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Houston, TX, United States

Li N.,AM Biotechnologies, LLC | Nguyen H.H.,University of California at Los Angeles | Byrom M.,University of Texas at Austin | Ellington A.D.,University of Texas at Austin
PLoS ONE | Year: 2011

Aptamers continue to receive interest as potential therapeutic agents for the treatment of diseases, including cancer. In order to determine whether aptamers might eventually prove to be as useful as other clinical biopolymers, such as antibodies, we selected aptamers against an important clinical target, human epidermal growth factor receptor (hEGFR). The initial selection yielded only a single clone that could bind to hEGFR, but further mutation and optimization yielded a family of tight-binding aptamers. One of the selected aptamers, E07, bound tightly to the wild-type receptor (Kd = 2.4 nM). This aptamer can compete with EGF for binding, binds to a novel epitope on EGFR, and also binds a deletion mutant, EGFRvIII, that is commonly found in breast and lung cancers, and especially in grade IV glioblastoma multiforme, a cancer which has for the most part proved unresponsive to current therapies. The aptamer binds to cells expressing EGFR, blocks receptor autophosphorylation, and prevents proliferation of tumor cells in three-dimensional matrices. In short, the aptamer is a promising candidate for further development as an anti-tumor therapeutic. In addition, Aptamer E07 is readily internalized into EGFR-expressing cells, raising the possibility that it might be used to escort other anti-tumor or contrast agents. © 2011 Li et al. Source


Patent
AM Biotechnologies, LLC and Board Of Regents Of The University Of Texas System | Date: 2013-10-10

Provided herein are methods for a novel bead-based next-generation X-aptamer selection scheme that extends aptamer technology to include X-modified bases, thus resulting in X-aptamers, at any position along the sequence because the aptamers are chemically synthesized via a split-pool scheme on individual beads. Also provides are application to a wide range of commonly used DNA modifications, including, but not limited to, monothioate and dithioate backbone substitutions. This new class of aptamer allows chemical modifications introduced to any of the bases in the aptamer sequence as well as the phosphate backbones and can be extended to other carbohydrate-based systems.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 180.12K | Year: 2012

DESCRIPTION (provided by applicant): Functional RNA molecules such as aptamers, siRNAs, miRNAs, and related compounds have enormous potential as human therapeutics and as tools for elucidating gene regulation in vivo. To reach this potential, such molecules must be highly potent and highly nuclease resistant. Unmodified RNAs typically do not meet these requirements. A variety of chemical modifications have been explored to overcome these limitations. In particular, some success has been achieved in varioussystems using 2'-F-ribose and phosphorothioate backbone modifications, alone or in combination. However, further improvements are highly desirable. In addition, phosphorothioate modifications are chiral, resulting in two distinct isomers at each backbonesubstitution. Thus, there is a need for improved chemical modifications that can be incorporated into functional RNAs. AM Biotechnologies will address these critical issues by developing 2'-F-ribonucleoside thiophosphoramidites (2'-F-thioamidites) to enable synthesis of phosphorodithioate 2'-F-RNA (PS2-2'-F- RNA). We have previously shown that PS2 modifications at selected backbone positions of DNA aptamers enhance binding affinity to target proteins without loss of specificity. In addition, selected PS2 modifications in siRNAs significantly improve gene silencing activities. Thus, selected PS2-2'-F-RNA modifications will significantly increase binding affinity and potency of 2'-F-RNA aptamers, and will offer new avenues for synthesis of highly potent siRNAs. PS2-2'-F-RNAs will also be achiral at phosphorus, eliminating the variable biochemical, biophysical, and biological properties of diastereomeric phosphorothioate substituted RNAs. This Phase I project will: 1) develop the chemistry to produce four 2'-F-thioamidites (ABz, CBz, GIbu and U); 2) optimize the synthesis of PS2-2'-F-RNAs; 3) evaluate the effects of PS2-2'-F modifications on the binding affinity of a model 2'-F-RNA aptamer; and 4) evaluate the effects of PS2-2'-F modifications on the gene silencing activities of siRNAs targeting 2-secretase. In Phase II, AM will (a) scale reagent production up to commercial quantities and purity; (b) optimize a robust protocol for synthesis of PS2-2'-F-RNA; (c) evaluate the effects of PS2-2'-F modifications on aptamers and siRNA activity in vivo; and (d) fully characterize the pharmacokinetic properties of PS2-2'-F-RNA. AM in Phase II may also offer for sale limited quantities of research-grade reagents for market beta testing. Upon successful completion of PhaseII, AM will work with its existing industry partners to commercialize the 2'-F-thioamidites and enable the entire life science community to use these unique reagents in developing improved high-potency RNA drugs for a wide variety of human disease applications. PUBLIC HEALTH RELEVANCE: Functional RNA molecules such as aptamers and siRNAs have exciting potential as therapeutics in areas such as viral infections, cancer, genetic disorders, and neurological diseases. However, these potential RNA drugsrequire chemical modifications to achieve the necessary potency and stability. AM Biotechnologies (AM) will develop 2'-F-ribonucleoside thiophosphoramidite reagents that will allow the life science community to produce high potency, highly stable phosphorodithioate 2'-F-RNA-based drugs. The unique reagents that AM will develop under this project could have a profound impact on public health.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 156.73K | Year: 2011

