Summary

Yizhou Dong’s group at Ohio State University describe a small molecule/mRNA lipid nanoparticle combination therapy for triple negative breast cancer (TNBC). TNBC represents about 15% of all breast cancers. The Dong lab’s approach involved delivering a conventional chemotherapy drug, paclitaxel (PTX), as well as an mRNA encoding tumour-suppressor gene p53. They found that having both p53 mRNA and PTX together worked better than either alone in terms of reducing tumour volume and survival in an orthotopic mouse model.

Mechanisms of Action
PTX is a conventional chemotherapeutic agent sold under the brand name Taxol for first line therapy of numerous cancers. Like other taxanes such as docetaxel and cabazitaxel, PTX binds to proteins in microtubules to inhibit cell division. PTX is generally cytotoxic and has numerous known side effects such as vomiting and hair loss. Albumin-based nanoparticle formulations of PTX are available under the trade name Abraxane to alter the biodistribution of PTX. 

The p53 pathway has been identified as an important signalling pathway, as numerous cancers involve genetic and epigenetic changes to p53 and associated pathways of regulation. Normally p53 is dormant, but can be activated through numerous pathways in response to cellular stresses to inhibit growth of abnormal cells. Once activated, p53 is transcribed and acts to modulate cell cycle arrest, apoptosis, or DNA repair among others. In breast cancer, abnormal p53 signalling is associated with worse clinical outcomes.

Formulation Approach
To deliver both the mRNA and PTX, the Dong lab first created an amino lipid conjugated to PTX. They then used their previously-described orthogonal array optimization approach to find a formulation best suited to deliver p53 mRNA in vitro before scaling the lead formulation up for in vivo testing. In vitro screening involved testing an array of 16 formulations for delivery of a reporter gene (GFP) in MDA-MB-231 cells. GFP fluorescence was quantified by flow cytometry. The top performing formulation was chosen for in vivo studies in an orthotopic mouse model. LNPs were made on the NanoAssemblr platform by mixing mRNA in buffer with an ethanol solution containing the lipid mix.

Findings and Conclusions
The combination formulation was tested against PTX formulated with Cremophor EL (solubility enhancer), a p53-mRNA-LNP, and a PTX-conjugated LNP with a control mRNA. The p53-mRNA/PTX combination therapy outperformed the other approaches, leading to significantly smaller tumours and longer survival. This study is a promising proof of principle for combination approaches to treat challenging cancers. With numerous available small molecule drugs as well as an expanding list of tumour associated genetic targets, there is a wealth of possible combinations. NanoAssemblr technology allows researchers to efficiently explore a large number of formulations at the discovery phase, and to scale and further optimize lead formulations through preclinical development and beyond. Furthermore, the reproducibility of NanoAssemblr formulations provide confidence that changes in the outcomes can be attributed to changes in the formulation, thus allowing rational formulation design and optimization of innovative treatment approaches. 

Abstract

Triple-negative breast cancer (TNBC) is one type of the most aggressive breast cancers with poor prognosis. It is of great urgency to develop new therapeutics for treating TNBC. Based on current treatment guideline and genetic information of TNBC, a combinational therapy platform integrating chemotherapy drugs and mRNA encoding tumor suppressor proteins may become an efficacious strategy. In this study, we developed paclitaxel amino lipid (PAL) derived nanoparticles (NPs) to incorporate both chemotherapy drugs and P53 mRNA. The PAL P53 mRNA NPs showed superior properties compared to Abraxane® and Lipusu® used in the clinic including high paclitaxel loading capacity (24 wt.%, calculated by paclitaxel in PAL), PAL encapsulation efficiency (94.7% ± 6.8%) and mRNA encapsulation efficiency (88.7% ± 0.7%). Meanwhile, these NPs displayed synergetic cytotoxicity of paclitaxel and P53 mRNA in cultured TNBC cells. More importantly, we demonstrated in vivo anti-tumor efficacy of PAL P53 mRNA NPs in an orthotopic TNBC mouse model. Overall, these chemotherapy drugs derived mRNA NPs provide a new platform to integrate chemotherapy and personalized medicine using tumor genetic information, and therefore represent a promising approach for TNBC treatment.