bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1182: Heparin-binding epidermal growth factor-like growth factor is a pro-differentiating factor in neuroblastoma
Authors: Gaviglio Angela L. , Blobe Gerard C.
Published: July 2016

Neuroblastoma is the most common cancer in infancy. Current therapies are only modestly effective; patients with high-risk disease have less than a 50% chance of survival. High-risk neuroblastoma is characterized by undifferentiated neuroblasts and low Schwannian stroma content. The tumor stroma contributes to the suppression of tumor growth by releasing soluble proteins, including heparan sulfate proteoglycans (HSPGs), which act to promote neuroblast differentiation.

Here we identify heparin-binding epidermal growth factor-like growth factor (HBEGF) as a potent pro-differentiating factor in neuroblastoma. Using microarray analysis, HBEGF mRNA expression is decreased in neuroblastoma compared to benign disease, correlating to an increase in mortality. HBEGF protein is expressed only in stromal compartments of tumor specimens, with tissue from high-stage disease having decreased stroma and HBEGF. Soluble HBEGF increased neuroblast differentiation and decreased proliferation, while HBEGF knockdown decreased neuroblast differentiation. In patient samples, HBEGF expression correlates with SOX10, a neural crest differentiation marker, and the cell cycle inhibitor, CDKN1A. Alternatively, HBEGF negatively correlates with ASCL1, a primitive neuroectodermal marker, and the cell cycle promoter CCND1. HSPGs, including TβRIII, GPC1, GPC3, and SDC3 further promote the differentiating effects of HBEGF treatment by forming a complex with the epidermal growth factor receptor (EGFR) via glycosaminoglycan modifications, leading to activation of the Erk1/2 pathway and upregulation of the inhibitor of DNA binding transcription factor, whose expression correlates with HBEGF in patient samples. Inhibition of EGFR with erlotinib, lapatinib, and gefitinib diminish HBEGF-induced differentiation.

These data identify a novel member of the pro-differentiating secretome and support the use of heparan sulfate mimetics in neuroblastoma, while cautioning against the use of EGFR inhibitors for neuroblastoma treatment.

Citation Format: Angela L. Gaviglio, Gerard C. Blobe. Heparin-binding epidermal growth factor-like growth factor is a pro-differentiating factor in neuroblastoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1182.
Abstract 1196: Combination of neuregulin with EGFR activation signatures predict activity of the anti-ErbB3 antibody KTN3379 in SCCHN
Authors: Ligon Gwenda F. , Lillquist Jay S. , Seibel Scott B. , Wallweber Jerry , Neumeister Veronique , Rimm David L. , LaVallee Theresa M. , Alvarado Diego
Published: July 2016

ErbB3, the kinase-dead member of the EGFR/ErbB family, has been implicated as an oncogenic driver in a number of solid tumor types, including squamous cell carcinoma of the head and neck (SCCHN). However, clinical trials have demonstrated limited responses with certain single agent anti-ErbB3-directed therapies, suggesting that multiple ErbB inhibition and/or biomarker-directed patient selection may be beneficial. The anti-EGFR antibody cetuximab has demonstrated improved overall survival in SCCHN, but the overall response rate is relatively low. Several preclinical studies have suggested that pan-ErbB inhibition may result in improved clinical benefit and additionally, may overcome cetuximab resistance. Here we show that ErbB3 and its ligand neuregulin (NRG) are widely expressed in SCCHN, and that ErbB2, but not EGFR, drives ErbB3 activation in the presence of NRG. Although NRG is required for ErbB3 activation, it is not sufficient to fully predict activity of the anti-ErbB3 antibody KTN3379 in SCCHN models. Metadata analysis of human SCCHN tumors revealed that NRG expression is significantly associated with expression of EGFR ligands amphiregulin (AREG) (P<0.0001; R2 = 0.33) and transforming growth factor α (TGFα) (P<0.0001; R2 = 0.25) but not other EGFR ligands. We demonstrate using NRG-positive cell lines that KTN3379 antitumor activity correlates with the level of secreted AREG and TGFα. Using a proximity-based assay for EGFR homodimerization as a surrogate assay for ligand (AREG or TGFα)-induced EGFR activation, we show that high EGFR homodimer levels correlate with KTN3379 antitumor activity in NRG-positive SCCHN cell lines and primary tumor models in vitro and in vivo. These preclinical studies provide the molecular rationale for combination treatment with KTN3379 and cetuximab in SCCHN patients. We propose that an enrichment strategy for NRG expression and EGFR activation signature may result in better clinical activity in response to treatment with KTN3379 and cetuximab in SCCHN patients.

