bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 721: Development of a therapeutic HPV vaccine based on mass spectrometry-verified target epitopes
Authors: Kruse Sebastian , Bonsack Maria , Becker Sara , Mohan Nitya , Steinbach Alina , Hoppe Stephanie , Blatnik Renata , Riemer Angelika B.
Published: August 2018

For rational therapeutic vaccine design, detailed knowledge about target epitopes that are truly presented on cancer cells is essential. Many potential tumor-specific epitopes, such as viral epitopes or mutation-derived neoepitopes, are presented at low abundance. Thus, detection of these epitopes remains a challenge. We have developed a high-sensitivity targeted mass spectrometry (MS) approach for direct detection of low-abundant epitopes. We used human papillomavirus (HPV) as a model system, as high-risk HPVs cause over 600,000 cervical, anogenital and oropharyngeal cancer cases per year. Moreover, the HPV oncoproteins E6 and E7 are essential for the induction and maintenance of the malignant phenotype, and thus are ideal targets for immunotherapy.

Potential HPV16 E6 and E7 target epitopes were predicted in silico and their HLA-binding verified in cellular binding assays. HLA-peptide complexes were immunoprecipitated from HPV16-positive cancer cells and the purified peptides were analyzed by MS. Identified HPV epitopes were assessed for immunogenicity in vitro with PBMC from healthy donors and in vivo in the MHC-humanized mouse model A2.DR1.

The binding assays resulted in the identification of known and novel HPV16-derived HLA-binding peptides. Furthermore, we used this data to formulate rules on how to optimally use epitope prediction tools to increase the chances of predicting true HLA ligands. 14 HLA-A2-restricted HPV16 epitopes were detected on the surface of CaSki and 866 cervical cancer cells, 9 of which were novel. 13 out of the 14 MS-detected peptides were immunogenic in ELISpot assays. Vaccination of A2.DR1 mice with one selected epitope induced high numbers of specific cytotoxic T-cells, and caused anti-tumor effects in a newly developed HPV16 E6/E7-expressing A2.DR1-compatible tumor model.

We conclude that our epitope detection and validation approach is suitable for validating even low-abundant candidate epitopes to be true immunotherapy targets. MS-detection of epitopes appears to be a good predictor of immunogenicity, which makes this approach an attractive platform for determining target epitopes for therapeutic cancer vaccines.

Citation Format: Sebastian Kruse, Maria Bonsack, Sara Becker, Nitya Mohan, Alina Steinbach, Stephanie Hoppe, Renata Blatnik, Angelika B. Riemer. Development of a therapeutic HPV vaccine based on mass spectrometry-verified target epitopes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 721.
Abstract 723: Targeting phosphorylated p53 to elicit tumor-reactive T helper responses against head and neck squamous cell carcinoma
Authors: Ohara Kenzo , Kumai Takumi , Hayashi Tatsuya , Ohkuri Takayuki , Bandoh Nobuyuki , Kobayashi Hiroya , Harabuchi Yasuaki
Published: August 2018

The p53 is a DNA-binding and transcription activation domain containing tumor suppressor that upregulates growth arrest and apoptosis-related genes. In response to DNA damage, p53 is phosphorylated at multiple sites including serine 33 or 37. Additionally, abnormal phosphorylation of cellular proteins is a hallmark of malignant transformation. The human immune system is capable of distinguishing between the normal components and these posttranslational modifications. Therefore, the existence of posttranslational modifications increases the diversity of potential immune reactions to a determinant antigen. We identified phosphorylated peptide epitopes from tumor protein p53 that could elicit effective T helper-cell responses. Two epitope peptides, p5322-41/Phospho-S33 and p5322-41/Phospho-S37, induced T helper responses against tumor cells expressing phosphorylated p53 protein. Moreover, cisplatin and doxorubicin augmented the responses of such CD4+ T cells via upregulation of phosphorylated p53. Importantly, we elucidated the presence of peripheral blood lymphocytes specific for these peptide epitopes in the PBMCs of head and neck squamous cell cancer patients. These results predict that epitopes p5322-41/Phospho-S33 and p5322-41/Phospho-S37 are immunogenic for anti-tumor responses and could be beneficial for cancer immunotherapy.

Citation Format: Kenzo Ohara, Takumi Kumai, Tatsuya Hayashi, Takayuki Ohkuri, Nobuyuki Bandoh, Hiroya Kobayashi, Yasuaki Harabuchi. Targeting phosphorylated p53 to elicit tumor-reactive T helper responses against head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 723.
Abstract 725: Targeting the skin microenvironment for effective cancer vaccines
Authors: You Jack R. , Hao Rong , Falo Louis D. , Chen Guo , Hao Xingxing , Carey Cara , You Zhaoyang , Falo Louis D.
Published: August 2018

