bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 2341: In situ labelling identifies a novel landscape of nuclear CDK2 substrates
Authors: Clurman Bruce E. , Chi Yong 
Published: July 2017

Abstract
Cellular signal transduction pathways rely heavily on protein kinase-mediated phosphorylation events. Comprehensive identification of the physiologic targets of protein kinases is key to understanding their functions and elucidating cellular signalling pathways. Various direct and indirect approaches, combined with advanced mass spectrometry technologies, have made it possible to identify many candidate targets of a given kinase, but have also yielded many false-positive identifications, largely due to technical limitations. Identifying direct and physiological kinase targets thus remains a difficult task. We developed a mass spectrometry-based in situ approach that utilizes substrate thiophosphorylation and ATP analog-sensitive CDK2 (AS-CDK2) to identify over one hundred potential cyclin-dependent kinase 2 (CDK2) substrate in intact nuclei. More than one-third of these candidates are known CDK substrates, indicating significant enrichment for physiologic CDK2 substrates, and we also we identified and validated new CDK2 substrates. Importantly, many of these new CDK2 substrates are chromatin-associated proteins with roles in histone modification, DNA replication, and DNA repair, which likely reflects our use of AS-CDK2 within its normal nuclear context, and regulated by endogenous cyclins. These methods should be broadly applicable to the study of other kinases with complex substrate networks.
Citation Format: Bruce E. Clurman, Yong Chi. In situ labelling identifies a novel landscape of nuclear CDK2 substrates [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2341. doi:10.1158/1538-7445.AM2017-2341
Abstract 235: Probiotics ameliorate Porphyromonas gingivalis-promoted pancreatic cancer progression in oncogenic Kras transgenic mice
Authors: Jan Ming-Shiou , Chen Wan-Ting , Chen Yu-Jen , Lin Chia-Wei , Chang Wen-Wei , Tsai Chung-Hung , Peng Jia-Shiou , Hsu Li-Jin 
Published: July 2017

Abstract
Periodontal disease has been shown to play important roles in the pathogenesis of many chronic diseases. Recent epidemiological studies and microbiome analysis have suggested that periodontal disease may be associated with the progression of pancreatic cancer. Our recent study has demonstrated that oral-smear of a periodontitis pathogen Porphyromonas gingivalis (P. gingivalis) significantly induced pancreatic cancer deterioration in the KrasG12D/-Pdx-1-Cre transgenic mouse model. Higher expression levels of epithelial-mesenchymal transition (EMT) markers, such as N-Cadherin, αSMA, SNAIL-1, Vimentin and ZEB1, were observed in P. gingivalis-treated KrasG12D/-Pdx-1-Cre mice than the untreated controls. In an attempt to develop a prophylactic treatment for pancreatic cancer, the mice were fed with a combination of two probiotics, Lactobacillus reuteri and Lactobacillus paracasei, for four consecutive weeks after oral-smear of P. gingivalis. These probiotic strains have been demonstrated to have the efficacy of either anti-inflammation or enhancement of cell-mediated immunity, respectively. Our results demonstrated that oral administration of the probiotics significantly suppressed oncogenic Kras-induced pancreatic cancer development and reversed P. gingivalis-stimulated cancer deterioration in mice. Significantly reduced expression of the aforementioned EMT markers in pancreatic cancer tissues was observed in KrasG12D/-Pdx-1-Cre transgenic mice treated with the probiotics. Our results suggest that the microbiota profile is very important for pancreatic cancer development. The manipulation of microbiota for enhancing antitumor immunity may be an emerging strategy for cancer prevention.
Citation Format: Ming-Shiou Jan, Wan-Ting Chen, Yu-Jen Chen, Chia-Wei Lin, Wen-Wei Chang, Chung-Hung Tsai, Jia-Shiou Peng, Li-Jin Hsu. Probiotics ameliorate Porphyromonas gingivalis-promoted pancreatic cancer progression in oncogenic Kras transgenic mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 235. doi:10.1158/1538-7445.AM2017-235
Abstract 2350: In vivo E2F reporting on efficacious dosing schedules of MEK plus CDK4/6 inhibition in melanoma
Authors: Teh Jessica , Nikbakht Neda , Purwin Timothy , Chervoneva Inna , Patel Prem , Davies Michael , Aplin Andrew 
Published: July 2017