Citation Format: Gwenda F. Ligon, Jay S. Lillquist, Scott B. Seibel, Jerry Wallweber, Veronique Neumeister, David L. Rimm, Theresa M. LaVallee, Diego Alvarado. Combination of neuregulin with EGFR activation signatures predict activity of the anti-ErbB3 antibody KTN3379 in SCCHN. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1196.
Abstract 1211: Killing cancer one cell at a time: Development and characterization of a novel antibody drug conjugate
Authors: Xiao Michael , Chu Roger , Pagaduan Jayson , Robison Richard A. , O’Neill Kim L.
Published: July 2016

Thymidine Kinase 1 (TK1) is a protein that we have established to be present on the surface of cancer cells and absent on the surface of normal cells. Various methods have been used to confirm the presence of TK1 using localization methods such as scanning electron microscopy gold labeling, fluorescence imaging, and flow cytometry analysis. In this project, we explored the potential of using the TK1 marker as a therapeutic target by conjugating an anti-TK1 antibody to the toxin saporin, a ribosome inhibiting protein derived from the seeds of Saponaria officinalis. The toxin is widely used in the biological and neurological fields due to its stability and adaptability. An additional benefit to using this particular toxin is the inability of saporin to transverse the cell membrane without an antibody or other agent mediating internalization into the cell. We expressed a cysteine mutated form of saporin in the BL21(DE3) E. coli strain. Saporin from the expressed protein was subsequently isolated with a cation exchange cellulose column before verification on a non-reducing SDS-PAGE gel. A single band was observed at approximately 30 kD, indicating purity of the protein. After thiolation of the amine groups in the anti-TK1 antibody, the cysteine residues on the mutated saporin toxin was linked to the antibody through the formation of disulfide bonds. Further purification of the conjugate was achieved by eliminating unconjugated saporin and antibody, and high molecular weight aggregates using a Sephacryl S-200 column. Over 90% cancer cell death was observed in the Burkitt's Lymphoma Raji cell line when exposed to conjugate concentrations of 10 nM. Complete cell death was observed with exposure to 100 nM concentrations. In addition, there was a significant difference between the saporin conjugated treated and untreated samples (p < 0.001). Further tests are underway to determine the effectiveness of the conjugate in other cancer cell lines. These promising results suggest the use of saporin in an effective antibody drug conjugate therapeutic that could influence the way we treat cancer: a drug that will selectively target cancer cells.

Citation Format: Michael Xiao, Roger Chu, Jayson Pagaduan, Richard A. Robison, Kim L. O’Neill. Killing cancer one cell at a time: Development and characterization of a novel antibody drug conjugate. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1211.
Abstract 1218: A novel synergistic antibody pair targeting non-overlapping epitopes of MET effectively inhibits MET-driven cancer models
Authors: Grandal Michael M. , Poulsen Thomas T. , Koefoed Klaus , Eriksen Karsten W. , Dahlman Anna , Conrotto Paolo , Bouquin Thomas , Jacobsen Helle , Horak Ivan D. , Kragh Michael , Lantto Johan , Pedersen Mikkel W.
Published: July 2016

The receptor tyrosine kinase MET (Hepatocyte Growth Factor Receptor, HGFR) has been associated with development and progression of a range of human tumors due to its regulation of cell proliferation, migration, invasion, and angiogenesis. A subset of human tumors, particularly of lung or gastric origin, appear to have a primary dependency on MET, which is driven by alterations, such as MET-gene amplification, MET-exon 14 deletion, activating mutations, or autocrine HGF production. Furthermore, MET-amplification has been reported as a key mechanism of de novo resistance to EGFR targeting agents.

Sym015 is a mixture of two humanized monoclonal antibodies against non-overlapping epitopes on MET. The specific pair of antibodies was identified in a high-throughput cell based screen searching for antibody mixtures with superior growth inhibitory activity against four MET-dependent cell lines. Both antibodies bind the SEMA domain of MET with high affinity. Synergistic activity was confirmed by dose-response curves in several MET-dependent cell lines and cell line and patient-derived xenograft models. Mechanistically, Sym015 blocks HGF binding to MET, induces MET internalization and degradation effectively diminishing MET oncogenic signaling. Sym015 also induces higher levels of antibody-dependent cell-mediated cytotoxicity (ADCC) in vitro compared to individual mAbs.

In conclusion, Sym015, a novel monoclonal antibody mixture against MET, shows strong inhibitory activity against MET driven cell lines and xenografts due to a combined effect of MET degradation and secondary effector functions. These data support initiation of clinical trials.

Citation Format: Michael M. Grandal, Thomas T. Poulsen, Klaus Koefoed, Karsten W. Eriksen, Anna Dahlman, Paolo Conrotto, Thomas Bouquin, Helle Jacobsen, Ivan D. Horak, Michael Kragh, Johan Lantto, Mikkel W. Pedersen. A novel synergistic antibody pair targeting non-overlapping epitopes of MET effectively inhibits MET-driven cancer models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1218.
Abstract 1231: Novel chromene analogs as small-molecule microtubule destabilizers for the treatment of chemo-resistant ovarian cancer
Authors: Kulshrestha Arpita , Ibrahim Safaa A. , Katara Gajendra K. , Patil Renukadevi , Patil Shivaputra , Beaman Kenneth D.
Published: July 2016

