Publications by authors named "Rebecca Levin"

32 Publications

Community-engaged research to develop a Chicago violence research agenda and recommendations to support future community engagement.

Inj Epidemiol 2021 Sep 13;8(Suppl 1):39. Epub 2021 Sep 13.

Ann & Robert H. Lurie Children's Hospital of Chicago, 225 E. Chicago Avenue, Chicago, IL, 60611, USA.

Background: Chicago has a history of gun violence with some neighborhoods, particularly Black and Brown communities, being disproportionately affected and Black male youth experiencing an even more disparate impact. Too often, violence prevention research is developed and carried out with little or no input from the people living in the most affected communities. The objective of the Community-Academic Collaboration to Prevent Violence in Chicago (CACPVC) was to bring together individuals from impacted communities with academic researchers and other community stakeholders to discuss violence and co-create a research agenda that addresses topics of mutual concern, and recommendations for engaging stakeholders including community members and organizations and funders in violence and violence prevention research.

Methods: From 2014 to 2015, community members and organizations from seven defined regions across Chicago were recruited to participate. An organization network gathering was held in each region for researchers, funders, and community organization representatives to discuss violence prevention. Open community forums then took place in each community. Violence data by region was shared followed by facilitated group discussions that were recorded by youth scribes. Notes were thematically coded, grouped, and compiled after which a list of topics was refined by the CACPVC Work Group, allowing for investigator triangulation. A survey was disseminated to community stakeholders to prioritize the topics.

Results: Seven network gatherings (127 attendees) and community forums (133 attendees) were held. Topic areas identified during the gatherings and forums included root causes/cycle of violence, racism and bias/structural violence, trajectory of violence, protective factors and nonviolence, geographic pattern change, violence prevention strategies, youth, family factors, community factors, school, police, gangs/street organizations, and media and public perceptions. Recommendations to support community engagement were grouped as role of research in reducing violence, role of community in violence research, relationships and respect, academic-community communication, financial considerations, training, practical considerations, research design, sharing results, communication about and use of data, and recommendations for funders.

Conclusions: The violence research agenda will be used to inform community-engaged violence prevention research. The recommendations for community engagement provide a resource for researchers about topics to consider to meaningfully engage community members in future research.
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http://dx.doi.org/10.1186/s40621-021-00335-9DOI Listing
September 2021

A 2-Year-Old Girl with Multisystem Inflammatory Syndrome in Children (MIS-C).

R I Med J (2013) 2021 May 3;104(4):49-52. Epub 2021 May 3.

Associate Professor of Emergency Medicine, Clinician Educator; Associate Professor of Pediatrics, Clinician Educator, Alpert Medical School of Brown University, Providence, RI.

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May 2021

Outbreak of COVID-19 and interventions in a large jail - Cook County, IL, United States, 2020.

Am J Infect Control 2021 09 2;49(9):1129-1135. Epub 2021 Apr 2.

Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, GA. Electronic address:

Background: Correctional and detention facilities are disproportionately affected by COVID-19 due to shared space, contact between staff and detained persons, and movement within facilities. On March 18, 2020, Cook County Jail, one of the United States' largest, identified its first suspected case of COVID-19 in a detained person.

Methods: This analysis includes SARS-CoV-2 cases confirmed by molecular detection among detained persons and Cook County Sheriff's Office staff. We examined occurrence of symptomatic cases in each building and proportions of asymptomatic detained persons testing positive, and timing of interventions including social distancing, mask use, and expanded testing and show outbreak trajectory in the jail compared to case counts in Chicago.

Results: During March 1-April 30, 907 symptomatic and asymptomatic cases of SARS-CoV-2 infection were detected among detained persons (n = 628) and staff (n = 279). Among asymptomatic detained persons in quarantine, 23.6% tested positive. Programmatic activity and visitation stopped March 9, cells were converted into single occupancy beginning March 26, and universal masking was implemented for staff (April 2) and detained persons (April 13). Cases at the jail declined while cases in Chicago increased.

Discussion/conclusions: Aggressive intervention strategies coupled with widespread diagnostic testing of detained and staff populations can limit introduction and mitigate transmission of SARS-CoV-2 infection in correctional and detention facilities.
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http://dx.doi.org/10.1016/j.ajic.2021.03.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8016534PMC
September 2021

COVID-19 in Pediatric Patients: Observations from the Initial Phase of the Global Pandemic in Rhode Island.

R I Med J (2013) 2021 Feb 1;104(1):55-60. Epub 2021 Feb 1.

Department of Emergency Medicine, Department of Pediatrics, Warren Alpert Medical School of Brown University, Providence, RI.

Objective: To describe characteristics of children undergoing SARS-CoV-2 testing during the initial wave of infections in Rhode Island.

Methods: This is a descriptive study of 729 children tested for SARS-CoV-2 at four emergency departments April 9 to May 7, 2020 in Rhode Island. Demographic information and symptoms were cataloged for those tested.

Results: 81 (11%) children tested positive for SARS-CoV-2. 94% of positive children were symptomatic. 74% of positive cases had constitutional symptoms and 72% had upper respiratory symptoms. While only 34% of those tested were Hispanic, 68% of the SARS-CoV-2- positive cases occurred in Hispanic children.

Conclusion: This study details the pediatric population's experience during the first wave of the pandemic in Rhode Island. It could inform testing allocation strategies in healthcare settings. It also highlights vulnerable populations in need of further public health support in our state.
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February 2021

Identification of Presymptomatic and Asymptomatic Cases Using Cohort-Based Testing Approaches at a Large Correctional Facility-Chicago, Illinois, USA, May 2020.

Clin Infect Dis 2021 03;72(5):e128-e135

Cermak Health Services, Chicago, Illinois, USA.

