylation of a conserved tyrosine-19 on Cdc28 (Human CDK1). We find that the SDL interaction between Cks1 and the mitotic entry checkpoint requires both Swe1 and the Cdc28 tyrosine-19 residue. The mitotic exit network is a signaling cascade that controls post-anaphase inhibition of CDK activity necessary for progression into G1. Since the polo-like kinase Cdc5 (Human PLK1) functions in both the mitotic entry and mitotic exit pathways, we investigated the effect of targeting PLK1 in breast cancers with varying expression of CKS1b. We analyzed published datasets and found that growth inhibition resulting from PLK1 knockdown was strongly correlated with increased CKS1b expression levels in breast cancer cell lines [Marcotte, R., Brown, K., Suarez, F., & Sayad, A. (2012). Cancer Discovery, 2:172]. We independently confirmed sensitivity to PLK1 knockdown in 8 breast cancer cell lines in which 3 of 4 sensitive lines had high CKS1b expression and 3 of 4 insensitive lines had low CKS1b expression. We modified CKS1b expression in hMEC cells and showed that increased expression enhanced apoptosis upon PLK1 knockdown while decreased CKS1b expression abrogated sensitivity. Thus identification of a yeast synthetic dosage lethal (SDL) interactions identifies a conserved interactions that may be targeted to kill cancer cells.

Citation Format: Robert J. Reid, Xing Du, Ivana Sunjevaric, Vinayak Rayannavar, John Dittmar, Matthew Maurer, Rodney Rothstein. A yeast synthetic lethal screen identifies a conserved interaction between PLK1 and CKS1b affecting cancer cell viability. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 693. doi:10.1158/1538-7445.AM2015-693
Abstract 708A: Suppression of glycolysis by retinoic acid sensitizes hepatocellular carcinoma cells to apoptosis induced by sorafenib via AMPK activation
Authors: Shiota Goshi , Shimizu Hiroki , Kanki Keita
Published: August 2015

Aims: Hepatocellular carcinoma (HCC) is the fifth cause of cancer death all over the world and its prognosis is very poor. To improve the prognosis of HCC patients, a novel therapy including chemotherapy against HCC should be developed. The strategies to enhance the efficacy of anticancer drugs are required. Sorafenib is the only drug to prolong overall survival of the patients with HCC, however, the outcome is not still satisfactory. In this study, we investigated the effects of combination treatment of anticancer drugs and retinoids,which has been known to show antitumor activity, in HCC cells. Methods: Cytotoxicity of combination treatment of retinoids (all-trans retinoic acid (ATRA), NIK-333 and Am80) and anticancer drugs (sorafenib, adriamycin, cisplatin, mitomycin C and 5-FU) was examined by cell viability assay. The effect on energy metabolic pathway, such as AMPK activation, intracellular ATP level, and the expression of glycolytic and TCA cycle genes were examined in HepG2 cells treated with sorafenib alone or in combination with retinoids. Induction of apoptosis was evaluated in the cells treated with the combination treatment of sorafenib and ATRA. Results: ATRA enhanced the cytotoxic effects of all 5 anticancer drugs tested, being much more effective than NIK-333 andAm80.AMPK activation, reduction of ATP content, and downregulation of glycolytic genes GLUT-1 and LDHA were observed in the cells treated with ATRA alone or in combination with sorafenib. In the combination treatment, increased apoptosis which was followed by activation of p38MAPK and JNK, upregulation and translocation of Bax to mitochondria, and activation of caspase-3 was observed. Conclusion: ATRA is useful for enhancing the cytotoxicity of antitumor drugs for HCC by regulating energy metabolism in HCC cells.