DESCRIPTION (provided by applicant): Functional RNA molecules such as aptamers, siRNAs, miRNAs, and related compounds have enormous potential as human therapeutics and as tools for elucidating gene regulation in vivo. To reach this potential, such molecules must be highly potent and highly nuclease resistant. Unmodified RNAs typically do not meet these requirements. A variety of chemical modifications have been explored to overcome these limitations. In particular, some success has been achieved in varioussystems using 2'-O-methyl-ribose and phosphorothioate backbone modifications, alone or in combination. However, further improvements are highly desirable. In addition, phosphorothioate modifications are chiral, resulting in two distinct isomers at each backbone substitution. Thus, there is a need for improved chemical modifications that can be incorporated into functional RNAs. AM Biotechnologies will address these critical issues by developing 2'-O-methyl-ribonucleoside thiophosphoramidites (2'-OMe-thioamidites) to enable synthesis of phosphorodithioate 2'-OMe-RNA (PS2-2'- OMe-RNA). We have previously shown that PS2 modifications at selected backbone positions of DNA aptamers enhance binding affinity to target proteins without loss of specificity. Similarly, selected PS2 modifications in siRNAs significantly improve gene silencing activities. Thus, selected PS2-2'-OMe-RNA modifications will significantly increase binding affinity and potency of 2'-OMe-RNA aptamers, and will offer new avenues for synthesis of highly potent siRNAs. PS2-2'-OMe-RNAs will also be achiral at phosphorus, eliminating the variable biochemical, biophysical, and biological properties of diastereomeric phosphorothioate substituted RNAs. This Phase I project will: 1) develop the chemistry to produce four 2'-OMe-thioamidites (ABz, CBz, GIbu and U); 2) optimize the synthesis of PS2-2'-OMe-RNAs; 3) evaluate the effects of PS2-2'-OMe modifications on the binding affinity of a model RNA aptamer; and 4) evaluate the effects of PS2-2'-OMe modifications on the gene silencing activities of siRNAs targeting 2-secretase. In Phase II, AM will (a) scale reagent production up to commercial quantities and purity; (b) optimize a robust protocol for synthesis of PS2-2'OMe-RNA; (c) evaluate the effects ofPS2-2'-OMe modifications on aptamers and siRNA activity in vivo; and (d) fully characterize the pharmacokinetic properties of PS2-2'-OMe-RNA. AM in Phase II may also offer for sale limited quantities of research-grade reagents for market beta testing. Uponsuccessful completion of Phase II, AM will work with its existing industry partners to commercialize the 2'-OMe-thioamidites and enable the entire life science community to use these unique reagents in developing improved high-potency RNA drugs for a widevariety of human disease applications. PUBLIC HEALTH RELEVANCE: Functional RNA molecules such as aptamers and siRNAs have exciting potential as therapeutics for viral infections, cancer, genetic disorders, and neurological diseases. However, thesepotential RNA drugs require chemical modifications to achieve the necessary potency and stability. AM Biotechnologies (AM) will develop 2'-O-methyl-ribonucleoside thiophosphoramidite reagents that will allow the life science community to produce high potency, highly stable phosphorodithioate 2'-O-methyl-RNA-based drugs. The unique reagents that AM will develop under this project could have a profound impact on public health.


Grant
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase II | Award Amount: 700.62K | Year: 2015

DESCRIPTION provided by applicant The X Aptamer Selection Kit XASK makes aptamers much more accessible to the life science market The low priced kit greatly minimizes the expertise and equipment required to perform aptamer selections thus enabling virtually any scientist with basic laboratory skills to rapidly develop his or her own synthetic next generation X Aptamer XA affinity reagents The Phase I SBIR successfully answered several fundamental questions associated with the bead based selection technology underlying the XASK It also conclusively demonstrated the feasibility of the kit as a commercial product Several independent users successfully selected X Aptamers using XASK prototypes which far exceeded the anticipated outcome from Phase I X Aptamers are chemically modified DNA affinity agents that utilize amino acid functional groups and even small molecules to enhance interaction with targets XAs routinely exhibit nanomolar to picomolar binding affinity as well as excellent specificity making them a promising synthetic alternative to antibodies AM Biotech has developed a rapid single cycle XA discovery process that is not based on SELEX The process is performed in two easy steps using bead based oligonucleotide libraries and standard laboratory equipment that enable packaging the discovery processes into XASK The Phase II SBIR will focus on enhancing the quality of the critically important bead based libraries and optimizing the bead based selection process to improve the success rate to andgt The XASK is a revolutionary product that will disrupt the aptamer market and will begin to make inroads into the antibody market AM Biotech believes that the XASK and X Aptamers are poised for strong market penetration that will help to pull aptamers into many commercial applications PUBLIC HEALTH RELEVANCE The X Aptamer Selection Kit XASK is a revolutionary product that minimizes the expertise and equipment required to perform aptamer selections which will make aptamers much more accessible to the life science market Synthetic affinity molecules like aptamers have many desirable characteristics that could solve numerous problems that are associated with the antibodies that they are intended to replace This is expected to lead to better diagnostic tests and to novel drug candidates for a wide range of diseases and conditions

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