Investigational Drug Steering Committee Team Science Meeting 11 Jan, - - PowerPoint PPT Presentation

Investigational Drug Steering Committee Team Science Meeting 11 Jan, 2013 11 Jan, 2013

This work was performed under the auspices of the U.S. Department of Energy by Lawrence Livermore National Laboratory under contract Livermore National Laboratory under contract DE‐AC52‐07NA27344. Lawrence Livermore National Security, LLC

• Is a multi‐sponsor FFRDC, managed by NNSA
• Our role…
• Provide enduring focus on issues in national security
• Leverage multidisciplinary capabilities

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Laser Sciences & Isotopic Sciences (National Ignition Facility)

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High Performance Computing

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Analytical and Measurement Sciences Analytical and Measurement Sciences

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Material Sciences

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Advanced Engineering, Instrumentation & Diagnostics

Lawrence Livermore National Laboratory

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Biosciences

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1073 551 18 633

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• Multidisciplinary teams including bioscientists engineers computational

Multidisciplinary teams including bioscientists, engineers, computational scientists, physicists, chemists . . .

• Working at the interface of engineering, materials, chemical and

g g g, , biological sciences

• Detection & diagnostics – detection and surveillance platforms, fit‐for‐

purpose arrays…

• High precision measurement/Instrumentation— Accelerator Mass

S i l l l i f b i i & Spectrometry, single molecule spectroscopy, microfabrication & engineering

• Computational biology

Massively parallel computing resources and

• Computational biology—Massively parallel computing resources and

biocomputing expertise

• Expertise in medical devices, assay development, genomics, biological

Lawrence Livermore National Laboratory

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Expertise in medical devices, assay development, genomics, biological /chemical threats, and countermeasures

• The original mission was to investigate the effects of ionizing radiation on humans
• Focus on goal driven research, developing technology that is coupled to solving

problems of national significance in health and the environment

1970’s 1980’s 1990’s 2000’s

FISH Chromosome Automated Cell S ti Human Genome Project High Performance Radiation Eff

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Chromosome Painting Sorting Project Performance Simulations Effects

Target discovery Lead generation Lead optimization Preclinical development Phase 0

Now: 15 years Our goal: 5 years

validation generation development

BSL/3 Testing Pathway Characterization AMS Microdosing 3D Organ Culture C t ti l Rapid sample prep & biomarker t Computational Prediction measurement lipid Apolipoprotein functionalization NLP’s for Delivery, Imaging &

Lawrence Livermore National Laboratory

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Lawrence Livermore National Laboratory

6 NLP s for Delivery, Imaging & Functional Studies Structural Characterization

Computational Analysis and Simulation Computational Analysis and Simulation

DNA sequence Protein structure

Homology based

Protein sequence and regulation

T A

• ter

Protein/enzyme function

protein structure prediction Sequence Annotation

Q

T C A G T A

Promo age

Molecular simulations

Y R

C G T T A C

Messa

Organism Pathway

• Expt. data

integration Network g simulations y simulations analysis

Bacterial communities & l i ll l i Bacteria d ll Metabolic pathways & l t t k Multi‐protein & multicellular organisms and cells & regulatory networks machines

HPC is Being Used to Design Affinity Reagents for HPC is Being Used to Design Affinity Reagents for Use in Detection Assays and Therapeutics Use in Detection Assays and Therapeutics

Utilizes DNA signature d i l

Use in Detection Assays and Therapeutics Use in Detection Assays and Therapeutics

and protein structural data to identify target sites.

• More robust detection

Longer shelf life

• Longer shelf‐life
• Better treatments

Protein structure information may allow developing synthetic ligands as interventions

Lawrence Livermore National Laboratory

developing synthetic ligands as interventions

• r detectors.

Why?

Fixed-target support

LCLS detector

Developing Advanced Methods for Structural Developing Advanced Methods for Structural Characterization with XFELs to improve Models Characterization with XFELs to improve Models

Why?