Citation Format: Goshi Shiota, Hiroki Shimizu, Keita Kanki. Suppression of glycolysis by retinoic acid sensitizes hepatocellular carcinoma cells to apoptosis induced by sorafenib via AMPK activation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 708A. doi:10.1158/1538-7445.AM2015-708A
Abstract 749: Both amplification and protein expression are required to predict FGFR tyrosine kinase inhibitor sensitivity in lung cancer
Authors: Ebi Hiromichi , Kotani Hiroshi , Yano Seiji
Published: August 2015

FGFR1 amplification has been suggested to be a driver oncogene in squamous and small cell lung carcinoma. However, early clinical trials show that response rate of FGFR tyrosine kinase inhibitors (FGFR-TKIs) is 20-30% in patients with FGFR1 amplified lung cancer. To elucidate the reason for de novo resistance to FGFR-TKI, six FGFR1 amplified lung cancer cell lines (NCI-H1581, DMS-114, NCI-H520, NCI-H1703, HCC95, and Calu-3) were analyzed. These cells were confirmed to have FGFR1 amplification by FISH analysis. Cell viability assay showed only 2 of six cell lines, NCI-H1581 and DMS-114, are sensitive to a FGFR-TKI, BGJ-398. Among BGJ-398 insensitive cells, NCI-H1703, HCC95, and Calu-3 cells express low mRNA and protein level of FGFR1 even in the presence of FGFR1 amplification. In addition, while BGJ-398 downregulates ERK phosphorylation in FGFR1 protein expressing cells, the drug had no effect on ERK signaling in BGJ-398 resistant cells with low FGFR1 protein expression. Importantly, other driver oncogenes were found in these BGJ-398 insensitive cell lines. PDGFRa amplification in H1703 cells and HER2 amplification in Calu-3 cells were reported previously. In this study, we have also found ERBB3 ligand NRG1 was overexpressed in HCC95 cells. Knockdown of NRG1 suppressed cell growth and combination of lapatinib with BGJ-398 led to cell death. To further determine whether FGFR1 protein expression is enough for the sensitivity to FGFR-TKIs, lung cancer cell lines were screened to FGFR-TKIs, BGJ-398 and PD-173034. This screen identified that all FGFR-TKI sensitive cell lines harbor FGFR1 amplification with high protein expression. These results suggest FGFR-TKIs should be used to patients harboring both FGFR1 amplification and protein expression. In case tumors with FGFR1 amplification, but expressing low FGFR1 protein, another driver oncogene may coexist and targeting both FGFR1 and these coexisting driver oncogenes are required to treat these tumors.

Citation Format: Hiromichi Ebi, Hiroshi Kotani, Seiji Yano. Both amplification and protein expression are required to predict FGFR tyrosine kinase inhibitor sensitivity in lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 749. doi:10.1158/1538-7445.AM2015-749
Abstract 750: Overcoming resistance to anti-EGFR therapy in non-small cell lung cancer with a T-type Ca2+ channels inhibitor
Authors: Dziegielewska Barbara , Gray Lloyd S. , Dziegielewski Jaroslaw
Published: August 2015

This study's aim was to define a novel molecular target and to develop an effective agent capable of overcoming intrinsic and acquired resistance to anti-EGFR therapy. Despite introduction of new therapies, lung cancer is a major cause of cancer death. One of its characteristics is the expression and activation of epidermal growth factor receptor (EGFR). Activating mutations in EGFR are present in approximately 20% of patients with non-small cell lung cancer (NSCLC), mostly in never-smokers. While most patients initially respond to treatment with anti-EGFR therapy, essentially all patients develop acquired drug resistance. T-type calcium channels, are present in lung cancer and support cell proliferation. Importantly, their expression often coincides with abnormal expression or mutations in EGFR. In this study, we investigated the effects of T-type channel inhibition on NSCLC survival and resistance to targeted therapy in vitro. We used a panel of cell lines differing in EGFR mutation status and sensitivity to EGFR tyrosine kinase inhibitors (TKIs). Our results show that T-type channel inhibitors alone arrest cells in the G1/G0 phase of the cell cycle and induce cell death. The treatment reduces activity of both PI3K/AKT/mTOR and JAK2/STAT5 pathways, often upregulated as a mechanism of acquired resistance to TKIs. Importantly, combined treatment with TKI, gefitinib, and T-type channels inhibitor, mibefradil, resulted in synergistic growth inhibition. Our results demonstrate a connection between T-type Ca2+ channels and aberrant EGFR activity in NSCLC, and provide the rationale for future use of two targeted therapies together. Since T-type calcium channel inhibitors are already undergoing clinical trials, our discoveries could be quickly translated into clinical practice.