Background: Coronavirus disease 2019 (COVID-19) continues to cause significant morbidity and mortality worldwide. Correctional and detention facilities are at high risk of experiencing outbreaks. We aimed to evaluate cohort-based testing among detained persons exposed to laboratory-confirmed cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in order to identify presymptomatic and asymptomatic cases.

Methods: During 1-19 May 2020, 2 testing strategies were implemented in 12 tiers or housing units of the Cook County Jail, Chicago, Illinois. Detained persons were approached to participate in serial testing (n = 137) and offered tests at 3 time points over 14 days (day 1, days 3-5, and days 13-14). The second group was offered a single test and interview at the end of a 14-day quarantine period (day 14 group) (n = 87).

Results: 224 detained persons were approached for participation and, of these, 194 (87%) participated in ≥1 interview and 172 (77%) had ≥1 test. Of the 172 tested, 19 were positive for SARS-CoV-2. In the serial testing group, 17 (89%) new cases were detected, 16 (84%) on day 1, 1 (5%) on days 3-5, and none on days 13-14; in the day 14 group, 2 (11%) cases were identified. More than half (12/19; 63%) of the newly identified cases were presymptomatic or asymptomatic.

Conclusions: Our findings highlight the utility of cohort-based testing promptly after initiating quarantine within a housing tier. Cohort-based testing efforts identified new SARS-CoV-2 asymptomatic and presymptomatic infections that may have been missed by symptom screening alone.
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http://dx.doi.org/10.1093/cid/ciaa1802DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7799274PMC
March 2021

Network Characteristics and Visualization of COVID-19 Outbreak in a Large Detention Facility in the United States - Cook County, Illinois, 2020.

MMWR Morb Mortal Wkly Rep 2020 Nov 6;69(44):1625-1630. Epub 2020 Nov 6.

Correctional and detention facilities have been disproportionately affected by coronavirus disease 2019 (COVID-19) because of shared space and movement of staff members and detained persons within facilities (1,2). During March 1-April 30, 2020, at Cook County Jail in Chicago, Illinois, >900 COVID-19 cases were diagnosed across all 10 housing divisions, representing 13 unique buildings. Movement within the jail was examined through network analyses and visualization, a field that examines elements within a network and the connections between them. This methodology has been used to supplement contact tracing investigations for tuberculosis and to understand how social networks contribute to transmission of sexually transmitted infections (3-5). Movements and connections of 5,884 persons (3,843 [65%] detained persons and 2,041 [35%] staff members) at the jail during March 1-April 30 were analyzed. A total of 472 (12.3%) COVID-19 cases were identified among detained persons and 198 (9.7%) among staff members. Among 103,701 shared-shift connections among staff members, 1.4% occurred between persons with COVID-19, a percentage that is significantly higher than the expected 0.9% by random occurrence alone (p<0.001), suggesting that additional transmission occurred within this group. The observed connections among detained persons with COVID-19 were significantly lower than expected (1.0% versus 1.1%, p<0.001) when considering only the housing units in which initial transmission occurred, suggesting that the systematic isolation of persons with COVID-19 is effective at limiting transmission. A network-informed approach can identify likely points of high transmission, allowing for interventions to reduce transmission targeted at these groups or locations, such as by reducing convening of staff members, closing breakrooms, and cessation of contact sports.
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http://dx.doi.org/10.15585/mmwr.mm6944a3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7643900PMC
November 2020

Unbiased Proteomic Profiling Uncovers a Targetable GNAS/PKA/PP2A Axis in Small Cell Lung Cancer Stem Cells.

Cancer Cell 2020 07 11;38(1):129-143.e7. Epub 2020 Jun 11.

Department of Pediatrics, Stanford University, 265 Campus Drive, Stanford, CA 94305-5457, USA; Department of Genetics, Stanford University, Stanford, CA 94305, USA. Electronic address:

Using unbiased kinase profiling, we identified protein kinase A (PKA) as an active kinase in small cell lung cancer (SCLC). Inhibition of PKA activity genetically, or pharmacologically by activation of the PP2A phosphatase, suppresses SCLC expansion in culture and in vivo. Conversely, GNAS (G-protein α subunit), a PKA activator that is genetically activated in a small subset of human SCLC, promotes SCLC development. Phosphoproteomic analyses identified many PKA substrates and mechanisms of action. In particular, PKA activity is required for the propagation of SCLC stem cells in transplantation studies. Broad proteomic analysis of recalcitrant cancers has the potential to uncover targetable signaling networks, such as the GNAS/PKA/PP2A axis in SCLC.
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http://dx.doi.org/10.1016/j.ccell.2020.05.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7363571PMC
July 2020

p38γ MAPK contributes to left ventricular remodeling after pathologic stress and disinhibits calpain through phosphorylation of calpastatin.

FASEB J 2019 12 22;33(12):13131-13144. Epub 2019 Oct 22.

School of Cardiovascular Medicine and Science, British Heart Foundation (BHF) Centre, King's College London, London, United Kingdom.

Despite the high and preferential expression of p38γ MAPK in the myocardium, little is known about its function in the heart. The aim of the current study was to elucidate the physiologic and biochemical roles of p38γ in the heart. Expression and subcellular localization of p38 isoforms was determined in mouse hearts. Comparisons of the cardiac function and structure of wild-type and p38γ knockout (KO) mice at baseline and after abdominal aortic banding demonstrated that KO mice developed less ventricular hypertrophy and that contractile function is better preserved. To identify potential substrates of p38γ, we generated an analog-sensitive mutant to affinity tag endogenous myocardial proteins. Among other proteins, this technique identified calpastatin as a direct p38γ substrate. Moreover, phosphorylation of calpastatin by p38γ impaired its ability to inhibit the protease, calpain. We have identified p38γ as an important determinant of the progression of pathologic cardiac hypertrophy after aortic banding in mice. In addition, we have identified calpastatin, among other substrates, as a novel direct target of p38γ that may contribute to the protection observed in p38γKO mice.-Loonat, A. A., Martin, E. D., Sarafraz-Shekary, N., Tilgner, K., Hertz, N. T., Levin, R., Shokat, K. M., Burlingame, A. L., Arabacilar, P., Uddin, S., Thomas, M., Marber, M. S., Clark, J. E. p38γ MAPK contributes to left ventricular remodeling after pathologic stress and disinhibits calpain through phosphorylation of calpastatin.
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http://dx.doi.org/10.1096/fj.201701545RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6894093PMC
December 2019

Phosphoregulation of the oncogenic protein regulator of cytokinesis 1 (PRC1) by the atypical CDK16/CCNY complex.

