Publications by authors named "Nicholas Conley"

16 Publications

  • Page 1 of 1

A Pooled Case-only Analysis of Reproductive Risk Factors and Breast Cancer Subtype Among Black Women in the Southeastern United States.

Cancer Epidemiol Biomarkers Prev 2021 Jul 4;30(7):1416-1423. Epub 2021 May 4.

Vanderbilt-Ingram Cancer Center, Nashville, Tennessee.

Background: We investigated the association between reproductive risk factors and breast cancer subtype in Black women. On the basis of the previous literature, we hypothesized that the relative prevalence of specific breast cancer subtypes might differ according to reproductive factors.

Methods: We conducted a pooled analysis of 2,188 (591 premenopausal, 1,597 postmenopausal) Black women with a primary diagnosis of breast cancer from four studies in the southeastern United States. Breast cancers were classified by clinical subtype. Case-only polytomous logistic regression models were used to estimate ORs and 95% confidence intervals (CI) for HER2 and triple-negative breast cancer (TNBC) status in relation to estrogen receptor-positive (ER)/HER2 status (referent) for reproductive risk factors.

Results: Relative to women who had ER/HER2 tumors, women who were age 19-24 years at first birth (OR, 1.78; 95% CI, 1.22-2.59) were more likely to have TNBC. Parous women were less likely to be diagnosed with HER2 breast cancer and more likely to be diagnosed with TNBC relative to ER/HER2 breast cancer. Postmenopausal parous women who breastfed were less likely to have TNBC [OR, 0.65 (95% CI, 0.43-0.99)].

Conclusions: This large pooled study of Black women with breast cancer revealed etiologic heterogeneity among breast cancer subtypes.

Impact: Black parous women who do not breastfeed are more likely to be diagnosed with TNBC, which has a worse prognosis, than with ER/HER2 breast cancer.
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http://dx.doi.org/10.1158/1055-9965.EPI-20-1784DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8254754PMC
July 2021

The Property-Based Practical Applications and Solutions of Genetically Encoded Acetylcholine and Monoamine Sensors.

J Neurosci 2021 03 24;41(11):2318-2328. Epub 2021 Feb 24.

Department of Pharmacology, University of Virginia School of Medicine, Charlottesville, Virginia 22908.

Neuromodulatory communication among various neurons and non-neuronal cells mediates myriad physiological and pathologic processes, yet defining regulatory and functional features of neuromodulatory transmission remains challenging because of limitations of available monitoring tools. Recently developed genetically encoded neuromodulatory transmitter sensors, when combined with superresolution and/or deconvolution microscopy, allow the first visualization of neuromodulatory transmission with nanoscale or microscale spatiotemporal resolution. and experiments have validated several high-performing sensors to have the qualities necessary for demarcating fundamental synaptic properties of neuromodulatory transmission, and initial analysis has unveiled unexpected fine control and precision of neuromodulation. These new findings underscore the importance of synaptic dynamics in synapse-, subcellular-, and circuit-specific neuromodulation, as well as the prospect of genetically encoded transmitter sensors in expanding our knowledge of various behaviors and diseases, including Alzheimer's disease, sleeping disorders, tumorigenesis, and many others.
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http://dx.doi.org/10.1523/JNEUROSCI.1062-19.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7984589PMC
March 2021

High School Students as Citizen Scientists to Decrease Radon Exposure.

Int J Environ Res Public Health 2020 12 8;17(24). Epub 2020 Dec 8.

Center for Environmental Genetics, University of Cincinnati, Cincinnati, OH 45267, USA.

Residents in rural Kentucky (KY) and suburban Ohio (OH) expressed concerns about radon exposure and lung cancer. Although 85% of lung cancer cases are caused by tobacco smoke, radon exposure accounts for 10-15% of lung cancer cases. Academic and community members from the University of KY and the University of Cincinnati developed and pilot-tested a family-centered, youth-engaged home radon testing toolkit. The radon toolkit included radon information, and how to test, interpret, and report back findings. We educated youth as citizen scientists and their teachers in human subjects protection and home radon testing using the toolkit in the classroom. Youth citizen scientists explained the study to their parents and obtained informed consent. One hundred students were trained in human subjects protection, 27 had parental permission to be citizen scientists, and 18 homeowners completed surveys. Radon values ranged from < 14.8 Bq/m to 277.5 Bq/m. Youth were interested and engaged in citizen science and this family-centered, school-based project provided a unique opportunity to further the healthy housing and quality education components of the Sustainable Development Goals for 2030. Further research is needed to test the impact of student-led, family-centered citizen science projects in environmental health as part of school curricula.
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http://dx.doi.org/10.3390/ijerph17249178DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763282PMC
December 2020

The Paraventricular Hypothalamus Regulates Satiety and Prevents Obesity via Two Genetically Distinct Circuits.

