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An integrated nano-reagent for cancer diagnosis and treatment and its preparation method and application

A reagent and cancer technology, applied in the field of cancer diagnosis and treatment integrated nano reagents and its preparation, to achieve the effect of SERS signal enhancement

Active Publication Date: 2022-07-29
NANJING UNIV OF POSTS & TELECOMM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, methods to achieve intercellular miRNA-triggered DNAzyme-based dual gene silencing, especially an all-in-one reagent for synergistic enhancement of target-triggered nanoparticle cascade assembly and DNAzyme amplification for cancer diagnosis and therapy have not yet been reported.

Method used

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  • An integrated nano-reagent for cancer diagnosis and treatment and its preparation method and application
  • An integrated nano-reagent for cancer diagnosis and treatment and its preparation method and application
  • An integrated nano-reagent for cancer diagnosis and treatment and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] Example 1: Preparation of miRNA-responsive cancer diagnosis and treatment integrated reagents

[0076] 1. Preparation of AuNP-Y

[0077] 1) Mix equal amounts (50 μM, 10 μL) of Y-motif ssDNA-A, Y-motif ssDNA-B and Y-motif ssDNA-C (i.e. Ya, Yb and Yc, respectively) in 50 μL PBS buffer, Then annealing is performed to assemble the Y-shaped DNA structure, that is, the Y-motif;

[0078] 2) After incubating 50 μL solution containing the above 10 μM Y-motif with 500 μL gold nanoparticles (0.3 nM), namely AuNPs at room temperature for 12 h, carry out aging treatment, that is, adding sodium chloride solution four times, each time At 30 min intervals, the final sodium chloride concentration was 0.2 M, and the mixture was gently shaken at room temperature for 12 h;

[0079] 3) Finally, wash with PBS three times by centrifugation at a rotational speed of 7000 rpm, and the time of each centrifugation is 20 min; then resuspend the precipitate in 50 μL of PBS to obtain AuNP-Y, and ...

Embodiment 2

[0086] Example 2: Reaction mechanism characterization of miRNA-responsive cancer diagnosis and treatment integrated reagents

[0087] like figure 1 Shown is the working principle of the miRNA-responsive cancer diagnosis and treatment integrated reagent of the present invention. In order to verify the correctness of the mechanism, a 10% PAGE gel was used for gel electrophoresis experiments, and 5 μL of DNA samples were mixed with 1 μL of 6×DNA loading buffer. After 80 min of operation at 80 V, imaging was performed.

[0088] figure 2 The formation of Y-motif and its specific response to miRNA-106a and ATP were characterized by 10% PAGE in . The DNA samples correspond specifically to DNA samples in different lanes, namely: M: DNA marker; Lane 1: Ya; Lane 2: Yb; Lane 3: Yc; Lane 4: Ya, Yb and Yc in equal amounts Hybridization (Y); Lane 5: miRNA-106a; Lane 6: Equivalent hybridization of Y with miRNA-106a; Lane 7: Equivalent hybridization of Yb with Yc; Lane 8: L; Lane 9: c-J...

Embodiment 3

[0093] Example 3: Feasibility verification of miRNA-triggered AuNP networked nanostructure assembly and SERS sensing

[0094] An equal amount (10 μL) of AuNP-Y and AuNP-D in Example 1 was mixed with 80 μL PBS containing 0.5 nM miRNA-106a, and incubated at 37°C with gentle shaking for 2 h to form target-induced AuNP networked nanoparticles. structure. The AuNP network nanostructures were characterized by absorption, dynamic light scattering (DLS) and scanning electron microscopy (SEM).

[0095] like Figure 4A As shown, the surface plasmon resonance (SPR) peak of AuNPs was red-shifted from 524 nm to 526.4 nm by the modification of Y-motifs. Similarly, through the modification of dsDNA linker and DTNB, the SPR of AuNPs was red-shifted to 526 nm, which indicated that AuNP-Ds were successfully prepared.

[0096] like Figure 4B The DLS characterization shown, the hydrodynamic diameters of AuNP-Y and AuNP-D are about 33 nm and 31.6 nm, respectively, and the SEM images show th...

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Abstract

The invention discloses an integrated nano-reagent for cancer diagnosis and treatment, including a miRNA identification probe prepared by modifying a Y-shaped DNA structure on the surface of AuNP, and a miRNA identification probe prepared by modifying a double-stranded DNA linker and a Raman molecule on AuNP For the obtained SERS probe, the invention also discloses the preparation method and application of the above-mentioned integrated reagent for cancer diagnosis and treatment. The integrated nano-reagent for cancer diagnosis and treatment of the present invention, driven by ATP in cancer cells, the specially designed Y-shaped DNA structure and double-stranded DNA linker on AuNP will undergo conformational transformation triggered by miRNA-106a, releasing miRNA- 106a and cyclic amplification, triggering the SERS signal to significantly enhance the recognition of cancer cells; at the same time, the DNAzyme generated and amplified by the conformational switch can catalyze the cleavage of Survivin mRNA and c-Jun mRNA, realizing effective dual gene silencing therapy.

Description

technical field [0001] The invention belongs to the technical field of functional nano-probes, and in particular relates to an integrated nano-reagent for cancer diagnosis and treatment and a preparation method and application thereof. Background technique [0002] The invention of a diagnostic and therapeutic integrated reagent with both diagnostic and therapeutic functions is an important way to achieve synergistic treatment of cancer with high efficiency and low toxicity, and is of great significance for early accurate diagnosis and effective treatment of cancer. The integrated reagent for realizing image-guided cancer therapy provides a powerful means for precise identification of cancer cells, further precise treatment, and timely monitoring of treatment effects. All-in-one reagents that can autonomously initiate treatment modalities after accurate identification of cancer, the cascade process of detection and treatment, have received great attention this year. However...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886C12Q1/6816C12N15/11A61K47/52A61K31/713A61P35/00B82Y5/00B82Y30/00
CPCC12Q1/6886C12Q1/6816A61K47/52A61K31/713A61P35/00B82Y5/00B82Y30/00C12Q2600/158C12Q2600/178C12Q2600/136C12Q2525/207C12Q2563/137C12Q2563/155C12Q2565/632
Inventor 宋春元汪联辉董晨
Owner NANJING UNIV OF POSTS & TELECOMM
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