Ovarian cancer is the most lethal gynecological malignancy with a 5-year survival rate of less than 40%, primarily because of treatment failure due to emergence of chemo-resistance to standard agents. This indicates a dire need for new treatments to improve survival rates. The tubulin dynamics is a promising target for new chemotherapeutic agents. In continuation of our efforts, using medicinal chemistry approach, we recently identified small molecular inhibitors of tubulin polymerization called chromenes, as new ovarian anti-cancer agents. A set of chromene compounds were screened for anti-cancer effect on cisplatin sensitive (A2780, SKOV-3, TOV112D) and cisplatin resistant (OVCAR-3, A2780-cisR, TOV112D-cisR) ovarian cancer cells. In vitro cell viability was assessed by fluorescence based Alamar Blue assay. Our preliminary studies identified SP-6-27 (IC50 range: 0.102± 0.006 - 2.24±0.056 μM) and SP-6-37 (IC50 range: 0.258± 0.159 - 3.61±0.049 μM) as most potent chromene analogs. Both compounds exhibited a potent anticancer activity towards both cisplatin sensitive (SP-6-27 IC50: 0.14± 0.03 μM; SP-6-37 IC50: 1.35± 0.88 μM) and cisplatin resistant (SP-6-27 IC50: 0.81± 1.2 μM; SP-6-37 IC50: 2.09± 1.7 μM) cell lines. The compounds exhibited least cytotoxicity towards normal ovarian epithelial cells (SP-6-27- IC50: 83.35 ± 9.47 μM; SP-6-37- IC50: 75.77 ± 5.37 μM). Additionally, the analysis of apoptotic changes as determined by Annexin-V assay revealed an enhanced apoptosis in both cisplatin sensitive and resistant cells upon treatment with compounds SP-6-27 or SP-6-37. Together, the data demonstrates that ovarian cancer cell lines respond sensitively to SP-6-27 and SP-6-37, demonstrating that the chromene scaffold is effective in suppressing ovarian cancer cell growth. The findings indicate that the novel chromene analogs are a new class of chemotherapeutic agents that will offer advantages for the treatment of ovarian cancer.

Citation Format: Arpita Kulshrestha, Safaa A. Ibrahim, Gajendra K. Katara, Renukadevi Patil, Shivaputra Patil, Kenneth D. Beaman. Novel chromene analogs as small-molecule microtubule destabilizers for the treatment of chemo-resistant ovarian cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1231.
Abstract 1256: Multi-focal control of mitochondrial gene expression by oncogenic MYC provides effective therapeutic targets in cancer
Authors: Taylor Amanda , Adams Clare , Zhang Xiao-yong , Gennaro Victoria , Cotney Justin , Shadel Gerald , Cameron Craig , Eischen Christine , McMahon Steven
Published: July 2016

The MYC transcription factor is the most dramatically overexpressed gene product in human cancer. However, defining the MYC-driven transcriptome critical to malignant transformation has remained a challenge, in part because MYC controls several thousand genes. Consequently, little progress has been made in the identification of sensitive nodes downstream of MYC that might be targeted therapeutically in order to interrupt its oncogenic signaling. To overcome this impediment, we performed a screen for downstream MYC effector pathways that are selectively linked to MYC's functional properties. Via this strategy, we identified the mitochondrial RNA polymerase (POLRMT) as a direct downstream target of MYC. POLRMT induction by MYC regulates transcription of the mitochondrial genome, as well as mitochondrial DNA replication. Recent studies have shown that POLRMT also interacts with a mitochondrial ribosomal RNA methyltransferase TFB1M to regulate mitochondrial ribosome assembly. Notably, blocking induction of POLRMT (or TFB1M) by MYC converts the cellular response to MYC activation from enhanced cell cycle progression to apoptotic cell death. Of immediate clinical relevance, compounds blocking mitochondrial gene expression at any of several steps cause acute synthetic lethality with oncogenic levels of MYC. Among these compounds, the mitochondrial targeting antibiotic Tigecycline is FDA-approved and well-tolerated in patients. Tigecycline eliminates tumor formation and dramatically improves survival in our animal model of MYC-driven lymphoma. Evidence presented here demonstrates that uncoupling of MYC's nuclear and mitochondrial transcriptomes exposes a point of exquisite vulnerability in tumor cells. Although targeting MYC as a therapeutic strategy has proven to be challenging in the past, our identification of a new node of sensitivity in MYC-driven cancers offers a potential for broad therapeutic implications.

Citation Format: Amanda Taylor, Clare Adams, Xiao-yong Zhang, Victoria Gennaro, Justin Cotney, Gerald Shadel, Craig Cameron, Christine Eischen, Steven McMahon. Multi-focal control of mitochondrial gene expression by oncogenic MYC provides effective therapeutic targets in cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1256.
Abstract 1277: Neural crest-like gene FOXD1 plays a role in melanoma cell migration and invasion
Authors: Wu Huizi , Larribere Lionel , Weina Kasia , Knappe Nathalie , Gebhardt Christoffer , Umansky Viktor , Utikal Jochen
Published: July 2016

Melanoma is a highly aggressive skin cancer arising from transformed melanocytes. Despite tremendous efforts and considerable progress in clinical treatment, melanoma remains a deadly disease characterized by abundant treatment resistance due to high phenotype plasticity of melanoma cells. In order to evade therapy, melanoma cells can undergo phenotype switching from a differentiated state towards a dedifferentiated and more stem cell-like state. Recently, some melanoma cells with neural crest-like characteristics were shown to be able to sustain tumor growth and to give rise to distant metastases.

