Nano/micromotors for active and dynamic intracellular payload delivery

Patent metadata

Field	Value
Publication number	US-11007281-B2
Application number	US-201815939104-A
Country	US
Kind code	B2
Filing date	Mar 28, 2018
Priority date	Mar 28, 2017
Publication date	May 18, 2021
Grant date	May 18, 2021

Abstract

Methods, systems, and devices are disclosed for intracellular payload delivery by nanomotor structures. In some aspects, a nanomotor for intracellular payload delivery includes an asymmetric body having a concave cavity at one end of the nanowire body; a functionalization layer on an outer surface of the nanowire body; and a payload substance coupled to the nanomotor by the functionalization layer in a biologically active conformation, wherein the payload substance is attached to a portion of the functionalization layer or at least partially encapsulated within the functionalization layer, in which the nanomotor is operable to propel in a biological medium and into an intracellular region of a living cell to initiate an interaction of the biologically active payload substance with an intracellular constituent of the living cell.

First claim

What is claimed is:

1. A method for intracellular delivery of a compound to a living cell, comprising:
providing a plurality of nanomotors operable to propel in a medium comprising a cell, wherein each nanomotor of the plurality of nanomotors includes a functionalization layer on an outer surface of the nanomotor coupling a payload substance in a biologically active conformation to the nanomotor; and
propelling the nanomotors in the medium to cause at least some of the nanomotors to penetrate into an intracellular region of the cell;
wherein the payload substance within the intracellular region of the cell initiates an interaction of the biologically active payload substance with an intracellular constituent of the cell, and
wherein the biologically active payload substance includes a protein 9 (Cas9) nuclease complexed with a clustered regularly interspaced short palindromic repeats RNA (CRISPR RNA) and a trans-activating CRISPR RNA (tracrRNA) and fused in a single guided RNA (sgRNA) to cause a targeted gene alteration of a target gene of the cell.

2. The method of claim 1 , wherein the functionalization layer includes a disulfide linkage including cysteine functional groups that reversibly attach the Cas9-sgRNA complex to the nanomotor, and facilitate detachment of the Cas9-sgRNA in the intracellular region.

3. A method for intracellular delivery of a compound to a living cell, comprising:
providing a plurality of nanomotors operable to propel in a medium comprising a cell, wherein each nanomotor of the plurality of nanomotors includes a functionalization layer on an outer surface of the nanomotor coupling a payload substance in a biologically active conformation to the nanomotor; and
propelling the nanomotors in the medium to cause at least some of the nanomotors to penetrate into an intracellular region of the cell;
wherein the payload substance within the intracellular region of the cell initiates an interaction of the biologically active payload substance with an intracellular constituent of the cell, and
wherein the biologically active payload substance includes a protein configured to affect a cellular function of the cell.

4. The method of claim 3 , wherein the biologically active payload substance includes a caspase-3 enzyme to trigger apoptosis of the cell.

5. The method of claim 4 , wherein the functionalization layer includes a biocompatible pH-responsive polymer coating on the nanomotor that encapsulates caspase-3 enzyme in an active conformation and prevents release of the caspase-3 enzyme in the medium, and that dissolves in a neutral or alkaline pH environment above 5.5 pH.

6. A method for intracellular delivery of a compound to a living cell, comprising:
providing a plurality of nanomotors operable to propel in a medium comprising a cell, wherein each nanomotor of the plurality of nanomotors includes a functionalization layer on an outer surface of the nanomotor coupling a payload substance in a biologically active conformation to the nanomotor; and
propelling the nanomotors in the medium to cause at least some of the nanomotors to penetrate into an intracellular region of the cell;
wherein the payload substance within the intracellular region of the cell initiates an interaction of the biologically active payload substance with an intracellular constituent of the cell, and
wherein the biologically active payload substance includes a nucleotide sequence configured to affect a gene expression of a target gene of the cell having a complementary nucleotide sequence,
wherein the biologically active payload substance includes a small interfering ribonucleic acid (siRNA), and the functionalization layer includes a rolling circle amplification (RCA) nucleic acid strand that binds the siRNA.

7. The method of claim 6 , wherein the RCA nucleic acid strand is attached to a gold surface of the nanomotor via a gold-thiol interaction.

Assignees

• Univ California
• The Regents Of The Univ Of Californa

Inventors

• Wang Joseph
• Esteban-Fernández De Ávila Berta
• Chen Yi
• Angell Chava
• Soto Fernando
• Zhang Liangfang
• Hansen-Bruhn Malthe

Classifications

• A61K41/0028 Primary

  Disruption, e.g. by heat or ultrasounds, sonophysical or sonochemical activation, e.g. thermosensitive or heat-sensitive liposomes, disruption of calculi with a medicinal preparation and ultrasounds · CPC title

• B82Y15/00

  Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors · CPC title

• C12N2320/32

  Special delivery means, e.g. tissue-specific · CPC title

• C12N15/87

  Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation · CPC title

• C12N15/111

  General methods applicable to biologically active non-coding nucleic acids · CPC title