Citation Format: Barbara Dziegielewska, Lloyd S. Gray, Jaroslaw Dziegielewski. Overcoming resistance to anti-EGFR therapy in non-small cell lung cancer with a T-type Ca2+ channels inhibitor. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 750. doi:10.1158/1538-7445.AM2015-750
Abstract 782: Modulation of receptor tyrosine kinase signaling by cMET inhibitor, INC280, in cMETmut/amp gastric carcinoma cell line Hs746.T
Authors: Pinzon-Ortiz Maria C. , Rong Xianhui , Liang Jinsheng , Wang Hui Qin , Huang Alan , Schlegel Robert , Cao Z. Alexander
Published: August 2015

Aberrant Receptor Tyrosine Kinase (RTK) activity is one of the hallmarks of cancer. The genetic alternations of ABL and EGFR are classical examples of RTK-dependent oncogenesis. Recent data suggest that non mutated RTK signaling can form feedback pathways to serve as resistant mechanism against targeted therapies. Here, we examined the regulation of RTK signaling by a potent cMET inhibitor, INC280 (enzymatic EC50 against c-MET = 0.13 nM), in a cMET-dependent human gastric tumor model Hs746.T.

Hs746.T is a heavily cMET dependent cell line with both cMET amplication (12.9 copies) and mutation (G-T mutation at slice donor site to cause juxtamembrane domain deletion). This model is exquisitely sensitive to cMET inhibitor, INC280 in vitro (GI50 = 9 nM) and in vivo. The treatment of INC280 lead to significant inhibition of pcMET. Furthermore, the treatment of INC280 resulted in the suppression of pEGFR, though INC280 does not actively inhibit EGFR (enzymatic EC50 against EGFR > 10 microM). This result supports prior reports that cMET and EGFR can form physical complex, with cMET modulating EGFR activation. Consequently, INC280 inhibited downstream signaling nodes of pAKT, pS6RP, pMEK and pERK in Hs746.T.

To examine the RTK signaling underlying possible treatment relapse, we treated Hs746.T cells chronically in vitro under increasing concentrations of INC280. After 8 weeks of treatment, Hs746.T-R cells were generated that grew at 600 nM of INC280. RTK signaling was examined in Hs746.T-R cells under treatment of INC280. As expected, cMET signaling was significantly curtailed in Hs746.T-R cells, as was pEGFR. Interestingly, pPDGFRb was elevated in Hs746.T-R, compared to the parental cell line. We hypothesize that the increased signaling through PDGFRb may serve as resistance mechanism to INC280 in Hs746.T-R cells.

In summary, we have examined RTK signaling in the cMET-dependent Hs746.T cell line. Our finding supported the notion of RTK cross-talk between cMET and EGFR in this setting. Additionally, our data reveal that additional RTK signaling (PDGFRb) can be induced under chronic treatment of INC280 to possibly mediate resistance to cMET inhibition.

Citation Format: Maria C. Pinzon-Ortiz, Xianhui Rong, Jinsheng Liang, Hui Qin Wang, Alan Huang, Robert Schlegel, Z. Alexander Cao. Modulation of receptor tyrosine kinase signaling by cMET inhibitor, INC280, in cMETmut/amp gastric carcinoma cell line Hs746.T. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 782. doi:10.1158/1538-7445.AM2015-782
Abstract 802: The inactivation of PP2A caused by the replacement of its regulatory subunit plays a crucial role in the hemangiomagenesis
Authors: Xu Qin , Xie Furong
Published: August 2015