Exp Mol Med 2019 04 16;51(4):1-17. Epub 2019 Apr 16.

Faculty of Medicine and Health Sciences, International University of Catalonia, 08195, Sant Cugat del Vallès, Barcelona, Spain.

CDK16 (also known as PCTAIRE1 or PCTK1) is an atypical member of the cyclin-dependent kinase (CDK) family that forms an active complex with cyclin Y (CCNY). Although both proteins have been recently implicated in cancer pathogenesis, it is still unclear how the CDK16/CCNY complex exerts its biological activity. To understand the CDK16/CCNY network, we used complementary proteomic approaches to identify potential substrates of this complex. We identified several candidates implicating the CDK16/CCNY complex in cytoskeletal dynamics, and we focused on the microtubule-associated protein regulator of cytokinesis (PRC1), an essential protein for cell division that organizes antiparallel microtubules and whose deregulation may drive genomic instability in cancer. Using analog-sensitive (AS) CDK16 generated by CRISPR-Cas9 mutagenesis in 293T cells, we found that specific inhibition of CDK16 induces PRC1 dephosphorylation at Thr481 and delocalization to the nucleus during interphase. The observation that CDK16 inhibition and PRC1 downregulation exhibit epistatic effects on cell viability confirms that these proteins can act through a single pathway. In conclusion, we identified PRC1 as the first substrate of the CDK16/CCNY complex and demonstrated that the proliferative function of CDK16 is mediated by PRC1 phosphorylation. As CDK16 is emerging as a critical node in cancer, our study reveals novel potential therapeutic targets.
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http://dx.doi.org/10.1038/s12276-019-0242-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6467995PMC
April 2019

Refusal of Vitamin K Injection: Survey of the Current Literature and Practical Tips for Pediatricians.

Pediatr Ann 2018 Aug;47(8):e334-e338

Vitamin K refusal and associated sequelae of vitamin K deficiency bleed (VKDB) in the newborn period is becoming a more common occurrence. We present six recent cases from a 4-month period in 2017 of parent refusal of vitamin K and describe the reasons for refusal and the clinical outcomes of these infants. There have been a number of case reports citing the rising incidence of VKDB and the reasons why parents refuse. However, there is a gap in the literature and clinical practice guidelines describing how a physician should approach a refusal in the hospital and in the office, and the need to report a refusal to child welfare. In addition, we describe a scenario in which the caregivers provide a religious reason for refusal of vitamin K that, to the best of our knowledge, has yet to be cited in the literature. [Pediatr Ann. 2018;47(8):e334-e338.].
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http://dx.doi.org/10.3928/19382359-20180709-02DOI Listing
August 2018

Kinome rewiring reveals AURKA limits PI3K-pathway inhibitor efficacy in breast cancer.

Nat Chem Biol 2018 08 25;14(8):768-777. Epub 2018 Jun 25.

Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, CA, USA.

Dysregulation of the PI3K-AKT-mTOR signaling network is a prominent feature of breast cancers. However, clinical responses to drugs targeting this pathway have been modest, possibly because of dynamic changes in cellular signaling that drive resistance and limit drug efficacy. Using a quantitative chemoproteomics approach, we mapped kinome dynamics in response to inhibitors of this pathway and identified signaling changes that correlate with drug sensitivity. Maintenance of AURKA after drug treatment was associated with resistance in breast cancer models. Incomplete inhibition of AURKA was a common source of therapy failure, and combinations of PI3K, AKT or mTOR inhibitors with the AURKA inhibitor MLN8237 were highly synergistic and durably suppressed mTOR signaling, resulting in apoptosis and tumor regression in vivo. This signaling map identifies survival factors whose presence limits the efficacy of targeted therapies and reveals new drug combinations that may unlock the full potential of PI3K-AKT-mTOR pathway inhibitors in breast cancer.
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http://dx.doi.org/10.1038/s41589-018-0081-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6051919PMC
August 2018

An Optimized Chromatographic Strategy for Multiplexing In Parallel Reaction Monitoring Mass Spectrometry: Insights from Quantitation of Activated Kinases.

Mol Cell Proteomics 2017 02 11;16(2):265-277. Epub 2016 Dec 11.

§Department of Pharmaceutical Chemistry, University of California San Francisco, San Francisco, California.

Reliable quantitation of protein abundances in defined sets of cellular proteins is critical to numerous biological applications. Traditional immunodetection-based methods are limited by the quality and availability of specific antibodies, especially for site-specific post-translational modifications. Targeted proteomic methods, including the recently developed parallel reaction monitoring (PRM) mass spectrometry, have enabled accurate quantitative measurements of up to a few hundred specific target peptides. However, the degree of practical multiplexing in label-free PRM workflows remains a significant limitation for the technique. Here we present a strategy for significantly increasing multiplexing in label-free PRM that takes advantage of the superior separation characteristics and retention time stability of meter-scale monolithic silica-C18 column-based chromatography. We show the utility of the approach in quantifying kinase abundances downstream of previously developed active kinase enrichment methodology based on multidrug inhibitor beads. We examine kinase activation dynamics in response to three different MAP kinase inhibitors in colorectal carcinoma cells and demonstrate reliable quantitation of over 800 target peptides from over 150 kinases in a single label-free PRM run. The kinase activity profiles obtained from these analyses reveal compensatory activation of TGF-β family receptors as a response to MAPK blockade. The gains achieved using this label-free PRM multiplexing strategy will benefit a wide array of biological applications.
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http://dx.doi.org/10.1074/mcp.M116.058172DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5294213PMC
February 2017

Discovery of new substrates of the elongation factor-2 kinase suggests a broader role in the cellular nutrient response.

