Oligonucleotide Chemistry Research Articles

A Sensitive Assay System To Test Antisense Oligonucleotides for Splice Suppression Therapy in the Mouse Liver

We have previously demonstrated the efficacy of antisense therapy for splicing defects in cellular models of metabolic diseases, suppressing the use of cryptic splice sites or pseudoexon insertions. To date, no animal models with these defects are available. Here, we propose exon skipping of the phenylalanine hydroxylase (Pah) gene expressed in liver and kidney to generate systemic hyperphenylalaninemia in mice as a sensitive in vivo assay to test splice suppression. Systemic elevation of blood L-Phe can be quantified using tandem MS/MS. Exon 11 and/or 12 skipping for the normal PAH gene was validated in hepatoma cells for comparing two oligonucleotide chemistries, morpholinos and locked nucleic acids. Subsequently, Vivo-morpholinos (VMO) were tested in wild-type and in phenotypically normal Pahenu2/+ heterozygous mice to target exon 11 and/or 12 of the murine Pah gene using different VMO dosing, mode of injection and treatment regimes. Consecutive intravenous injections of VMO resulted in transient hyperphenylalaninemia correlating with complete exon skipping and absence of PAH protein and enzyme activity. Sustained effect required repeated injection of VMOs. Our results provide not only a sensitive in vivo assay to test for splice-modulating antisense oligonucleotides, but also a simple method to generate murine models for genetic liver diseases.

Splicing therapy for neuromuscular disease

Duchenne muscular dystrophy (DMD) and spinal muscular atrophy (SMA) are two of the most common inherited neuromuscular diseases in humans. Both conditions are fatal and no clinically available treatments are able to significantly alter disease course in either case. However, by manipulation of pre-mRNA splicing using antisense oligonucleotides, defective transcripts from the DMD gene and from the SMN2 gene in SMA can be modified to once again produce protein and restore function. A large number of in vitro and in vivo studies have validated the applicability of this approach and an increasing number of preliminary clinical trials have either been completed or are under way. Several different oligonucleotide chemistries can be used for this purpose and various strategies are being developed to facilitate increased delivery efficiency and prolonged therapeutic effect. As these novel therapeutic compounds start to enter the clinical arena, attention must also be drawn to the question of how best to facilitate the clinical development of such personalised genetic therapies and how best to implement their provision.

Development of bis-locked nucleic acid (bisLNA) oligonucleotides for efficient invasion of supercoiled duplex DNA

In spite of the many developments in synthetic oligonucleotide (ON) chemistry and design, invasion into double-stranded DNA (DSI) under physiological salt and pH conditions remains a challenge. In this work, we provide a new ON tool based on locked nucleic acids (LNAs), designed for strand invasion into duplex DNA (DSI). We thus report on the development of a clamp type of LNA ON—bisLNA—with capacity to bind and invade into supercoiled double-stranded DNA. The bisLNA links a triplex-forming, Hoogsteen-binding, targeting arm with a strand-invading Watson–Crick binding arm. Optimization was carried out by varying the number and location of LNA nucleotides and the length of the triplex-forming versus strand-invading arms. Single-strand regions in target duplex DNA were mapped using chemical probing. By combining design and increase in LNA content, it was possible to achieve a 100-fold increase in potency with 30% DSI at 450 nM using a bisLNA to plasmid ratio of only 21:1. Although this first conceptual report does not address the utility of bisLNA for the targeting of DNA in a chromosomal context, it shows bisLNA as a promising candidate for interfering also with cellular genes.

Design and Analysis of Effects of Triplet Repeat Oligonucleotides in Cell Models for Myotonic Dystrophy

Myotonic dystrophy type 1 (DM1) is caused by DM protein kinase (DMPK) transcripts containing an expanded (CUG)n repeat. Antisense oligonucleotide (AON)-mediated suppression of these mutant RNAs is considered a promising therapeutic strategy for this severe disorder. Earlier, we identified a 2′-O-methyl (2′-OMe) phosphorothioate (PT)–modified (CAG)7 oligo (PS58), which selectively silences mutant DMPK transcripts through recognition of the abnormally long (CUG)n tract. We present here a comprehensive collection of triplet repeat AONs and found that oligo length and nucleotide chemistry are important determinants for activity. For significant reduction of expanded DMPK mRNAs, a minimal length of five triplets was required. 2′-O,4′-C-ethylene-bridged nucleic acid (ENA)–modified AONs appeared not effective, probably due to lack of nuclear internalization. Selectivity for products from the expanded DMPK allele in patient myoblasts, an important requirement to minimize unwanted side effects, appeared also dependent on AON chemistry. In particular, RNase-H–dependent (CAG)n AONs did not show (CUG)n length specificity. We provide evidence that degradation of long DMPK transcripts induced by PS58-type AONs is an RNase-H independent process, does not involve oligo-intrinsic RNase activity nor does it interfere with splicing of DMPK transcripts. Our collection of triplet repeat AONs forms an important resource for further development of a safe therapy for DM1 and other unstable microsatellite diseases.