Neuron 2019 05 14;102(3):653-667.e6. Epub 2019 Mar 14.

Division of Endocrinology, Diabetes, and Metabolism, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02215, USA. Electronic address:

SIM1-expressing paraventricular hypothalamus (PVH) neurons are key regulators of energy balance. Within the PVH population, melanocortin-4 receptor-expressing (PVH) neurons are known to regulate satiety and bodyweight, yet they account for only half of PVH neuron-mediated regulation. Here we report that PVH prodynorphin-expressing (PVH) neurons, which notably lack MC4Rs, function independently and additively with PVH neurons to account for the totality of PVH neuron-mediated satiety. Moreover, PVH neurons are necessary for prevention of obesity in an independent but equipotent manner to PVH neurons. While PVH and PVH neurons both project to the parabrachial complex (PB), they synaptically engage distinct efferent nodes, the pre-locus coeruleus (pLC), and central lateral parabrachial nucleus (cLPBN), respectively. PB-projecting PVH neurons, like PVH neurons, receive input from interoceptive ARC neurons, respond to caloric state, and are sufficient and necessary to control food intake. This expands the CNS satiety circuitry to include two non-overlapping PVH to hindbrain circuits.
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http://dx.doi.org/10.1016/j.neuron.2019.02.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6508999PMC
May 2019

Developmental phosphoproteomics identifies the kinase CK2 as a driver of Hedgehog signaling and a therapeutic target in medulloblastoma.

Sci Signal 2018 09 11;11(547). Epub 2018 Sep 11.

Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.

A major limitation of targeted cancer therapy is the rapid emergence of drug resistance, which often arises through mutations at or downstream of the drug target or through intrinsic resistance of subpopulations of tumor cells. Medulloblastoma (MB), the most common pediatric brain tumor, is no exception, and MBs that are driven by sonic hedgehog (SHH) signaling are particularly aggressive and drug-resistant. To find new drug targets and therapeutics for MB that may be less susceptible to common resistance mechanisms, we used a developmental phosphoproteomics approach in murine granule neuron precursors (GNPs), the developmental cell of origin of MB. The protein kinase CK2 emerged as a driver of hundreds of phosphorylation events during the proliferative, MB-like stage of GNP growth, including the phosphorylation of three of the eight proteins commonly amplified in MB. CK2 was critical to the stabilization and activity of the transcription factor GLI2, a late downstream effector in SHH signaling. CK2 inhibitors decreased the viability of primary SHH-type MB patient cells in culture and blocked the growth of murine MB tumors that were resistant to currently available Hh inhibitors, thereby extending the survival of tumor-bearing mice. Because of structural interactions, one CK2 inhibitor (CX-4945) inhibited both wild-type and mutant CK2, indicating that this drug may avoid at least one common mode of acquired resistance. These findings suggest that CK2 inhibitors may be effective for treating patients with MB and show how phosphoproteomics may be used to gain insight into developmental biology and pathology.
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http://dx.doi.org/10.1126/scisignal.aau5147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6475502PMC
September 2018

Neuropilin-2 contributes to tumorigenicity in a mouse model of Hedgehog pathway medulloblastoma.

J Neurooncol 2013 Nov 12;115(2):161-8. Epub 2013 Sep 12.

Department of Neurosurgery, Stanford University School of Medicine, Stanford, CA, USA,

The Hedgehog (Hh) signaling pathway has been implicated in the most common childhood brain tumor, medulloblastoma (MB). Given the toxicity of post-surgical treatments for MB, continued need exists for new, targeted therapies. Based upon our finding that Neuropilin (Nrp) transmembrane proteins are required for Hh signal transduction, we investigated the role of Nrp in MB cells. Cultured cells derived from a mouse Ptch (+/-) ;LacZ MB (Med1-MB), effectively modeled the Hh pathway-related subcategory of human MBs in vitro. Med1-MB cells maintained constitutively active Hh target gene transcription, and consistently formed tumors within one month after injection into mouse cerebella. The proliferation rate of Med1-MBs in culture was dependent upon Nrp2, while reducing Nrp1 function had little effect. Knockdown of Nrp2 prior to cell implantation significantly increased mouse survival, compared to transfection with a non-targeting siRNA. Knocking down Nrp2 specifically in MB cells avoided any direct effect on tumor vascularization. Nrp2 should be further investigated as a potential target for adjuvant therapy in patients with MB.
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http://dx.doi.org/10.1007/s11060-013-1216-1DOI Listing
November 2013

A selenium analogue of firefly D-luciferin with red-shifted bioluminescence emission.