• Allows 2D crystallography
• Membrane proteins in a native

environment!

• Reduced sample consumption

Fixed-target support

LCLS detector

• Reduced sample consumption
• 100s µg vs 10s to 100s mg
• Room temperature protein structures!
• Possibly more biologically

relevant Apply sample

LCLS beam

relevant

3D crystallography results from Thorough characterization of mechanical damage pathways and implications for structure determination proteins involved in putative virulence pathways to atomic resolution determination

Lawrence Livermore National Laboratory

• Administration of low doses (/100th

(/ the anticipated therapeutic dose) to healthy volunteers

• Many benefits including :
• Reduced animal testing
• Reduced drug synthesis
• l

f

• Early assessment of

efficacy/safety

• Difficult without extremely sensitive

Lawrence Livermore National Laboratory

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• Difficult without extremely sensitive

methods of analysis

T f t t th t

• Type of mass spectrometer that

analyzes ions at very high energies through use of a linear acceleration stage

V l 1 l d d

0.0007

g

• Measures isotope ratios

Volunteer 1 – selected data

0.0005 0.0006

Run 2a Run 2b Run 3a Run 2c n (ng/ml)

• Analysis of attomole quantities
• f chemical or biological entities

in µg ‐ mg sized samples with

0 0002 0.0003 0.0004

Run 3b Run 3c sma concentration

in µg mg sized samples with high precision

• Directly determines drug safety

0.0000 0.0001 0.0002

Plas

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• Directly determines drug safety

& relevance in humans

4 8 12 16 20 24

• 0.0001

Time (hours)

Microfluidic chip

Piezoelectric transducer LE channel LE gel electrode TE l l t d sample channel TE channel TE gel electrode

gel electrode edges

focusing of fluorescein

Lawrence Livermore National Laboratory 12

edges

focusing of fluorescein Maxim Shusteff, LLNL

A ib d A Blood Blood serum A ib d A Blood Blood serum Blood Blood serum

Next generation device uses simple microfluidics for smart phone‐based diagnostics Passive Lateral flow array assay

Antibody Array Antibody Array serum serum sample sample Cancer Cancer Antigen Antigen Antibody Array Antibody Array serum serum sample sample Cancer Cancer Antigen Antigen serum serum sample sample Cancer Cancer Antigen Antigen

for smart phone based diagnostics

8”

Porous Membrane Porous Membrane Wicking block Wicking block Conjugate Pad Conjugate Pad containing antibody containing antibody-

• reporter complex

reporter complex Porous Membrane Porous Membrane Wicking block Wicking block Conjugate Pad Conjugate Pad containing antibody containing antibody-

• reporter complex

reporter complex

8” 4”

1 ug, 100 ng, 10 ng. 1 ng

Lawrence Livermore National Laboratory 13

Hand‐held detector for array‐based assays

1st Principal Component for the 4 Biomarkers

25000 15000 20000 25000

Metastatic

5000 10000 10000

• 5000

Early onset Controls

• 15000
• 10000

50 100 150 200

4 biomarkers for breast cancer / n = 40 people per group

Lawrence Livermore National Laboratory

4 biomarkers for breast cancer / n = 40 people per group

Rao, R., et al., (In preparation) Passive-flow breast cancer screens capable of discriminating between normal from metastatic breast cancer in serum.

i l i

5,964 microbial species l b

• 3,368 viral species
• 2,223 bacterial species
• 136 fungi

387,156 total probes

• Probes are 50‐65 bases long
• Unique regions from viral and

A Microbial Detection Array (MDA) for Viral and Bacterial Detection.

bacterial sequences used

• >15‐50 probes per each

sequence

Gardner SN, Jaing CJ, McLoughlin KS, Slezak TR. BMC Genomics 2010, 11:668doi:10.1186/1471‐2164‐11‐668, published Nov 25, 2010. http://www.biomedcentral.com/1471‐2164/11/668.

