Table Of ContentWeihong Tan · Xiaohong Fang E ditors
Aptamers
Selected by
Cell-SELEX for
Theranostics
Aptamers Selected by Cell-SELEX
for Theranostics
Weihong Tan Xiaohong Fang
(cid:129)
Editors
Aptamers Selected
by Cell-SELEX
for Theranostics
123
Editors
WeihongTan Xiaohong Fang
Molecular Science andBiomedicine Instituteof Chemistry
Laboratory,StateKey Laboratoryfor ChineseAcademy ofSciences
Chemo/Bio-Sensingand Chemometrics Beijing
Collegeof Chemistryand Chemical China
Engineering
HunanUniversity
Changsha
China
and
Department of Chemistry
Universityof Florida
Gainesville, FL
USA
ISBN 978-3-662-46225-6 ISBN 978-3-662-46226-3 (eBook)
DOI 10.1007/978-3-662-46226-3
LibraryofCongressControlNumber:2015931245
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Foreword
One of the great disappointments in chemistry over the past half-century has been
thefailureoftheorytoallowchemiststodesignmoleculesthatbindspecificallyin
aqueous solution to other molecules. This is, of course, the “medicinal chemistry
problem”; a classical pharmaceutical is simply a molecule that binds to a protein
target with a therapeutically interesting affinity, say a dissociation constant of less
than one nanomolar. While heuristic processes are available to sort through hun-
dreds of lead molecules to get dozens of hits that might generate single drug
candidates ready for clinical trials, these rely on only broad theoretical concepts
and, more frequently, chemical intuition, rarely on constructively detailed design.
Even the three-dimensional molecular models for protein target that are now often
routinethankstomoderncrystallographyhavenotdeliveredadefinitivesolutionto
this problem. The complexities of molecular interactions as well as the challenges
of modeling the solvent in which they must occur continue to defeat the largest
computers and the best theory.
The inability of theory to support explicit molecular design stands in stark
contrast to the ability of natural biology to deliver molecules that bind other mol-
ecules,oftenwithexquisitespecificityandenormousaffinity.Naturalbiologydoes
not, of course, exploit explicit design. Rather, the binding molecules that nature
delivers come from random variation followed by natural selection, where specif-
ically tight binding enhances the fitness of the host organism. The binding mole-
cules are often proteins and, in the case of antibodies, a protein scaffold that has
evolvedtosupportrandomvariationinabindingpocket.Here,naturalselectionfor
tightly binding antibodies occurs within a single organism. However, a primary
antibodylibraryofperhaps100millionspeciesissufficient,followingmutation,to
generatebinderswithsub-nanomolaraffinities,morethanenoughtosupportfitness.
Itwasagainstthisbackdropoftheoreticaldisappointmentthatinthelate1980s,
syntheticbiologistsdecidedtotrytodolaboratoryinvitroevolution(LIVE).Here,
nucleic acids, not proteins, were to provide the framework from which selective
binding molecules were to be delivered “on demand”.
As Jack Szostak, one of the developers of this technology was later to write, it
was “an idea whose time had come”. Not long before, Thomas Cech, Sidney
v
vi Foreword
Altman, and others had found specific examples where natural biology used RNA
molecules to catalyze reactions under physiological conditions. A number of these
wereinvolvedintheprocessingofRNAmoleculesthatwereusedbytheribosome
tobiosynthesizeproteins.Theribosomeitselfwassuspectedtobe,andlatershown
to be, a catalytic RNA molecule. Further, an analysis of the commonalities of
organisms all cross Earth suggested that we had all descended from an “RNA
world”, a biosphere where the only genetically encoded biological catalysts were
themselvesRNAmolecules.Indeed,somehadsuggestedthattheRNAworldhada
sufficient number of RNA catalysts to support a complex metabolism.
IflibrariesofRNAcouldleadtocatalyticRNAmoleculesondemand,surely(it
was thought) that they could solve a simpler problem: creating binding RNA
molecules on demand. Accordingly, Larry Gold, Andrew Ellington, and Jack
Szostak himself undertook to develop the technology that would create binding
moleculesstartingwithRNAlibraries.GeraldJoyceandothersthenfollowedwith
efforts to do the same thing where DNA was the matrix.