Hemangioma is the most frequent vascular tumor of early childhood. However, the mechanism leading to their development, progression and maintenance has remained unclear. In this study, we generated an endothelial-specific PyMT-expressing transgenic (Tg) mouse (Tie2-PyMT) that spontaneously develop hemangioma. Using Tie2-PyMT transgenic mice and bEnd.3 cells, an PyMT induced mouse brain endothelial cell line idicates that the direct expression of PyMT gene in vascular endothelial cells leads to rapid cell proliferation and hemangioma development. An interaction between the core AC dimer of protein phosphatase 2A (PP2A) and PyMT, but not B regulatory subunit was verified both in hemangioma endothelial cells of Tie2-PyMT Tg mice and bEnd.3 cells by immunoprecipitation. Whereas in endothelial cells of Tg negative mice and PyMT silenced bEnd.3 cells, the B regulatory subunit binds to the core AC dimer of PP2A. PyMT binding to PP2A led to the inactivation of PP2A phosphatases and subsequently activates its downstream ERK1/2 and AKT pathway, which may be responsible for the proliferative effect of PyMT on vascular endothelial cells. Moreover, the effect of PyMT on PP2A phosphatases activity could be rescued by Okadaic acid (OA), a specific inhibitor of PP2A. Treamtment of Tie2-PyMT transgenic mice with the PP2A activator, FTY720, significantly delayed hemangiomagenesis in this model. Furthermore, PP2A complex composition and its phosphatases activity were also examined in patient hemangioma specimens. Consisten with the animal model, a decreased PP2A phosphatases activity and the dissociation of B subunit from the PP2A core dimer were observed. These findings suggest that the inactivation of PP2A caused by the replacement of its regulatory subunit plays a crucial role in the hemangiomagenesis.

Citation Format: Qin Xu, Furong Xie. The inactivation of PP2A caused by the replacement of its regulatory subunit plays a crucial role in the hemangiomagenesis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 802. doi:10.1158/1538-7445.AM2015-802
Abstract 81: Effect of HSP90 inhibition on the gene expression profile of MDM2 transfected LNCaP-MST prostate cancer cells
Authors: Venkatesan Thiagarajan , Alaseem Ali , Alhazzani Khalid , Rathinavelu Appu
Published: August 2015

The heat shock protein 90 (HSP90) is a major molecular chaperone of the cell that appears to be a master regulator of the stability and activity of multiple oncoproteins such as HER2, Akt, Bcr-Abl, c-Kit, EGFR and BRAF. Recent approaches have revealed that most of the target proteins of HSP90 are protein kinases or transcription factors which play important roles in the cellular carcinogenesis. The HSP90 also plays an important role in the regulation of cell cycle, proliferation, vascularization, invasion and metastasis of cancer cells, as well as in maintaining the stability and functions of several carcinogenic proteins involved in altering the signal transduction pathways of apoptosis. The 17-allylamino-17-demethoxygeldanamycin (17-AAG) is a well known inhibitor of HSP90 and it was speculated that, this inhibitor may have particular value in the antitumor and angiostatic therapies. Hence, we were trying to determine the expression profile of the genes in the angiogenesis pathway following treatment of MDM2 overexpressing LNCaP-MST prostate cancer cells with 17-AAG. In the LNCaP-MST cells, the expression of genes such as TNF-α, MMP9 and TGFB1 were found to be significantly up-regulated compared to the LNCaP cells. Hence, MDM2 overexpression was suspected as the primary cause of these increases. However, after treating the LNCaP-MST cells with 10 μm 17-AAG for 24 hrs, several of these genes were significantly altered compared to the untreated LNCaP-MST cells. In particular, the 17-AAG treatment significantly down-regulated the expression of TNF-α, TGFB1, ID1, IL8 and F3 which are involved in the regulation of angiogenesis and tumor growth. The heat map and scatterplot analysis clearly confirmed the alterations in the expression levels of these genes following 17-AAG treatment. Cluster analysis further indicated that the gene expression patterns are positively correlated with the anti-angiogenic effects of 17-AAG treatment in LNCaP-MST cells compared to untreated cells. Our results offer convincing evidence to suggest that the inhibition of HSP90 can control both pro-angiogenic and cell proliferative signals in LNCaP-MST cells. (The financial support from the Royal Dames of Cancer Research Inc., Ft. Lauderdale, Florida is gratefully acknowledged).