Enabled by Bioinformatics

• Expertise in DNA signatures

and biostatistics

• More than 20 collaborations with academia, government

d h ti l i

• More than 20 collaborations with academia, government

d h ti l i

Lawrence Livermore National Laboratory

• Large Cluster computing used

agency and pharmaceutical companies

• Multiple licensing discussions underway.

agency and pharmaceutical companies

• Multiple licensing discussions underway.

Non-Hodgkin’s lymphoma Non Hodgkin s lymphoma, malignancy of bone marrow-derived cells which results in unregulated replication and expansion of these

• Drs. Ralph Greene and Mingyi Chen; Postdoc Joe Tellez

p p cells in lymphoid tissue and beyond.

• Analyze viruses and/or bacteria in control lymphoid tissues, indolent lymphomas and

aggressive lymphomas

• Identify unique groups of commensal and pathogenic microbial species that might

y q g p p g p g be associated with each sample group

• Compare the relative abundances of microbial families between normal vs indolent

vs progressive lymphomas to determine whether higher abundance of certain

Lawrence Livermore National Laboratory

p g y p g microbes in tissues correlates with lymphomas and the progression of lymphomas.

• Dr. Crystal Jaing, LLNL

• Facilitate strain‐collection

access via LLNL collaborations with sequencing centers

• Conduct shift/drift and genetic

engineering detection studies g g

• Protein structure function

pipeline

Genomics Validated assays Bioinformatics

Lawrence Livermore National Laboratory

• Eliminate long ramp times by porous media flow
• Allows PCR to be governed by diffusion and enzyme processivity
• Optical (real‐time) detection
• Architecture can be ruggedized and miniaturized
• Target <10 minutes STA ‐ including sample prep & detection

Disposable plastic insert for PCR Fl idi f i

Lawrence Livermore National Laboratory

Disposable plastic insert for PCR (5μL wells) Fluidics system for convective heat transfer

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• Dr. Reginald Beer, LLNL

Enables Studies of:

Microfluidic Cell Culture Platform Microfluidic Cell Culture Platform

• Host Cell‐Pathogen interactions
• Viral Evolution, anti‐infective

resistance… h d h l

Cells in P f i Ch b

• Biochemistry and Physiology
• Drug safety

Perfusion Chamber Micro‐Fluidic Cell Culture Chip

• 16 devices per chip
• Each device consists of 16 addressable chambers
• Each chamber has ~150 pL, total Device ~5 nL

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Collaborations with the University of California, Collaborations with the University of California, Davis Comprehensive Cancer Center Davis Comprehensive Cancer Center p

NLPs for imaging, immune modulation and drug delivery Accelerator Mass Spectrometer for High density microarrays Chemotherapy/diagnostic development g y y for viral oncology Optical Biopsy Endoscopy Proton radiotherapy Computational Biology for Rational Drug Design

• We hire people with the personality that fits the team environment

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Training begins with Postdoctoral scientists g g

• Team work is a key part of performance management

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We reward teams as much as we do individuals

• We administratively organize to allow fluid movement of SME’s among

programs/projects

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We use a matrix environment where every person fits in a discipline organization which is responsible for career management and is also assigned to a program/project where that subject matter expertise is

• ca ee

a age e t a d s a so ass g ed to a p og a /p oject e e t at subject atte e pe t se s needed

• Facilities are organized to ensure interaction among disciplines

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Multi disciplinary teams are either housed together p y g

• Career management considers the team environment

Lawrence Livermore National Laboratory

• Enthusiastic Leadership

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Needs to be well understood responsibilities

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Passion for the project

• Communication, communication and more communication..
• Working towards a common vision

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Group developed understanding of the problem, critical path, challenges

• Setting goals and understanding requirements

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Everyone needs to have the same understanding

• Understanding strengths and weaknesses of the team

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Leadership needs to understand this