Here, we use the technique of directed differentiation of human induced pluripotent stem cells (hiPSCs) to characterize the human neural crest cell population in vitro and to identify potential genes that are reactivated during melanoma progression. We differentiated hiPSCs towards melanocytes via the neural crest stage. Expression analyses revealed an overexpression of the Forkhead Box D1 (FOXD1) transcription factor in neural crest cells and importantly, increased FOXD1 expression was also found in melanoma cells compared to normal human melanocytes. Interestingly, in invasive and less proliferative melanoma cells, FOXD1 expression was elevated compared to highly proliferative and less motile cells, indicating a potential role of this factor in motility acquisition. To gain deeper insight into the expression pattern of FOXD1 in clinical samples, a protein expression analysis using tissue microarrays was performed determining FOXD1 expression in melanocytic nevi, primary and metastatic melanoma samples. Surprisingly, in benign melanocytic nevi, FOXD1 was dominantly localized in the nucleus whereas a shift towards a cytoplasmic localization was observed in both, primary and metastatic melanomas. Loss of FOXD1 gene in melanoma cells also revealed the importance of this gene in melanoma cell migration and invasion. Further characterization of FOXD1 and its role in melanoma progression may help to understand the phenotypic plasticity of melanoma cells, which is required for overcoming resistance in anti-melanoma therapy.

In conclusion, FOXD1 is overexpressed and aberrantly localized in malignant melanoma cells compared to melanocytes. Our data indicate its function in melanoma migration and invasion. FOXD1 might be an interesting target in melanoma therapy.

Citation Format: Huizi Wu, Lionel Larribere, Kasia Weina, Nathalie Knappe, Christoffer Gebhardt, Viktor Umansky, Jochen Utikal. Neural crest-like gene FOXD1 plays a role in melanoma cell migration and invasion. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1277.
Abstract 1298: CD40L-CD40 signaling on B-cell lymphoma response to BTK inhibitors
Authors: Sun Zhijian , Luo Lusong
Published: July 2016

The aberrant activation of B cells receptor (BCR) signaling plays an essential role in the pathogenesis of B-cell lymphomas. Targeting BCR signaling by inhibiting Bruton's tyrosine kinase (BTK) has been shown to be a successful strategy in treating B-cell lymphomas including mantle-cell lymphoma (MCL) and chronic lymphocytic leukemia (CLL). Ibrutinib, an oral, covalent BTK inhibitor achieved high overall response rate (ORR) and prolonged duration of responses in CLL and MCL patients. In contrast, ibrutinib was reported to have much lower response rate in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) and the responses in DLBCL was restricted to the activated B-cell (ABC) subtype. These differences highlight the need to understand the bypass signaling/pathway to BTK inhibition. In this study, we identified CD40L-CD40 pathway as one of the potential bypass pathways to BTK inhibitory therapy. We showed that CD40 was consistently expressed in mature B cells, whilst CD40L expression levels were heterogenic in tissue microarray (TMA) with 150 lymphoma samples. In these samples, circa 57% of lymphoma tissues have medium or strong level of CD40L expression in TMA, indicating the existence of CD40L-CD40 signaling in human lymphoma tissues. Consistent with the response rate of ibrutinib in patients, in 68 non-GCB type of DLBCL, in which aberrant activation of NF-kB pathway is important for the pathogenesis, about 60% samples showed CD40L expression. Co-culture of Rec-1 or TMD8 cells with HEK293/CD40L attenuated the anti-proliferation activity of ibrutinib in vitro. This observation was also confirmed with recombinant soluble CD40L protein. Detailed mechanistic analysis revealed that CD40L-CD40 signaling activated the NF-κB and MAPK pathway and offset the inhibition of ibrutinib. Additionally, activating NF-κB pathway with PKC agonist abolished the cell killing effects of ibrutinib. These data suggest CD40L may be used as a potential biomarker for studying clinical responses to BTK inhibitors. Additionally, our results support combination strategies of BTK inhibitors with agents blocking CD40 or downstream NF-κB and MAPK pathway in patient population where CD40L-CD40 pathway is activated.

Citation Format: Zhijian Sun, Lusong Luo. CD40L-CD40 signaling on B-cell lymphoma response to BTK inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1298.
Abstract 1326: 1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranoside a novel c-Myc inhibitor induces GNMT expression and apoptosis in hepatocellular carcinoma
Authors: Kant Rajni , Yen Chia Hung , Lu Chung Kuang , Tung Chien Yi , Chen Yi-Ming
Published: July 2016

1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranoside (PGG) possesses potent anticancer activities. We identified PGG as GNMT inducer by using GNMT based drug screening platform. In this study we aimed to find the mechanism of action of PGG in hepatocellular carcinoma (HCC). We used microarray analysis to identify the differently expressed genes induced by PGG. Microarray experiment showed that c-Myc was involved in all detected pathways in enrichment analysis. PGG suppressed c-Myc mRNA and protein expression in Huh7 and Hep G2 cells. Knocked-down of c-Myc resulted in significant induction of GNMT promoter activity and endogenous GNMT mRNA expression. Furthermore, overexpression of c-Myc significantly inhibited GNMT promoter activity and endogenous GNMT protein expression. Moreover, PGG not only inhibited c-Myc mRNA expression but also induced degradation of c-Myc protein in Huh7 cells. Finally we found that PGG induces apoptosis in Huh7 cells. Taken together, PGG is a novel c-Myc inhibitor and induces GNMT through c- Myc modulation in HCC. Our results also suggest that PGG may potentially be a good drug candidate for HCC and c-Myc overexpressing cancers.