Abstract
Pharmacological targeting of cyclin dependent kinases 4 and 6 (CDK4/6) could represent a viable therapeutic option in combination with BRAF and/or MEK inhibitors in different genetic subsets of melanoma. Indeed, continuous and concurrent dosing of MEK inhibitor (MEKi) plus a CDK4/6 inhibitor (CDK4/6i) leads to melanoma regressions in in vivo models and delays the onset of MEKi resistance. Current scheduling in breast cancer patients for the CDK4/6i, palbociclib, is intermittent due to adverse effects resulting in neutropenia. It is unclear what the most efficacious schedules are of palbociclib plus a MEKi. Utilizing an E2F reporter system, we sought to analyze the efficacy of different CDK4/6i plus MEKi schedules in a quantitative and temporal manner. Intermittent dosing (3 weeks on/1 week off) of both CDK4/6i and MEKi combination therapy resulted in tolerant tumors and rapid reactivation of E2F activity during drug holiday. Continuous MEKi with intermittent CDK4/6i led to more complete responses as compared to continuous CDK4/6i with intermittent MEKi. Weight loss of mice was also evident in the continuous CDK4/6i plus intermittent MEKi arm suggesting adverse events related to continuous CDK inhibition. Importantly, functional proteomic analysis revealed distinct mechanisms of acquired resistance/drug tolerance that arose from the three scheduling arms. Here, we report that upregulation of S6 phosphorylation is associated with acquired resistance to MEK plus CDK4/6 inhibition and can be overcome with an mTOR inhibitor. Taken together, in vivo reporting allows for quantitative measurement of pathway activity associated with inhibitor resistance and can be utilized to optimize combination schedules to improve the therapeutic index in patients.
Citation Format: Jessica Teh, Neda Nikbakht, Timothy Purwin, Inna Chervoneva, Prem Patel, Michael Davies, Andrew Aplin. In vivo E2F reporting on efficacious dosing schedules of MEK plus CDK4/6 inhibition in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2350. doi:10.1158/1538-7445.AM2017-2350
Abstract 2351: The novel potential of palbociclib (CDK4/6 inhibitor) in the treatment of triple-negative breast cancer
Authors: Asano Yuka , Kashiwagi Shinichiro , Goto Wataru , Takahashi Katsuyuki , Takashima Tsutomu , Noda Satoru , Onoda Naoyoshi , Tomita Shuhei , Hirakawa Kosei , Ohira Masaichi 
Published: July 2017

Abstract
Background: The effectiveness of palbociclib (CDK4/6 inhibitor) for estrogen receptor positive breast cancer has been demonstrated by large-scale clinical studies, with the drug garnering attention as a key drug for breast cancer subtypes with endocrine sensitivity in the future. According to PALOMA-3 trial, palbociclib has been demonstrated to contribute to the extension of progression-free survival in patients with advanced hormone receptor-positive and HER2-negative metastatic breast cancer after endocrine therapy. On the other hand, in the case of triple-negative breast cancer (TNBC), luminal AR (LAR) related to androgen signaling is believed to have endocrine activity. Previous clinical data revealed that palbociclib shows high sensitivity in luminal breast cancer cell lines with endocrine activity, with effectiveness also expected in LAR. In this study, we created TNBC cell lines that forcibly express AR and examined the effectiveness of palbociclib for TNBC.
Materials and Methods: MCF-7 and T-47D were used as luminal breast cancer cell lines, while MDA-MB-231 and BT-549 were used as TNBC cell lines. In addition, we created TNBC cell lines that forcibly express AR, called AR-MDA-MB-231, by the transfection of pEGFP-C1-AR Plasmid Vector using Lipofectamine® 3000 Reagent. We confirmed the expression of AR by qRT-PCR as well as Western blotting and examined the impact of palbociclib on proliferation as well as apoptosis of breast cancer cell lines.
Results: AR was found to have been expressed only in luminal breast cancer cell lines but not TNBC cell lines. It was confirmed that AR was expressed in AR-MDA-MB-231 which are stable cell lines with the properties of LAR. In a CCK assay, palbociclib showed high sensitivity in AR-MDA-MB-231 as in luminal breast cancer cell lines. Furthermore, in an apoptosis assay using FACS and cell cycle assay, apoptosis was induced in AR-MDA-MB-231 and cell cycle arrest at the G1S check point was confirmed.
Conclusion: palbociclib (CDK4/6 inhibitor) showed effectiveness for TNBC cell lines that compulsively express AR, suggesting it may be one treatment option for TNBC in the future.
Note: This abstract was not presented at the meeting.
Citation Format: Yuka Asano, Shinichiro Kashiwagi, Wataru Goto, Katsuyuki Takahashi, Tsutomu Takashima, Satoru Noda, Naoyoshi Onoda, Shuhei Tomita, Kosei Hirakawa, Masaichi Ohira. The novel potential of palbociclib (CDK4/6 inhibitor) in the treatment of triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2351. doi:10.1158/1538-7445.AM2017-2351
Abstract 2360: Regulation of the MAPK phosphatase MKP-1 by the E3 ligase GP78 in cancer cells
Authors: Kho Dhonghyo , Wu Gen Sheng 
Published: July 2017