Quality Aspects of Oligonucleotide Drug Development: Specifications for Active Pharmaceutical Ingredients

This article, which is the first in a planned series intended to address chemistry, manufacturing, and control (CMC) aspects of therapeutic oligonucleotides, examines the topic of specifications for active pharmaceutical ingredients (APIs). The authors attempt to present basic scientific considerations for the broadest range of oligonucleotide APIs. Tests and analytical methods suitable for the control of single- and double-stranded oligonucleotide APIs and conjugated oligonucleotide APIs are discussed.

TricycloDNA-modified oligo-2′-deoxyribonucleotides reduce scavenger receptor B1 mRNA in hepatic and extra-hepatic tissues—a comparative study of oligonucleotide length, design and chemistry

We report the evaluation of 20-, 18-, 16- and 14-mer phosphorothioate (PS)-modified tricycloDNA (tcDNA) gapmer antisense oligonucleotides (ASOs) in Tm, cell culture and animal experiments and compare them to their gap-matched 20-mer 2′-O-methoxyethyl (MOE) and 14-mer 2′,4′-constrained ethyl (cEt) counterparts. The sequence-matched 20-mer tcDNA and MOE ASOs showed similar Tm and activity in cell culture under free-uptake and cationic lipid-mediated transfection conditions, while the 18-, 16- and 14-mer tcDNA ASOs were moderate to significantly less active. These observations were recapitulated in the animal experiments where the 20-mer tcDNA ASO formulated in saline showed excellent activity (ED50 3.9 mg/kg) for reducing SR-B1 mRNA in liver. The tcDNA 20-mer ASO also showed better activity than the MOE 20-mer in several extra-hepatic tissues such as kidney, heart, diaphragm, lung, fat, gastrocnemius and quadriceps. Interestingly, the 14-mer cEt ASO showed the best activity in the animal experiments despite significantly lower Tm and 5-fold reduced activity in cell culture relative to the 20-mer tcDNA and MOE-modified ASOs. Our experiments establish tcDNA as a useful modification for antisense therapeutics and highlight the role of chemical modifications in influencing ASO pharmacology and pharmacokinetic properties in animals.

Synthetic Gene Transfer Vectors II: Back to the Future.

The discovery of RNA interference has given a new lease on life to both the chemistry of oligonucleotides and chemical approaches for the intracellular delivery of nucleic acids. In particular, delivery of siRNA, whether in vitro for screening and target validation purposes or in humans as a new class of drugs, may revolutionize our approach to therapy. Their impact could equal that of the bioproduction and various uses of monoclonal antibodies today. Unfortunately, global pharmaceutical companies again seem to be waiting to buy the next Genentech or Genzyme of gene silencing rather than investing research and development into this promising area of research. Gene silencing encounters barriers similar to gene addition and hence may benefit from the extra decade of experience brought by gene therapy. "Chemical" transfection of cells in culture has become routine, and this Account discusses some of the reasons this success has not extended to nonviral gene therapy trials, most of which do not progress beyond the phase 2 stage. The author also discusses a (much debated) mechanism of nucleic acid cell entry and subsequent release of the polycationic particles into the cytoplasm. Both topics should be useful to those interested in delivery of siRNA. The move from gene therapy toward siRNA as an oligonucleotide-based therapy strategy provides a much wider range of druggable targets. Even though these molecules are a hundredfold smaller than a gene, they are delivered via similar cellular mechanisms. Their complexes with cationic polymers are less stable than those with a higher number of phosphate groups, which may be compensated by siRNA concatemerization or by chemical conjugation with the cationic carrier. Thus chemistry is again desperately needed.