Skin, the most accessible organ, harbors keratinocytes and immunocytes that contribute to both innate and adaptive immunity, rendering it an ideal anatomic target for immunization. Keratinocytes can drive skin inflammation by producing pro-inflammatory mediators that promote cutaneous immune responses. Thus, targeting accessible keratinocytes to express transgenic antigens and increase production of pro-inflammatory mediators is a salient and attractive approach to vaccination, but remains a critical challenge to achieve. Toward this goal, we evaluated the effects of transient overexpression of the stress response factor x-box binding protein 1 (XBP1) in keratinocytes on the skin microenvironment. In vitro, keratinocytes genetically engineered to overexpress XBP1 increased production of both co-transduced exogenous antigen and pro-inflammatory mediators. In vivo, overexpression of XBP1 and transgenic antigen in keratinocytes induced both protective and therapeutic antitumor immunity that was associated with enhanced cutaneous expression of pro-inflammatory mediators and co-delivered antigen, increased skin-infiltrating immunocytes including dendritic cells (DCs), and the induction of durable antigen-specific CD8+ T cells and skin-resident memory CD8+ T cells. Importantly, in human skin, correlative effects were observed: transient overexpression of XBP1 in skin triggered the expression of genes linked to a pro-immunogenic microenvironment, increased the expression of the co-delivered antigen, and promoted the activation and IFN-γ-producing T cell-stimulating function of skin DCs. These findings provide novel insights into the rational design of cancer vaccines with improved immunogenicity through strategic modulation of the skin microenvironment.

Citation Format: Jack R. You, Rong Hao, Louis D. Falo, Guo Chen, Xingxing Hao, Cara Carey, Zhaoyang You, Louis D. Falo. Targeting the skin microenvironment for effective cancer vaccines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 725.
Abstract 731: A minigene platform to validate novel immunogenic peptides arising from somatic mutations as therapeutic cancer vaccines
Authors: Shi Xiaoshan , Kode Vasumathi , Lee Kayla , Chaudhuri Amit , Chakraborty Papia
Published: August 2018

Cancer vaccines have shown promise in treating late-stage cancers that have failed traditional therapies. Somatic mutations provide a rich source of potential cancer vaccines with minimal T-cell tolerance. We have built a robust screening assay to test somatic mutations for CD8+ T cell activation in a T cell-dendritic cell co-culture assay with synthetic peptides added from outside. Our screening assay revealed that non-mutated wild-type peptides in many instances induced T-cell activation, although their HLA binding affinity was weak. We suspect that addition of high concentration of the peptides from outside may overcome their weak HLA-binding property, allowing them to bind HLA and activate T cells. To reduce the bias of artificial HLA presentation, we have built a minigene platform to screen pairs of wild-type and mutant peptides to assess their immunogenicity. We applied our proprietary cancer vaccine prediction platform to a set of somatic single-nucleotide variants (SNVs) and indels and selected immunogenic peptides by a combination of features that include TCR binding, HLA binding, gene expression and proteasomal processing. Predicted mutant immunogenic peptides and their wild-type counterpart were cloned as minigenes and expressed in HLA restricted antigen-presenting cells (APCs). The minigene-expressing APCs were co-cultured with allogenic CD8+ T cells and activation was monitored by the combined expression of IFN-gamma, TNF-alpha and granzyme-A (CTL phenotype of T cells) by flow cytometry. Our screening assay reveals that a significant number of indels are not presented when they are expressed as a longer peptide because of the stability of the transcripts. However, their presentation and immunogenicity are preserved when they are expressed as short 9-mer peptides with engineered proteasomal cleavage sites. Our study also reveals a range of HLA-binding affinities for peptide presentation. In conclusion, our analysis demonstrates that the two approaches for investigating immunogenicity of peptides—minigene approach and external addition of peptide approach—have differential utilities for testing and validating the immunogenicity of indels and SNVs.

Citation Format: Xiaoshan Shi, Vasumathi Kode, Kayla Lee, Amit Chaudhuri, Papia Chakraborty. A minigene platform to validate novel immunogenic peptides arising from somatic mutations as therapeutic cancer vaccines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 731.
Abstract 745: Development of a novel ron receptor targeted antibody-drug conjugates using cysteine bridging technology for potential treatment of pancreatic cancer
Authors: Suthe Sreedhar Reddy , Yao Hang-Ping , Trippier Paul C. , Wang Ming-Hai
Published: August 2018

Therapeutics targeting known oncoproteins have been applied for pancreatic cancer treatment but clinical outcomes are not promising. Hence, there is an urgent need to identify novel targets and develop effective drugs to improve pancreatic cancer therapeutic index. Antibody-drug conjugates (ADC) represent a promising class of drugs for targeted cancer therapy. Here we developed a novel ADC targeting RON receptor tyrosine kinase for potential pancreatic cancer treatment. To this end, we have synthesized a bis-alkylating linker (BL), attached to a lysosomal protease-cleavable dipeptide with payload Monomethyl auristatin E (MMAE). The BL-MMAE was then conjugated to Zt/g4 (anti-RON mAb) through cysteine bridging technology to produce Zt/g4-BL-MMAE with a homogeneous conjugation profile and an antibody to drug ratio of 1:4. Zt/g4-BL-MMAE showed significant improvement in drug conjugation homogeneity and serum stability over conventional ADCs prepared through maleimide based linkers. In pancreatic cancer cell lines overexpressing RON, Zt/g4-BL-MMAE specifically targeted RON-expressing tumor cells and was effective in rapid induction of cell surface RON endocytosis. Functional analysis revealed that Zt/g4-BL-MMAE caused cell cycle arrest at G2/M phase, reduction of cell viability and subsequently resulted in massive cell death. The calculated IC50 is in the range of 1 to 2 µg/ml. We conclude that Zt/g4-BL-MMAE is a novel anti-RON ADC with excellent conjugation profile, serum stability, and selective cytotoxicity for pancreatic cancer cells. This work provides a pharmaceutical opportunity for evaluating potentials of RON-targeted ADCs in pancreatic cancer treatment in the future.