Abstract
Mitogen-activated protein kinase (MAPK) phosphatase 1 (DUSP1/MKP-1) is a member of the threonine-tyrosine dual-specificity phosphatase family that is endogenous inhibitor for extracellular signal-regulated protein kinases (ERKs), p38 MAPKs, and JNKs signaling. MKP-1 plays a key role in determining the magnitude and duration of MAPK activation in response to extracellular stimuli. In human cancers, abnormal expression of DUSP1 is associated with tumor development, chemoresistance and prognosis. However, the mechanism by which MKP-1 is regulated is not fully understood. Here, we show that GP78, an E3 ligase, plays an important role in regulation of MKP-1 protein levels. Specifically, we found that MKP-1 interacts with GP78 both in vitro and in vivo. Their interaction occurs between the N-terminal, Rhodanese domain of MKP-1 and C-terminal E3 ligase domain of GP78. We also found that K230, 280, and 289 residues on MKP-1 are responsible for its ubiquitination. Importantly, this interaction causes MKP-1 ubiquitination and subsequent degradation. Furthermore, we showed by overexpression and knockdown studies that GP78 downregulates the level of MKP-1 protein in cancer cells. Therefore, we identify a novel mechanism by which GP78 controls MKP-1 protein levels and suggest that this regulatory mechanism may play an important in cancer.
Citation Format: Dhonghyo Kho, Gen Sheng Wu. Regulation of the MAPK phosphatase MKP-1 by the E3 ligase GP78 in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2360. doi:10.1158/1538-7445.AM2017-2360
Abstract 2362: Mechanistic insights into phosphoinositide 3-kinase α
Authors: Maheshwari Sweta 
Published: July 2017

Abstract
Phosphoinositide 3-kinases (PI3Ks) are at the center of mTOR pathway that plays key roles in regulating cell survival, proliferation, protein synthesis and vesicle trafficking. Dysregulation of the mTOR pathway has been implicated in various diseases such as cancer, diabetes, arthritis, inflammation and respiratory illnesses. PI3Ks have dual kinase specificity with a lipid kinase activity that phosphorylates the 3’-hydroxyl of phosphoinositides, and a protein kinase activity including auto-phosphorylation. In this study, we explored the roles of residues of the catalytic domain and regulatory subunit of human PI3Kα in the lipid and protein phosphorylation. We used site-directed mutagenesis and kinetic assays to precisely map the molecular basis of substrate recognition and catalysis in PI3Kα. Kinetic analysis of K776, located in the P-loop of PI3Kα, reveals that a lysine at this position is essential for the recognition of the lipid and ATP substrates. This same residue plays an important role in the auto-phosphorylation of the enzyme. Replacement of histidine residues–H936 and H917 in the activation and catalytic loops respectively with alanine residues results in a dramatic change in the kinetics of PI3Kα. While H936A inactivates the lipid kinase activity without affecting auto-phosphorylation, H917A abolishes both the lipid and protein kinase activities of PI3Kα. On the basis of kinetic and structural analysis, we proposed possible mechanistic roles of these critical residues in catalysis.
Citation Format: Sweta Maheshwari. Mechanistic insights into phosphoinositide 3-kinase α [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2362. doi:10.1158/1538-7445.AM2017-2362
Abstract 2364: Src-like adaptor protein 2 negatively regulates FLT3 and KIT downstream signaling
Authors: Moharram Sausan A. , Rönnstrand Lars , Kazi Julhash U. 
Published: July 2017

