HEPAMUT- The first European project focused on HCC mutanome for neoantigen discovery. Background and rationale: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. Identification and immunological validation of specific mutated neo-antigens may help at improving therapeutic outcome in HCC patients.
Hypothesis: The hypothesis is that identification and immunological validation of HCC-specific mutated neo-antigens will be critical for developing immunotherapy strategies for better clinical outcome in HCC patients.
Aims: The primary aim is the identification and immunological validation of mutated neo-antigens specific to HCC. Specific aims will be: 1) evaluate the mutational rate in HCC and predict the presentation of neo-epitopes by HLA-A2*01 allele; 2) assess the frequency of specific T cells to such mutant epitopes in HCC patients, before and after treatment with checkpoint inhibitors (CI); 3) validate the immunogenicity of neo-epitopes in an HLA-transgenic mice and their therapeutic effect in a PDX animal model; 4) identify mutated full-length proteins on the surface of HCC cancer cells; 5) develop MAbs to such mutated proteins and validate their specificity in a PDX animal model. Potential impact: HCC specific mutated neo-antigens will provide a source of immunogens for immunotherapies to be used alone or in combination with HCC specific shared wild-type antigens identified within the ongoing FP7-funded HEPAVAC project (Coordinator L. Buonaguro).
Phase I-II clinical trials of VIREXXA
Endometrial cancer is one of the most common invasive gynecologic cancers. In Europe the estimated incidence was 28.3 per 10.000 and estimated mortality 6.8 per 10.000 in 2006. Currently no therapy of recurrent or metastatic endometrial carcinoma is available to patients.
Growth of the uterine endometrium is controlled by estrogen and progesterone. Endometrial carcinogenesis is related to estrogen overexposure that is not modulated by the differentiating effects of progesterone and the role of progesterone in the glandular epithelium of the endometrium is primarily to induce cellular differentiation and to antagonize estrogen-mediated cell proliferation. The biological functions of progesterone are mediated through progesterone receptors, which function as ligand-responsive transcription factors in the nucleus.
VIREXXA is known as a safe low-molecular interferon inducer. Now VIREXXA is proposed for the adjunctive use together with progestins in treatment of stage IV endometrial cancer. As was shown in preclinical studies VIREXXA modifies progestin and estrogen receptors level.
In the clinical trial performed in 2000-2002 17 of 21 PR-negative patients with endometrial cancer became PR-positive following short course VIREXXA or VIREXXA + progestin treatment.
Mechanism studies and preclinical development of CD443MUT
CD44-3MUT is a new human recombinant protein drug candidate which has an inhibitory effect on angiogenesis. Angiogenesis is an essential process in the growth and progress of solid tumors.
The aim of the project is to carry out pre-clinical research of a new anti-cancer drug candidate, define its mechanism of action, and to develop novel forms of the molecule.
To improve the stability and the plasma half-life of CD44-3MUT an optimized modification protocol for N-terminal conjugation with 20 kDa polyethylene glycol (PEG) has been developed. PEG-ylated protein versions testing in in vivo half life in rat serum using ELISA based assay showed significantly extended half life compared to unmodified protein. Therapeutic efficacy of modified protein versions will be tested in mouse s.c. tumor xenograft model. Based on half-life and functional assays optimal CD44-3MUT-X kDa PEG version will be chosen for further GLP-production.
CD44-3MUT inhibits cell proliferation in endothelial cells. CD44-3MUT treatment causes significant inhibition of cyclin D1 and p27Kip1 (cdk inhibitor) protein levels in quiescent serum starved endothelial cells entering cell cycle G1 phase. CCCR’s previous research has shown that CD44-3MUT is rapidly endocytosed by endothelial cells by binding intermediate filament protein vimentin on endothelial cell surface. Vimentin mediates CD44-3MUT endocytosis, as endothelial cells isolated from vimentin knock-out mice have lost this function.
Identification and characterizing inhibitors of Wnt and AKT pathways
This project is focused on the identification and validation of inhibitors of protein-protein interactions, which regulating the pathways playing major role in cancer formation and maintenance. By using a FRET-based small molecule screen we have previously identified an inhibitor of the p53-Hdm2 interaction. Hdm2 is the major antagonist of the tumor suppressor protein p53 and is overexpressed in a variety of tumors inactivating the p53 function.