Citation Format: Sreedhar Reddy Suthe, Hang-Ping Yao, Paul C. Trippier, Ming-Hai Wang. Development of a novel ron receptor targeted antibody-drug conjugates using cysteine bridging technology for potential treatment of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 745.
Abstract 750: Modular coiled-coil masking domains for tumor-specific antibody activation
Authors: Trang Vivian H. , Zhang Xinqun , Dominguez Melissa M. , Zeng Weiping , Cochran Julia H. , Stone Ivan J. , Senter Peter D. , Levengood Matthew R.
Published: August 2018

The power of monoclonal antibody therapeutics arises from their remarkable selectivity for a particular antigen. However, antibody therapies can still be limited by serious side effects that result from target-mediated toxicity when the antigen is also expressed on non-diseased tissue. An emerging concept in the field of therapeutic antibodies is to restrict antibody binding in healthy tissues while empowering the antibody at sites of disease. This has previously been accomplished by fusing masking groups to the antibody through cleavable sequences that can be activated upon hydrolysis by disease-associated proteases. Here, we present a novel approach to mask antibodies utilizing a variety of leucine zipper coiled-coil peptides fused to the N-termini of antibody heavy and light chains. An optimized heterodimeric coiled-coil masking domain was identified that significantly impaired the binding of multiple antibodies to their cognate receptors with no antibody-specific optimization required. Protease-cleavable linkers were introduced between the coiled-coil peptides and antibody termini, and protease-catalyzed removal of the coiled-coil domains restored antibody binding affinity to that of the unmasked parent antibody. The coiled-coil mask did not impair production of antibody-drug conjugates (ADCs) of varying drug-to-antibody ratios, and masked antibodies and ADCs had similar stability and pharmacokinetic profiles to unmasked counterparts. However, coiled-coil masked ADCs and antibodies displayed decreased functional activity in biological assays, commensurate with the decrease in affinity afforded by masking, and this activity could be restored upon cleavage of the mask. Lastly, coiled-coil masked antibodies and ADCs had been tested in vivo and had demonstrated improved safety benefits as well as equivalent antitumor activity compared to the unmasked counterparts. Coiled-coil masked antibodies represent an advance in the field of antibody pro-drugging, as this modular domain can be rapidly applied to an array of therapeutic antibodies and ADCs.

Citation Format: Vivian H. Trang, Xinqun Zhang, Melissa M. Dominguez, Weiping Zeng, Julia H. Cochran, Ivan J. Stone, Peter D. Senter, Matthew R. Levengood. Modular coiled-coil masking domains for tumor-specific antibody activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 750.
Abstract 761: The role of PI3 kinase pathway in the the skin of Sezary syndrome
Authors: Cristofoletti Cristina , Bresin Antonella , Picozza Mario , Picchio Maria Cristina , Passarelli Francesca , Monzo Francesca , Citterich Mauro Helmer , Monopoli Alessandro , Benucci Roberto , Cantonetti Maria , Frezzolini Alessandra , Scala Enrico , Caprini Elisabetta , Russo Giandomenico , Narducci Maria Grazia
Published: August 2018

Sezary Syndrome (SS) is an aggressive variant of cutaneous T cell lymphoma (CTCL) characterized by an accumulation of malignant T lymphocytes - the Sezary (SS) cells - in the skin, lymph nodes and blood. Until now, most studies on the characterization of this lymphoma have been performed on circulating SS cells because of the easy availability of tumor material from this compartment. For this reason, the role of skin microenvironment has not been characterized although it might play a fundamental role in the pathogenesis and in the disease progression of this neoplasia. According to this view, we recently demonstrated that PTEN downregulation, due to genetic loss and microRNA deregulation, commonly observed in SS patients, mainly activates AKT in skin-resident respect to blood-derived SS cells. This data underline how microenvironment signals amplify genetic abnormalities. Based on these observations, we investigated the effect of the skin microenvironment on SS cells in vivo. Comparing skin and blood-derived SS cells concurrently obtained from SS patients, we demonstrate that skin milieu promotes proliferation and PI3K/AKT/mTORC1 activation, a pathway that we prove to have multiple copy number (CN) alterations in our cohort of forty-three patients, thus indicating these genetic lesions as valid therapeutic targets to treat SS. At the end, we also demonstrate that the progressive increase of these CN alterations is associated to the shortening of overall survival of SS patients.

Citation Format: Cristina Cristofoletti, Antonella Bresin, Mario Picozza, Maria Cristina Picchio, Francesca Passarelli, Francesca Monzo, Mauro Helmer Citterich, Alessandro Monopoli, Roberto Benucci, Maria Cantonetti, Alessandra Frezzolini, Enrico Scala, Elisabetta Caprini, Giandomenico Russo, Maria Grazia Narducci. The role of PI3 kinase pathway in the the skin of Sezary syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 761.
Abstract 768: Identification of signal transduction pathways specific for mutant KRas-dependent tumors
Authors: Puli Oorvashi Roy , Yang Hua , Fang Bin , Jiang Hong Yuan (Rays) , Koomen John , Sebti Said M.
Published: August 2018

Targeting KRas directly has proven difficult. Therefore, identifying vulnerabilities specific for mutant KRas tumors is an important alternative approach to target mutant KRas-driven tumors. Towards this goal, we used phosphoproteomics to dissect key differences in signaling pathways between mutant KRas-dependent and mutant KRas-independent human cancer cells. Mass spectrometry analysis led to the identification of 25 proteins with significantly higher (up to 410 fold) phosphorylation levels in mutant KRas-dependent cells. Among these, 25 proteins- 8, 8 and 3 were tyrosine (Y), Serine (S) and Threonine (T) phosphorylated, and 6 were phosphorylated on both S and T. Presently, CRISPR-Cas9 knockout and siRNA depletion studies are ongoing to identify which of these 25 hits are required for the survival of mutant KRas-dependent human cancer cells. These studies can lead to identifying signaling circuits that can be targeted to treat mutant KRas-driven cancers.

