Childhood Cancer Research, Incidence, Survival

Childhood cancer is the second leading cause of death in children under 20 years of age in the United States.  Although mortality rates for childhood cancer have declined by 67% over the course of the last few decades, an estimated 10,380 new cases and 1,250 cancer deaths are expected to occur among children in the USA in 2015 alone. These days, children in the USA benefit from substantial progress in therapeutics and clinical trials, leading to an overall 80% survival rate for the first 5 years after diagnosis; many also suffer, however, from  short- and long-term side effects of their treatments. A longitudinal cohort study of child cancer survivors in the United States demonstrated, for example, that over 75% developed a chronic health condition that was linked to the treatment itself, and half of these cases became life threatening.

Recurrence presents the central challenge in the field of childhood cancer. The Childhood Cancer Survivor Study (CCSS) indicates that recurrence is the leading cause of death from 5 to 20 years following diagnosis. In this cohort study, monitoring 14,359 patients under 21 years old diagnosed from 1970 to 1986, it was shown that 10.89% had a recurrence within the first 5 years and 4.4-6.3% developed cancer within 5 to 20 years after the primary diagnosis (late relapse). Addressing the issue of recurrence is seen as especially pivotal considering that only 49% of the patients with recurrence survive, compared to 92% of the patients with no recurrence. Among all pediatric cancers, Ewing sarcoma and astrocytoma have the highest rates of recurrence, suggesting the way in which we are very much in need of a better understanding of heterogeneous cancer cell populations at the single cell level.  

Childhood sarcomas are rare, malignant tumors that affect bone and soft tissue and constitute approximately 10% of all types of cancer found in children. One of the challenges associated with the treatment of childhood sarcomas is the fact that they are comprised of heterogeneous cell populations that are difficult to classify. It has even been suggested that this is one of the principal reasons why therapy has not yet been more successful, especially at preventing recurrence in sarcomas. Thus, it is essential to understand the phenotypic and genetic characteristics of sarcomas so as to be able to target diverse sub-populations of cancer cells in such a way as to prevent recurrence.

Sarcomagenisis is a process in which different sarcomas form through genetic and gene expression dis-regulations and sarcomas fall into one of two major classes: (a) sarcomas with genetic alterations and usually simple karyotypes, including fusion genes that result from reciprocal translocations and specific point mutations and (b) sarcomas with nonspecific genetic alterations and complex unbalanced karyotypes. This study is focused on ES which falls into the first category. ES is the second most aggressive bone cancer found in children and young adults after osteosarcoma. It has been characterized genetically by a reciprocal translocation between EWS-ETS resulting in a functional protein, EWS-FLI1, an onco-protein that exists in ~85-90% of all ES cases. Tumor initiation in ES is known to involve EWS-FLI1 expression in a supportive microenvironment, transcriptionally dis-regulating more than 1000 different genes involved in differentiation, proliferation, survival and apoptosis. But full malignant transformation is an independent event downstream of EWS-FLI1 formation that requires both molecular and cellular changes leading to clonal selection and expansion through evolutionary processes over time that generally result in secondary changes such as epigenetic deregulations (instability), altered RNA splicing, aberrant regulation of mi-RNAs, enhanced proliferation, avoidance of cell senescence, altered cytoskeleton and cell adhesion, inhibition of cell death, and abnormal maintenance of embryonic development programs in response to various stimuli such as stresses, growth factors, and environmental changes.

