CORDIS - EU research results

Nanomedicine and Hadrontherapy

Final Report Summary - NANOHAPY (Nanomedicine and Hadrontherapy)

Summary description of the project objectives
Cancer, the second most common cause of death after cardiovascular diseases, is a major European health concern. To cure the cancer, around 50% of patients receive external radiotherapy, based on X-rays and γ–rays radiations. The main limitation of radiotherapies comes from the sensitivity of healthy tissues surrounding the tumors. Approaches that enhance radiosensitivity within tumors relative to normal tissue therefore have the potential to provide a major positive impact on patients.
Fast ions (mostly carbon and protons) used as ionizing radiations enable high localization of the radiation effects into a well-defined volume (the Bragg Peak). As a result, the dose delivered to healthy tissues is lower compared with photon radiation therapy. However, due to the beam tuning to cover the whole tumor, the dose deposited on the path of the beam in front of the tumour enhances. This remains a high limitation of the hadrontherapy treatments.

It is thus of great challenge to improve the performances of the hadrontherapy by concentrating the effects of ion radiations into the tumor cells. In this perspective, we have proposed a new strategy where nanomedicine is combined to the hadrontherapy. Hence, our main activity consists of testing and characterizing the properties of nanoagents to improve the strategy and help developing new targeted nanoagents that can amplify the effects of fast ions in the tumors.

The project “NanoHapy” has arisen in this frame of development and understanding of nanoagents actions for the hadrontherapy. It has been created in the period when it became crucial to study the biological mechanisms that are involved in nanoparticles actions upon irradiation of tumors.
This project has been thus dedicated to characterization of metal-based nanoparticles (NPs) interaction with cancer cells and elucidation of their biological effects upon irradiation. It has been composed of two research objectives (RO).

• RO1: The characterization of the effects of the NPs interacting with the cells (uptake, localization, toxicity) to select the best NPs and to better understand and control the internalization of NPs into the cells, prior to the irradiation.

• RO2: The determination of the NPs induced biological effects under ion radiation at molecular and cellular scales.

Description of the work performed since the beginning of the project
As mentioned above The Marie Curie postdoctoral researcher (L. Stefancikova, L. S.) came to the host laboratory (Nanomedecine and Hadrontherapy group headed by S. Lacombe, ISMO, Orsay, France) at the moment when it was crucial to enrich the research on metal-based NPs by biological approaches. With her background in molecular and cellular biology and 7 years’ experience in cancer research, she came to search, find, install and optimize new methods to introduce the interdisciplinary group (physics, chemistry) towards the biological world. For this purpose she didn´t hesitate to start new collaborations or involve her former laboratory in part of the research.
Regarding the research on the project itself she has proceeded as follows:
Together with medical doctors and experienced scientists L. S. has selected cancer cell lines as suitable probes for NPs effects evaluation. In the project, she has privileged three high-Z atoms based NPs: gold NPs (AuNPs), gadolinium-based NPs (GdBN) and platinum NPs (PtNPs). She has tested the cytotoxicity of these NPs in cancer and healthy cell lines using approaches to reveal both short and long-term NPs cytotoxic effects. L.S. together with collaborating labs, has quantified NPs in several cell lines by complementary approaches. Furthermore, the fellow has performed various microscopy technics to localize the NPs in the cells. Next, the kinetics of internalization and expulsion of NPs to and out of the cells has been targeted.
L. S. has studied the NPs effects upon irradiation by hadrons and photons on molecular (molecular probe) and cellular scale (living cells). She has tested the biological perturbations induced in the cells irradiated in presence of NPs such as: cell viability and cell death, cell cycle changes, protein expression changes, DNA damage induction and repair and finally real time changes in crucial cell parameters.

Description of the main results achieved so far
The selected results will be mentioned for the two ROs separately:
Three independent technics observing both short-term and long-term toxicity of NPs show no evidences of cytotoxicity of AuNPs, PtNPs and GdBN, respectively. More detailed analysis of higher concentrations uncovered the maximum concentration with no toxic impact on human cells.
NPs feature Gaussian distribution in human cancer cells which suggest intercellular variability in the NPs concentration among the cells. Comparing the fluorescence intensities in several cancer cell lines and human non-cancer cells uncovered great differences in NPs concentrations between the cell lines.
The NPs internalize with unique efficiency and pattern in selected human cancer cell lines and non-cancerous cells. In all cells tested, these NPs were localized in the cell cytoplasm out of cell nuclei.

The NPs significantly increase the number of DSBs on DNA plasmids as molecular probes, this show their radiation effect enhancing potential. Clear radiosensitisation by NPs has been detected also in-vitro for gamma rays and fast carbon ions (C6+) with greater effect when using ions.
Cell viability tested after irradiation of cancer cells with or without NPs show decrease of viability for cells undertaken the combined treatment. The NPs effects in the cells are NPs-type, irradiation-type and cell type specific. Detailed analyses are required to understand which cellular background is favourable for NPs use in clinics.

Introduction and optimization of biological technics by L. S. initiated a new research focus in the host laboratory which continues even after the end of the project. Similarly, the new collaborations that she has launched continue to be vital and intense.

L. S. has obtained a postdoc position in a well-known research institute in France (Institut de Genetique Humaine, IGH, Montpellier and is now preparing a proposal to couple the project on understanding of biological effects of metal-based NPs with her new project dedicated to the resistance of tumors to the therapies.

Final results and their potential impact and use
The results described above showed importance to test different NPs on several cancer cell lines as the effect of NPs is not equal. The NPs when combined with irradiation can significantly increase the effects of radiotherapy and result in better targeting of tumors. Clear understanding of molecular changes in cells that are responsible for tumor resistance to even combined treatment with NPs are needed. Personalized approach will be required in clinics.

The Marie Curie IEF project has opened new perspectives in horizon of understanding of metal-based NPs actions upon radiation treatment. Thanks to the research carried out during the project we have understood the necessity to dig deeper to the molecular mechanisms of cells response to the combined treatment involving NPs. The arising project of the fellow in her current laboratory promises to continue in this direction.

The results of the research carried out during the project have been presented by the fellow on 5 international conferences, two of which as invited talks. The results have also been described in three first author papers (one published, one submitted and one in preparations) and five co-authored articles (one already published and 4 in preparations).

1) Stefancikova L et al., Cancer Nanotechnology 2014, 5:6.
2) Schlathölter T et al., International Journal of Nanomedicine, 2016,11:1549-1556.
3) Stefancikova L et al., Journal of Nanobiotechnology, minor revisions