Citation Format: Oorvashi Roy Puli, Hua Yang, Bin Fang, Hong Yuan (Rays) Jiang, John Koomen, Said M. Sebti. Identification of signal transduction pathways specific for mutant KRas-dependent tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 768.
Abstract 773: A heterochromatin gene signature unveils HP1α mediating neuroendocrine prostate cancer development and aggressiveness
Authors: Ci Xinpei , Hao Jun , Dong Xin , Choi Stephen Y. , Xue Hui , Qu Sifeng , Fazli Ladan , Crea Francesco , Ong Christopher , Zoubeidi Amina , He Housheng H. , Gleave Martin E. , Collins Colin C. , Lin Dong , Wang Yuzhuo
Published: August 2018

Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer (PCa) arising mostly from adenocarcinoma via NE transdifferentiation following androgen deprivation therapy. However, mechanisms contributing to both NEPC development and its aggressiveness remain elusive. In light of the fact that hyperchromatic nuclei is a distinguishing histopathological feature of NEPC, we identified 36 heterochromatin-related genes that are significantly enriched in NEPC by transcriptomic analyses of our patient-derived xenograft (PDX) models, multiple clinical cohorts, and genetically engineered mouse models. Longitudinal analysis using our unique, first-in-field PDX model of adenocarcinoma-to-NEPC transdifferentiation revealed that, among those 36 heterochromatin-related genes, heterochromatin protein 1α (HP1α) expression is increased early and steadily during NEPC development and remain elevated in the developed NEPC tumor. Its elevated expression is further confirmed in multiple PDX and clinical NEPC samples. Functional studies showed that HP1α knockdown in the NCI-H660 NEPC cell line dramatically inhibits proliferation, completely ablates colony formation, and induces apoptotic cell death, ultimately leading to tumor growth arrest. Its ectopic expression significantly promotes NE transdifferentiation in adenocarcinoma cells subjected to androgen deprivation treatment. Mechanistically, HP1α reduces expression of androgen receptor (AR) and RE1 silencing transcription factor (REST), two crucial transcription factors silenced in NEPC, and enriches the repressive histone mark trimethylation of histone H3 at Lys9 (H3K9me3) on their respective gene promoters. These observations indicate a novel mechanism underlying NEPC development mediated by abnormally expressed heterochromatin genes, with HP1α as an early functional mediator and a potential therapeutic target for NEPC prevention and/or management.

Citation Format: Xinpei Ci, Jun Hao, Xin Dong, Stephen Y. Choi, Hui Xue, Sifeng Qu, Ladan Fazli, Francesco Crea, Christopher Ong, Amina Zoubeidi, Housheng H. He, Martin E. Gleave, Colin C. Collins, Dong Lin, Yuzhuo Wang. A heterochromatin gene signature unveils HP1α mediating neuroendocrine prostate cancer development and aggressiveness [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 773.
Abstract 806: KI2011 is a novel potent FLT3 tyrosine kinase inhibitor in acute myeloid leukemia cells with FLT3-mutation
Authors: Ryu Hwani , Song Jie-Young , Ahn Jiyeon
Published: August 2018

More than 30% of patients with acute myeloid leukemia (AML) harbor a mutation with internal tandem duplication (ITD) in the juxtamembrane domain and point mutations in the tyrosine kinase domain of FMS-like tyrosine kinase-3 (FLT3), which results in aberrant activation FLT3 by auto-phosphorylation, and confers poor prognosis. To develop novel FLT3 inhibitors, ten novel oxazole amine derivatives were designed and synthesized based on FLT3 protein structure. Among the synthesized small molecule compounds, KI2011 exhibited selectively inhibitory activities of the class III receptor tyrosine kinases, especially FLT3, wile-type, FLT3-D835Y mutant and FLT3-internal tandem duplication (ITD) mutant isoforms with IC50 values of 25, 9, and 1.2 nM, respectively, determined by an in vitro kinase activity assay. The compound decreased FLT3 phosphorylation and its downstream signaling in MV4-11 cells harboring FLT3-ITD mutation. Treatment of KI2011 selectively inhibited cell proliferation and induced apoptosis of MV411 cells in a concentration-dependent manner, but not in K562 (FLT3 negative) cells. In addition, we found that KI2011 enhanced radiosensitivity of MV4-11 cells. The results suggest that KI2011 is a novel potent FLT3 inhibitor that may be further developed in the preclinical studies as a therapeutic agent in AML treatments.