Abstract
Type III receptor tyrosine kinases (RTKs) including FLT3 and KIT play a major role in cell differentiation, proliferation, and survival of hematopoietic stem cells. FLT3-ITD and KIT-D816V mutations are the most common oncogenic mutations in FLT3 and KIT found in hematological cancers. These mutations lead to constitutive activation of proliferative and survival signals. Tyrosine kinase inhibitors (TKIs) in combination with chemotherapy display promising results in a clinical setting, but patients develop resistant disease after short-term treatment,. Hence, proteins that regulate the activity of RTKs can be alternative targets for patients carrying these mutations. Activation of FLT3 and KIT results in phosphorylation on several tyrosine residues that recruit SH2 domain-containing signaling proteins. In this study we identified Src-like adaptor protein 2 (SLAP2) as a potent FLT3 and KIT interacting protein. The interaction requires an intact SH2 domain of SLAP2 as well as phosphorylation of the receptor. Overexpression of SLAP2 in murine proB Ba/F3 cells inhibited oncogenic FLT3-ITD-mediated cell proliferation and colony formation. SLAP2 displayed a similar inhibitory potential in cells expressing KIT-D816V. SLAP2 partially blocked phosphorylation of several FLT3 and KIT downstream signaling proteins such as AKT, ERK and p38. Moreover, SLAP2 expression inhibited FLT3-ITD-mediated STAT5 phosphorylation and KIT-D816V-mediated STAT3 phosphorylation. SLAP2 expression significantly accelerated ubiquitination-mediated degradation of FLT3 and KIT. Collectively, these data suggest that SLAP2 negatively regulates FLT3 and KIT signaling and therefore, modulation of SLAP2 expression levels may become a potential target for anticancer therapy.
Citation Format: Sausan A. Moharram, Lars Rönnstrand, Julhash U. Kazi. Src-like adaptor protein 2 negatively regulates FLT3 and KIT downstream signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2364. doi:10.1158/1538-7445.AM2017-2364
Abstract 2371: Oncogenic transformation of FLT3-ITD is partially dependent on tyrosine 842 in the activation loop
Authors: Kazi Julhash U. , Rönnstrand Lars 
Published: July 2017

Abstract
Acute myeloid leukemia (AML) is a heterogeneous disease of the blood. About 30 % of AML patients carry an oncogenic mutant of the type III receptor tyrosine kinase FLT3. Among the various mutations in FLT3, the internal tandem duplication (ITD) mutations in the juxtamembrane domain are the most common type of mutation, while point mutations in the kinase domain also occur. Although oncogenic FLT3 mutants are constitutively active, the wild-type receptor needs its ligand (FLT3 ligand, FL) for activation. Ligand binding to the receptor induces dimerization, auto-phosphorylation, and activation of the receptor. Tyrosine phosphorylation sites on FLT3 constitute docking sites for interacting proteins that transduce pro-survival and proliferative signals. The so-called activation loop tyrosine residue (in FLT3, Y842) is well conserved in all receptor tyrosine kinases and long been known essential for the activity of some but not all receptor tyrosine kinases. Recently we have shown that activation loop tyrosine residue in KIT is not essential for its activation but plays an important role in receptor stabilization and downstream signaling. However, so far the role of FLT3 activation loop tyrosine residue remains unknown. We generated 32D cell lines carrying a Y-to-F mutant (Y842F) of FLT3 in either the wild-type or ITD background to study the role of this tyrosine residue. We show that the Y842 residue is not necessary for FLT3 activation or stability but is required for oncogenic transformation. We observed that Y842F mutation results in reduced cell viability, proliferation and in vitro colony formation in semi-solid medium. In addition, cells expressing the Y842F mutant in FLT3-ITD background display a significant reduction in tumor volume and weight in a xenograft model in nude mice. Gene set enrichment analysis (GESA) shows that mutation of Y842 causes suppression of anti-apoptotic genes. Furthermore, we demonstrated that phosphorylated Y842 is one of the binding sites of the protein tyrosine phosphataseSHP2, which is required for activation of signaling through the RAS/ERK pathway. Taken together, our data suggest that activation loop tyrosine residue in FLT3 plays an important role in the cellular transformation which is regulated by RAS/ERK signaling.
Citation Format: Julhash U. Kazi, Lars Rönnstrand. Oncogenic transformation of FLT3-ITD is partially dependent on tyrosine 842 in the activation loop [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2371. doi:10.1158/1538-7445.AM2017-2371
Abstract 2379: Nuclear lemur tyrosine kinase-2 regulates RNA polymerase II dependent transcription in prostate cancer
Authors: Shah Kalpit , Foley Justin , Nickerson Michael L. , Dean Michael , Bradbury Neil 
Published: July 2017