Har Ghobind Khorana (1922-2011)

Har Ghobind Khorana (1922-2011)

Natural Antisense Makes Sense for Gene-specific Activation in Brain

The upregulation of specific genes in vivo has been an elusive goal for gene therapy when compared with the wide repertoire of methods available to silence genes or modify mRNA splicing patterns. In the latest issue of Nature Biotechnology, Modarresi and colleagues1 accomplished in vivo upregulation of brain-derived neurotrophic factor (BDNF), a relevant therapeutic target for a number of neurodegenerative diseases. Rather than using small molecules or microRNA inhibitors, which could lead to activation of off-target genes, Modarresi et al.1 upregulated BDNF by inhibiting a natural antisense transcript (NAT) in response to the local delivery of oligonucleotides to the central nervous systems of mice.

Upon the tightrope in prostate cancer: two acrobats on the same tightrope to cross the finishline

Prostate cancer is a multifactorial, multistep progressive disorder that is undruggable to date because of stumbling blocks in the standardization of therapy. It is triggered by a broad range of proteins, signaling networks and DNA damage response modulators. It is becoming increasingly apparent that DNA repair mediators have split personalities, as they are instrumental in suppressing and promoting carcinogenesis. In this article, we discuss on post-transcriptional processing of regulators of DNA damage response, and how DNA repair proteins trigger shuttling of androgen receptor. Substantial fraction of information has been added into the existing literature of ATM biology; however, the particular area of post-transcriptional processing errors and gene therapy for reprogramming of ATM has been left unaddressed in prostate cancer. It is therefore noteworthy that the facet of targeting strategy, antisense morpholino oligonucleotides chemistry, and systematic delivery of AOs has promising outlook in splice-targeted antisense-mediated therapy.

Cationic Polymer Based Nanocarriers for Delivery of Therapeutic Nucleic Acids

The design and development of nucleic acids based therapeutics for the treatment of diseases arising from genetic abnormalities has made significant progress over the past few years. Advances in synthetic oligonucleotide chemistry resulted in synthesis of nucleic acids that are relatively stable in in vivo environments. However, cellular targeting and intracellular delivery of nucleic acids still remains a challenge. Further, development of nucleic acids based therapeutics depends on the progress of safe and effective carriers for systemic administration. Cationic vectors with diminished cytotoxicity and enhanced efficacy are rapidly emerging as systems of choice. These vectors protect nucleic acids from enzymatic degradation by forming condensed complexes along with targeted tissue and cellular delivery. During the past few years, myriad of reports have appeared reporting delivery of nucleic acids mediated by nanocarriers. This review provides an overview of various nanocarriers employed for in vitro and in vivo delivery of therapeutically relevant nucleic acids e.g., DNA, siRNA, LNA, PNA etc.

Abstract SY31-02: Locked nucleic acid (LNA)-modified antisense oligonucleotides as anticancer agents: Using high-affinity antisense molecules in the laboratory and in the clinic

Abstract The inhibition of cellular processes associated with the malignant pheonotype is the goal many oncolytics. In the antisense oligonucleotide (ASO) approach, Watson and Crick base-pairing rules serve as the basis of rational drug design to create single-stranded oligonucleotides that are complementary and bind to RNAs critical for the malignant phenotype. The targets for antisense approaches can be mRNAs that encode for disease related proteins like those that control tumor growth, cell division, or survival. More recently, RNA targets in oncology have been expanded to include microRNAs. A successful oncolytic oligonucleotide must effectively bind and inhibit a target mRNA or miRNA that is critical for the malignant transformation or survival. Target identification presents the same challenge in antisense therapeutics as any other class of oncolytics, but drug design is based the known sequence of the target RNA. The first clinical trials to use antisense oligonucleotides in oncology produced only modest efficacy in clinical trials, but the therapeutic potential of these initial antisense drugs may have been hampered by their low stability and low binding affinity. Advancements in oligonucleotide chemistry have produced oligonucleotides with increased stability and increased affinities. This talk will focus on the use of oligonucleotides with the locked nucleic acid (LNA) modification. In these antisense constructs, some of the nucleotides in the sequence have a modification of the ribose sugar that includes a methylene bridge between the 2’ and 4’ positions. This bridge functions to lock the ribose into a (C3’-endo) configuration: a configuration that is optimized for binding to its cognate nucleotide. LNA-modified oligonucleotides bind to their target RNAs with higher affinities than most other oligonucleotide chemistries. One measure of affinity is the melting temperature (Tm) for binding to its complementary sequence. For each LNA-modified nucleotide added to an ASO sequence, Tm can increase Tm by over 5°. For example, an LNA-modified oligonucleotide currently in clinical development, miravirsen (SPC3649), with 9 LNA-modified nucleotides has a Tm of approximately 80° demonstrating that LNA-modified oligonucleotides have sufficient affinities to bind to and inhibit RNA function. Affinities like these have translated into increase potency for the inhibition of target RNAs in nonclinical models and should yield therapeutic benefit more robust than those seen with earlier generations of oligonucleotide therapeutics. Other drug-like properties of antisense therapeutics have also been improved by including LNA-modified nucleotides. For example, the pharmacokinetic properties of antisense oligonucleotides support their use in clinical medicine. Antisense oligonucleotides are single stranded and bind to proteins in circulation and on cell surfaces. After parenteral administration, antisense oligonucleotide are bound first to plasma proteins, limiting glomerular filtration, and then later the antisense oligonucleotides appear bound to cell surface (proteins), allowing them to be internalized in cells. Delivery into cells and tissues is achieved without the need for formulations more complex than just aqueous solutions. LNA modifications increase the metabolic stability in both circulation and inside of cells resulting in stable tissue concentrations and prolonged activities. This long tissue residence and stability, allows for constant exposure in tumor cells with dosing as infrequent as weekly or fortnightly. Again, this represents an improvement over antisense drugs used in previous oncology trials. Using in vitro and in vivo laboratory models it has been possible to demonstrate that treatment with LNA-modified oligonucleotides produces reductions in target gene expression and ultimately tumor growth that are sequence-, concentration-, and duration-of-therapy-dependent. Taken together the properties of LNA-modified oligonucleotides make them excellent candidates for inhibiting key RNA targets and at this time there are multiple LNA-modified oligonucleotides in clinical trials in oncology. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr SY31-02. doi:10.1158/1538-7445.AM2011-SY31-02