Citation Format: Hwani Ryu, Jie-Young Song, Jiyeon Ahn. KI2011 is a novel potent FLT3 tyrosine kinase inhibitor in acute myeloid leukemia cells with FLT3-mutation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 806.
Abstract 815: New approach for old target: W0101 antibody drug conjugate effectively inhibits tumor growth in preclinical models of IGF-1R overexpressing solid tumors
Authors: Akla Barbara , Loukili Noureddine , Robert Alain , Beau-Larvor Charlotte , Malissard Martine , Haeuw Jean-Francois , Beck Alain , Perez Michel , Dreyfus Cyrille , Pavlyuk Mariya , Chetaille Eric , Corvaia Nathalie
Published: August 2018

Insulin-like growth factor type 1 receptor (IGF-1R) has been recognized for decades for its role in tumorigenesis and growth1. In addition, overexpression of this receptor has been largely documented in numerous tumor tissues. However, so far, previous therapeutic approaches based on monoclonal antibodies and tyrosine kinase inhibitors failed to show clinical benefit in the overall patient population2. Here we disclose for the first time a novel IGF-1R targeted Antibody Drug Conjugate (ADC), W0101, designed to deliver highly potent cytotoxic drugs selectively to IGF-1R over-expressing tumor cells.

The naked antibody for our ADC was selected on the basis of its specific binding properties to IGF-1R compared to insulin receptor (IR), and for its internalization properties.

After coupling of a proprietary auristatin derivative to the naked antibody, neither the binding nor the internalization properties of W0101 evaluated by FACS analysis, were modified.

Interestingly, the capacity of W0101 to induce cytotoxicity was evaluated in vitro in various cell lines and was demonstrated to be independent of effector functions.

Mouse efficacy studies with various doses and schedules of administration were conducted in several models expressing different level of IGF-1Rin order to determine their sensitivity to W0101. The sensitivity of W0101 was correlated to the expression level of IGF-1R evaluated by IHC. And in a 3+ MCF-7 breast cancer model, a single injection of W0101 at 3mg/kg led to more than 90% tumor growth inhibition (TGI), associated with a modulation of the IGF-1R expression on the tumor cells (IHC study).

The first in-human trial of W0101 is currently on going to address clinical safety. We believe that W0101 will bring a new therapeutic option for patients that overexpress IGF-IR.

1Pollak M. Insulin and insulin-like growth factor signaling in neoplasia. Nat Rev Cancer, 2008,8:915-928.

2Corvaia N, Beck A, Caussanel V, Goetsch L. Insulin-like growth factor receptor type I as a target for cancer therapy. Frontiers in Bioscience, Scholar, 5, 439-450, January 1, 2013

Citation Format: Barbara Akla, Noureddine Loukili, Alain Robert, Charlotte Beau-Larvor, Martine Malissard, Jean-Francois Haeuw, Alain Beck, Michel Perez, Cyrille Dreyfus, Mariya Pavlyuk, Eric Chetaille, Nathalie Corvaia. New approach for old target: W0101 antibody drug conjugate effectively inhibits tumor growth in preclinical models of IGF-1R overexpressing solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 815.
Abstract 82: Epigenetic reader ZMYND8 bridges BRD4 and hypoxia-inducible factors to mediate breast cancer progression and metastasis
Authors: Chen Yan , Zhang Bo , Bao Lei , Jin Lai , Yang Mingming , Peng Yan , Wang Jennifer , Wang Chenliang , Zou Xuan , Wang Yingfei , Luo Weibo
Published: August 2018

Metastatic breast cancer has high rates of relapse and mortality. The absence of definitive prognostic biomarker and effective therapeutic target is the major obstacle to end this aggressive disease. Epigenetic dysregulation plays a crucial role in breast cancer metastasis. However, the mechanism by which epigenetic dysregulation stimulates the hypoxia-mediated breast cancer progression and metastasis remains unknown.

Based on luciferase reporter assays and analysis of expression changes of 720 epigenetic genes in three microarray datasets, we identified epigenetic reader ZMYND8 as a novel hypoxia-inducible factor (HIF) target gene. Human breast cancer tissue microarray data indicated that ZMYND8 is highly expressed in invasive breast tumors and this overexpression is significantly correlated with poor clinical outcomes in patients with breast cancer. ZMYND8 depletion dramatically attenuates the tumorigenic potential of breast cancer cells in colony formation, migration and invasion in vitro, and suppresses breast tumor growth and metastasis in mice. To elucidate the underlying mechanism, we performed co-immunoprecipitation, chromatin immunoprecipitation and quantitative PCR assays and found that ZMYND8 interacts with HIF complex and acetyl lysine 16 of histone H3 at the hypoxia inducible elements (HREs) to provoke the expression of HIF target genes LOX, AGR2, AQP1 and VEGFA. ZMYND8 controls breast cancer growth and metastasis through HIF in mice. We further found that p300 binds and acetylates ZMYND8 in breast cancer cells. Acetylated ZMYND8 interacts with and recruits BRD4 to the HREs to stimulate RNA polymerase II phosphorylation, thereby promoting transcriptional elongation of HIF target genes. ZMYND8 acetylation is necessary and sufficient for breast tumor growth and metastasis in vitro and in mice. Together, p300-ZMYND8-BRD4-HIF axis is critical for breast cancer progression and metastasis.

In summary, ZMYND8 represents a positive feedback mechanism that amplifies HIF-mediated breast cancer progression and metastasis, and provides a potential epigenetic target for the prognosis and treatment of breast cancer.

