Apparatus and method for selectively ranking sequences for antisense targeting
AssigneeBoard of Regents, The University of Texas System
InventorsDonald M. Gray, Christopher L. Clark
Life of this patent
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Abstract
The invention provides methods of identifying a nucleic acid molecule having a sequence capable of targeting a gene of interest, and hence nucleic acid molecules useful in gene theraphy and disease treatment. A system and method for selecting antisense DNA oligonucleotide sequences that will form the most stable DNA:RNA hybrids within a given target mRNA sequence is provided. An apparatus capable of identifying a subject nucleic acid sequence capable of targeting a gene of interest is further provided. The apparatus includes the use of a software program and a computing unit. Methods for controlling gene expression are also provided.
Claims40
A method for identifying a nucleic acid molecule having a sequence capable of targeting a gene of interest comprising:
- (a) a first database comprising a list of stability values for independent combinations of nearest-neighbor nucleic acid base pairs;
- (b) a computing unit comprising:
- a means for inputting data comprising a nearest-neighbor nucleic acid pair, N(x), data list, defining a nucleic acid sequence of interest to be targeted to provide a second database;
- and
- (c) a program capable of processing said first and said second database to provide a nearest-neighbor nucleic acid base pair, N(x), comparison and provide a stability value of a nucleic acid sequence capable of targeting the gene of interest.
A method for identifying nucleic acid sequences for antisense oligonucleotide targeting comprising the steps of:
- storing a plurality of operating instructions in a memory unit of a control computer;
- and
- operating said control computer pursuant to said plurality of operating instructions for:
- accessing a list of values for independent combinations of nearest-neighbor nucleic acid base pairs;
- obtaining a nucleic acid sequence;
- accepting a nucleic acid segment length value;
- comparing each nucleic acid segment within said nucleic acid sequence with said list of values for independent combinations of nearest-neighbor nucleic acid base pairs;
- and
- identifying nucleic acid sequences for antisense oligonucleotide targeting by determining a hybrid duplex stability value for each contiguous nucleic acid segment of said length within said nucleic acid sequence.
A compiler for identifying nucleic acid sequences for antisense oligonucleotide targeting comprising:
- a first plurality of binary values for obtaining a nucleic acid sequence in a first data format;
- a second plurality of binary values containing a list of stability values for independent combinations of nearest-neighbor nucleic acid base pairs in a second data format;
- a third plurality of binary values for comparing said nucleic acid sequence with said list of stability values for independent combinations of nearest-neighbor nucleic acid base pairs, N(x), to obtain hybrid duplex stability values;
- and
- a fourth plurality of binary values for electronically routing said hybrid duplex stability values to an output device.
A method for selectively ranking antisense oligonucleotides for inhibitory efficiency comprising the steps of:
- determining the fractions of nearest-neighbor base pair types in each target sequence zone RNA:ASO-DNA hybrid sequence;
- substituting nearest-neighbor base pair fractions into formulas to determine the fractions of each of a series of 13 independent nearest-neighbor base pair combinations;
- and
- determining a hybrid duplex stability value for each target sequence zone RNA:DNA hybrid sequence wherein the duplex stability values for said target sequence zones are ordered to produce a ranking for inhibitory efficiency.
A method for selectively ranking antisense oligonucleotides for hybridization efficiency comprising the steps of:
- determining the fractions of nearest-neighbor base pair types in each target sequence zone RNA:ASO-DNA hybrid sequence;
- substituting nearest-neighbor nucleic acid base pair fractions into formulas to determine the fractions of each of a series of 13 independent nearest-neighbor base pair combinations;
- and
- adding the products of the fractions which have been multiplied by the stability values of the independent combination to determine a hybrid stability value for each target sequence zone;
- wherein the stability values for said target sequence zones are ordered to produce a ranking for hybridization efficiency.
