DNA Nanotechnology for Cancer Diagnosis and Therapy.

Cancer is one of the leading causes of mortality worldwide, because of the lack of accurate diagnostic tools for the early stages of cancer. Thus, early diagnosis, which provides important information for a timely therapy of cancer, is of great significance for controlling the development of the disease and the proliferation of cancer cells and for improving the survival rates of patients. To achieve the goals of early diagnosis and timely therapy of cancer, DNA nanotechnology may be effective, since it has emerged as a valid technique for the fabrication of various nanoscale structures and devices. The resultant DNA-based nanoscale structures and devices show extraordinary performance in cancer diagnosis, owing to their predictable secondary structures, small sizes, and high biocompatibility and programmability. In particular, the rapid development of DNA nanotechnologies, such as molecular assembly technologies, endows DNA-based nanomaterials with more functionalization and intellectualization. Here, we summarize recent progress made in the development of DNA nanotechnology for the fabrication of functional and intelligent nanomaterials and highlight the prospects of this technology in cancer diagnosis and therapy.

Prospective observational study on biomarkers of response in pancreatic ductal adenocarcinoma.

Adjuvant chemotherapy benefits patients with resected pancreatic ductal adenocarcinoma (PDAC), but the compromised physical state of post-operative patients can hinder compliance. Biomarkers that identify candidates for prompt adjuvant therapy are needed. In this prospective observational study, 1,171 patients with PDAC who underwent pancreatectomy were enrolled and extensively followed-up. Proteomic profiling of 191 patient samples unveiled clinically relevant functional protein modules. A proteomics-level prognostic risk model was established for PDAC, with its utility further validated using a publicly available external cohort. More importantly, through an interaction effect regression analysis leveraging both clinical and proteomic datasets, we discovered two biomarkers (NDUFB8 and CEMIP2), indicative of the overall sensitivity of patients with PDAC to adjuvant chemotherapy. The biomarkers were validated through immunohistochemistry on an internal cohort of 386 patients. Rigorous validation extended to two external multicentic cohorts-a French multicentric cohort (230 patients) and a cohort from two grade-A tertiary hospitals in China (466 patients)-enhancing the robustness and generalizability of our findings. Moreover, experimental validation through functional assays was conducted on PDAC cell lines and patient-derived organoids. In summary, our cohort-scale integration of clinical and proteomic data demonstrates the potential of proteomics-guided prognosis and biomarker-aided adjuvant chemotherapy for PDAC.

Smartly responsive DNA-miRNA hybrids packaged in exosomes for synergistic enhancement of cancer cell apoptosis.

Endogenous and exogenous tumor-related microRNAs (miRNAs) are considered promising tumor biomarkers and tumor therapeutic agents. In this work, we propose a miRNA self-responsive drug delivery system (miR-SR DDS), which enables the association between endogenous and exogenous miRNAs, so as to achieve a smart responsive and synergistic drug delivery. The miR-SR DDS consists of DNA-miRNA hybrids of let-7a and the complementary DNA of miR-155, which was packaged in exosomes. In response to the overexpressed miR-155 in breast cancer cells, the hybrids disintegrate and release let-7a and the complementary DNA of miR-155 to inhibit the expression of HMGA1 and relieve the inhibition of SOX1, respectively. Under the dual-targeted gene regulation, results show that the growth, migration and invasion of breast cancer cells can be synergistically inhibited through the Wnt/ß-catenin signaling pathway. The concept and successful practice of the miR-SR DDS can be used as a reference for the development of miRNA drugs.

A proximity-exponential hybridization chain reaction (PEHCR) and its application for nondestructive analysis of membrane protein-protein interactions on living cells.