Citation Format: Yan Chen, Bo Zhang, Lei Bao, Lai Jin, Mingming Yang, Yan Peng, Jennifer Wang, Chenliang Wang, Xuan Zou, Yingfei Wang, Weibo Luo. Epigenetic reader ZMYND8 bridges BRD4 and hypoxia-inducible factors to mediate breast cancer progression and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 82.
Abstract 822: Predicting response to kinase inhibitors based on tumor genomics in NF1-related MPNSTs
Authors: Grit Jamie , Pridgeon Matt , Tovar Elizabeth , Madaj Zachary , Peacock Jackie , Wulfkuhle Julia , Petricoin Emanuel , Graveel Carrie , Steensma Matt
Published: August 2018

Malignant Peripheral Nerve Sheath Tumors (MPNSTs) are aggressive, chemoresistant sarcomas that develop in patients with the genetic disorder Neurofibromatosis Type 1 (NF1). Loss of TRP53 and amplification of the receptor tyrosine kinase (RTK) MET is observed 25-50% of cases. To investigate the role of MET and p53 in MPNST progression, we developed three MPNST mouse models: an NF1 null and MET overexpression MPNST model (NF1-MET), an NF1 and TRP53 heterozygous knockout model (NF1-p53), and a NF1 null model (NF1). Using orthotopic tumorgrafts from these models, we tested the efficacy of MET (capmatinib) and MEK (trametinib) inhibition on MPNST growth. Trametinib moderately inhibited tumor growth in all of the models; however, combined inhibition of MEK and MET was highly effective in both the NF1-MET and NF1 models. NF1-p53 tumors were the least responsive to combined inhibition and upregulated AKT signaling in response to single agent therapy. These data suggest that alternate RTK signaling pathways downstream of MET may compensate for MEK inhibition, and that p53 loss promotes kinome reprogramming in response to targeted inhibition in MPNSTs. To determine the effects of MEK, MET, PI3K, and mTOR inhibition on kinome signaling, we generated MPNST cell lines from each of our MPNST models. Western blot analysis revealed that NF1-MET MPNST cells are highly sensitive to MET inhibition, NF1-p53 MPNST cells maintain ERK and AKT activity in the presence of MET inhibition, and NF1-p53 MPNST cells uniquely upregulate MEK signaling in response to PI3K/mTOR inhibition. These results suggest p53 plays a unique role in kinome reprograming and resistance to kinase inhibitors. Currently, we are determining the effects of combined kinase inhibition on proliferation of MPNST cells and utilizing reverse phase protein arrays to evaluate the effect of MET and MEK inhibition on the phospho-proteome in our MPNST models. By understanding these complex signaling networks we will identify combinations of targeted therapies that can be used to effectively treat MPNSTs.

Citation Format: Jamie Grit, Matt Pridgeon, Elizabeth Tovar, Zachary Madaj, Jackie Peacock, Julia Wulfkuhle, Emanuel Petricoin, Carrie Graveel, Matt Steensma. Predicting response to kinase inhibitors based on tumor genomics in NF1-related MPNSTs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 822.
Abstract 826: Th1 cytokines and EGFR inhibition: A combinatorial therapeutic strategy in TNBC
Authors: Basu Amrita , Kodumudi Krithika , Wiener Doris , Czerniecki Brian
Published: August 2018

Oncodrivers are a promising target in novel breast cancer therapy development, owing to oncogene addiction of breast cancer cells to sustain their malignancy. We have previously shown progressive loss of CD4+ T-helper type 1 response to oncodrivers in triple negative breast cancer (TNBC) patients. TNBC is marked by lack of cell surface ER, PR and Her2 receptors and is the most aggressive subtype of breast cancer. Lack of therapeutic targets, resistance to existing hormone therapy, metastasis and poor survival in TNBC patients underline the need for novel TNBC therapy development. In this study, we investigated the effect of EGFR inhibitors (cetuximab, erlotinib and lapatinib) alone or combination with Th1 cytokines (IFN-γ and TNF-α) on growth and proliferation of MDA-MB-231 and MDA-MB-468 TNBC cells. We observed decrease in EGFR expression in TNBC cells when treated with IFN-γ alone and in combination with EGFR inhibitors. In addition, IFN-γ alone, in combination with TNF-α and all three EGFR inhibitors, markedly increased STAT1 phosphorylation, indicating suppression of growth and proliferation in TNBC cells. Decreased STAT3 phosphorylation by combination treatment in TNBC cells may induce apoptosis and inhibit proliferation further. Combination of cetuximab and IFN-γ increased cleaved caspase-3 expression in MDA-MB-231, but not in MDA-MB-468 TNBC cells. EGFR inhibition and Th1 cytokine treatment showed combination treatment resulted in severe cell loss and morphological alteration, while Th1 cytokines alone did not induce significant senescence in MDA-MB-231 cells in senescence associated β-galactosidase assay. We are currently investigating activation status of other signaling pathways in TNBC cells, following Th1 cytokines treatment with EGFR inhibition. Our data suggests a combinatorial treatment approach, including DC1 vaccination to elicit Th1 immune response and inhibition of EGFR oncodriver, may lead to an effective and novel therapeutic strategy for triple negative breast cancer.