Citation Format: Rajni Kant, Chia Hung Yen, Chung Kuang Lu, Chien Yi Tung, Yi-Ming Chen. 1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranoside a novel c-Myc inhibitor induces GNMT expression and apoptosis in hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1326.
Abstract 1329: Intravenous delivery of Erlotinib-loaded PLA-PEG nanoparticles for treatment of GB
Authors: Hartman Lauren K. , Dhruv Harshil D. , Householder Kyle T. , Roth Jeffrey E. , Schorzman Allison N. , Zamboni William , Sirianni Rachel , Berens Michael E.
Published: July 2016

Amplification, overexpression, and mutation of the epidermal growth factor receptor (EGFR) gene represent common genetic signature abnormalities (>50%) in primary glioblastomas (GBs) that arise de novo. Although activated EGFR signaling represents the most common genetic abnormality in GB, targeted therapeutics against EGFR, such as Erlotinib and Gefitinib, have found very limited clinical success mainly because of restricted access to the tumor cells behind the blood brain barrier. Nanoparticles, such as those composed of poly(lactic acid)-poly(glycolic acid) (PLA-PEG), may be capable of overcoming delivery barriers to expand the armamentarium of drugs useful against GB. PLA-PEG is one of only two polymer NP formulations in phase II clinical trial for intravenous administration (NCT01812746). In this study, we tested the ability of Erlotinib-loaded PLA-PEG nanoparticles to inhibit the EGFR signaling pathway in short-term cultures of GB patient derived xenograft (PDX) models. We also tested the ability of systemically administered PLA-PEG nanoparticles to deliver Erlotinib payload to intracranial GB PDX cells by pharmacokinetic (PK) analysis. Potency of Erlotinib-loaded PLA-PEG was determined in four PDX models (GBM6, GBM12, GBM39, and GBM59) with known EGFR and PTEN status using immunoblot analysis and CellTiterGlo® assay. Our results demonstrate that Erlotinib PLA-PEG inhibits EGFR signaling in all PDX models tested with potency equal to free Erlotininb (EC50 range 4.9 - 16.8 μM). Pharmacokinetic measurements were done in plasma and brain tissue samples (tumor-bearing hemisphere and contralateral hemisphere were analyzed separately) collected from intracranial GBM12 tumor bearing mice at 5 min, 1 hr, 6hr, and 24 hr after intravenous (IV) dose of free Erlotinib at 7.5 mg/Kg and Erlotininb PLA-PEG at 13.125 mg/Kg. PLA-PEG-mediated brain penetration (AUCplasm:AUCBrain) of Erlotinib is ∼7.5%. Using orthotopic PDX implants, perivascular/intratumoral penetration by PLA-PEG is being measured. Additional improvements in PLA-PEG to heighten uptake into brain, as well as tumor-specific targeting and enriched payload density will foster improved clinical utility of EGFR targeted therapies against GB. Supported by a grant from the Ben & Catherine Ivy Foundation.

Citation Format: Lauren K. Hartman, Harshil D. Dhruv, Kyle T. Householder, Jeffrey E. Roth, Allison N. Schorzman, William Zamboni, Rachel Sirianni, Michael E. Berens. Intravenous delivery of Erlotinib-loaded PLA-PEG nanoparticles for treatment of GB. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1329.
Abstract 1337: Exploration of 2-aryl fused thiophene analogs as selective inhibitors of MEK5
Authors: Flaherty Patrick T. , Shah Dhruv , Gupta Mohit , Wright Thomas , Cavanaugh Jane E. , Burow Matthew E.
Published: July 2016

In several cancers intermediate enzymes in the MAPK cascades are up-regulated and may represent adaptive changes giving these cancers a competitive metabolic edge over normal cells. Additionally these alterations may induce resistance to previously useful chemotherapeutic agents. The enzyme MEK5, a component of the MEK5/ERK5 cascade, is up-regulated in several cancer types including triple negative breast cancer and prostrate cancers. Our group has previously described the synthesis and testing of anthranillic acid derived compounds and their ring constrained analogs. To explore chemical functional group tolerance and preference on the central arene ring by MEK5, a series of fused-thiophene ring variations, obtainable by the Gewald reaction were prepared and tested. These compounds were examined at 10 μM with MDA-MB-231 cells treated with EGF to induce MEK1/2 and MEK5 phosphorylation. Western blot analysis using IR-tagged antibodies against pERK1/2, pERK5, ERK1/2, or ERK5 was used to determine total and relative inhibition of MEK-mediated phosphorylation of respective pERK products. Incubation with PD0325901 resulted in full suppression of both pERK1/2 and pERK5. For the variations explored, several novel compounds were identified that resulted in no inhibition in the formation of pERK1/2 and 70% inhibition of pERK5. This establishes a new structural class of selective Type III inhibitors of MEK5 that are more amenable to structural variation, facilitating additional structure activity relationship studies.