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Collaboration

• Mentoring
• Rewarding success for the team

Lawrence Livermore National Laboratory

• Strong Management support

• Most of us were not trained to work in teams
• Keeping communication and interaction among all team members

going g g

• Walking the line between individual needs and project needs
• Reward team success as often as we do individual
• Reward team success as often as we do individual
• We emphasize long-term, high risk projects that are large but

support individual

Lawrence Livermore National Laboratory

Lawrence Livermore National Laboratory

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Work at the interface of the physical chemical biological and Work at the interface of the physical chemical biological and

• Characterize key biochemical and molecular networks and enable

Work at the interface of the physical, chemical, biological and Work at the interface of the physical, chemical, biological and engineering sciences to solve important problems in public health engineering sciences to solve important problems in public health y simulation of biological networks, definition of new biomarkers, and understanding of disease processes

• Develop advanced deployable instrumentation for measurement of

indicators of pathogens, early disease detection, and toxic chemical exposure

• Develop and translate technologies that will accelerate the preclinical

development phases of drug development and more accurately predict safety and efficacy in humans safety and efficacy in humans

• Collaborate to quickly discover and deploy new countermeasures

(assays devices and therapeutics)

Lawrence Livermore National Laboratory

(assays, devices and therapeutics)

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Programmatic and National Programmatic and National Impact Impact Foundational S&T Foundational S&T p

BioWatch Threat Agent Science and Novel Assays Handheld PCR NARAC/IMAAC Joint Enabling Bioinformatics & Micro arrays Joint Genome Institute Micro‐arrays Bio‐Instrumentation Biodefense Knowledge Center Forensic Science Center Computational Biology & Molecular Modeling Lawrence Livermore National Laboratory 26 Forensic Science Center

Tumor visualization lipid Nanodisc with attached telodendrimer chemistry Cryo EM to image 20 nM disc

300 450 600

RFU

A li i Telodendrimer

25 150 300

Apolipoprotein

• Novel cell‐free methods for the rapid assembly of apolipoproteins into <25 nm for in vitro

and in vivo biochemistry.

• Telodendrimer chemistry to specifically tune the size and monodispersity of the nanodisc.

The telodendrimer also allows a platform for the attachment of targeting peptides labels The telodendrimer also allows a platform for the attachment of targeting peptides, labels and immunomodulatory drugs.

• These disc have shown to be a valuable resource for the production and solubilization of

membrane bound proteins. They have also proven useful for in vivo delivery of drugs and

Lawrence Livermore National Laboratory

membrane bound proteins. They have also proven useful for in vivo delivery of drugs and imaging reagents.

• Surface of virus exclusively displays E protein

Model antigen ‐ West Nile virus envelop protein (E): In vivo studies (Swiss Webster mice) Serology Mortality/ bidi y p y p

• His‐tagged, truncated E (trE) conjugated to NiNLPs

α‐trE Ab Production

Single i.p. inoculation (2.5µg)

Serology WNV

challenge

morbidity assessment

Survival

Fischer NO et al (2010) Bioconjugate Chem. 21, 1018‐22

Diluent n=15 NiNLP 5

Lawrence Livermore National Laboratory 28

NiNLP n=5 trE n=15 trE:NiNLP n=15

• Dr. Nicholas Fischer, LLNL

Out of hospital diagnosis for traumatic chest injuries

Chest x ray of a pneumothorax. [Ref. Educational

Early device Concept

Computing for Health Technologies teaching web site, Michigan State University] Note the pleural line visible

• n the patient’s right side (image left side).

LLNL in collaboration with Electrosonics Medical, Inc, Detroit Sinai‐Grace Hospital, and Detroit Receiving Hospital

Currently CRADA 2007 R&D 100 Award, 8 d l b C i d Lawrence Livermore National Laboratory 2008 Federal Laboratory Consortium Award

John Chang, LLNL