Citation Format: Amrita Basu, Krithika Kodumudi, Doris Wiener, Brian Czerniecki. Th1 cytokines and EGFR inhibition: A combinatorial therapeutic strategy in TNBC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 826.
Abstract 836: ISU104, a fully human anti-ErbB3 antibody, overcomes acquired cetuximab resistance
Authors: Kim Miyoung , Hur Youngmi , Hong Mirim , Sohn Youngsoo , Shin Kum-Joo , Kim Kyungyong , Hong Seung-Beom , Shin Kum-Joo , Bae Donggoo
Published: August 2018

ErbB3 is noted as one of major causes of acquired cetuximab resistance in colorectal and head and neck cancers. ErbB3 causes activation of alternative signaling pathways that bypass the original target and sustained PI3K/AKT activation, and these are associated with cetuximab resistance. We confirmed the induced ErbB3 activation by the cetuximab treatment in FaDu head and neck squamous-cell carcinoma (HNSCC) xenograft model. Immunoblot analysis was shown that twice weekly 10 mg/kg of cetuximab treatment upregulated ErbB3 expression and phosphorylation even though tumor growth was well controlled. To investigate whether ErbB3 activation by ligand, heregulin (HRG,) might induce cetuximab resistance, two cetuximab-sensitive colorectal cancer cell lines, DiFi and LIM1215, were treated 0-25 ng/mL of HRG. HRG induced dose-dependent cetuximab resistance in cell proliferation assay, and this was reversed via ISU104 treatment. To evaluate if ISU104 could overcome resistance to cetuximab in vivo, acquired cetuximab-resistant FaDu xenograft model was established. Tumors that had acquired resistance to 5 mg/kg of cetuximab treatment were significantly regressed by replaced treatment of ISU104 alone (10 mg/kg) or combination treatment of ISU104 and cetuximab, while mice continued on cetuximab only showed uncontrolled tumor growth like vehicle-treated group. Our results suggest that ISU104 effectively overcomes cetuximab resistance and may provide clinical benefit to cetuximab-resistant patients.

Citation Format: Miyoung Kim, Youngmi Hur, Mirim Hong, Youngsoo Sohn, Kum-Joo Shin, Kyungyong Kim, Seung-Beom Hong, Kum-Joo Shin, Donggoo Bae. ISU104, a fully human anti-ErbB3 antibody, overcomes acquired cetuximab resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 836.
Abstract 837A: Combining CRISPR/Cas9 mutagenesis and a small-molecule inhibitor to probe the function of MELK in cancer
Authors: Giuliano Christopher J. , Lin Ann , Smith Joan C. , Palladino Ann C. , Sheltzer Jason M.
Published: August 2018

The Maternal Embryonic Leucine Zipper Kinase (MELK) has been identified as a promising therapeutic target in multiple cancer types. MELK over-expression is associated with aggressive disease, and MELK has been implicated in numerous cancer-related processes, including chemotherapy resistance, stem cell renewal, and tumor growth. Based on these preclinical results, the MELK inhibitor OTS167 is currently being tested as a novel chemotherapy agent in several clinical trials. Here, we report that cancer cell lines harboring CRISPR/Cas9-induced null mutations in MELK exhibit wild-type growth in vitro, under environmental stress, in the presence of cytotoxic chemotherapies, and in vivo. Furthermore, MELK-knockout lines remain sensitive to OTS167, suggesting that this drug blocks cell division through an off-target mechanism. By combining our MELK-knockout clones with a new highly-specific MELK inhibitor, we further demonstrate that the acute inhibition of MELK results in no specific anti-proliferative phenotype. Analysis of gene expression data from cancer patients identifies MELK expression as a correlate of mitotic activity, explaining its association with poor prognosis. In total, our results demonstrate the power of CRISPR/Cas9-based genetic approaches to investigate cancer drug targets, and call into question the rationale for treating patients with anti-MELK monotherapies.

Citation Format: Christopher J. Giuliano, Ann Lin, Joan C. Smith, Ann C. Palladino, Jason M. Sheltzer. Combining CRISPR/Cas9 mutagenesis and a small-molecule inhibitor to probe the function of MELK in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 837A.
Abstract 841: Protection of normal tissues by DIM (3,3'-Diindolylmethane) during radiation therapy
Authors: Chen Renxiang , Li Lijun , Lin Loretta Y. , Fornace Albert J. , Li Heng-Hong
Published: August 2018

96 Normal 0 false false false EN-US ZH-CN X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-fareast-language:EN-US;} Despite advances in precision targeting of tumors by radiation therapy, local recurrence is still a serious problem due to the inability to sometimes deliver a sufficiently high dose of radiation to achieve tumor control without unacceptable late normal tissue and organ toxicity. DIM (3,3'-diindolylmethane), a commercially available dietary supplement, has long been studied as a cancer prevention and anti-tumor agent. We are investigating DIM's differential effects after radiation in tumor and normal tissues using a mouse model approach. We monitored tumor growth after localized radiation in a tumor engraftment model of MDA-MB-231 human breast carcinoma cells using immunocompromised NSG mice. The fractionated regimen consists of a total dose of 24 Gy in four fractions (6 Gy per fraction given every other day) with administration of DIM or vehicle at one hour prior to each fraction. DIM showed no significant effect on the growth of irradiated tumors, i.e. DIM did not affect the treatment efficacy of radiation on tumors. On the other hand, we are investigating the molecular mechanism of DIM's radio-protective effects in normal tissues. Our transcriptomic data indicate that DIM treatment increases expression of some stress-responsive genes without causing DNA damage. Results to date indicate that DIM might delay radiation-induced cell cycle arrest and apoptosis. The ultimate goal is to develop DIM as a clinical radioprotector in order to improve the therapeutic index by allowing higher doses of radiation to improve local tumor control by reducing late dose-limiting normal tissue toxicity by radiation.