Citation Format: Thiagarajan Venkatesan, Ali Alaseem, Khalid Alhazzani, Appu Rathinavelu. Effect of HSP90 inhibition on the gene expression profile of MDM2 transfected LNCaP-MST prostate cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 81. doi:10.1158/1538-7445.AM2015-81
Abstract 824: A novel mechanism of areca nut-induced oral cancer is through the conversion of cancer stem cells to acquire malignant phenotypes
Authors: Li Yi-Chen , Cheng Ann-Joy
Published: August 2015

Areca nut is one of the most commonly used psychoactive substance worldwide. It is has been identified as a primary carcinogen of oral cancer in Southeast Asia. Previous studies to examine the effects of this carcinogen often used short-term and high-dose treatment of area nut extract (ANE) as a research model. To elucidate the molecular effects of areca nut and mimic patient chronic exposure situation, we established four areca nut extract (ANE)-trained oral cancer sublines by treating cells with low-dose of ANE (IC30, dose for 30% growth inhibition) for 2 months. Without evidence of cellular senescence, these sublines exhibited significant increases in invasive ability, along with the up-regulation of MMP1 and the reversed expression of epithelial-mesenchymal transition markers. These sublines also showed more resistant to chemotherapeutics and irradiation, which was associated with the over-expression of ABCG2 transporters, increased production of anti-oxidative enzymes, and insensitive to ROS induction. Furthermore, these chronically treated cells possessed characteristics of cancer stem cells, as indicated by enrichment of CD24-/CD44+ population, enhancing spheroid cell formation, and increasing expressions of stemness associated molecules, including Slug, Twist1, Sox9, and Grp78. Thus, we concluded that chronic areca nut exposure induces a pleotropic effect on stemness regulators that facilitates the conversion of cancer stem cells to acquire malignant transformation in oral cancer. This novel finding provides valuable knowledge on areca nut-induced oral carcinogenesis.

Citation Format: Yi-Chen Li, Ann-Joy Cheng. A novel mechanism of areca nut-induced oral cancer is through the conversion of cancer stem cells to acquire malignant phenotypes. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 824. doi:10.1158/1538-7445.AM2015-824
Abstract 826: DNA adducts induced by in vitro activation of diesel and biodiesel exhaust extracts
Authors: Ross Jeffrey A. , Nelson Garret B. , Mutlu Esrta , Warren Sarah H. , Peggy Matthews P. , Gilmour M. Ian , DeMarini David M.
Published: August 2015

Petroleum diesel exhaust is a complex mixture containing many probable and known carcinogens. The development of biodiesel and biodiesel blends offers a renewable alternative to petroleum diesel, but few data are available concerning the carcinogenic potential of biodiesel exhausts. We compared the formation of covalent DNA adducts by the in vitro metabolic activation of organic extracts of diesel exhaust particles (DEP)from petroleum diesel and biodiesel. Two different petroleum diesel DEPs were examined (C-DEP and B0), as well as one biodiesel DEP (B100), and DEP resulting from combustion of a blend of 20% B100 and 80% B0 (B20), which is representative of commercially available biodiesel. Oxidative activation was performed in the presence of calf thymus DNA (ctDNA) using rat liver microsomal fractions with required cofactors (S9), and nitroreductive activation was performed using xanthine oxidase (XO) and hypoxanthine in the presence of ctDNA. The modified DNA was hydrolyzed and analyzed by 32P-postlabeling using either butanol extraction or nuclease P1 pre-enrichment. Multiple DNA adducts were produced with chromatographic mobilities consistent with polycyclic aromatic hydrocarbon (PAH) and nitro-PAH adducts. The types and quantities of DNA adducts produced from two independent petroleum diesel DEPs were similar, with evidence of formation of both PAH- and nitroPAH- derived adducts. In contrast, the biodiesel DEP B100 induced higher total levels of DNA adducts in both activation systems. The lowest levels of DNA adduct formation were observed with B20 DEP. These results suggest that the DEP from available biodiesel blends (B20) pose lower risk for induction of DNA damage than petroleum diesel.