Angew Chem Int Ed Engl 2012 Apr 17;51(14):3350-3. Epub 2012 Feb 17.

Department of Chemistry, Stanford University, CA 94305-5080, USA.

A selenium analogue of amino-D-luciferin, aminoseleno-D-luciferin, is synthesized and shown to be a competent substrate for the firefly luciferase enzyme. It has a red-shifted bioluminescence emission maximum at 600 nm and is suitable for bioluminescence imaging studies in living subjects.
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http://dx.doi.org/10.1002/anie.201105653DOI Listing
April 2012

Sensing cooperativity in ATP hydrolysis for single multisubunit enzymes in solution.

Proc Natl Acad Sci U S A 2011 Oct 6;108(41):16962-7. Epub 2011 Sep 6.

Department of Chemistry, Stanford University, Stanford, CA 94305, USA.

In order to operate in a coordinated fashion, multisubunit enzymes use cooperative interactions intrinsic to their enzymatic cycle, but this process remains poorly understood. Accordingly, ATP number distributions in various hydrolyzed states have been obtained for single copies of the mammalian double-ring multisubunit chaperonin TRiC/CCT in free solution using the emission from chaperonin-bound fluorescent nucleotides and closed-loop feedback trapping provided by an Anti-Brownian ELectrokinetic trap. Observations of the 16-subunit complexes as ADP molecules are dissociating shows a peak in the bound ADP number distribution at 8 ADP, whose height falls over time with little shift in the position of the peak, indicating a highly cooperative ADP release process which would be difficult to observe by ensemble-averaged methods. When AlFx is added to produce ATP hydrolysis transition state mimics (ADP·AlFx) locked to the complex, the peak at 8 nucleotides dominates for all but the lowest incubation concentrations. Although ensemble averages of the single-molecule data show agreement with standard cooperativity models, surprisingly, the observed number distributions depart from standard models, illustrating the value of these single-molecule observations in constraining the mechanism of cooperativity. While a complete alternative microscopic model cannot be defined at present, the addition of subunit-occupancy-dependent cooperativity in hydrolysis yields distributions consistent with the data.
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http://dx.doi.org/10.1073/pnas.1112244108DOI Listing
October 2011

Molecules and methods for super-resolution imaging.

Methods Enzymol 2010 ;475:27-59

Department of Chemistry, Stanford University, Stanford, California, USA.

By looking at a fluorescently labeled structure one molecule at a time, it is possible to side-step the optical diffraction limit and obtain "super-resolution" images of small nanostructures. In the Moerner Lab, we seek to develop both molecules and methods to extend super-resolution fluorescence imaging. Methodologies and protocols for designing and characterizing fluorophores with switchable fluorescence required for super-resolution imaging are reported. These fluorophores include azido-DCDHF molecules, covalently linked Cy3-Cy5 dimers, and also the first example of a photoswitchable fluorescent protein, enhanced yellow fluorescent protein (EYFP). The imaging of protein superstructures in living Caulobacter crescentus bacteria is used as an example of the power of super-resolution imaging by single-molecule photoswitching to extract information beyond the diffraction limit. Finally, a new method is described for obtaining three-dimensional super-resolution information using a double-helix point-spread function.
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http://dx.doi.org/10.1016/S0076-6879(10)75002-3DOI Listing
October 2010

Genetic and functional assessment of the role of the rs13431652-A and rs573225-A alleles in the G6PC2 promoter that are strongly associated with elevated fasting glucose levels.

Diabetes 2010 Oct 9;59(10):2662-71. Epub 2010 Jul 9.

CNRS-UMR-8199, Institut Pasteur de Lille, Lille, France.

Objective: Genome-wide association studies have identified a single nucleotide polymorphism (SNP), rs560887, located in a G6PC2 intron that is highly correlated with variations in fasting plasma glucose (FPG). G6PC2 encodes an islet-specific glucose-6-phosphatase catalytic subunit. This study examines the contribution of two G6PC2 promoter SNPs, rs13431652 and rs573225, to the association signal.