Though a variety of methods have been developed for the analysis of membrane protein-protein interactions (PPIs), amplified, dynamic and nondestructive analysis in situ is always a challenge. To address this issue, here we develop a method called proximity-exponential hybridization chain reaction (PEHCR). In our strategy, when two membrane proteins approach due to interaction, they will draw their respective oligonucleotide-labeled antibodies together. The proximity of the oligonucleotides thereafter triggers a well-designed enzyme-free exponential hybridization chain reaction, which can output amplified fluorescence imaging signals. As a model, analysis of EGFR-HER2 interactions under the regulation of different activators and inhibitors is achieved. Owing to the superior signal amplification performance, we are able to clearly observe the membrane PPIs by using a common fluorescence microscope. Furthermore, unlike the existing proximity techniques that require enzymes, our enzyme-free strategy avoids the need to use a specific buffer suitable for enzyme catalysis and can be run directly in cell liquid media to maximize the physiological activity of the cells. So, dynamic analysis of membrane PPIs on living cells is achieved, and the cells, after the analysis, are still alive and are available for other usage. The successful implementation of this work enriches the toolbox for the study of membrane PPIs especially on those heterogeneous cell populations with small amount.

An all-in-one homogeneous DNA walking nanomachine and its application for intracellular analysis of miRNA.

DNA walker is a powerful type of DNA nanomachine that can produce amplified signals during the "burnt-bridge"-like walking process. Despite their successful application in extracellular bioanalysis, the heterogeneity of the existing DNA walkers makes it difficult to guarantee the consistency of the results during the analysis of different cells. Methods: Here, an all-in-one homogeneous DNA walking nanomachine is reported that can be delivered into living cells for intracellular bioanalysis of miRNA without auxiliary materials. Results: This DNA walking nanomachine is constructed of gold nanoparticles on which two types of interrelated DNA tracks are assembled. The target miRNA, cancer-related miR-21, can be captured by one of the tracks (track 1) and then walk to the other track (track 2), releasing the hybrid of track 1 and track 2 from the nanoparticle to produce a signal. The walking process can proceed in a cyclic 1-2-1-2 manner and thereby produce amplified signals. Thus, sensitive imaging of the miRNA in situ can be achieved. Conclusion: Benefiting from the homogeneity of the detection system, the method can be applied for intracellular analysis without interference induced by the fluctuations of stimuli or accessorial contents.

An electrochemical biosensor integrating immunoassay and enzyme activity analysis for accurate detection of active human apurinic/apyrimidinic endonuclease 1.

A novel electrochemical biosensing method that can take into account both immunoassay and enzyme activity analysis was reported in this work for determination of the enzymatically active human apurinic/apyrimidinic endonuclease 1 (APE1). The basic principle is to design and construct a DNA catalytic hairpin assembly (CHA) triggered by APE1 catalysis in enzyme activity analysis, and the assembled DNAs are labeled with electrochemically active CdS and PbS quantum dots to output electrochemical signals. In this system, the signal generation needs to satisfy both the conditions of immunological recognition and enzymatic catalysis, providing a basis for accurate analysis of active APE1. Results show that this method can reflect the regulation of the enzyme activity and can also distinguish APE1 from its isozymes with the same enzyme activity. The concept and successful implementation of this integrated system will contribute to the research and application of APE1 in biomedicine, and provide a reference for the accurate analysis of other enzymes.

Triplex DNA Nanoswitch for pH-Sensitive Release of Multiple Cancer Drugs.

A DNA-based stimulus-responsive drug delivery system for synergetic cancer therapy has been developed. The system is built on a triplex-DNA nanoswitch capable of precisely responding to pH variations in the range of ∼5.0-7.0. In extracellular neutral pH space, the DNA nanoswitch keeps a linear conformation, immobilizing multiple therapeutics such as small molecules and antisense compounds simultaneously. Following targeted cancer cell uptake via endocytosis, the nanoswitch inside acidic intracellular compartments goes through a conformational change from linear to triplex, leading to smart release of the therapeutic combination. This stimuli-responsive drug delivery system does not rely on artificial responsive materials, making it biocompatible. Furthermore, it enables simultaneous delivery of multiple therapeutics for enhanced efficacy. Using tumor-bearing mouse models, we show efficient gene silencing and significant inhibition of tumor growth upon intravenous administration of the smart nanoswitch, providing opportunities for combinatorial cancer therapy.