FIELD OF THE INVENTION

The present invention relates to a method for the preparation of an immunogenic lysate from mesothelioma tumor cells, to such a lysate and to dendritic cells loaded with the lysate. Furthermore, the invention provides a pharmaceutical composition comprising such lysate or dendritic cells. Moreover, the invention further provides the use of the lysate, the dendritic cells or the pharmaceutical composition in the prevention or treatment of mesothelioma.

BACKGROUND OF THE INVENTION

Malignant mesothelioma of the pleural cavity is a highly lethal neoplasm. With median survival durations of 9-15 months from the onset of symptoms, the prognosis is poor. To date, there is no standard curative therapy for malignant pleural mesothelioma (hereinafter: mesothelioma). Surgical approaches such as pleurectomy and extrapleural pneumonectomy result in high local recurrence rates and questionable survival benefit. The only treatment approved by the US Food and Drug Administration (FDA) is pemetrexed (Alimta)/cisplatin and has a median survival benefit of three months. Because of the limited success of current treatments, novel therapeutic regimens are urgently needed. The potential to harness the potency and the specificity of the immune system underlies the growing interest in cancer immunotherapy. One approach to activate the patient's immune system uses dendritic cells to present tumor-associated antigens and thereby generating anti-tumor specific responses.

Dendritic cells are highly mobile and extremely potent antigen presenting cells located at strategic places where the body comes in contact with its environment. In these locations they pick up antigens and transport them to the secondary lymphoid organs where they instruct and control activation of natural killer cells, B and T-lymphocytes, and efficiently activate them against the antigens [1]. This property makes them attractive candidates for use in therapeutic strategies against cancer [2-3]. Dendritic cells can be generated in large numbers in vitro; bypassing antigen presentation deficiencies caused by the immune suppressing tumor environment [4-7], and can subsequently be injected in a mature state to induce anti-tumoral responses [8-11].

Dendritic cell based immunotherapy has been found to induce protective and anti-tumor immunity in experimental animals and in some cancers in humans e.g. renal cell carcinoma, melanoma, glioma and lung cancer [12]. This has also been proven for mesothelioma. It has particularly been found that dendritic cell based immunotherapy had a beneficial effect on survival and reducing tumor growth in a mouse model for malignant mesothelioma [13].

Based on these preclinical findings in laboratory animals, a clinical trial was initiated to address the safety and feasibility of pulsed dendritic cells to induce tumor-specific cytotoxic T-cell responses in mesothelioma patients [14]. Ten patients were treated with chemotherapy (pemetrexed/cisplatin) followed by three vaccinations of autologous tumor lysate-loaded monocyte-derived dendritic cells. Chemotherapy was given prior to dendritic cell vaccination in order to reduce tumor load, thereby potentially augmenting the efficacy of the vaccination, as was earlier found in mice. From this study it became clear that injection of autologous tumor lysate-pulsed dendritic cells in mesothelioma patients after chemotherapy was safe and well tolerated.

However, using autologous tumor cell lysate to load dendritic cells has certain theoretical and practical disadvantages. A key problem associated with the autologous approach is that the number of tumor cells from resected tumor material of thoracic malignancies, in particular mesothelioma (either pleural fluid or biopsy) is very limited. As an example, in the first human clinical trial with dendritic cell based immunotherapy in mesothelioma, most patients were excluded from participation [14]. From a total number of 57 mesothelioma patients only ten patients were able to provide enough tumor material, although all patients had high tumor burden on CT/MRI scan.

Furthermore, autologous tumor material obtained from patients suffering from mesothelioma is very divers in total tumor amount and “contamination” with other cells. This results in highly variable tumor cell lysates making standardization extremely difficult. When such tumor material is loaded onto autologous dendritic cells different outcomes of the phenotype and stimulatory capacity are awaited. Moreover, preparing the autologous tumor material for each individual patient is time consuming and laborious. Also, quality tests are needed on each patient's lysate batch, making the procedure of dendritic cell based immunotherapy with autologous tumor lysate very expensive.

Another limitation associated with the use of autologous mesothelioma tumor cells, is the difficulty of in vitro manipulation of tumor derived cells. Different approaches to optimally prepare tumor cell derivatives (apoptotic or necrotic fragments), increasing the immunogenicity or enrichment by protein fractionation, and other important steps to increase efficacy, cannot be performed directly on autologous tumor material. Culturing autologous cells ex-vivo, until the number of cells is adequate for manipulation, is not an option for reasons that even short term culture of mesothelioma cells mostly failed to produce cell lines. Another reason why using autologous tumor cells is not feasible is the fact that relatively long culture periods are needed, during which the patient's disease progresses further.

In the prior art it has been suggested to use apoptotic allogeneic mesothelioma tumor cells for loading autologous dendritic cells. In this regard reference is made to the article of Ebstein et al, 2003, in the American Journal of Respiration and Critical Care Medicine. In this article intact apoptotic allogeneic mesothelioma tumor cells derived from one cell line are used in an in vitro study wherein autologous dendritic cells are loaded with said intact apoptotic tumor cells.

Although it was shown in this study that dendritic cells fed with apoptotic heat shock protein over-expressing mesothelioma tumor cells induced a cytotoxic T cell response, the method and cells used by Ebstein are not clinically useable. The cells used by Ebstein originate from one cell-line, consequently the antigens provided to the dendritic cells will be relatively limited and would thus seriously reduce it clinical use. This is also exemplified by the fact that the cell-lines used only show a positive result if the tumor cells have been subjected to a prior heat-shock treatment. More importantly, Ebstein uses intact tumor cells which have only been treated with UV-B to induce apoptosis. Since many cells may survive such an UV-B treatment, they are unsuitable for administration to patients. In this regard reference is made to Chalmers A. H. et al., 1976, in Cancer Research and to Salucci S. et al., 2013, in International Journal of Molecular Sciences.

Hence, a need remains for the provision of a safe and reliable method for treating patients suffering from mesothelioma and for the provision of medicaments and pharmaceutical compositions for use in the prevention or treatment of mesothelioma.