Comparison of different miR-21 inhibitor chemistries in a cardiac disease model

We would like to comment on a recent study that analyzed the role of microRNA-21 (miR-21) in a mouse model of cardiac disease (1). Using miR-21–deficient mice and novel, very short, 8-nucleotide anti–miR-21 oligonucleotides, the authors failed to detect any modulation of pressure overload-induced myocardial hypertrophy and fibrosis and concluded that miR-21 plays no role in cardiac disease. In contrast, we and others reported that inhibition of miR-21 with highly specific, 22- and 15-nucleotide-long anti–miR-21 oligonucleotides effectively inhibits myocardial and pulmonary fibrosis (2, 3). While genetic deletion of a target may lead to compensation during development and is often different from pharmacological inhibition of this target in the adult organism, the discrepancy between the therapeutic trials using long versus short 8-mer oligonucleo­tides is striking. We therefore carried out a direct head-to-head comparison of three different oligonucleotide chemistries (Figure ​(Figure1A)1A) in the same model of pressure overload-induced cardiac hypertrophy (transaortic constriction [TAC]). The two 22-mer oligonucleotides were complementary to the full-length miR-21, while the 8-mer was complementary to nucleotides 2 to 9 of miR-21, locked nucleic acid modified (LNA modified), and identical to the oligonucleotide used in the report by Patrick et al. (Figure ​(Figure1A). 1A). Figure 1 Comparison of different miR-21 oligonucleotide inhibitors.

Locked vs. unlocked nucleic acids (LNA vs. UNA): contrasting structures work towards common therapeutic goals

Oligonucleotide chemistry has been developed greatly over the past three decades, with many advances in increasing nuclease resistance, enhancing duplex stability and assisting with cellular uptake. Locked nucleic acid (LNA) is a structurally rigid modification that increases the binding affinity of a modified-oligonucleotide. In contrast, unlocked nucleic acid (UNA) is a highly flexible modification, which can be used to modulate duplex characteristics. In this tutorial review, we will compare the synthetic routes to both of these modifications, contrast the structural features, examine the hybridization properties of LNA and UNA modified duplexes, and discuss how they have been applied within biotechnology and drug research. LNA has found widespread use in antisense oligonucleotide technology, where it can stabilize interactions with target RNA and protect from cellular nucleases. The newly emerging field of siRNAs has made use of LNA and, recently, also UNA. These modifications are able to increase double-stranded RNA stability in serum and decrease off-target effects seen with conventional siRNAs. LNA and UNA are also emerging as versatile modifications for aptamers. Their application to known aptamer structures has opened up the possibility of future selection of LNA-modified aptamers. Each of these oligonucleotide technologies has the potential to become a new type of therapy to treat a wide variety of diseases, and LNA and UNA will no doubt play a part in future developments of therapeutic and diagnostic oligonucleotides.

