Retinoblastoma (Rb) is the most common eye cancer in children and an important cause of childhood cancer death worldwide. In developing countries, most children continue to die of Rb because of lack of access to appropriate health screening and care, or because of the reluctance in some cultures to subject the child to the only life-saving measure for advanced disease – removal of the affected eye(s). Even in developed countries where survival rates are high due to the availability of intraarterial and intra-ocular chemotherapy and laser treatments, severe visual impairment is the norm, and eye removal frequently remains necessary. These persistent challenges are due in large part to a lack of targeted therapies for Rb. Indeed, there is not a single therapy for Rb currently in use that was designed to target a molecular determinant specific to this cancer. The initiating event in most Rb tumors is the complete loss of the RB1 gene, the first tumor suppressor to be discovered in 1986. Paradoxically, however, this monumental discovery in cancer biology has not resulted in effective targeted therapies for Rb itself. Further, RB1 mutations alone do not explain the vast diversity of clinical manifestations seen in these pediatric patients.

Our laboratory has developed human based laboratory platforms and animal models that allow us to explore the early developmental and cellular events leading to Rb tumor formation, the molecular signaling pathways that govern progression of nascent Rb tumors, and the clonal evolution of chemoresistant clones within this cancer.

Using a novel human stem cell-derived retinal organoid platform in combination with primary Rb cell lines established in the laboratory, we have discovered a molecular switch that allows early Rb tumors to undergo hypoxic adaptation and grow in the oxygen-depleted environments of the eye. This switch is mediated by the loss of regulation of a protein – ESRRG – which is normally under tight regulation by RB1 and BCOR, the two most mutated genes in these tumors. This discovery was published in 2022 in the journal Science Advances (https://doi.org/10.1126/sciadv.abm8466) and constitutes a viable target for therapeutic intervention to specifically address Rb tumor biology.

Rb tumors are exquisitely adept at undergoing chemotherapy resistance, which constitutes the major treatment failure mode in these pediatric tumors and makes eye removal an urgent necessity. Using our laboratory platforms and novel single-cell genetic barcoding techniques, we have mapped the evolution of Rb cell clones capable of developing resistance to commonly used chemotherapeutic drugs in this disease. In our article published in Cancers in 2022 (https://doi.org/10.3390/cancers14194966), we identify an epigenetic reprogramming event around specific intracellular pathways which confer resistance to chemotherapy in Rb cells. We plan to expand on this discovery on the development of adjuvant treatments to prevent the development of chemoresistance and improving the effectiveness and outcome of current mainstay therapies in this pediatric patient population.

One of the challenges of studying Rb tumors is that they originate while babies are still in their mother’s womb, preventing the study of very early molecular events in the formation of these tumors. Similarly, no animal model has been developed which recapitulates the human manifestations and behavior of this disease, further complicating the development of target therapeutic strategies. The development of our human stem cell derived retinal organoid platform, and the insights it has given us into the cellular and molecular events that contribute to Rb tumor formation during the development of the retina has enabled us to embark on a project to develop the first-ever human relevant mouse model of Retinoblastoma. By targeting specific progenitor cell pools in development, we are developing a mouse model which produces retinoblastoma tumors with the correct morphologic and phenotypic features seen in human Rb tumors. This model will allow for the investigation and intervention in this tumor type at all stages of disease, benefitting all children suffering from the disease.

Adenoid cystic carcinoma (ACC) is a rare but highly lethal cancer that originates in the secretory glands of the head and neck. Despite radical surgeries such as eye and eye socket removal followed by extensive chemotherapy and radiation, long term survival for this cancer type remains dismal, with less than 20% of patient survival at 10 years, and virtually no patients surviving beyond 20 years regardless of treatment. Amongst the challenges in managing and treating this tumor are the lack of molecular information surrounding its initiation and behavior (due to its rarity), its propensity to invade nerves and infiltrate the brain, and its very slow growth rate. In fact, most ACC patients will develop metastatic disease several years out, sometime decades, after primary treatment. This means that an ACC diagnoses results in a life-long sentence of screening and worrying about when and where the disease will reappear. Our laboratory has sought to tackle some of these challenges by studying the molecular biology of the disease, by developing novel molecular assays to detect metastases earlier than currently possible, and by designing advanced biological therapeutic strategies to address metastatic disease.

Using the most advanced sequencing techniques with spatial resolution combined with Mass Spectrometry we have identified and reported on the molecular signatures specific to ACC cells in two separate 2022 publications in the journals Data in Brief and Cancers (https://doi.org/10.3390/cancers15123211). These data allow us to elaborate therapeutic and pharmacological interventions that specifically target ACC biology, hopefully improving our ability to manage these tumors, and the overall survivability of patients diagnosed with ACC.

Spatially resolved genetic sequencing technology has allowed us to discover the mechanism behind ACCs behavior of invading nerves and spreading to the brain. In a study to be published soon, we demonstrate that ACC tumors reactive early events in gland formation and innervation to express the nerve growth factor receptor (NGFR, also known as p75NTR) which allows ACC cells to adhere to nerves and migrate along the nerve path. This discovery can lead to the use of p75NTR blocking agents to prevent brain involvement and improve our ability to remove the tumors entirely.

As stated above, almost all ACC patients will develop metastatic disease during their lifetime. The most common sites for metastases to appear are the liver and lungs, but they can unfortunately be discovered in almost any site of the body. Moreover, these metastases can take up to 20 years to resurface, leaving ACC patients with a life sentence of costly and cumbersome routine medical imaging studies to scan for these metastatic lesions. In the laboratory, we have harnessed our molecular understanding of these tumors in the development of a simple blood test that can sensitively and rapidly inform us of the presence of metastatic disease. We have evidence showing that our blood test can detect metastases even before they can be seen on medical imaging scans (micro-metastases), improving our chances of early intervention and better survival outcomes for these patients. We are now gearing up for the deployment and approval of our blood test to become routine medical care for this patient population.

Regrettably, once metastases are detected in ACC patients, there is little that can be done to improve survival outcomes. There is currently no approved treatment for metastatic ACC, and current treatment strategies involving chemotherapy, radiation, and immunotherapy have all proven ineffective. In our laboratory, we have decided to fight biology with biology by developing a novel type of virus that can infect and kill ACC cells, in a detrimental gene therapy paradigm. We have engineered common harmless viruses to infect only carcinoma type cells, and leveraged a genomic feature specific to ACC cells to drive the expression of a gene that will cause cell death. This technique effectively turns a specific tumor trait into a vulnerability driving its demise. We are currently exploring the use of these oncolytic viruses in preclinical models of ACC and hope to eventually be able treat metastatic ACC disease using this approach.