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Distributed Drug Discovery (D3): Education of Undergraduate Students in Drug Discovery Research Directed toward Neglected Diseases
Ryan Denton, Milata M. Abraham, Gregory G. Anderson , Jon M. Carnahan, Courtney E. Glos, Richard W. Harper, Jonathan M. LaCombe, Kathleen A. Marrs, Matthew D. Phillips, J. Geno Samaritoni, Martin J. O'Donnell and William L. Scott, Indiana University Purdue University Indianapolis; Alexus T. Copes, Howard University, DC; Jacob D. Durrant, University of California, San Diego
Presenter: William Scott
(Project funded by National Science Foundation Grant: NSF/DUE-1140602)
Our Goal: Engage Undergraduates in FundamentalResearch and Scientific Habits of Thought in the
Context of Neglected Disease Drug Discovery
What students learn:
• Integrated expertise in chemistry, computational analysis and biological evaluation required for drug discovery
• The critical importance of reproducibility and controls to produce reliable data for use in the scientific process
• To use validated data to make and test their own predictions, solving problems with no predetermined solution
• The concept and powerful application of combinatorial chemistry in both nature and the lab
Students’ Real-World Challenge:Neglected Disease Drug Discovery
Potential malariapatient (nytimes.com)
Cystic fibrosis patient(photos.oregonlive.com)
Afghan leishmaniasispatient (www.rawa.org)
Disease
Treat
Screen (Test)
Acquire moleculesto test
Moleculesfrom Nature
Moleculesfrom synthesis
Scale up production
Identifybest
molecule
Learn fromtest results
Deliverto patient
The Drug Discovery Process
ChemicalSynthesis
AnalyticalChemistry
HumanitarianConcerns
Reproducibility
Informatics Biology andBiochemistry
ComputationalChemistry
Education
NeglectedDisease
DrugDiscovery
Distributed Drug Discovery (D3): The Motivation
Proposedpotential
drug leads
Moleculesfor
screening
Moleculesscreened
LargeD3 virtual catalogs
Stage Two Stage Three Stage Four
Student Computation Many Student Chemists Many Student Biologists
Distribute Combine Distribute Combine Distribute CombineComputationalEnumeration
Stage One
ComputationalAnalysis
ChemicalSynthesis
BiologicalScreening
D3 DirectedBasic
ChemistryResearch
Distributed Problem Solving Process
Grand Plan for Distributed Drug Discovery (D3):Decentralized Interdisciplinary Science Education
Coupled to Neglected Disease Research
Keys to Enabling Authentic Undergraduate ResearchIn Drug Discovery:
• Strong lab foundation from undergrad-faculty research
• Undergraduate lab-proven procedures that permit the use of variable inputs: a) Combinatorial chemistry b) Solid-Phase chemistry
• Synthetic chemistry closely coupled to computational workflows and robust biological assays
• Easily accessible centralized database for distributed student decision making
• Low cost equipment and materials
Simple, Inexpensive Solid-Phase Equipment
Bill-Board 6-pack Bill-Board Drain Tray Collection Vial Rack
FmocNH
R1
O
OH
X
R2
O
OH
HN
R1
O
O
O
R2
Protected Amino Acid R1
1) Wash2) Cleave link to resin
1) Remove Fmoc protecting group2) Wash
Solid-Phase Resin: key to doingmultiple small-scale reactions with simple filtration work-up
R1
FmocNH
O
O
H2N
R1
O
O
Fmoc = Nitrogen protecting group
HN
R1
O
OH
O
R2
Substitute anyof 20 different
amino acids here
4 to 51) Couple R2CO2H2) Wash
1
3
4 5
6
2
Many (e.g. 20 x 100 = 2000)acylated amino acid combinations 6
(3 commercially available startingmaterial supplied to students)(Substitute many
[100's]different carboxylic
acids here)
3 to 4
Combinatorial Example: Modification of Solid-Phase SynthesisProvides Many Possibilities from Simple “Recipe” (D3 Lab 2)
• Scaffolds for enumerated virtual library
• SP07 - SP13 ~780 unique compounds synthesizedby students
• Subset sentto NIH Small Molecule Repository
NO
O
Ph
Ph
NO
O
Ph
Ph
W
W
FmocNHO
O
R1
W
NN
OCl
R3
B
HN
OH
O
R1
R2
O
HN
OMe
O
R1
R2
O
HN
OH
O
R1
R2
O
HN
N
O
R1
R2
O
R3
H
D3 Lab 2: The combinatorial synthesis of acylated natural amino acids
D3 Lab 3: The combinatorial synthesis of acylated unnatural amino acid methyl esters
D3 Lab 4: The combinatorial synthesis of acylated unnatural amino acid amides
D3 Lab 1: The combinatorial synthesis of acylated unnatural amino acids
Cl100 R1 x 100 R2 x 100 R3 x 2 enant.=
2,000,000+ possible compounds
NO
O
Ph
Ph
W H2NOH
O
R1
D3 Lab 6: The combinatorial synthesis of unnatural amino acids (with acylated controls)
100 R1 x 100 R2 x 2 enant.=
20,000+ possible compounds
100 R1 x 100 R2 =
10,000+ possible compounds
100 R1 x 100 R2 x 2 enant.=
20,000+ possible compounds
100 R1 x 2 enant.=
200+ possible compounds
D3 Lab 5: The combinatorial synthesis of acylated unnatural amino acid primary amides