Inhibitors of Platelet Adhesion

Platelet aggregation, in fundamental terms, is considered a biological end point that contributes to the occurrence of clinical events among patients with advanced atherosclerotic coronary artery disease. Acute coronary syndrome, including non–ST elevation myocardial infarction (NSTEMI) and ST elevation myocardial infarction (STEMI), accounts for upward of 733 000 hospital admissions yearly in the United States.1 The primary pathophysiological mechanism responsible for the majority of acute coronary syndromes is endothelial plaque disruption with and subsequent platelet adhesion, activation, and thrombus formation.2 The end result is formation of thrombus within a coronary artery, leading to subtotal vessel occlusion with NSTEMI and complete occlusion of the artery with STEMI. In this review, we provide a contemporary view of platelet adhesion as a highly coordinated and teleologically conserved process achieved by surface receptors, protein ligands, and matrix proteins operating at the platelet–subendothelium interface. We also discuss drugs in development, including monoclonal antibodies, inhibitory peptides, and oligonucleotides; preclinical data; and, where available, clinical trial results, highlighting the potential translation of fundamental constructs in platelet biology to patient care. Platelets are derived from a hematopoietic bone marrow stem cell precursor.3 Through a highly conserved program of cellular differentiation, this stem cell precursor becomes a megakaryocyte at a rate of approximately 100 000 000 megakaryocytes produced per day.3 Subsequently, each individual megakaryocyte will give rise to approximately 500 platelets via additional developmental steps within the bone marrow that involve a proplatelet intermediate stage.3 Once released into circulation, a mature platelet has an expected life span of 7 to 10 days.3 Platelet maturation and development involve the expression of receptors on the platelet cell surface. These receptors facilitate platelet adhesion and activation, and they promote thrombus development through receptor–ligand interactions, with several ligands expressed on the surface of endothelial cells, within the …

The true RNA-seq

With a modified polymerase and optimized oligonucleotide chemistry, Helicos' single-molecule sequencer takes on RNA.

Oligonucleotides Containing 7‐Deaza‐2′‐deoxyinosine as Universal Nucleoside: Synthesis of 7‐Halogenated and 7‐Alkynylated Derivatives, Ambiguous Base Pairing, and Dye Functionalization by the Alkyne–Azide ‘Click’ Reaction

AbstractOligonucleotides containing 7‐deaza‐2′‐deoxyinosine derivatives bearing 7‐halogen substituents or 7‐alkynyl groups were prepared. For this, the phosphoramidites 2b–2g containing 7‐substituted 7‐deaza‐2′‐deoxyinosine analogues 1b–1g were synthesized (Scheme 2). Hybridization experiments with modified oligonucleotides demonstrate that all 2′‐deoxyinosine derivatives show ambiguous base pairing, as 2′‐deoxyinosine does. The duplex stability decreases in the order Cd>Ad>Td>Gd when 2b–2g pair with these canonical nucleosides (Table 6). The self‐complementary duplexes 5′‐d(F7c7I‐C)6, d(Br7c7I‐C)6, and d(I7c7I‐C)6 are more stable than the parent duplex d(c7I‐C)6 (Table 7). An oligonucleotide containing the octa‐1,7‐diyn‐1‐yl derivative 1g, i.e., 27, was functionalized with the nonfluorescent 3‐azido‐7‐hydroxycoumarin (28) by the Huisgen–Sharpless–Meldal cycloaddition ‘click’ reaction to afford the highly fluorescent oligonucleotide conjugate 29 (Scheme 3). Consequently, oligonucleotides incorporating the derivative 1g bearing a terminal CC bond show a number of favorable properties: i) it is possible to activate them by labeling with reporter molecules employing the ‘click’ chemistry. ii) Space demanding residues introduced in the 7‐position of the 7‐deazapurine base does not interfere with duplex structure and stability (Table 8). iii) The ambiguous pairing character of the nucleobase makes them universal probes for numerous applications in oligonucleotide chemistry, molecular biology, and nanobiotechnology.

DNA hybridization detection by porous silicon-based DNA microarray in conjugation with infrared microspectroscopy

A method is described for the label-free detection of DNA hybridization on porous silicon (por-Si), based upon the pairing of oligonucleotide chemistry and standard silicon nanotechnology. Por-Si with a pore diameter of approximately 30 nm was used to immobilize probe DNA. Infrared microspectroscopy was employed to monitor the hybridization of probe-DNA immobilized on pore surfaces with its complementary DNA (target-DNA). The immobilization of probe DNA on por-Si facilitates hybridization detection for a small sensing area (approximately 50×50 μm2) with a high detection efficiency. In this study, we fabricated a porous silicon-based DNA microarray (por-Si-microarray) using photolithographic and Si anodizing techniques. We demonstrate that DNA hybridization can be detected on a por-Si-microarray through the analysis of infrared absorption spectral profiles in the region where the vibration modes of the bases appear. This present approach demonstrates that por-Si-microarray in conjugation with infrared microspectroscopy has potential application in DNA sensing chips.