Citation Format: Renxiang Chen, Lijun Li, Loretta Y. Lin, Albert J. Fornace, Heng-Hong Li. Protection of normal tissues by DIM (3,3'-Diindolylmethane) during radiation therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 841.
Abstract 857: Metal oxide nanoparticles as adjuvant for radiation therapy
Authors: Wiesmann Nadine , Kluenker Martin , Tremel Wolfgang , Brieger Juergen
Published: August 2018

Background: Radiation therapy comprises a fundamental component of modern tumor treatment. Unfortunately, its success is limited by the development of radiation resistances. The emerging field of nanotechnology offers great opportunities for diagnosing, imaging, as well as treating cancer. Metal oxide nanoparticles in particular zinc oxide nanoparticles (ZnO-NP) have been shown to display a selective cytotoxic effect on tumor cells via a yet unknown mechanism. Most likely the generation of reactive oxygen species (ROS), breakdown of mitochondria and DNA damage are involved. The success of radiation therapy equally relies on the generation of ROS, which develop their cytotoxic potential by damaging DNA, proteins and membranes in a fashion that exceeds the repair capacity of tumor cells. Thus, our aim was to evaluate the applicability of ZnO-NP as radiosensitizer to restore the response of tumor cells to radiation therapy.

Methods: An AlamarBlueTM assay was used to evaluate the cellular metabolic activity (CMA) of A549 adenocarcinoma cells after treatment with ZnO-NP. To assess cellular viability and cell death a flow cytometric apoptosis assay was applied and genotoxicity of ZnO-NP was analysed by γH2AX foci analysis. The performance of ZnO-NP as radiosensitizer was assessed by a colony formation assay (CFA).

Results: We were able to show that ZnO-NP-mediated cytotoxicity is conveyed by dissolved Zn2+ ions as well as by the particles themselves. Treatment with ZnO-NP resulted in double-strand breaks (DSB) of the DNA measured by γH2AX foci analysis. The apoptosis assay could show induction of apoptosis in A549 cells after treatment with 100 µg/ml ZnO-NP for four hours. The colony formation assay showed that irradiation with 2 or 4 Gray, according to dosages that are typically applied in clinical settings, in combination with ZnO-NP treatment could enhance tumor cell death and reduce clonogenic survival. This revealed that ZnO-NP could improve the success of radiation therapy.

Conclusions: Our study proves that ZnO-NP exert a cytotoxic and genotoxic effect on human tumor cells. Additionally, we could show, that ZnO-NP are able to serve as radiosensitizer. Combined treatment of human tumor cells with ZnO-NP and irradiation resulted in reduction of tumor cell survival. Since nanoparticles can in principle be targeted selectively to tumor tissue, ZnO-NP could be used to specifically sensitize tumor cells for irradiation while sparing healthy tissue. All in all, the study shows that ZnO-NPs could be a promising antitumor agent.

Citation Format: Nadine Wiesmann, Martin Kluenker, Wolfgang Tremel, Juergen Brieger. Metal oxide nanoparticles as adjuvant for radiation therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 857.
Abstract 893: Hyperthermia regulates both SLC22A16 expression and ABCG2 expression via ROS production to enhance the cytotoxicity of doxorubicin
Authors: Matsui Hirofumi
Published: August 2018

Introduction: Hyperthermia (HT) is a non-invasive cancer therapy. Treatment temperature between 41°C to 44°C has no cytotoxic damage in normal cells, however shows cytotoxicity in cancer cells because of the underdeveloped vascular system. HT often used with other cancer therapy such as radiation therapy and chemotherapy. However mechanism of synergistic effect using these therapies remains unclear. Compared to 37°C, 42°C is mild heat stress for cells, thus superoxide anion is released from tissue. Superoxide anion is produced by mitochondrial electron transport chain. Reactive oxygen species (ROS), produced by mild heat stress, can be released from mitochondria. We have previously reported that ATP-binding cassette sub-family G member 2 (ABCG2) expression was suppressed by increasing mitochondrial ROS, andinduction of the cancer specific porphyrin accumulation. ABCG2 is a transporter of doxorubicin (DOX), therefore we hypothesized that synergistic effect of HT and chemotherapy would be induced by down-regulation of ABCG2 expression via intracellular ROS increase. In this study, we investigated if cytotoxic effect of breast cancer cell using DOX can be enhance by HT via intracellular ROS increase. Materials and methods: The murine breast cancer cell line, 4T1E was incubated at 37°C or 42°C for 1h. Intracellular ROS generation after HT treatment was detected by electron spin resonance (ESR). Twenty four hours after HT treatment, cells were incubated in medium containing 0, 0.1 and 1 μM DOX for 24 h. Cell viability was measured using the Cell Counting Kit 8, a water-soluble tetrazolium-8 based colorimetric assay. ABCG2 expression in whole cells was analyzed by Western blotting. Results and discussion: ESR signal peak with HT treatment became high as compared to without HT treatment, indicating intracellular ROS level was increased by HT treatment. Cell viability and ABCG2 expression were decreased by DOX exposure and by HT treatment. The enhancement of HT treatment effect by DOX is considered to be result of down-regulation of ABCG2 expression by ROS. Conclusion: HT treatment involved intracellular ROS production and down-regulated the expression of ABCG2 protein. HT treatment also enhanced the cell damage by DOX.

Citation Format: Hirofumi Matsui. Hyperthermia regulates both SLC22A16 expression and ABCG2 expression via ROS production to enhance the cytotoxicity of doxorubicin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 893.