Tumor Cell–Targeted Delivery of Nanoconjugated Oligonucleotides in Composite Spheroids

Abstract

Standard tissue culture has often been a poor model for predicting the efficacy of anti-cancer agents including oligonucleotides. In contrast to the simplicity of monolayer tissue cultures, a tumor mass includes tightly packed tumor cells, tortuous blood vessels, high levels of extracellular matrix, and stromal cells that support the tumor. These complexities pose a challenge for delivering therapeutic agents throughout the tumor, with many drugs limited to cells proximal to the vasculature. Multicellular tumor spheroids are superior to traditional monolayer cell culture for the assessment of cancer drug delivery, since they possess many of the characteristics of metastatic tumor foci. However, homogeneous spheroids comprised solely of tumor cells do not account for some of the key aspects of metastatic tumors, particularly the interaction with host cells such as fibroblasts. Further, homogeneous culture does not allow for the assessment of targeted delivery to tumor versus host cells. Here we have evaluated delivery of targeted and untargeted oligonucleotide nanoconjugates and of oligonucleotide polyplexes in both homogeneous and composite tumor spheroids. We find that inclusion of fibroblasts in the spheroids reduces delivery efficacy of the polyplexes. In contrast, targeted multivalent RGD-oligonucleotide nanoconjugates were able to effectively discriminate between melanoma cells and fibroblasts, thus providing tumor-selective uptake and pharmacological effects.

Collective cancer cell invasion requires RNA accumulation at the invasive front

“…However, currently, efficient targeting and sustained effects can be attained when targeting cells of the central nervous system ( 57 , 58 ). Methodologies to achieve tumor-specific delivery, in combination with chemistries ensuring efficient uptake and sustained action ( 59 – 61 ), will facilitate the exploration of altering subcellular RNA distributions as a therapeutic approach.…”

A novel pancreatic tumour and stellate cell 3D co-culture spheroid model

“…Extending MCTS and other matrix 3D models to allow for studies of tumour-stromal crosstalk, there are now e.g. tumour spheroid models of cancer cells and fibroblasts in lung cancer and cervical carcinoma [21], melanoma [22], breast [23,24], colorectal [25,26], liver [27] as well as pancreatic cancer [28]. Although more complex models with multiple cell lines are desirable to answer certain types of questions, there is still a need for simpler co-culture models with only two cell types, where direct cell cross-talk can be more easily investigated.…”

Bioconjugates for targeted delivery of therapeutic oligonucleotides

“…Both methods produced small NPs with uniform and monodispersed size distribution at a diameter of 12-13nm, which may be a favorable size for long circulation time [31, 80]. The intermediate sized HSA-ON conjugates could penetrate deeply and distribute throughout 3-D tumor spheroids (Fig 4), whereas the conventional NPs with sizes over 300 nm could only deliver to the cells on the surface of the tumor spheroids [31, 81, 82]. Utilization of HSA as carrier enabled displaying multiple (10-15) targeting ligands on the surface of the NPs, and this dramatically enhanced receptor-specific cellular delivery of ONs in integrin-expressing cancer cells (Fig 4).…”