ES comprise approximately 1% of all pediatric cancers and despite an overall 70% advance in controlling the disease, this form of cancer has the highest rate of relapse among all (childhood) cancers and metastatic ES is heavily associated with poor therapy outcome (<20%). It has been more than 20 years since EWS-FLI1 has been identified as the master regulator driving ES in children and young adults and scientists in the area of pediatric cancer research have at least been successful in significantly improving our understanding of the main signaling pathways disrupted in ES; and studies have identified direct and indirect interaction of molecular components of EWS-FLI. But, the fact remains that this form of cancer in children has the highest level of recurrence and the overall therapeutic outcome has remained un-changed. Study of the regulation of oncogenic hubs downstream of EWS– FLI1 fusion indicates an ability to drive cancer through various mechanisms besides transcriptional activation, such as epigenetics, bi-switching cell-cycle regulation by E2F3. ES has a relatively stable genetic background and this suggest the effect of epigenetic mechanisms leading to oncogenic addiction in ES. Supportive evidence suggests that Vorinostat-mediated HDAC inhibition disrupts the growth of Ewing sarcoma cells in tissue culture, and the ability of those cells to form colonies in soft-agar assays. A study conducted by S. Lessnick has shown EWS/FLI1 co-immunoprecipitats with HDAC2 and HDAC3, but not with HDAC1. A polycomb gene, BMI-1 over expression, has been associated with anchorage independent growth which indirectly modulates cell adhesion pathways and cell proliferation. In ES there is a correlation between a low therapy outcome and the number of mutations. In biopsies, the most common kind of mutation identified was a C to T transition, which was linked to the common event of deamination of methylated cytosines.(8) Cancer cell dormancy, drug resistance and tumor heterogeneity are among the most challenging areas of ongoing investigation in the struggle to overcome the tenacity of these cells, which could lead to better therapeutic results and less recurrence.

Heterogeneous sub-populations of ES have to be understood in light of the way in which cancer cells with micro-metastatic properties, cell cycle heterogeneity, and epigenetic transition states are likely to result in the survival of at least some cancer cells, transitioning through different states of evolution and promoting various functions in the tumor. Different molecular phenotypes have been reported at different stages of cancer progression in response to various niches and stressors, such as cytotoxic agents. The central goal is to identify key regulators that serve to switch cancer states, where cells remain quiescent, with low metabolic and proliferating activity, which may have similar characteristics to the response of cells to treatment. In this way we could stratify subpopulations and identify drivers of transition states to sensitize our current therapies in accordance with the identification of novel targets for new therapeutics. This study will help us to better understand the evolutionary mechanisms leading to signaling paths and molecular modifications which ES cells acquire to adapt to new environmental needs, including metastasizing and dormancy in response to stress.

Recent studies suggest that a solid tumor contains bystander or quiescent cells that help to maintain tumor function despite external perturbations. This adds to speculation that these are cells have an advantage for survival and the ability to multi-task according to their environmental needs, leading to heterogeneous populations, disease progression, drug resistance and relapse. Most patients show drug resistance in the face of disease recurrence and single cell analysis is the only strategy that shows promise for understanding important and novel predictive factors for chemo response and relapse. Over expression of Her2/ERBB2 has been correlated with drug resistance (cisplatin, doxorubicin, and ifosfamid) in OS and ES. One of the mechanisms proposed turns on how Her2 confers to low response rate by mediating reduced cell proliferation. Over expression of EWS-FLI1 has also been associated with resistance to chemotherapy. MiRNAs regulating the cell cycle and RB leading to cell cycle arrest have been shown to confer chemoresistance.

I hypothesize that in Ewing Sarcoma EWS-FLI1 contributes to cell cycle heterogeneity through a developmental-proliferative network, and that understanding gene expression patterns associated with dormancy at the single cell level may elucidate novel genetic and epigenetic, evolutionary alterations leading to cancer survival and fitness in response to various stressors and cytotoxic agents. Pediatric cancer patients are at a high risk for side effects and recurrence as a result of limited information about their cancer during diagnosis and treatment. This study aims to address the challenges faced by our current therapeutic initiatives in the face of ES and ways to prevent cancer recurrence to improve children’s post-treatment lives. Lower proliferating cells with stemness properties are among the cells least affected by chemotherapies and are accepted to be the major cause of relapse. This research attempts to understand novel predictive regulators leading to cell-cycle heterogeneity and to better understand the underlying algorithm and mechanisms (network) of tumor sub-evolution at the level of the single cell. This could serve to enhance the understanding of health care providers of the disease and our ability to treat patients in a personalized manner, laboring to overcome challenges posed by heterogeneous functional properties, such as cell-cycle heterogeneity, leading to cancer relapse and drug resistance in ES.

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