Triple-negative breast cancer is one of the most lethal cancers. Despite advances with the introduction of poly (ADP-ribose) polymerase 1 inhibitors (PARPi) for patients with BRCA1/2 mutations, therapeutic resistance, early relapse and poor survival are still a reality. In this way, liposomes emerged as a versatile and widely studied platform for the delivery of biologically active molecules due to their ability to enhance the therapeutic efficacy of these agents and significantly minimize side effects.

A new liposomal formulation containing a synergistic combination of a PARPi and a copper complex, called LUV/CBP-01N-PARPi, was evaluated for its antitumor potential in an in vivo chorioallantoic membrane (CAM) model using chicken embryos bearing TNBC.

Previously, an in vivo toxicity assay was conducted using the CAM model of chicken embryos, in which windows were opened in the shells of fertile chicken eggs on the 7th day of incubation. The membrane was removed from the underlying CAM vascular membrane and the treatments were inserted into the window and CAM. For LUV/CBP-01N-PARPi, five serial dilutions were used. Treatments with free CBP-01N and PARPi and with the combination of both (CBP-01N-PARPi) were also included. Treatments with the dilution vehicles PBS 1 × and dimethyl sulfoxide were also carried out. In each group, five eggs were included. After 8 days, the live embryos were euthanized, weighed and macroscopic lesions were evaluated. For the inhibition assay, a xenograft was performed using human triple-negative breast cancer cells (MDA-MB-231) on the CAM. Matrigel and the LUV/CBP-01N-PARPi treatment were inoculated. Treatments with control LUV, 1 × PBS, DMSO and CBP-01N-PARPi/free were also included. For each group, 10 eggs were separated, and, on the 14th day, they were opened to evaluate the size of the tumor mass. The entire procedure with Gallus gallus domesticus embryos was authorized by the Ethics Committee on the Use of Animals (CEUA), under Protocol n° 23117.081885/2023-22.

In the toxicity assay, survival, weight loss, and malformations in the internal organs of the embryo were evaluated. Starting from the fourth dilution, 80% survival was achieved and only one hepatic alteration was observed, which was then selected for the antitumor assay. In the treatments with PBS, DMSO, and control LUV, tumor masses with prominent vascularization were observed. In the treatment with CBP-01N-PARPi-free the tumor was formed but with a smaller volume compared to the controls. However, the most pronounced effect was observed in the treatment with LUV/CBP-01N-PARPi, where a small, dark, and friable mass was present. These findings suggest that the nanostructured system enhances the release of the synergistic agents, potentiating the antitumorigenic effect. Controlling the progression of TNBC cells is important to inhibit tumor progression processes such as metastasis.

LUV/CBP-01N-PARPi proved to be promising in inhibiting the progression of TNBC cells. Additional assays are being conducted to validate the antitumor effect. However, the potential of LUV/CBP-01N-PARPi as an innovative strategy for the treatment of TNBC is undeniable.