In either case, perhaps 100 trillion different DNA or RNA (collectively xNA)
species were synthesized in a library. The library was then presented to a target
receptor, in an experimental architecture that allowed the receptor to extract from
these libraries specific binding xNA species. Then, the special ability of nucleic
acids to direct their own replication would allow amplification to follow. A few
binding molecules would have descendants by the polymerase chain reaction.
Perhaps with some mutation, laboratory selection and evolution would produce
higher affinity binding molecules by a process analogous to the maturation of
antibody affinity. Perhaps xNA “aptamers” (as they came to be called) could have
an affinity that would rival the affinity of antibodies.
Of course, this was easier said than done, and a rich literature emerged in the
followingquartercenturyattemptingtoachievethisgoal.Muchofthisliteratureis
represented by the chapters in this book. However, these chapters take the next
steps in many dimensions. For example, rather than aptamers that bind to simple
protein or small molecule targets, the practitioners who contribute to this volume
haveincludedcellsasthemajortargetsforbindingmoleculesondemand.Thefield
of “Cell SELEX” is today exploding.
Thisbookisfocusedoncell-targetedselection,andiseditedbythepeoplewho
firstmadeitworkconvincingly.Inthisrespect,ithasthecharacterofamonograph,
withtheeditorsbeingtheco-authorsofmanychapters,andwithstillotherchapters
being written by those who were trained in the editor’s laboratories.
Some of the unique features of this book include its description of the cell-
SELEX process itself, with potential applications in molecular medicine being
linked in a logical and coherent way. This reflects the design of the book chapters
by the editors to allow individual authors to make contributions that blend coher-
ently with the other contributions. Thus, the product contrasts with many edited
books where each set of authors has written a self-standing chapter that need not
blend with the other chapters.
In the chapters on medical applications, the visionary question is addressed:
How should we use cell-targeted aptamers? Binding is often insufficient to meet a
Foreword vii
particular biomedical goal. One often also wants to capture, signal, or create
downstream action. This book has numerous chapters that meet this challenge,
including labeling, immobilization, and signaling that actually use the binding
molecules created “on demand”. The book contains individual chapters on each.
My own contributions to this field have been minimal. In December 1985, just
before I moved my laboratory to Switzerland to pursue these goals, I had dinner
with Jack Szostak, who told me of his plans to do directed evolution offunctional
RNAlibraries.MycommenttoJackwasbasedonachemist’sperception.Ipointed
out to Jack that unlike proteins, which have a rich collection offunctional groups,
nucleic acids have very few of the moieties that are needed to do catalysis or
binding. Further, with only four nucleotides, xNA as delivered to us by prebiotic
chemistry and subsequent evolution, has too few folding motifs, too much con-
formational ambiguity, and too little structural diversity to have any hope of
rivaling antibodies as a matrix to support binding molecules on demand.
However,IexplainedthateveningtoJack,therewasasolutiontothisproblem,
recorded in a witnessed notebook on November 14. All one needed to do was
rearrange the hydrogen bonding moieties on the nucleobases to give different
hydrogenbondingpatterns. Ifonedidso,onecouldcreate12different nucleotides
that should be able to form six independently replicable nucleobase pairs. Some
of these could carry functional groups similar to those found on proteins. In short,
this was the invention of a single biopolymer that had the replicability of nucleic
acids and the functional group diversity of proteins.
Jack’sresponsewasprescient.“Steve,”Irememberhimsaying,“itwilltakeyou
10years tomake thesemoleculesandanother10yearstogetpolymerasestoeven
acceptthem”inaPCRreaction.Hewasapproximatelycorrect.Onlylastyearwere
we finally able to do LIVE with an AEGIS alphabet.
Nevertheless, I am delighted to see a chapter by Liqin Zhang in this volume,
showing the first examples where an artificially expanded genetic system has been
usedtocreate bindingmoleculesondemand.Catalysts arenowfollowing,andthe
deficienciesoftheDarwiniansystempresentedtousbynaturemightnowbesolved
bymoleculardesign,notexplicitlyforeveryspecifictarget-receptorinteraction,but
ratherbythedesignofasystemthatismoreevolvableandthusbetterabletocreate
functional species.