This is an abstract or a proposed presentation and does not necessarily reflect EPA policy.

Citation Format: Jeffrey A. Ross, Garret B. Nelson, Esrta Mutlu, Sarah H. Warren, Matthews P. Peggy, M. Ian Gilmour, David M. DeMarini. DNA adducts induced by in vitro activation of diesel and biodiesel exhaust extracts. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 826. doi:10.1158/1538-7445.AM2015-826
Abstract 829: Circulating immune/inflammation markers in Chinese workers occupationally exposed to formaldehyde
Authors: Lan Qing , Seow Wei Jie , Zhang Luoping , Vermeulen Roel , Tang Xiaojiang , Hu Wei , Bassig Bryan A. , Ji Zhiying , Shiels Meredith S. , Kemp Troy J. , Shen Min , Qiu Chuangyi , Reiss Boris , Freeman Laura Beane , Blair Aaron , Kim Christopher , Guo Weihong , Wen Cuiju , Li Laiyu , Pinto Ligia A. , Huang Hanlin , Smith Martyn T. , Hildesheim Allan , Rothman Nathaniel
Published: August 2015

Background: Formaldehyde has been proposed to be a human myeloid leukemogen. However, the exact mechanism for the basis for this link is still debated. We aimed to evaluate whether circulating immune/inflammation markers were different in workers occupationally exposed to formaldehyde.

Methods: Using a multiplexed bead-based assay, we measured serum levels of 38 immune/inflammation markers in a cross-sectional study of 43 formaldehyde-exposed and 51 unexposed factory workers in Guangdong, China. Linear regression models adjusting for potential confounders were used to compare marker levels in exposed and unexposed workers.

Results: We found significantly lower circulating levels of four markers among exposed factory workers compared to unexposed controls that remained significant after adjusting for multiple comparisons, including thymus and activation regulated chemokine (TARC;52pg/ml in exposed versus 75pg/ml in controls,P = 0.0015), TNF-related apoptosis-inducing ligand (TRAIL;11.7pg/ml in exposed versus 22.1pg/ml in controls,P = 0.011), C-reactive protein (CRP;797.3ng/ml in exposed versus 3269.1ng/ml in controls,P = 0.013) and chemokine (C-X-C motif) ligand 11 (CXCL11;36.2pg/ml in exposed versus 48.4pg/ml in controls,P = 0.017), suggesting immunosuppression among formaldehyde-exposed workers.

Conclusions: Our findings are consistent with recently emerging understanding that immunosuppression might be associated with myeloid diseases. These findings, if replicated in a larger study, may provide insights into the mechanism by which formaldehyde promotes leukemogenesis.

Citation Format: Qing Lan, Wei Jie Seow, Luoping Zhang, Roel Vermeulen, Xiaojiang Tang, Wei Hu, Bryan A. Bassig, Zhiying Ji, Meredith S. Shiels, Troy J. Kemp, Min Shen, Chuangyi Qiu, Boris Reiss, Laura Beane Freeman, Aaron Blair, Christopher Kim, Weihong Guo, Cuiju Wen, Laiyu Li, Ligia A. Pinto, Hanlin Huang, Martyn T. Smith, Allan Hildesheim, Nathaniel Rothman. Circulating immune/inflammation markers in Chinese workers occupationally exposed to formaldehyde. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 829. doi:10.1158/1538-7445.AM2015-829
Abstract 858: Periodontal bone loss and risk of epithelial ovarian cancer in the Nurses’ Health Study
Authors: Babic Ana , Poole Elizabeth M. , Terry Kathryn L. , Cramer Daniel W. , Teles Ricardo P. , Tworoger Shelley S.
Published: August 2015