Cell Signal 2017 01 17;29:78-83. Epub 2016 Oct 17.

Howard Hughes Medical Institute and Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA. Electronic address:

Elongation Factor-2 Kinase (eEF2K) in an unusual mammalian enzyme that has one known substrate, elongation factor-2. It belongs to a class of kinases, called alpha kinases, that has little sequence identity to the >500 conventional protein kinases, but performs the same reaction and has similar catalytic residues. The phosphorylation of eEF2 blocks translation elongation, which is thought to be critical to regulating cellular energy usage. Here we report a system for discovering new substrates of alpha kinases and identify the first new substrates of eEF2K including AMPK and alpha4, and determine a sequence motif for the kinase that shows a requirement for threonine residues as the target of phosphorylation. These new substrates suggest that eEF2K has a more diverse role in regulating cellular energy usage that involves multiple pathways and regulatory feedback.
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http://dx.doi.org/10.1016/j.cellsig.2016.10.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5138099PMC
January 2017

Innate immunity kinase TAK1 phosphorylates Rab1 on a hotspot for posttranslational modifications by host and pathogen.

Proc Natl Acad Sci U S A 2016 08 1;113(33):E4776-83. Epub 2016 Aug 1.

Department of Microbiology and Immunology, University of California, San Francisco, CA 94143

TGF-β activated kinase 1 (TAK1) is a critical signaling hub responsible for translating antigen binding signals to immune receptors for the activation of the AP-1 and NF-κB master transcriptional programs. Despite its importance, known substrates of TAK1 are limited to kinases of the MAPK and IKK families and include no direct effectors of biochemical processes. Here, we identify over 200 substrates of TAK1 using a chemical genetic kinase strategy. We validate phosphorylation of the dynamic switch II region of GTPase Rab1, a mediator of endoplasmic reticulum to Golgi vesicular transport, at T75 to be regulated by TAK1 in vivo. TAK1 preferentially phosphorylates the inactive (GDP-bound) state of Rab1. Phosphorylation of Rab1 disrupts interaction with GDP dissociation inhibitor 1 (GDI1), but not guanine exchange factor (GEF) or GTPase-activating protein (GAP) enzymes, and is exclusive to membrane-localized Rab1, suggesting phosphorylation may stimulate Rab1 membrane association. Furthermore, we found phosphorylation of Rab1 at T75 to be essential for Rab1 function. Previous studies established that the pathogen Legionella pneumophila is capable of hijacking Rab1 function through posttranslational modifications of the switch II region. Here, we present evidence that Rab1 is regulated by the host in a similar fashion, and that the innate immunity kinase TAK1 and Legionella effectors compete to regulate Rab1 by switch II modifications during infection.
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http://dx.doi.org/10.1073/pnas.1608355113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4995946PMC
August 2016

Synthetic Lethal Targeting of ARID1A-Mutant Ovarian Clear Cell Tumors with Dasatinib.

Mol Cancer Ther 2016 07 30;15(7):1472-84. Epub 2016 Jun 30.

The CRUK Gene Function Laboratory, The Institute of Cancer Research, London, United Kingdom. Breakthrough Breast Cancer Research Centre, The Institute of Cancer Research, London, United Kingdom.

New targeted approaches to ovarian clear cell carcinomas (OCCC) are needed, given the limited treatment options in this disease and the poor response to standard chemotherapy. Using a series of high-throughput cell-based drug screens in OCCC tumor cell models, we have identified a synthetic lethal (SL) interaction between the kinase inhibitor dasatinib and a key driver in OCCC, ARID1A mutation. Imposing ARID1A deficiency upon a variety of human or mouse cells induced dasatinib sensitivity, both in vitro and in vivo, suggesting that this is a robust synthetic lethal interaction. The sensitivity of ARID1A-deficient cells to dasatinib was associated with G1-S cell-cycle arrest and was dependent upon both p21 and Rb. Using focused siRNA screens and kinase profiling, we showed that ARID1A-mutant OCCC tumor cells are addicted to the dasatinib target YES1. This suggests that dasatinib merits investigation for the treatment of patients with ARID1A-mutant OCCC. Mol Cancer Ther; 15(7); 1472-84. ©2016 AACR.
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http://dx.doi.org/10.1158/1535-7163.MCT-15-0554DOI Listing
July 2016

Isocitrate Dehydrogenase Mutations Confer Dasatinib Hypersensitivity and SRC Dependence in Intrahepatic Cholangiocarcinoma.

Cancer Discov 2016 07 26;6(7):727-39. Epub 2016 May 26.

Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, Massachusetts.

Unlabelled: Intrahepatic cholangiocarcinoma (ICC) is an aggressive liver bile duct malignancy exhibiting frequent isocitrate dehydrogenase (IDH1/IDH2) mutations. Through a high-throughput drug screen of a large panel of cancer cell lines, including 17 biliary tract cancers, we found that IDH mutant (IDHm) ICC cells demonstrate a striking response to the multikinase inhibitor dasatinib, with the highest sensitivity among 682 solid tumor cell lines. Using unbiased proteomics to capture the activated kinome and CRISPR/Cas9-based genome editing to introduce dasatinib-resistant "gatekeeper" mutant kinases, we identified SRC as a critical dasatinib target in IDHm ICC. Importantly, dasatinib-treated IDHm xenografts exhibited pronounced apoptosis and tumor regression. Our results show that IDHm ICC cells have a unique dependency on SRC and suggest that dasatinib may have therapeutic benefit against IDHm ICC. Moreover, these proteomic and genome-editing strategies provide a systematic and broadly applicable approach to define targets of kinase inhibitors underlying drug responsiveness.