Dystrophin expression in the mdx mouse after localised and systemic administration of a morpholino antisense oligonucleotide

Duchenne and Becker muscular dystrophies are allelic disorders arising from mutations in the dystrophin gene. Duchenne muscular dystrophy is characterised by an absence of functional protein, while Becker muscular dystrophy is usually caused by in-frame deletions allowing synthesis of some functional protein. Treatment options are limited, and we are investigating the potential of transcript manipulation to overcome disease-causing mutations. Antisense oligonucleotides have been used to induce specific exon removal during processing of the dystrophin primary transcript and thereby by-pass protein-truncating mutations. The antisense oligonucleotide chemistry most widely used to alter pre-mRNA processing is 2'-O-methyl-modified bases on a phosphorothioate backbone. The present studies evaluate 2'-O-methylphosphorothioate, peptide nucleic acid and morpholino antisense oligonucleotides in the mdx mouse model of muscular dystrophy, which has a nonsense mutation in exon 23 of the dystrophin gene. We demonstrate dystrophin expression in mdx mouse tissues after localised and systemic delivery of a morpholino antisense oligonucleotide designed to target the dystrophin exon 23 donor splice site. The stability of the morpholino structural type, and the fact that it can be delivered to muscle in the absence of a delivery reagent, render this compound eminently suitable for consideration for therapeutic exon skipping to address dystrophin mutations.

Tone Reversal of an AFM Lateral Force Image Due to Hybridization of Oligonucleotides Immobilized on Polymers

The immobilization and molecular conformation of oligonucleotides on surfaces is critical to DNA-based microdevices, such as biosensors, microand nanoarrays, and lab-ona-chip devices. Traditionally, these devices use glass or silicon as the base material, which have well characterized surface chemistry. However, the increasing demand for DNAbased microdevices requires low-cost, easily processable materials that could be suitable for disposable devices. The use of polymers, which are the logical choice from the manufacturing point of view, is however challenged by two sets of fundamental problems. Firstly, as polymer bulk properties (relevant to manufacturing) need to be decoupled from surface properties (relevant to biomolecule immobilization), additional surface functionalization is usually needed for covalent binding of oligonucleotides. Plasma processing, which changes only the very top surface of the polymer, has been widely used for polymer processing but not for DNA-based microdevices. The second, less apparent, challenge is the complexity and dynamic character of the polymer surface compared with, for example, glass. Consequently, the key performance criteria of oligonucleotide immobilization on surfaces, i.e., a high density of bound target molecule and favorable molecular conformation, have to be reconsidered with respect to the interaction of oligonucleotide chains, themselves of a quasi-polymeric nature, with the polymer surface. Atomic force microscopy (AFM) is commonly used for the mapping of nanotopography, but also of spatial distribution of oligonucleotides chemistry and hydrophobicity, and the local mechanical properties of polymers. AFM has also been used to visualize individual DNA molecules and, recently, to fabricate DNA nanoarrays. This communication reports on the use of two very different polymers, both good candidates for the fabrication of bio-microdevices, that is, polycarbonate (PC) and cycloolefin copolymer (COC), for probing the interaction between oligonucleotides during immobilization and hybridization. This communication also reports on the use of AFM, and in particular the lateral force (LF), as a means to detect the immobilization and hybridization of oligonucleotides on polymers. Although the model polymers have similar thermomechanical properties (glass transition temperature, Tg, of 136 8C and 145 8C, for COC and PC, respectively), their different chemistry translates into very different processes induced by plasma treatment, which are confined to the top 200 nm. Firstly, the [total O]/[total C] atomic ratio increases from 15.5% to 21.5% for COC samples, and decreases from 45.2% to 26.7% for PC samples. Secondly, the [oxygen-bound carbon (CxOy)]/[total C] ratio increases for COC from 15.5% to 21.5% over the full duration of plasma processing (up to 5 min), but decreases dramatically for PC during the first 20 s of plasma treatment (from 45% to 29%) and further decreases to 26% at the end of the process (Figure 1). These data suggest that the opening of the