Research Design And Methods: We genotyped 9,532 normal FPG participants (FPG <6.1 mmol/l) for three G6PC2 SNPs, rs13431652 (distal promoter), rs573225 (proximal promoter), rs560887 (3rd intron). We used regression analyses adjusted for age, sex, and BMI to assess the association with FPG and haplotype analyses to assess comparative SNP contributions. Fusion gene and gel retardation analyses characterized the effect of rs13431652 and rs573225 on G6PC2 promoter activity and transcription factor binding.

Results: Genetic analyses provide evidence for a strong contribution of the promoter SNPs to FPG variability at the G6PC2 locus (rs13431652: β = 0.075, P = 3.6 × 10(-35); rs573225 β = 0.073 P = 3.6 × 10(-34)), in addition to rs560887 (β = 0.071, P = 1.2 × 10(-31)). The rs13431652-A and rs573225-A alleles promote increased NF-Y and Foxa2 binding, respectively. The rs13431652-A allele is associated with increased FPG and elevated promoter activity, consistent with the function of G6PC2 in pancreatic islets. In contrast, the rs573225-A allele is associated with elevated FPG but reduced promoter activity.

Conclusions: Genetic and in situ functional data support a potential role for rs13431652, but not rs573225, as a causative SNP linking G6PC2 to variations in FPG, though a causative role for rs573225 in vivo cannot be ruled out.
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http://dx.doi.org/10.2337/db10-0389DOI Listing
October 2010

Azido push-pull fluorogens photoactivate to produce bright fluorescent labels.

J Phys Chem B 2010 Nov 27;114(45):14157-67. Epub 2009 Oct 27.

Department of Chemistry, Stanford University, Stanford, California 94305-5080, USA.

Dark azido push-pull chromophores have the ability to be photoactivated to produce bright fluorescent labels suitable for single-molecule imaging. Upon illumination, the aryl azide functionality in the fluorogens participates in a photochemical conversion to an aryl amine, thus restoring charge-transfer absorption and fluorescence. Previously, we reported that one compound, DCDHF-V-P-azide, was photoactivatable. Here, we demonstrate that the azide-to-amine photoactivation process is generally applicable to a variety of push-pull chromophores, and we characterize the photophysical parameters including photoconversion quantum yield, photostability, and turn-on ratio. Azido push-pull fluorogens provide a new class of photoactivatable single-molecule probes for fluorescent labeling and super-resolution microscopy. Lastly, we demonstrate that photoactivated push-pull dyes can insert into bonds of nearby biomolecules, simultaneously forming a covalent bond and becoming fluorescent (fluorogenic photoaffinity labeling).
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http://dx.doi.org/10.1021/jp907080rDOI Listing
November 2010

DCDHF fluorophores for single-molecule imaging in cells.

Chemphyschem 2009 Jan;10(1):55-65

Department of Chemistry, Stanford University, Stanford, California 94305, USA.

There is a persistent need for small-molecule fluorescent labels optimized for single-molecule imaging in the cellular environment. Application of these labels comes with a set of strict requirements: strong absorption, efficient and stable emission, water solubility and membrane permeability, low background emission, and red-shifted absorption to avoid cell autofluorescence. We have designed and characterized several fluorophores, termed "DCDHF" fluorophores, for use in live-cell imaging based on the push-pull design: an amine donor group and a 2-dicyanomethylene-3-cyano-2,5-dihydrofuran (DCDHF) acceptor group, separated by a pi-rich conjugated network. In general, the DCDHF fluorophores are comparatively photostable, sensitive to local environment, and their chemistries and photophysics are tunable to optimize absorption wavelength, membrane affinity, and solubility. Especially valuable are fluorophores with sophisticated photophysics for applications requiring additional facets of control, such as photoactivation. For example, we have reengineered a red-emitting DCDHF fluorophore so that it is dark until photoactivated with a short burst of low-intensity violet light. This molecule and its relatives provide a new class of bright photoactivatable small-molecule fluorophores, which are needed for super-resolution imaging schemes that require active control (here turning-on) of single-molecule emission.
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http://dx.doi.org/10.1002/cphc.200800581DOI Listing
January 2009

Cy3-Cy5 covalent heterodimers for single-molecule photoswitching.

J Phys Chem B 2008 Sep 28;112(38):11878-80. Epub 2008 Aug 28.

