Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Science

Program

Microbiology and Immunology

Supervisor

Maleki, Saman

2nd Supervisor

Koropatnick, James

Joint Supervisor

Abstract

Immunotherapy has improved patient outcomes in advanced or metastatic settings across a number of cancers. Patients with tumours deficient in the DNA mismatch repair (DNA-MMR) pathway often show high response rates to immune checkpoint inhibitors (ICIs) with a rise in immune surveillance. However, little is known about the immune sensitization effects of inducing DNA- MMR-deficiency in low tumour mutational burden (TMB) cancers, such as ICI refractory neuroblastoma. In addition, the dynamic T-cell profile that results from such a DNA-MMR inactivation, and whether this may confer a therapeutic benefit, is poorly understood. Here we used CRISPR/CAS9 genome editing technology to knock out (KO) MLH1, a crucial molecule in the DNA-MMR pathway, in mouse neuroblastoma (neuro-2a) cells – a low TMB pediatric cancer refractory to ICIs – to induce MMR deficiency. To analyze tumour growth inhibition in response to ICIs and immunophenotype the tumour-infiltrating lymphocytes (TILs), tumours with intact or induced MMR deficiency were injected subcutaneously into immunocompetent mice. Tumour growth were measured after treatment with anti-PD1 antibodies and TILs were analyzed for activation, exhaustion, and effector markers, allowing for an in-depth flow cytometric analysis of T-cell subsets in these tumours. Our study shows that inducing MMR-deficiency induces a robust anti-tumour response in a low TMB cancer. Moreover, this sensitization was accompanied by specific phenotypic changes of T-cells in response to anti-PD1 therapy in the tumour.

Summary for Lay Audience

Immunotherapy is a new form of cancer treatment that utilizes the immune system to recognize and kill cancer cells. In this thesis, I focus on immune checkpoint inhibitors (ICIs), which is a form of cancer immunotherapy that binds to “immune checkpoints” found on immune cells. These immune checkpoints typically serve to tamper down the immune response in order to prevent a harmful overactivation of the immune system. ICIs therefore bind to immune checkpoint molecules, blocking them from exerting their function, which then allows for immune cells to become re-invigorated and target cancer cells. Over the past few years, clinical trials have shown that a number of patients that are treated with immunotherapy have a long-term response to treatment. Moreover, responders to ICIs often have a very high number of mutations in their tumours. A subset of cancer patients that show a favorable response to immune checkpoint inhibitors are cancer patients that have a deficiency in their ability to repair DNA mistakes, which results in a large number of mutations in their tumour. Unfortunately, many cancer patients with a low number of tumour mutations do not respond to ICIs. In this thesis, I studied the effect of inducing a deficiency of DNA repair in a tumour with a low number of mutations and hypothesized that this would result in an increase in mutations in the tumour and a better response to ICIs.

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