Abstract
The Androgen Receptor (AR), a DNA-binding transcription factor plays a key role in the development and maintenance of prostate epithelia by modulating expression of growth-promoting genes. AR has been proposed to regulate gene expression by enhancing the efficiency of RNA polymerase II (RNAPII) dependent transcription elongation. Dysregulation of this AR-dependent transcriptional activity has been implicated not only in prostate cancer but also androgen-independent prostate cancer. Recently, we showed that protein expression of Lemur Tyrosine Kianse-2 (LMTK2), a serine-threonine kinase is downregulated in prostate cancer. Importantly, our study ascribed a novel role for LMTK2, as a negative regulator of AR-dependent transcriptional activity in prostate epithelial cells. However, the mechanism through which LMTK2 is able to regulate AR remains to be determined. Here, we show that LMTK2 is not only present in cytoplasmic fraction but also, within the nucleus of mammalian cells where it co-localizes with AR, newly transcribed mRNA and with RNA polymerase II (RNAPII). Interestingly, our data reveals that LMTK2 colocalizes with elongating RNAPII, phosphorylated on serine 2 of the carboxyl-terminal domain. This presents an interesting possibility that LMTK2 might be able to regulate transcription of AR-dependent genes by interacting with the elongating RNAPII and transcription & epigenetic factors. As such, our study potentially identifies LMTK2 as an important player regulating the AR-RNAPII mediated transcription machinery in mammalian cells.
Citation Format: Kalpit Shah, Justin Foley, Michael L. Nickerson, Michael Dean, Neil Bradbury. Nuclear lemur tyrosine kinase-2 regulates RNA polymerase II dependent transcription in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2379. doi:10.1158/1538-7445.AM2017-2379
Abstract 2380: Insertion mutations in the tyrosine kinase domain of FLT3 display a higher oncogenic potential than the D835Y mutation in acute myeloid leukemia
Authors: Marhäll Alissa , Fischer Thomas , Heidel Florian , Kazi Julhash U. , Rönnstrand Lars 
Published: July 2017

Abstract
Acute myeloid leukemia (AML) remains the most common form of acute leukemia among adults and accounts for a large number of deaths. Mutation in FMS-like tyrosine kinase 3 (FLT3) is one of the most prevalent factor in this heterogeneous disease. The major mutations in FLT3 can be categories as internal tandem duplications (ITD) and point mutations. Recent studies suggest that ITDs are subdivided in to two groups depending on their location: the juxtamembrane domain (ITD-JM) and the tyrosine kinase domain (ITD-TDK). Although, ITD-JM has been characterized well the ITD-TKD has not yet been studied well due to its recent discovery. For this reason, we compared ITD mutations in TKD and JM, as well as the most frequently occurring point mutation located in the TKD, D835Y. The purpose of this study was to understand whether it is the mutation’s nature or location that plays the driving role in leukemogenesis. We used a cytokine-dependent mouse pro-B cell line, BaF3, to overexpress different FLT3 mutants. We first examined the colony formation capacity in semisolid cytokine- and serum-free medium. The assay resulted in indistinguishable number and size of colonies for both ITD-JM and ITD-TKD, while D835Y-TKD transfected cells failed to form colonies suggesting that the ITD-TKD mutations have stronger transforming potential than other TKD mutations. In addition to colony formation assays, cell proliferation and survival was significantly higher in ITD-TKD expressing cells compared to cells expressing D835Y-TKD. Finally, we showed that phosphorylation of STAT5 and AKT is increased in ITD-TKD, while other FLT3 downstream signaling remained unaffected. All together, our data suggest that ITD-TKD displays higher oncogenic potential than other TKD mutants.
Citation Format: Alissa Marhäll, Thomas Fischer, Florian Heidel, Julhash U. Kazi, Lars Rönnstrand. Insertion mutations in the tyrosine kinase domain of FLT3 display a higher oncogenic potential than the D835Y mutation in acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2380. doi:10.1158/1538-7445.AM2017-2380