Significance: IDH mutations define a distinct subtype of ICC, a malignancy that is largely refractory to current therapies. Our work demonstrates that IDHm ICC cells are hypersensitive to dasatinib and critically dependent on SRC activity for survival and proliferation, pointing to new therapeutic strategies against these cancers. Cancer Discov; 6(7); 727-39. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 681.
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http://dx.doi.org/10.1158/2159-8290.CD-15-1442DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5458737PMC
July 2016

P-TEFb regulation of transcription termination factor Xrn2 revealed by a chemical genetic screen for Cdk9 substrates.

Genes Dev 2016 Jan;30(1):117-31

Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, New York 10029, USA;

The transcription cycle of RNA polymerase II (Pol II) is regulated at discrete transition points by cyclin-dependent kinases (CDKs). Positive transcription elongation factor b (P-TEFb), a complex of Cdk9 and cyclin T1, promotes release of paused Pol II into elongation, but the precise mechanisms and targets of Cdk9 action remain largely unknown. Here, by a chemical genetic strategy, we identified ∼ 100 putative substrates of human P-TEFb, which were enriched for proteins implicated in transcription and RNA catabolism. Among the RNA processing factors phosphorylated by Cdk9 was the 5'-to-3' "torpedo" exoribonuclease Xrn2, required in transcription termination by Pol II, which we validated as a bona fide P-TEFb substrate in vivo and in vitro. Phosphorylation by Cdk9 or phosphomimetic substitution of its target residue, Thr439, enhanced enzymatic activity of Xrn2 on synthetic substrates in vitro. Conversely, inhibition or depletion of Cdk9 or mutation of Xrn2-Thr439 to a nonphosphorylatable Ala residue caused phenotypes consistent with inefficient termination in human cells: impaired Xrn2 chromatin localization and increased readthrough transcription of endogenous genes. Therefore, in addition to its role in elongation, P-TEFb regulates termination by promoting chromatin recruitment and activation of a cotranscriptional RNA processing enzyme, Xrn2.
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http://dx.doi.org/10.1101/gad.269589.115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4701974PMC
January 2016

Identification of AMPK Phosphorylation Sites Reveals a Network of Proteins Involved in Cell Invasion and Facilitates Large-Scale Substrate Prediction.

Cell Metab 2015 Nov 8;22(5):907-21. Epub 2015 Oct 8.

Cancer Biology Program, Stanford University, Stanford, CA 94305, USA; Department of Genetics, Stanford University, Stanford, CA 94305, USA; Glenn Laboratories for the Biology of Aging, Stanford, CA 94305, USA. Electronic address:

AMP-activated protein kinase (AMPK) is a central energy gauge that regulates metabolism and has been increasingly involved in non-metabolic processes and diseases. However, AMPK's direct substrates in non-metabolic contexts are largely unknown. To better understand the AMPK network, we use a chemical genetics screen coupled to a peptide capture approach in whole cells, resulting in identification of direct AMPK phosphorylation sites. Interestingly, the high-confidence AMPK substrates contain many proteins involved in cell motility, adhesion, and invasion. AMPK phosphorylation of the RHOA guanine nucleotide exchange factor NET1A inhibits extracellular matrix degradation, an early step in cell invasion. The identification of direct AMPK phosphorylation sites also facilitates large-scale prediction of AMPK substrates. We provide an AMPK motif matrix and a pipeline to predict additional AMPK substrates from quantitative phosphoproteomics datasets. As AMPK is emerging as a critical node in aging and pathological processes, our study identifies potential targets for therapeutic strategies.
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http://dx.doi.org/10.1016/j.cmet.2015.09.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4635044PMC
November 2015

Determinants of adherence to diabetes medications: findings from a large pharmacy claims database.

Diabetes Care 2015 Apr 8;38(4):604-9. Epub 2015 Jan 8.

Research and Evaluation Analytics, LLC, Ringwood, NJ.

Objective: Adults with diabetes typically take multiple medications for hyperglycemia, diabetes-associated conditions, and other comorbidities. Medication adherence is associated with improved outcomes, including reduced health care costs, hospitalization, and mortality. We conducted a retrospective analysis of a large pharmacy claims database to examine patient, medication, and prescriber factors associated with adherence to antidiabetic medications.

Research Design And Methods: We extracted data on a cohort of >200,000 patients who were treated for diabetes with noninsulin medications in the second half of 2010 and had continuous prescription benefits eligibility through 2011. Adherence was defined as a medication possession ratio ≥ 0.8. We used a modified adherence measure that accounted for switching therapies. Logistic regression analysis was performed to determine factors independently associated with adherence.

Results: Sixty-nine percent of patients were adherent. Adherence was independently associated with older age, male sex, higher education, higher income, use of mail order versus retail pharmacies, primary care versus nonendocrinology specialist prescribers, higher daily total pill burden, and lower out-of-pocket costs. Patients who were new to diabetes therapy were significantly less likely to be adherent.

Conclusions: Several demographic, clinical, and potentially modifiable system-level factors were associated with adherence to antidiabetic medications. Patients typically perceived to be healthy (those who are younger, new to diabetes, and on few other medications) may be at risk for nonadherence. For all patients, efforts to reduce out-of-pocket costs and encourage use of mail order pharmacies may result in higher adherence.
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http://dx.doi.org/10.2337/dc14-2098DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370331PMC
April 2015

Linking tumor mutations to drug responses via a quantitative chemical-genetic interaction map.

Cancer Discov 2015 02 12;5(2):154-67. Epub 2014 Dec 12.