Citation Format: Patrick T. Flaherty, Dhruv Shah, Mohit Gupta, Thomas Wright, Jane E. Cavanaugh, Matthew E. Burow. Exploration of 2-aryl fused thiophene analogs as selective inhibitors of MEK5. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1337.
Abstract 1344: Novel targeting all-in-one nanoporphyrin platform against bladder cancer
Authors: Li Yuanpei , Lin Tzu-yin , Pan Chongxian , Lam Kit S.
Published: July 2016

We have developed an extremely versatile and highly innovative theranostic nanoporphyrin (NP) platform for the integration of a broad range of clinical relevant imaging and therapeutic functions, including magnetic resonance imaging (MRI), positron emission tomography, fluorescence optical tomography, photodynamic therapy, photothermal therapy and targeted drug delivery. NPs exhibit architecture-dependent fluorescence- and magnetic resonance- properties, which could significantly increase the sensitivity of optical imaging and MRI for tumor detection. We recently integrated the NP platform into our previously reported bladder cancer targeted nanoparticles that were coated with a bladder cancer-specific ligand named PLZ4, resulting in a novel bladder cancer targeting and multifunctional PLZ4-nanoporphyrin (PNP) platform. PNP selectively delivered photosensitizer and chemotherapy into bladder cancer cells 30-50 times more than to adjacent normal urothelial cells, compared to only 2-3 times with reported photosensitizer, 5-aminolevulinic acid (ALA). Photodynamic therapy with PNP generated reactive oxygen species was over 100 times more potent than 5-ALA in vitro. PNP-mediated intravesical photo-therapy completely eliminated orthotopic patient-derived bladder cancer xenografts (PDXs). Image-guided photodynamic and photothermal therapies synergized with targeted chemotherapy of doxorubicin and significantly prolonged overall survival of mice carrying PDXs. This uniquely engineered targeting PNP platform has tremendous potential to improve the management of bladder cancer in clinic.

Citation Format: Yuanpei Li, Tzu-yin Lin, Chongxian Pan, Kit S. Lam. Novel targeting all-in-one nanoporphyrin platform against bladder cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1344.
Abstract 1358: Structure-based drug design of 3-site binding, high affinity inhibitors of S100B in malignant melanoma
Authors: Cavalier Michael C. , Wilder Paul T. , Luci Diane , Maloney David J. , Fang Lei , Ansari Mohd. Imran , MacKerell Alexander D. , Coop Andrew , Jadhav Ajit , Weber David J.
Published: July 2016

An especially predictive biomarker when used in combination with other diagnostic indicators, S100B is elevated in >90% of malignant melanoma (MM) patients and its protein level correlates directly with poor survival (<1 yr.) and relapse. For patients with low levels of S100B (5-10%), MM vaccines are uniquely effective. While this correlation is not fully understood, mechanistic studies show that S100B is capable of blocking p53 oligomerization, promoting p53 phosphorylation in its C-terminus, and contributing to p53 degradation in concert with hdm2. Correspondingly, lowering S100B levels via siRNA or via small molecule inhibitors restore p53 levels and its tumor suppressor activities including UV-activated apoptosis pathways found in normal melanocytes. Additionally, elevated S100B and overexpression of the receptor for advanced glycation end products (RAGE) confers a metastatic phenotype. Studies into how S100B affects tumor progression and metastasis are ongoing, novel S100B inhibitors (termed SBiXs) to ablate S100B-dependent effects in cancer are in development. Structure-activity relationship (SAR) studies are ongoing to improve the affinity and selectivity of SBiXs. These studies reveal that the binding surface of S100B can be divided into “hot spots” or “persistent sites” that contribute more energetically to the protein-protein interactions (PPI) than the surrounding protein surface. Described here are descriptions of SBiXs which bind these Sites (1, 2, and 3), details to the SAR studies involved in the chemical optimization of these ligands, and information into the tethering of these SBiXs into a single-molecule capable of binding all 3 sites simultaneously. Also provided is a discussion regarding how protein dynamics affect the on/off rates of SBiXs and the resulting apparent binding affinity. The results described here represent a Structure-based drug design approach to the development of PPI inhibitors nearing the threshold required for a useful therapeutic window.

Citation Format: Michael C. Cavalier, Paul T. Wilder, Diane Luci, David J. Maloney, Lei Fang, Mohd. Imran Ansari, Alexander D. MacKerell, Andrew Coop, Ajit Jadhav, David J. Weber. Structure-based drug design of 3-site binding, high affinity inhibitors of S100B in malignant melanoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1358.
Abstract 1386: Clinical research results for a NGS-based kit for targeted detection of clinically relevant gene rearrangements in lung tumor samples
Authors: Schageman Jeoffrey J. , Costa José Luis , Sheils Orla , Glassco John E. , Chi David , Sherlock Jon , Bishop John , Petraroli Rosella P. , Bramlett Kelli S.
Published: July 2016

In recent years, advances in next-generation sequencing (NGS) technologies have enabled faster and cheaper methods for uncovering the genetic basis of disease. For cancer, NGS based screening for known tumor subtypes may inform diagnosis and allow the clinician to tailor a specific therapeutic approach in the future. Here, we present the testing results of one such NGS based kit used to detect specific chromosomal translocations in retrospective non-small cell lung cancer (NSCLC) samples by targeting specific breakpoints in known fusion transcripts.