Currently, the project centers on the WNT and AKT signaling pathways, which play a major role in the maintenance of hepatocellular carcinoma (HCC). To enhance the sensitivity and specificity of the compound identification, a protein complementation assay (PCA)-based small molecule screen is set up with the aim to find the inhibitors of β-catenin-TCF4 and Akt1-PDK1 interactions. These play a definitive role in the regulation of the activity of the WNT and AKT signaling pathways correspondingly. Identified compounds will be tested in cultured HCC cells and in mouse HCC models in vivo.
Cell penetrating antibodies
Novel treatment possibilities against melanoma
Anticancer vaccines based on Plant Virus Coat Protein Virus Like Particles
The worldwide incidence of malignant melanoma is rising at a faster rate than of any other solid tumor. Due to limited efficacy of chemo- and radiation therapy, immunotherapy has become a major focus of investigational treatment of melanoma.
One of the major drawbacks of most currently available vaccines is their failure to induce cellular immunity against given antigens. In recent years, the use of plant virus-derived virus like particles (VLPs) as novel systems for the expression of foreign epitopes and for the development of new vaccines has triggered much interest. By genetically modifying these viruses, immunogenic peptides can be fused efficiently to the coat protein (CP) and exposed on the surfaces of the repetitively assembled plant VLPs. These VLPs can induce humoral and also cellular immunity against given antigens.
A novel antigen presentation system based on Potato A potyvirus (PVA) CP VLPs is being developed to present foreign epitopes from melanoma tumor associated antigens to the immune system. Our aim is to investigate whether PVA CP VLPs can serve as an effective carrier to induce (alone and/or in combination with selected immunostimulator(s)) B and T cells immune responses and can serve as a melanoma vaccine candidate.
Clinical development of a new drug candidate against rare leukemias
Novel chemically modified oligonucleotides as anti-cancer drugs
Tropomyosin related kinase (Trk) receptors TrkA and TrkB are receptors for neurotrophins nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF). Expression and function of these Trk receptors and their ligands is crucial for normal development and homeostasis of the mammalian nervous system. In addition, overexpression of the tyrosine kinase receptors TrkA and TrkB is associated with development of several cancers, including neuroblastoma, breast cancer, lung cancer, hepatocellular carcinoma, prostate cancer, Wilms’s tumor, pancreatic cancer, multiple myelomas and others.
The signals transmitted by activated TrkA and TrkB promote cellular proliferation, invasivity, metastasis, and resistance to anoikis (cell death induced by loss of cell adhesion) and chemotherapy in cancer cells accompanied by poor prognosis for cancer patients. In addition Trk receptors have critical roles in the development of tumor neovascularization and mediate pain response, including in cancer. Therefore inhibitors of TrkA and TrkB receptors could be powerful anticancer drugs.
Antisense technology is one of the most straightforward approaches for suppressing unwanted gene expression. We have recently developed a technology based on chemically modified oligonucleotides that are bound to organic nuclease. These oligonucleotides are therapeutically effective at significantly lower concentrations than standard antisense oligonucleotides or siRNAs. The aim of this project is to develop novel modified oligonucleotides specific for Trk receptors, as prototypes for new generation of antisense drugs.
Cellular immunotherapy based on activation of NK cells and T cells
The immunotherapy project is focused on the use of Natural Killer (NK) cells for immunotherapy of malignant diseases, in particular hematological diseases.
The project is addressing the questions of cell culture media optimization to reduce interdonor variability in NK cell in vitro expansion and optimize large scale culture of NK cells. We plan to release improved cell culture vessels and growth media for specific expansion of immunologically active lymphocytes.
In the gene therapy line of work, we have constructed mutated NK cell activation receptors with potential to retarget gene-modified NK cells. Work is under way to optimize the mutants and the gene transfer system for use in primary donor cells and test their function in vitro and in murine leukemia models.
We have planned a phase I/II clinical trial for using NK cells on Acute Myeloid Leukemia, Chronic Myeloid Leukemia and Multiple Myeloma.
The overall aim of the project is to develop cell-based immunotherapy for treatment of malignant diseases and facilitate its establishment in clinical practice.