University of California, San Francisco, San Francisco, California.

Unlabelled: There is an urgent need in oncology to link molecular aberrations in tumors with therapeutics that can be administered in a personalized fashion. One approach identifies synthetic-lethal genetic interactions or dependencies that cancer cells acquire in the presence of specific mutations. Using engineered isogenic cells, we generated a systematic and quantitative chemical-genetic interaction map that charts the influence of 51 aberrant cancer genes on 90 drug responses. The dataset strongly predicts drug responses found in cancer cell line collections, indicating that isogenic cells can model complex cellular contexts. Applying this dataset to triple-negative breast cancer, we report clinically actionable interactions with the MYC oncogene, including resistance to AKT-PI3K pathway inhibitors and an unexpected sensitivity to dasatinib through LYN inhibition in a synthetic lethal manner, providing new drug and biomarker pairs for clinical investigation. This scalable approach enables the prediction of drug responses from patient data and can accelerate the development of new genotype-directed therapies.

Significance: Determining how the plethora of genomic abnormalities that exist within a given tumor cell affects drug responses remains a major challenge in oncology. Here, we develop a new mapping approach to connect cancer genotypes to drug responses using engineered isogenic cell lines and demonstrate how the resulting dataset can guide clinical interrogation.
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http://dx.doi.org/10.1158/2159-8290.CD-14-0552DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4407699PMC
February 2015

Impact of specialty pharmacy on telaprevir-containing 3-drug hepatitis C regimen persistence.

J Manag Care Spec Pharm 2014 Dec;20(12):1227-34

One Express Way, 2W05, St. Louis, MO 63134.

Background: Although the recommended treatment of hepatitis C continues to evolve as newer and more effective medications are made available, hepatitis C drug regimens consisting of a 3-drug combination of a protease inhibitor, pegylated interferon, and ribavirin were recommended by the American Association for the Study of Liver Diseases for the HCV genotype I beginning in 2011. Although more effective than the earlier standard of care, these regimens have complex dosing schedules, prolonged duration, and deleterious side effects. It has been shown that patients tend to discontinue these regimens prematurely. Specialty pharmacies offer specialized care management programs to hepatitis C patients, consisting of such services as regularly scheduled patient counseling, assessing regimen appropriateness, monitoring treatment progress, scheduling refill reminders, and coordinating patient care with prescribers. The use of specialty pharmacies by hepatitis C patients may improve persistence on the 3-drug hepatitis C regimens.

Objective: To examine the association of pharmacy dispensing channel (specialty pharmacy or retail pharmacy) and hepatitis C regimen persistence among patients on a 3-drug hepatitis C regimen containing telaprevir, a widely used hepatitis C protease inhibitor. 

Methods: A retrospective, observational study was conducted using pharmacy claims data from a national pharmacy benefits manager for the period July 2011 to June 2013. Continuously eligible patients who started a new 3-drug regimen containing telaprevir were included in the study and followed for up to 12 months after the index hepatitis C claim. The study outcome was persistence to the 3-drug regimen at treatment week 24 (day 168), representing the completion of an important milestone in the regimen. Patients were defined as persistent if they filled 84 days' supply of telaprevir and 168 days' supply of pegylated interferon and ribavirin each, as required by the regimen protocol. Multivariate logistic regression was used to evaluate the association between dispensing channel and persistence, controlling for differences in demographics, medication burden, out-of-pocket spend per 30-day adjusted hepatitis C prescription, and average days' supply per unadjusted hepatitis C prescription.

Results: The final study sample consisted of 1,475 patients-1,182 in the specialty pharmacy group and 293 in the retail pharmacy group. A significantly greater proportion of patients were persistent to the 3-drug hepatitis C regimen containing telaprevir in specialty pharmacy, compared with retail pharmacy (56.0% vs. 39.9%, P  less than  0.001). After multivariate adjustment, patients in the specialty pharmacy group had 1.89 times greater odds of being persistent to 3-drug hepatitis C regimens containing telaprevir compared with patients in the retail group (95% CI=1.44-2.48).

Conclusions: Patients who used a specialty pharmacy offering refill reminders, care management, and care coordination with prescribers were significantly more likely to be persistent to 3-drug hepatitis C regimens, compared with patients using a retail pharmacy.
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http://dx.doi.org/10.18553/jmcp.2014.20.12.1227DOI Listing
December 2014

Cholesterol levels in fragile X syndrome.

Am J Med Genet A 2015 Feb 25;167A(2):379-84. Epub 2014 Nov 25.

Departments of Pediatrics, Rush University Medical Center, USA; Departments of Neurological Sciences, Rush University Medical Center, USA; Departments of Biochemistry, Rush University Medical Center, USA.

Fragile X syndrome (FXS) is associated with intellectual disability and behavioral dysfunction, including anxiety, ADHD symptoms, and autistic features. Although individuals with FXS are largely considered healthy and lifespan is not thought to be reduced, very little is known about the long-term medical health of adults with FXS and no systematically collected information is available on standard laboratory measures from metabolic screens. During the course of follow up of a large cohort of patients with FXS we noted that many patients had low cholesterol and high density lipoprotein (HDL) values and thus initiated a systematic chart review of all cholesterol values present in charts from a clinic cohort of over 500 patients with FXS. Total cholesterol (TC), low density lipoprotein (LDL) and HDL were all significantly reduced in males from the FXS cohort relative to age-adjusted population normative data. This finding has relevance for health monitoring in individuals with FXS, for treatments with cholesterol-lowering agents that have been proposed to target the underlying CNS disorder in FXS based on work in animal models, and for potential biomarker development in FXS.
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http://dx.doi.org/10.1002/ajmg.a.36850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5436717PMC
February 2015

Oncogene mimicry as a mechanism of primary resistance to BRAF inhibitors.