The included panel tested consists of a single primer pool containing amplicon designs to simultaneously screen for over 75 specific rearrangements involving the receptor tyrosine kinase (RTK) genes ALK, RET and ROS1 as well as NTRK1. The panel was compatible with formalin-fixed paraffin-embedded (FFPE) lung tumor research samples and achieved high-sensitivity down to 10 ng of RNA input. In addition, amplicon assays designed at the 5’ and 3’ ends the RTK genes provide non-specific evidence that a translocation exists in a sample by comparing expression imbalance between the two ends.

Testing was carried out at three external clinical research laboratories. In addition to positive and negative control samples, each site contributed FFPE lung tumor research samples for which ALK fusion status was known prior to NGS library preparation carried out using the Ion AmpliSeq™ workflow. For site-specific samples (n = 144, 16 samples per sequencing run), high concordance, sensitivity and specificity were measured at 97.2%, 90.5% and 98.4%, respectively.

Citation Format: Jeoffrey J. Schageman, José Luis Costa, Orla Sheils, John E. Glassco, David Chi, Jon Sherlock, John Bishop, Rosella P. Petraroli, Kelli S. Bramlett. Clinical research results for a NGS-based kit for targeted detection of clinically relevant gene rearrangements in lung tumor samples. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1386.
Abstract 142: Mutation and immune profiles in early-stage lung squamous cell carcinoma
Authors: Choi Murim , Kadara Humam , Zhang Jiexin , Parra Cuentas Edwin , Rodriguez Canales Jaime , Gaffney Stephen G. , Zhao Zi-Ming , Behrens Carmen , Fujimoto Junya , Chow Chi-Wan , Kalhor Neda , Moran Cesar , Rimm David , Swisher Stephen , Gibbons Don L. , Heymach John V. , Kaftan Edward , Townsend Jeffrey , Lynch Thomas J. , Schlessinger Joseph , Lee J. Jack , Lifton Richard , Herbst Roy S. , Wistuba Ignacio I.
Published: July 2016

PURPOSE: Lung squamous cell carcinoma (LUSC) accounts for 20-30% of non-small cell lung cancers (NSCLCs). There are limited treatment strategies for LUSC in part due to our inadequate understanding of the molecular underpinnings of the disease. We sought to perform whole-exome sequencing (WES), comprehensive immune profiling and clinicopathological analysis of early-stage LUSCs to increase our understanding of the pathobiology of this malignancy.

METHODS: Matched pairs of surgically resected stage I-III LUSCs and normal lung tissues (n = 108) were analyzed by WES. Immunohistochemistry and image analysis-based profiling of 10 immune markers was done on a subset of LUSCs (n = 91). Associations among mutations, immune markers and clinicopathological variables were statistically examined using ANOVA and Fisher tests. Cox proportional hazards regression models were used for statistical analysis of clinical outcome.

RESULTS: This early-stage LUSC cohort displayed an average of 209 exonic mutations per tumor. Fourteen genes exhibited significant enrichment for mutation: TP53, MLL2, PIK3CA, NFE2L2, CDH8, KEAP1, PTEN, ADCY8, PTPRT, CALCR, GRM8, FBXW7, RB1 and CDKN2A. Among mutated genes associated with poor recurrence-free survival, MLL2 mutations predicted poor prognosis in both TP53 mutant and wild type LUSCs. We also found that in treated patients, FBXW7 and KEAP1 mutations were associated with poor response to adjuvant therapy, particularly in TP53-mutant tumors. Analysis of mutations with immune markers revealed that ADCY8 and PIK3CA mutations were associated with markedly decreased tumoral PD-L1 expression, LUSCs with PIK3CA mutations exhibited elevated CD45ro levels and CDKN2A-mutant tumors displayed an up-regulated immune response.

CONCLUSION: Our findings pinpoint mutated genes that may impact clinical outcome as well as personalized strategies for targeted immunotherapies in early-stage LUSC.

Citation Format: Murim Choi, Humam Kadara, Jiexin Zhang, Edwin Parra Cuentas, Jaime Rodriguez Canales, Stephen G. Gaffney, Zi-Ming Zhao, Carmen Behrens, Junya Fujimoto, Chi-Wan Chow, Neda Kalhor, Cesar Moran, David Rimm, Stephen Swisher, Don L. Gibbons, John V. Heymach, Edward Kaftan, Jeffrey Townsend, Thomas J. Lynch, Joseph Schlessinger, J. Jack Lee, Richard Lifton, Roy S. Herbst, Ignacio I. Wistuba. Mutation and immune profiles in early-stage lung squamous cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 142.
Abstract 1441: Systems-level interrogation of resistance mechanisms to immunotherapy through pooled shRNA screens
Authors: Wang Zhe , Malu Shruti , Peng Weiyi , McKenzie Jodi , Mbofung Rina , Williams Leila , Seth Sahil , Heffernan Tim , Hwu Patrick
Published: July 2016

