Cellular delivery and activation of polypeptide-nucleic acid complexes
First Claim
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1. A method of delivering a polypeptide to a cell, comprising:
- (a) contacting said cell with, in any order or combination, said polypeptide, a nucleic acid, a fluorescent molecule, and a cellular delivery molecule; and
(b) treating said cell with a treatment that results in the dissociation of said polypeptide from one or more of said nucleic acid, said fluorescent molecule, and said cellular delivery molecule.
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Abstract
In general, the present invention provides compositions for the cellular delivery of nucleic acids, polypeptides and/or flourophores, molecular complexes comprising fluorescent molecules or moieties, nucleic acids and polypeptides, and methods of making and using such compositions. Light-activated dispersal of the complexes leads to the intracellular release of one or more nucleic acids and/or peptides from the compositions or complexes. The biological activities of nucleic acids, polypeptides and flourophores may be repressed within the complexes, and these activities are restored upon release from the complexes.
189 Citations
117 Claims
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1. A method of delivering a polypeptide to a cell, comprising:
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(a) contacting said cell with, in any order or combination, said polypeptide, a nucleic acid, a fluorescent molecule, and a cellular delivery molecule; and
(b) treating said cell with a treatment that results in the dissociation of said polypeptide from one or more of said nucleic acid, said fluorescent molecule, and said cellular delivery molecule. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27)
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28. A method of delivering a polypeptide to a cell, comprising:
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(a) contacting said cell with, in any order or combination, said polypeptide, a nucleic acid, a fluorescent molecule, a cellular delivery molecule, and a transfection agent; and
(b) treating said cell with a treatment that results in the dissociation of said polypeptide from one or more of said nucleic acid, said fluorescent molecule, and said cellular delivery molecule. - View Dependent Claims (29, 30)
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- 31. A kit comprising at least one fluorescent molecule and at least one cellular delivery molecule.
- 39. A kit comprising at least one transfection agent and at least one RNAi molecule.
- 41. A complex comprising a cellular delivery polypeptide and an agent that is desirably taken up by cells, wherein said cellular delivery polypeptide comprises a fluorescent moiety.
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61. A method of delivering a nucleic acid to a cell, comprising:
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(a) contacting said cell with, in any order or combination, said nucleic acid, a fluorescent molecule, and a cellular delivery molecule; and
(b) treating said cell with a treatment that results in the dissociation of said nucleic acid from one or both of said fluorescent molecule and said cellular delivery molecule. - View Dependent Claims (62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87)
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88. A method of delivering a nucleic acid to a cell, comprising:
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(a) contacting said cell with, in any order or combination, said nucleic acid, a fluorescent molecule, a cellular delivery molecule, and a transfection agent; and
(b) treating said cell with a treatment that results in the dissociation of said nucleic acid from one or more of said fluorescent molecule and said cellular delivery molecule. - View Dependent Claims (89, 90)
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91. A molecular complex comprising one or more nucleic acids, one or more fluorophores, and one or more cellular delivery polypeptides, wherein each cellular delivery polypeptide:
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(a) is m % basic amino acids, wherein m is from about 10% to 100%;
(b) comprises a sequence of n contiguous basic amino acids, wherein n is any whole integer between 2 and 50; and
(c) is not derived from a herpes simplex virus (HSV) protein. - View Dependent Claims (92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107)
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108. A method of determining a cellular response to a test compound comprising:
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(a) contacting a first cell with, in any order or combination, a first nucleic acid, a fluorescent molecule, and a cellular delivery molecule;
(b) contacting a second cell with, in any order or combination, a second nucleic acid, said fluorescent molecule, and said cellular delivery molecule;
(c) treating said cells with a treatment that results in the dissociation of said polypeptide from one or more of said nucleic acid, said fluorescent molecule, and said cellular delivery molecule;
(d) contacting said cells with said test compound, before (a);
during (a), (b) or (c);
between (a) and (b);
between (b) and (c); and
, additionally or alternatively, after (c);
(e) detecting a signal from said cells, wherein said signal corresponds to a cellular response; and
(f) comparing the signal from said first cell with the signal from said second cell. - View Dependent Claims (109, 110, 111, 112)
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113. A method of identifying a compound having a preselected activity or effect comprising:
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(a) contacting a first cell with, in any order or combination, a first nucleic acid, a fluorescent molecule, and a cellular delivery molecule;
(b) contacting a second cell with, in any order or combination, a second nucleic acid, a fluorescent molecule, and a cellular delivery molecule;
(c) treating said cells with a treatment that results in the dissociation of said polypeptide from one or more of said nucleic acid, said fluorescent molecule, and said cellular delivery molecule;
(d) contacting said cells with said test compound, before (a);
during (a), (b) or (c);
between (a) and (b);
between (b) and (c); and
, additionally or alternatively, after (c);
(e) detecting a signal from said cells, wherein said signal corresponds to a cellular response; and
(f) comparing the signal from said first cell with the signal from said second cell, wherein a difference in the signal from first cell from the signal from said second cell corresponds to said preselected activity or effect. - View Dependent Claims (114, 115, 116, 117)
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Specification