Cell Rep 2014 Aug 7;8(4):1037-48. Epub 2014 Aug 7.

Howard Hughes Medical Institute and Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA. Electronic address:

Despite the development of potent RAF/mitogen-activated protein kinase (MAPK) pathway inhibitors, only a fraction of BRAF-mutant patients benefit from treatment with these drugs. Using a combined chemogenomics and chemoproteomics approach, we identify drug-induced RAS-RAF-MEK complex formation in a subset of BRAF-mutant cancer cells characterized by primary resistance to vemurafenib. In these cells, autocrine interleukin-6 (IL-6) secretion may contribute to the primary resistance phenotype via induction of JAK/STAT3 and MAPK signaling. In a subset of cell lines, combined IL-6/MAPK inhibition is able to overcome primary resistance to BRAF-targeted therapy. Overall, we show that the signaling plasticity exerted by primary resistant BRAF-mutant cells is achieved by their ability to mimic signaling features of oncogenic RAS, a strategy that we term "oncogene mimicry." This model may guide future strategies for overcoming primary resistance observed in these tumors.
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http://dx.doi.org/10.1016/j.celrep.2014.07.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4294625PMC
August 2014

Trends in use of ezetimibe after the ENHANCE trial, 2007 through 2010.

JAMA Intern Med 2014 Sep;174(9):1486-93

Robert Wood Johnson Clinical Scholars Program, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut3Section of Health Policy and Administration, Department of Epidemiology and Public Health, Yale School of Public Hea.

Importance: Results from the Ezetimibe and Simvastatin in Hypercholesterolemia Enhances Atherosclerosis Regression (ENHANCE) trial, announced in January 2008, demonstrated that ezetimibe use lowered cholesterol levels but did not slow the progression of atherosclerosis.

Objective: To examine the association of this announcement with national patterns of ezetimibe prescribing, including medication initiation and discontinuation, as well as predictors of use.

Design, Setting, And Participants: Retrospective analysis of a national sample of adults 18 years or older who were continuously enrolled in plans of a large US pharmacy benefit manager from 2007 to 2010.

Main Outcomes And Measures: Lipid-lowering therapy prescription claims were categorized as ezetimibe-containing treatments or any other lipid-lowering agent. Initiation was defined as an ezetimibe claim without another in the prior 180 days; discontinuation, as an ezetimibe claim without another in the subsequent 180 days.

Results: From 2007 to 2010, 29.1% of the 10,597,296 continuously eligible adults obtained at least 1 lipid-lowering agent prescription. Among these adults, 17.8% were prescribed ezetimibe and 95.3% another lipid-lowering agent, predominantly statins. Ezetimibe use peaked in January 2008, when 2.5% of all adults were ezetimibe users, but declined to 1.8% by December 2010. The ENHANCE trial announcement was associated with a nonsignificant 0.16% fewer monthly ezetimibe users (P = .11) but a significant 0.14% more monthly monotherapy users and 0.30% fewer users of ezetimibe concomitant with other lipid-lowering agents (both P = .01). The ENHANCE trial was also associated with 0.44% fewer monthly ezetimibe initiations (P = .002) and 10.4% more monthly ezetimibe discontinuations (P < .001), particularly of ezetimibe monotherapy for both. More than half of adults who initiated ezetimibe use did so without first being prescribed another lipid-lowering agent, both before (50%-60%) and after (60%-70%) the trial. Those aged 50 to 64 years and those living in the East South Central US Census division were both more likely to initiate and less likely to discontinue ezetimibe after the ENHANCE trial.

Conclusions And Relevance: After announcement of the results of the ENHANCE trial, nearly 2% of all continuously enrolled adult beneficiaries within a large US pharmacy benefit manager used ezetimibe, although ezetimibe initiations declined and discontinuations increased.
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http://dx.doi.org/10.1001/jamainternmed.2014.3404DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4208827PMC
September 2014

Paediatrician knowledge, attitudes, and counselling patterns on teen driving.

Inj Prev 2012 Feb 23;18(1):10-5. Epub 2011 May 23.

Paediatric Hospitalist Division, Phoenix Children's Hospital, Phoenix, Arizona 85016, USA.

Background: Motor vehicle crashes (MVCs) are the leading cause of death among teenagers. Little is known about the content of US paediatrician counselling about teen driving.

Objective: To examine US paediatrician knowledge, attitudes, and counselling patterns regarding teen driving.

Methods: A random sample questionnaire was mailed to American Academy of Pediatrics members in 2009 (n=1606; response=875 (55%)). Analysis was limited to 596 paediatricians who provide adolescent checkups. Questions addressed counselling and attitudes towards roles in promoting safe driving. Logistic regression assessed the relationship between counselling topics and practice characteristics.

Results: Most (89%) respondents provide some counselling about driving. Two topics commonly discussed by paediatricians were seatbelts (87%) and alcohol use (82%). Less frequently discussed were: cell phones (47%), speeding (43%), and dangers of transporting teen passengers (41%). Topics rarely discussed were: night driving (21%), graduated driver licensing laws (13%), safe cars (9%), driver education (9%), fatigue (25%), and parental limit setting (23%). Only 10% ever recommend a parent-teen driver agreement. Paediatricians who had a patient injured or killed in an MVC were more likely to discuss night driving (OR=2.86). Physicians caring for a high proportion of adolescents (OR=1.83) or patients with private insurance (OR=1.85) counsel more about the risks of driving with teen passengers.

Conclusions: Paediatricians in the USA support counselling on teen driving during routine office visits, but omit many important risk factors. Few recommend parent-teen driver agreements. Methods that help clinicians efficiently and effectively counsel families about teen driving should be developed.
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http://dx.doi.org/10.1136/ip.2010.031112DOI Listing
February 2012

Global turnover of histone post-translational modifications and variants in human cells.