Periodontitis, a chronic inflammatory reaction to dysbiotic oral microbiota, is common in the adult population in the United States. It is associated with increased risk of several medical conditions, including cardiovascular diseases, and potentially with lung, oral and pancreatic cancer. One of the proposed mechanisms behind these associations is systemic inflammation, which has also been implied in ovarian cancer etiology. We investigated association between periodontal bone loss, a marker of periodontitis, and risk of epithelial ovarian cancer among 60,560 participants of the Nurses’ Health Study. We did not observe an increased risk of ovarian cancer among participants suffering from periodontal bone loss (HR = 0.86, 95% CI: 0.64-1.15). Association was unchanged after adjusting for numerous factors including smoking. Among women younger than 69 years periodontal bone loss was associated with a 40% (HR = 0.60, 95% CI: 0.36-0.98) decreased ovarian cancer risk, while there was no association in those older than 69 (HR = 1.09, 95% CI: 0.75-1.58), although this difference did not reach a statistical significance (p-heterogeneity = 0.06). There also was a suggestive decreased risk for serous tumors (HR = 0.76, 95% CI: 0.53-1.09). The number of natural teeth and number of root canals, which are other metrics of oral health, were not associated with ovarian cancer risk. Our results do not support an increased risk of ovarian cancer for women with periodontal bone loss; however, given the unexpected associations in younger women, further research is warranted.

Note: This abstract was not presented at the meeting.

Citation Format: Ana Babic, Elizabeth M. Poole, Kathryn L. Terry, Daniel W. Cramer, Ricardo P. Teles, Shelley S. Tworoger. Periodontal bone loss and risk of epithelial ovarian cancer in the Nurses’ Health Study. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 858. doi:10.1158/1538-7445.AM2015-858
Abstract 889: Mouse model for chemotherapy-induced alopecia with transplantation of human hair follicles onto immune deficient mouse
Authors: Kwon Ohsang , Yoon Ji-Seon , Choi Mira , Shin Chang Yup , Kim Kyu Han
Published: August 2015

Optimized research models of chemotherapy-induced alopecia (CIA) require for further understanding the pathogenesis and protection of CIA. The present study was aimed to develop a new mouse model for CIA by follicular unit transplantation of human hair follicles onto the immune-deficient mice. At 22 weeks after transplantation, single dose of cyclophosphamide (Cph) was treated in mice of Cph100 (100mg/kg) and Cph150 (150mg/kg) groups. On day 6 of treatment, hair follicles showed dystrophic anagen representing with swollen dermal papilla and ectopic melanin clumping in the hair bulb. In addition, up-regulated P53 expression and apoptotic matrix keratinocytes, and down-regulated expression of Ki67 and melanogenic protein in the hair bulb were also noted in both groupsin Cph100 and Cph150. Subsequently, hair follicles in Cph100 appeared to recover as anagen hair follicles after 9 days. Ki67 and melanogenic protein expression returned as the levels analogous to the control. On the other hands, hair follicles of Cph150 prematurely entered dystrophic catagen-telogen phase after 9 days of treatment. Ki67 and melanogenic protein expression in immunofluorescence staining were barely visible. Those findings indicate that this model represented two pathways of dystrophic anagen pathway in Cph100 and dystrophic catagen pathway in Cph150. Our model will be a useful tool for the further understanding of the impact of chemotherapy on the human hair follicle.

Citation Format: Ohsang Kwon, Ji-Seon Yoon, Mira Choi, Chang Yup Shin, Kyu Han Kim. Mouse model for chemotherapy-induced alopecia with transplantation of human hair follicles onto immune deficient mouse. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 889. doi:10.1158/1538-7445.AM2015-889