Despite the impressive clinical efficacy of immunotherapy in some patients, many still do not respond or progress following an initial response. The molecular mechanisms underlying the tumor resistance in those non-responders remain largely undefined. To address this issue, we set out to perform high-throughput unbiased pooled shRNA screens to identify critical genes that confer immune resistance. Patient-derived melanoma cells were transduced with barcoded pooled lentiviral shRNA libraries that targeted the human kinome, followed by exposure to cytotoxicity mediated by autologous tumor infiltrating lymphocytes. Tumor cells were then subject to genomic deep sequencing and integrated analysis to identify depleted barcodes and corresponding genes. One identified candidate of particular interest is Aurora Kinase A (AURKA), a cell cycle regulator that has previously been shown to contribute to tumorigenesis and correlate with poor prognosis for cancer patients. Importantly, our independent screening with a bioactive compound library also implicated Aurora Kinase as an immune resistance candidate against T cell immunotherapy. Further studies showed that suppression of Aurora kinase activity with a pan inhibitor - AMG900, exhibited a synergistic cytotoxic effect with tumor infiltrating lymphocyte-mediated killing on autologous tumor cells from patients. Furthermore, our Nanostring analysis of tumor biopsies from patients that received adoptive T cell therapy revealed significantly increased expression of AURKA in tumors from non-responding patient when compared with responder counterparts. These results have substantiated the validity of our pooled shRNA screening platform. Further investigations are ongoing to elucidate the underpinnings of Aurora kinase-mediated tumor resistance to immunotherapy and to functionally and physiologically validate other putative targets we have identified.

Citation Format: Zhe Wang, Shruti Malu, Weiyi Peng, Jodi McKenzie, Rina Mbofung, Leila Williams, Sahil Seth, Tim Heffernan, Patrick Hwu. Systems-level interrogation of resistance mechanisms to immunotherapy through pooled shRNA screens. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1441.
Abstract 1455: IL-4 derived from T follicular helper cells in tumor draining lymph nodes regulate myeloid cell properties and anti-tumor immunity
Authors: Shirota Hidekazu , Klinman Dennis M. , Ito Shuku-ei A. , Ishioka Chikashi
Published: July 2016

Recent findings show that immune cells constitute a large fraction of the tumor microenvironment and modulate tumor progression. Clinical data indicate that chronic inflammation is present at tumor sites and that IL-4 in particular is up-regulated. Current results demonstrate that T follicular helper (Tfh) cells arise in tumor draining lymph nodes where they produce an abundance of IL-4. Deletion of IL-4 expressing Tfh cells improves anti-tumor immunity, delays tumor growth and reduces the immunosuppressive activity of myeloid-derived suppressor cells (MDSC) and M2 macrophages. These findings suggest that IL-4 from Tfh cells impact anti-tumor immunity and constitute an attractive therapeutic target to reduce immune suppression in the tumor microenvironment and thus enhance the efficacy of cancer immunotherapy.

Citation Format: Hidekazu Shirota, Dennis M. Klinman, Shuku-ei A. Ito, Chikashi Ishioka. IL-4 derived from T follicular helper cells in tumor draining lymph nodes regulate myeloid cell properties and anti-tumor immunity. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1455.
Abstract 1462: Non-inflammatory role of ASC-dependent inflammasomes in promoting gastric tumourigenesis via IL-18
Authors: Jenkins Brendan , Deswaerte Virginie , West Alison , Nguyen Paul , Putoczki Tracy
Published: July 2016

Introduction

Gastric cancer is the third most lethal cancer worldwide and represents a growing number of cancers linked with inflammation. However, the identity of innate immune regulators with oncogenic potential in the host gastric mucosal epithelium remains obscure. We aim to identify the molecular basis by which specific pattern recognition receptors belonging to the inflammasome family promotes gastric tumorigenesis.

Experimental procedures

We have employed the gp130F/F gastric cancer mouse model coupled with mice lacking specific inflammasome-related genes. In addition, human gastric cancer tumor (and matched non-tumor) biopsies and cell lines were used. Immunohistochemistry on gastric tissue sections was used to assess the extent of cellular proliferation, apoptosis, inflammation and angiogenesis, and quantitative real-time PCR was used to measure the expression of genes of interest. Immunoblotting and ELISA were used to measure the activation and expression levels of inflammasome-related proteins.

Results

A causal role for ASC-dependent inflammasomes in gastric tumorigenesis was confirmed upon the genetic ablation of ASC in gp130F/F:Asc-/- mice, which resulted in a ∼50% reduction in tumor burden compared to parental gp130F/F mice. The suppressed gastric tumorigenesis in gp130F/F:Asc-/- mice was associated with increased gastric epithelial (tumor) cell apoptosis, as determined by elevated numbers of TUNEL and caspase-8 positive cells. Surprisingly however, no changes in the number and activation status of inflammatory cells were observed. The suppressed gastric tumorigenesis in gp130F/F:Asc-/- mice was also characterized by a gene signature comprising apoptotic-related genes. In gastric tumor tissue, IL-18, but not IL-1β, protein levels were augmented compared to matched non-tumour tissue, and genetic ablation of IL-18 in gp130F/F:IL-18-/- mice suppressed gastric tumorigenesis comparable to that observed upon ASC deficiency. The treatment of human gastric cancer cells with IL-18 also potentiated their growth compared to IL-1β.

Conclusions

Collectively, these data reveal that ASC-dependent inflammasomes and their downstream mediator IL-18 represent a novel oncogenic mechanism in gastric cancer.

Citation Format: Brendan Jenkins, Virginie Deswaerte, Alison West, Paul Nguyen, Tracy Putoczki. Non-inflammatory role of ASC-dependent inflammasomes in promoting gastric tumourigenesis via IL-18. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1462.