Epigenetics Chromatin 2010 Dec 6;3(1):22. Epub 2010 Dec 6.

Department of Molecular Biology, Princeton University, Princeton NJ, 08544, USA.

Background: Post-translational modifications (PTMs) on the N-terminal tails of histones and histone variants regulate distinct transcriptional states and nuclear events. Whereas the functional effects of specific PTMs are the current subject of intense investigation, most studies characterize histone PTMs/variants in a non-temporal fashion and very few studies have reported kinetic information about these histone forms. Previous studies have used radiolabeling, fluorescence microscopy and chromatin immunoprecipitation to determine rates of histone turnover, and have found interesting correlations between increased turnover and increased gene expression. Therefore, histone turnover is an understudied yet potentially important parameter that may contribute to epigenetic regulation. Understanding turnover in the context of histone modifications and sequence variants could provide valuable additional insight into the function of histone replacement.

Results: In this study, we measured the metabolic rate of labeled isotope incorporation into the histone proteins of HeLa cells by combining stable isotope labeling of amino acids in cell culture (SILAC) pulse experiments with quantitative mass spectrometry-based proteomics. In general, we found that most core histones have similar turnover rates, with the exception of the H2A variants, which exhibit a wider range of rates, potentially consistent with their epigenetic function. In addition, acetylated histones have a significantly faster turnover compared with general histone protein and methylated histones, although these rates vary considerably, depending on the site and overall degree of methylation. Histones containing transcriptionally active marks have been consistently found to have faster turnover rates than histones containing silent marks. Interestingly, the presence of both active and silent marks on the same peptide resulted in a slower turnover rate than either mark alone on that same peptide. Lastly, we observed little difference in the turnover between nearly all modified forms of the H3.1, H3.2 and H3.3 variants, with the notable exception that H3.2K36me2 has a faster turnover than this mark on the other H3 variants.

Conclusions: Quantitative proteomics provides complementary insight to previous work aimed at quantitatively measuring histone turnover, and our results suggest that turnover rates are dependent upon site-specific post-translational modifications and sequence variants.
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http://dx.doi.org/10.1186/1756-8935-3-22DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3004898PMC
December 2010

Choking prevention among young children.

Pediatr Ann 2010 Nov;39(11):721-4

Division of Safety and Health Promotion, American Academy of Pediatrics, Elk Grove Village, IL 60007, USA.

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http://dx.doi.org/10.3928/00904481-20101013-11DOI Listing
November 2010

In vivo residue-specific histone methylation dynamics.

J Biol Chem 2010 Jan 23;285(5):3341-50. Epub 2009 Nov 23.

Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.

Methylation of specific histone residues is capable of both gene activation and silencing. Despite vast work on the function of methylation, most studies either present a static snapshot of methylation or fail to assign kinetic information to specific residues. Using liquid chromatography-tandem mass spectrometry on a high-resolution mass spectrometer and heavy methyl-SILAC labeling, we studied site-specific histone lysine and arginine methylation dynamics. The detection of labeled intermediates within a methylation state revealed that mono-, di-, and trimethylated residues generally have progressively slower rates of formation. Furthermore, methylations associated with active genes have faster rates than methylations associated with silent genes. Finally, the presence of both an active and silencing mark on the same peptide results in a slower rate of methylation than the presence of either mark alone. Here we show that quantitative proteomic approaches such as this can determine the dynamics of multiple methylated residues, an understudied portion of histone biology.
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http://dx.doi.org/10.1074/jbc.M109.063784DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2823435PMC
January 2010

Effects of tango on functional mobility in Parkinson's disease: a preliminary study.

J Neurol Phys Ther 2007 Dec;31(4):173-9

Program in Physical Therapy, Washington University School of Medicine, St. Louis, Missouri, USA.

Recent research has shown that dance, specifically tango, may be an appropriate and effective strategy for ameliorating functional mobility deficits in people who are frail and elderly. Individuals with Parkinson's disease (PD) experience declines in functional mobility that may be even more pronounced than those experienced by frail elderly individuals without PD. The purpose of this study was to compare the effects of two movement programs: tango classes or exercise classes. Nineteen subjects with PD were randomly assigned to a tango group or a group exercise class representative of the current classes offered in our geographical area for individuals with PD. Subjects completed a total of 20 tango or exercise classes and were evaluated the week before and the week following the intervention. Both groups showed significant improvements in overall Unified Parkinson's Disease Rating Scale (UPDRS) score and nonsignificant improvements in self-reported Freezing of Gait. In addition, the tango group showed significant improvements on the Berg Balance Scale. The exercise group did not improve on this measure. Finally, the tango group showed a trend toward improvement on the Timed Up and Go test that was not observed in the exercise group. Future studies with a larger sample are needed to confirm and extend our observation that tango may be an effective intervention to target functional mobility deficits in individuals with PD.
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http://dx.doi.org/10.1097/NPT.0b013e31815ce78bDOI Listing
December 2007

Prefrontal Cortex, Emotion, and Approach/Withdrawal Motivation.

Soc Personal Psychol Compass 2008 Jan;2(1):135-153

University of Illinois at Urbana-Champaign.

This article provides a selective review of the literature and current theories regarding the role of prefrontal cortex, along with some other critical brain regions, in emotion and motivation. Seemingly contradictory findings have often appeared in this literature. Research attempting to resolve these contradictions has been the basis of new areas of growth and has led to more sophisticated understandings of emotional and motivational processes as well as neural networks associated with these processes. Progress has, in part, depended on methodological advances that allow for increased resolution in brain imaging. A number of issues are currently in play, among them the role of prefrontal cortex in emotional or motivational processes. This debate fosters research that will likely lead to further refinement of conceptualizations of emotion, motivation, and the neural processes associated with them.
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http://dx.doi.org/10.1111/j.1751-9004.2007.00064.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2889703PMC
January 2008
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