Background

Although traditional cancer treatments (e.g., chemotherapy, radiotherapy, and surgery) have been widely applied in clinical cancer therapy, they have many side effects (such as systemic toxicity or incomplete elimination), leading to poor therapeutic efficiency and easy recurrence and metastasis. In recent years, photothermal therapy (PTT) has been broadly investigated because of its advantages in deeper tissue penetration, higher spatial selectivity, and local tumor thermal ablation with less systemic toxicity and higher anti-tumor efficiency (Xiong et al., 2020; Zhou et al., 2021). However, due to the non-uniform tumor distribution of photothermal agents (PAs), PTT cannot kill all tumor cells (Yan et al., 2016). Therefore, it is urgent to develop a novel method to achieve uniform intra-tumor distribution (UITD) of PAs and high photothermal therapeutic efficiency.

Several size shrinkage strategies based on cross-linking covalent bonds have been developed to improve UITD of PAs (Li et al., 2016; Zhou et al., 2019; Wang et al., 2020). However, covalent bonds with high bond energy cannot be broken quickly and the synthesis of size-shrinkable nanosystems is complex, which easily results in differences in physicochemical characteristics between different batches. Because of the low hydrogen bond energy in the DNA double helix and the simplicity of DNA self-assembling process, DNA-assembled nanotheranostics may be a promising platform to achieve UITD of PAs.

In this protocol, we will introduce a DNA-assembled nanosystem, i.e., DNA-assembled GRS-DNA-CuS nanodandelion (as shown in Figure 1), which is composed of the core of a Gap-enhanced rod-shaped gold structure (termed as GRS) modified by single-stranded DNA (ssDNA), and the shell of ultrasmall copper sulfide (CuS) PAs modified by complementary single-stranded DNA (cssDNA) (Zhang et al., 2022). Based on the principle of Watson–Crick base pairing, GRS-DNA-CuS nanodandelions are prepared by cross-linking hydrogen bonds between GRS-ssDNA and CuS-cssDNA PAs. Under the first near-infrared (NIR) laser irradiation, hydrogen bonds in the GRS-DNA-CuS nanodandelions are explosively broken once above the DNA melting temperature, and the large-sized GRS-DNA-CuS nanodandelions are dissociated into GRS and ultrasmall CuS PAs within one minute (Figure 1). Both the rapid increase in local concentration of ultrasmall CuS PAs and the slight increase in intratumor temperature have effectively promoted UITD of CuS PAs. This UITD helps to eliminate the subcutaneous solid tumors under the second NIR light irradiation. In a word, we will present the detailed preparation process of DNA-assembled GRS-DNA-CuS nanodandelions and characterize their physicochemical and dissociation properties in order to provide a novel UITD-guided PTT strategy and advance their therapeutic and diagnostic applications in clinic.

Figure 1. Schematic illustration of the dissociation process of DNA-assembled GRS-DNA-CuS nanodandelions under 808 nm NIR laser irradiation. Left: 3D structure of DNA-assembled GRS-DNA-CuS nanodandelions. GRS: Gap-enhanced rod-shaped gold structure; CuS: copper sulfide.