A method for selectively ranking an antigene oligonucleotide sequence for inhibitory efficiency comprising the steps of:
- determining the fractions of nearest-neighbor base triplet types in each target sequence zone DNA:AGO-DNA triplex sequence;
- substituting nearest-neighbor triplet fractions into formulas to determine the fractions of each of three independent nearest-neighbor combinations of nucleic acid sequence triplet types selected from the group consisting of:
- poly[d(T):d(A):d(T)];
- poly[d(C+):d(G):d(C)];
- and poly[d(TC+):d(AG):d(CT)];
- and
- multiplying the fractions of the independent nearest-neighbor triplet combinations by the stability values of the combinations and adding the products to give a triplex stability ranking value for the target sequence zone DNA:AGO-DNA triplex nucleic acid sequence;
- wherein the results for various target sequence zone DNA sequences are ordered to produce a ranking for inhibitory efficiency.
A method for ranking an antigene oligonucleotide sequence for hybridization efficiency comprising the steps of:
- determining the fractions of nearest-neighbor base triplet types in each target sequence zone DNA:AGO-DNA triplex sequence;
- substituting nearest-neighbor triplet fractions into formulas to determine the fractions of each of three independent nearest-neighbor combinations of nucleic acid sequence triplet types selected from the group consisting of:
- poly[d(T):d(A):d(T)];
- poly[d(C+):d(G):d(C)];
- and poly[d(TC+):d(AG):d(CT)];
- and
- multiplying the fractions of the independent nearest-neighbor triplet combinations by the stability values of the combinations and adding the products to give a triplex stability ranking value for the target sequence zone DNA:AGO-DNA triplex nucleic acid sequence;
- wherein the results for various target sequences zone DNA sequences are ordered to produce a ranking for hybridization efficiency of said antigene oligonucleotide.
A method for controlling gene expression by selectively locating a target sequence zone comprising the steps of:
- obtaining a target sequence;
- determining the fractions of nearest-neighbor base pair or base triplet types in each subsequence target sequence zone within said target sequence;
- substituting nearest-neighbor nucleic acid base pair or base triplet fractions into formulas to determine the fractions of each of a series of independent nearest-neighbor base pair combinations;
- determining a duplex or triplex stability value by multiplying the fractions by the stability values of the independent combinations and adding the products;
- eliminating one or more subsequence target sequence zones from all possible subsequence target zones;
- and
- ordering the remaining stability values to produce a ranking of the remaining subsequence target sequence zones for efficiency of controlling gene expression.
A method for identifying nucleic acid sequences for antisense oligonucleotide targeting by selectively locating a target sequence zone comprising the steps of:
- obtaining a target sequence;
- determining the fractions of nearest-neighbor base pair and base pair-plus-end neighbor types in each subsequence target sequence zone of any length within said target sequence;
- substituting fractions of nucleic acid nearest-neighbor base pair or base pair-plus-end neighbor types into formulas to determine the fractions of each of a series of 20 independent nearest-neighbor combinations which take end effects into account selected from the group consisting of:
- EUAE/ETAE, EUUE/EAAE, EUCE/EGAE, EUGE/ECAE, EAAE/ETTE, EAUE/EATE, EACE/EGTE, EAGE/ECTE, ECAE/ETGE, ECUE/EAGE, ECCE/EGGE, ECGE/ECGE, EGAE/ETCE, EGUE/EACE, EGCE/EGCE, EGGE/ECCE, EUE/EAE, EAE/ETE, and ECE/EGE, EGE/ECE:
- determining a hybrid stability value for each subsequence target zone by multiplying the fractions by the stability values of the 20 independent nearest-neighbor combinations and adding the products;
- eliminating one or more subsequence target sequence zones from the possible subsequence target sequence zones;
- and
- ordering the remaining stability values to produce a ranking of the remaining subsequence target sequence zones to provide nucleic acid sequences for antisense oligonucleotide targeting.
Dependencies are inferred from the claim text.