A preclinical therapy model for childhood neuroectodermal tumours

Sammanfattning: Childhood cancers show fundamental differences to most common adult solid tumours in their cancer-causing genetics, cell biology and their local tissue microenvironment. Effective treatments will be attainable when the molecular events that are specific to childhood tumourigenesis are better understood. However, it is in this context critical to consider both species and developmental aspects when looking for the relevant signalling. An influence from the microenvironment on clonal dominance is likely contributing to the disparity between primary and metastatic tumours seen in many patients, as well as inter-tumour heterogeneity between patients with the same tumour type. In this thesis a novel concept is presented for preclinical studies of embryonic tumours in a recently described humanised model. Benign human experimental teratoma was generated in NOD SCID gamma (NSG) mice from diploid bona fide pluripotent stem cells (the PSCT model). An abundant presence of developing early neural components suggests this human embryonic model to be uniquely suited for in vivo transplantation studies of primitive neuroectodermal tumours originating early in life. Following transplantation of tumour cell lines (Paper I), or patient biopsy material (Paper II) the initial microcolonisation showed a tumour type specific tissue tropism in that growth was instigated into specific embryonic tissue compartments, constituting the preferred cellular context/niche supporting initial colonisation and expansion. The phenomenon of microcolonisation reflects in this context an ability of tumour cell clones to comply or adapt to new environments, a feature which has great impact on metastasis and clinical prognosis. Chemotherapy is widely accepted as part of first-line therapy for high-risk paediatric neuroblastoma. In a third study (Paper III) we have examined chemotherapy responsiveness/resistance of neuroblastoma in the PSCT model. Two well-characterized neuroblastoma cell lines were subjected to chemotherapy using the anthracycline drug doxorubicin and we report for the first time in situ studies performed in a human homologous embryonic in vivo microenvironment. Here we combined classical histopathology with high throughput single cell screening for preclinical drug evaluation. Using laser scanning cytometry for screening of chemotherapy-induced changes of nuclear DNA index (sub G1 fraction), in combination with immunohistochemistry and cytology for proliferation, apoptosis and cell death, we could demonstrate dose dependent and diversified responses to single-dose regimens of doxorubicin. Following a recurrent doxorubicin regimen (repeated administration with 48h interval), we observed the presence of an asynchronous response within individual tumour colonies, with cells exhibiting cytotoxic effects or enhanced proliferative index. The results illustrate the feasibility of the approach and are encouraging for clinically relevant studies of patient material regarding intra tumour heterogeneity and asynchronous response to therapy. Furthermore, the findings demonstrate several important advantages using the PSCT model compared to employing conventional preclinical in vivo models and lead us to propose that the use of this human embryonic microenvironment is a well-needed complement for preclinical in vivo studies of primitive neuroectodermal tumours originating early in life.