Steven A. Benner
Foundation for Applied Molecular Evolution and
The Westheimer Institute for Science and Technology
Gainesville, FL
Contents
1 Introduction to Aptamer and Cell-SELEX. . . . . . . . . . . . . . . . . . 1
Libo Zhao, Weihong Tan and Xiaohong Fang
2 Cell-SELEX: Aptamer Selection Against Whole Cells. . . . . . . . . . 13
Dihua Shangguan, Tao Bing and Nan Zhang
3 Unnatural Nucleic Acids for Aptamer Selection . . . . . . . . . . . . . . 35
Liqin Zhang
4 Cell-Specific Aptamer Characterization . . . . . . . . . . . . . . . . . . . . 67
Tao Chen, Cuichen Wu and Weihong Tan
5 Molecular Engineering to Enhance Aptamer Functionality. . . . . . 89
Da Han, Cuichen Wu and Weihong Tan
6 Aptamers-GuidedDNANanomedicineforCancerTheranostics.... 111
Guizhi Zhu, Liping Qiu, Hongmin Meng, Lei Mei and Weihong Tan
7 Properties of Nucleic Acid Amphiphiles and Their
Biomedical Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139
Haipeng Liu
8 Aptamer-Based Hydrogels and Their Applications. . . . . . . . . . . . 163
Chun-Hua Lu, Xiu-Juan Qi, Juan Li and Huang-Hao Yang
9 Cell-Specific Aptamers for Disease Profiling and Cell Sorting. . . . 197
Kwame Sefah, Joseph Phillips and Cuichen Wu
ix
x Contents
10 Using Cell-Specific Aptamer-Nanomaterial Conjugates
for Cancer Cell Detection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215
Zhi Zhu
11 Cell-Specific Aptamers for Molecular Imaging. . . . . . . . . . . . . . . 239
Jing Zheng, Chunmei Li and Ronghua Yang
12 Discovery of Biomarkers Using Aptamers Evolved
in Cell-SELEX Method. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 265
Prabodhika Mallikaratchy, Hasan Zumrut and Naznin Ara
13 Cell-Specific Aptamers for Targeted Therapy. . . . . . . . . . . . . . . . 301
Yue He, Andrea del Valle and Yu-Fen Huang
14 The Clinical Application of Aptamers: Future Challenges
and Prospects. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 339
Yanling Song, Huimin Zhang, Zhi Zhu and Chaoyong Yang
Contributors
Naznin Ara Department of Chemistry, Lehman College-City University of
New York, Bronx, NY, USA
TaoBing BeijingNationalLaboratoryforMolecularSciences,KeyLaboratoryof
Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese
Academy of Sciences, Beijing, People’s Republic of China
Tao Chen Genentech Inc., South San Francisco, CA, USA
Xiaohong Fang Beijing National Laboratory for Molecular Sciences, Key Labo-
ratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry,
Chinese Academy of Sciences, Beijing, China
Da Han Intel Corporation, Hillsboro, OR, USA; Departments of Chemistry
Physiology and Functional Genomics, Center for Research at the Bio/Nano Inter-
face, Shands Cancer Center, UF Genetics Institute and McKnight Brain Institute,
University of Florida, Gainesville, FL, USA
Yue He Department of Biomedical Engineering and Environmental Sciences,
National Tsing Hua University, Hsinchu, Taiwan, ROC
Yu-Fen Huang Department of Biomedical Engineering and Environmental Sci-
ences, National Tsing Hua University, Hsinchu, Taiwan, ROC
ChunmeiLi KeyLaboratoryofLuminescentandReal-TimeAnalyticalChemistry,
Ministry of Education, College of Pharmaceutical Sciences, Southwest University,
Chongqing,China;BeijingNationalLaboratoryforMolecularSciences,Instituteof
Chemistry, Chinese Academy of Sciences, Beijing,China
Juan Li The Key Lab of Analysis and Detection Technology for Food Safety
of the MOE, College of Chemistry, Fuzhou University, Fuzhou, China
HaipengLiu DepartmentofChemicalEngineeringandMaterialsScience,Wayne
State University, Detroit, MI, USA
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