Human defense peptides as novel agents against cancer

Each year 10.9 million people worldwide are diagnosed with cancer and there are 6.7 million deaths from this disease. It is estimated that there are 24.6 million people alive who have received a diagnosis of cancer in the last five years. Although much progress has been achieved concerning therapies in the last decades, surgery, chemotherapies or radiation are not successful in more than 50% of the cases and for those who survive, strong adverse reactions and the risk of reoccurrence of the disease represents a big problem. In this project we copy and enforce a weapon of the immune system against degenerated cells in the form of small cationic peptides that are important components of the innate immune system of all species. Some of these amphipathic peptides exhibit besides antibiotic, antifungal and antiviral also anti-tumor activity. To enhance this natural antitumor properties the membrane-active part of Lactoferrin is taken as a parent peptide and will be further optimized in its activity and selectivity towards selected cancer cell lines. Although the exact killing mechanism on cancer cells is not understood, it is supposed that peptide-lipid interactions leading to membrane disruption are responsible for rapid killing by peptides, hindering the development of resistance which can occur with conventional chemotherapeutic drugs. Another big advantage is the strong selectivity for cancerous cells (over healthy cells). This is provided by the interaction of the cationic peptides with cancer cells that expose the negatively charged phospholipid phosphatidylserine (PS) in the outer leaflet of their plasma membranes which in healthy cells only comprise neutral lipids. Peptide optimization in respect of distribution of charges and hydrophobicity will be achieved by structure-activity relationship studies on model membranes followed by efficacy studies on cell cultures of cancer cells of diverse selected cancer types carrying the PS marker. The aim of the project is to develop novel highly potent agents against cancer cells acting rapidly by membrane lysis without any adverse reactions or formation of resistance.

In the 21st century cancer is still a major problem and accounts for 13% of all deaths worldwide. Regarding therapy, much progress has been achieved but cancer is still not curative in more than 50% of the cases. Major problems of therapy are reoccurrence, (multi-) drug resistance, metastases and severe side effects due to inadequate specificity. A new strategy applied within this FWF-project uses host defense peptides like human lactoferricin (hLFcin) and derivatives thereof to specifically target the cancer cell membrane. Host defense peptides which are part of the innate immune system of many diverse species mostly comprise short stretches of amino acids and are cationic and amphipathic. These peptides target the negatively charged phospholipid phosphatidylserine (PS) which is only exposed in the outer leaflet of the cell membrane during malignant transformation, whereas the outer leaflet of the healthy cell membrane comprises the neutral phospholipids phosphatidylcholine (PC) and sphingomyelin (SM). In this study we could prove PS exposure of cancer cells of various cancer types of primary as well as of metastatic lesions, of cancer cell lines as well as of primary cancer cells. Furthermore we demonstrated that PS exposure correlates with stage of malignity. Our findings demonstrate that PS could serve as a uniform marker for cancer cells and as potent target for anticancer peptides. Therefore we tested several hLFcin derivatives, differing in content and length of primary sequence, number of positive charges, hydrophobicity and consequently secondary structure and efficacy, for their toxicity against cancer cells and their interaction with exposed PS. In vitro data nicely correlated with the model studies in which cancer cells are mimicked by PS- and non-cancer cells by PC-liposomes. Biophysical investigations proved that PS is the target for the studied active and cancer selective peptides. In vitro studies revealed different modes of action of the peptides. Some peptides reached their highest activity already after several minutes indicating cell death through membrane lysis, whereas others needed prolonged incubation times up to several hours supporting the idea of a killing mechanism through apoptosis. Interestingly, peptides performing a slow killing seem to be more specific than those who kill through lysis of the cell membrane. Additionally, secondary structure seems to play an important role for the cancer specificity.