Department of Chemistry, Stanford University, Stanford, California, USA.

Covalent heterodimers of the Cy3 and Cy5 fluorophores have been prepared from commercially available starting materials and characterized at the single-molecule level. This system behaves as a discrete molecular photoswitch, in which photoexcitation of the Cy5 results in fluorescence emission or, with a much lower probability, causes the Cy5 to enter into a long-lived, but metastable, dark state. Photoinduced recovery of the emissive Cy5 is achieved by very low intensity excitation (5 W cm(-2)) of the Cy3 fluorophore at a shorter wavelength. A similar system consisting of proximal, but not covalently linked, Cy3 and Cy5 has found application in stochastic optical reconstruction microscopy (STORM), a single-molecule localization-based technique for super-resolution imaging that requires photoswitching. The covalent Cy3-Cy5 heterodimers described herein eliminate the need for probabilistic methods of situating the Cy3 and Cy5 in close proximity to enable photoswitching. As proof of principle, these heterodimers have been applied to super-resolution imaging of the tubular stalk structures of live Caulobacter crescentus bacterial cells.
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http://dx.doi.org/10.1021/jp806698pDOI Listing
September 2008

A photoactivatable push-pull fluorophore for single-molecule imaging in live cells.

J Am Chem Soc 2008 Jul 24;130(29):9204-5. Epub 2008 Jun 24.

Department of Chemistry, Stanford University, Stanford, California 94305-5080, USA.

We have reengineered a red-emitting dicyanomethylenedihydrofuran push-pull fluorophore so that it is dark until photoactivated with a short burst of low-intensity violet light. Photoactivation of the dark fluorogen leads to conversion of an azide to an amine, which shifts the absorption to long wavelengths. After photoactivation, the fluorophore is bright and photostable enough to be imaged on the single-molecule level in living cells. This proof-of-principle demonstration provides a new class of bright photoactivatable fluorophores, as are needed for super-resolution imaging schemes that require active control of single molecule emission.
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http://dx.doi.org/10.1021/ja802883kDOI Listing
July 2008

Bulk and single-molecule characterization of an improved molecular beacon utilizing H-dimer excitonic behavior.

J Phys Chem B 2007 Jul 21;111(28):7929-31. Epub 2007 Jun 21.

Pairs of fluorophores in close proximity often show self-quenching of fluorescence by the well-known H-dimer mechanism. We use a pair of fluorophores in the new dicyanomethylenedihydrofuran (DCDHF) dye family in the design and characterization of a new fluorescent probe for nucleic acid detection, which we refer to as a self-quenched intramolecular dimer (SQuID) molecular beacon (MB). We obtain a quenching efficiency of 97.2%, higher than the only other reported value for a MB employing fluorophore self-quenching by H-dimer formation. Furthermore, the excellent single-molecule (SM) emitter characteristics of the DCDHF dyes allow observation of individual SQuID MB-target complexes immobilized on a surface, where the doubled SM emission intensity of our target-bound beacon ensures a higher signal-to-background ratio than conventional fluorophore-quencher MBs. Additional advantages of the SQuID MB are single-pot labeling, visible colorimetric detection of the target, and intrinsic single-molecule two-step photobleaching behavior, which offers a specific means of discriminating between functional MBs and spurious fluorescence.
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http://dx.doi.org/10.1021/jp073310dDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2663424PMC
July 2007

Toward nanometer-scale optical photolithography: utilizing the near-field of bowtie optical nanoantennas.

Nano Lett 2006 Mar;6(3):355-60

E.L. Ginzton Laboratory and Department of Chemistry, Stanford University, California 94305, USA.

Optically resonant metallic bowtie nanoantennas are utilized as fabrication tools for the first time, resulting in the production of polymer resist nanostructures <30 nm in diameter at record low incident multiphoton energy densities. The nanofabrication is accomplished via nonlinear photopolymerization, which is initiated by the enhanced, confined optical fields surrounding the nanoantenna. The position, size, and shape of the resist nanostructures directly correlate with rigorous finite-difference time-domain computations of the field distribution, providing a nanometer-scale measurement of the actual field confinement offered by single optical nanoantennas. In addition, the size of the photoresist regions yields strong upper bounds on photoacid diffusion and resist resolution in SU-8, demonstrating a technique that can be generalized to the study of many current and yet-to-be-developed photoresist systems.
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http://dx.doi.org/10.1021/nl052322cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1447673PMC
March 2006
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