NNH
O
Ph
B
HN
N
O
R1
R2
O
H
H
100 R1 x 100 R2 x 2 enant. =
20,000+ possible compoundsPh
Currently 70,000+ virtual compounds freely available on Collaborative Drug Discovery (CDD) Vault
Large D3 Biomimetic Virtual Catalogs Based on Solid-Phase AccessibleD3 Lab Procedures Fundamental to Enabling UndergraduateStudent Participation in Authentic Research
Student Process:
1. Choose (or are given) a biological target
2. Analyze existing data to develop a hypothesis based on molecular features associated with biological activity
3. Select, from large D3 enabled virtual catalogs, five new molecules to test their hypothesis
4. Synthesize and test, in replicated fashion, a control and the five new molecules
FmocNH
R1
O
OH
X
R2
O
OH
HN
R1
O
O
O
R2
Protected Amino Acid R1
1) Wash2) Cleave link to resin
1) Remove Fmoc protecting group2) Wash
Solid-Phase Resin: key to doingmultiple small-scale reactions with simple filtration work-up
R1
FmocNH
O
O
H2N
R1
O
O
Fmoc = Nitrogen protecting group
HN
R1
O
OH
O
R2
Substitute anyof 20 different
amino acids here
4 to 51) Couple R2CO2H2) Wash
1
3
4 5
6
2
Many (e.g. 20 x 100 = 2000)acylated amino acid combinations 6to populate D3 Lab 2 virtual catalog
(3 commercially available startingmaterial supplied to students)(Substitute many
[100's]different carboxylic
acids here)
3 to 4
Reminder: Synthetic Scheme for D3 Lab 2
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
2 3
4 5 6
HN
O
OH
O
1Student selects 1as a control
HN
O
OH
O
1
Cl
Cl
Cl
Cl
Cl
Plus five follow-up compounds from large D3 Lab 2 virtual catalog
What if 1 (From D3 Lab 2) Were a“Hit” in a Leishmania Assay?
(1 can be synthesizedby D3 Lab 2)
Step 2: Acylatedown columnswith nR2-CO2H
2 x 3 CombinatorialBill-Board products
B
A A1
B1
1 2
B2
A2
B3
A3
3
B1 B2 B3
A1 A2 A3Step 1: Introduce anddeprotect Fmoc protectedamino acid resins in rows
HN
O
OFmoc
HN
O
OFmoc
OH
O
OH
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
HN
O
OH
O
A1 A2 A3
B1 B2 B3
Combinatorial Bill-Board layoutfor student to test their hypothesis
HN
O
OH
O
"Hit" for followup work
Five new molecules proposed, accessible byD3 Lab 2 procedures
Cl
Cl
Cl
Cl
Cl
Cl
Cl
• All students get same reagent for Row A and Column 1: A1 (hit) now becomes the “control”• Different reagents for columns 2&3• (And for Row B too) = 5 new
cmpds• Each Bill-Board combinatorial arrangement is replicated in at least one other Bill-Board
Capping bottoms of reaction vessels Adding reagents to reaction vessels
Washing out reagents and by-products Celebrating success!
Biological Evaluation of D3 Student Compounds
I. By undergraduate D3 biology students(these will soon be D3 biology procedures)
• Agar disc plate assay for antibiotics• Assay for bacterial biofilm production and destruction (to treat cystic fibrosis)
II. By labs with more sophisticated equipment
• Malaria and Leishmania: National Institutes of Health (“NIH”)• Type II diabetes: IU medical school
Assessment
Performed by the IUPUI Center for Urban and Multicultural Education
• Quantitative (surveys, scored open response questions)• Qualitative (surveys, focus groups, interviews, lab observations)
Strengths (from Year One Report formative feedback)• interdisciplinary nature enhances openness to novelty• encourages autonomous learning & scientific imagination• humanitarian value promotes deepening of learning motivations
Challenges, Implementations, and Process of Change• students’ sense of competence• instructional adjustment attempting to create an optimal level of challenge
for student learning within time constraints• the difficulties of meeting multiple standards as well as tailoring the
program to fit individual student learning capacity
YOU & >1,250 PREVIOUS IUPUI C344 STUDENTS (Undergraduates Underlined, People Participating in Workshop Asterisked)
Chemistry Development (IUPUI) Computational Development*Marty O’Donnell *Jacob Durrant (UCSD)*Ryan Denton Collaborative Drug Discovery (CDD)Richard Harper ChemAxon*Geno Samaritoni*Shelby Colglazier Laboratory Management*Julian Dilley Waiping Kam (C344)Matt Fries Grady Chism (Biology)Matt PhillipsAlexus Copes (Howard) Analytical (Eli Lilly)DeMarcus Crews (Morehouse) Chris ReutterCalvin Mendel (Northwestern) Keith Burton
Veronika Cerninova (Palacky, Czech Rep.)Monika Tomanova (Palacky, Czech Rep.) CUME (Assessment)
Ying-Yi Chou48 Undergraduate Researchers John Houser34 Graduate Theses Robert Helfenbein 23 Postdoctoral Fellows
FundingBiology Development (IUPUI) National Science Foundation (NSF/DUE-1140602)*Kathy Marrs IUPUI Summer HBCU STEM Scholars ProgramGregory Anderson National Institutes of Health (RO1-GM28193)Jonathan LaCombeAnna BrennemanVictoria Rarity
