Monitoring tumor response to radiation therapy with longitudinal and multimodal molecular imaging

Purpose: Metabolic changes induced by radiation therapy (RT) provide a biological measure of tumor response to treatment beyond anatomic changes. In this preclinical study we investigated these changes through longitudinal monitoring of tumor response to varying doses of RT with 18F-Fluorodeoxyglucose Positron Emission Tomography (FDG-PET) and Computed Tomography (CT), as well as bioluminescence imaging (BLI). We sought to establish the metabolic alterations in irradiated tumors at early timepoints following treatment. Materials & Methods: Human HT-29 colon carcinoma cells were inoculated subcutaneously on both sides of the back of nude mice. Tumors were grown for two or three weeks. One tumor per animal was treated with 10 or 20 Gy of RT on day 0. The first ten mice were imaged with both FDG-PET and CT on day -1, 1, 3, 9, and 14. The other 10 mice were imaged with BLI once a week during tumor growth and on the same days as the PET-CT study. These findings were correlated with histology. Results: At the beginning of treatment tumors exhibited low FDG uptake. The volume of the treated tumors (TT) determined by CT images, remained stable or decreased (for dose of 20 Gy), whereas the untreated tumors (UT) continued growing. Surprisingly, 9 days after RT, the FDG-uptake increased in the TT and remained low in the UT. We observed a clear decrease in BLI signal in TTs appearing also 9 days after RT. Conclusion: We have applied preclinical imaging and radiotherapy methods to study tumor response to treatment. The surprising rise in FDG uptake at day 9 after RT could be the result of several mechanisms including in inflammation, that still remains to be determined with immunohistochemistry. This study raises the possibility of FDG PET producing false negatives for early response. The preliminary data reported here will be used to establish future technical and biological approaches towards monitoring cancer therapies

Targeting and Modulating Tumor-Associated Sites with Novel Nanoparticle-Based Immunotherapies

Laura Jeanbart

Cancer is a global disease and a leading cause of death worldwide. While surgery, radiotherapy and chemotherapy comprise the classical tools to eradicate tumors, they do not cure cancer, cannot prevent metastases, lead to side effects, and most importantly they do not instruct the patientʼs immune system to recognize and fight tumor cells. Since the discovery of the immune systemʼs implication in cancer, immunotherapies, which aim at using and educating the immune system to fight and reject cancer, have come to complement classical tumor-debulking methods. A major goal of immunotherapy is to induce or activate effector cytotoxic CD8+ T lymphocytes that can infiltrate and kill the tumor while overcoming immune suppressive mechanisms, which impede their efficacy. Moreover, cancer immunotherapies aim at inducing memory to tumor antigens to prevent relapse or metastases, which traditional therapies cannot do. Dendritic cell (DC) targeting and activation are key for inducing adaptive immunity. Our laboratory has developed synthetic nanoparticles (NPs) capable of draining through lymphatics to target skin-draining lymph nodes (LNs) and being phagocytosed by resident antigen-presenting cells (APCs) upon intradermal injection. We developed a reproducible method to conjugate tumor antigens to NPs and co-delivered them with CpG-conjugated NPs. We found that a NP vaccine composed of a single melanoma-derived peptide (TRP- 2180-188) led to dramatic tumor growth delay in melanoma and was more efficient than a NP vaccine containing several tumor antigens from whole tumor cell lysate (TL). These findings contradicted our hypothesis that immunization with TL might lead to a broader T cell response and hence improved therapeutic outcomes. Additionally, we found that the dose of CpG could modulate the magnitude of the therapeutic outcomes in both single epitope and multiple antigen based NP-vaccines. Considering the efficacy of the developed NP vaccines, we used them as a tool to ask a fundamental and extremely relevant question regarding the influence of vaccine administration site on clinical efficacy: is it therapeutically more beneficial to target a tumor- draining lymph node (tdLN), which is immune suppressed but tumor antigen-primed, or a non-tdLN, which is neither suppressed nor primed by the tumor? We found that targeting the tdLN with NP vaccines led to significantly enhanced therapeutic outcomes in two different tumor models over targeting a non-tdLN, and that efficacy relied on NP conjugation of antigen and adjuvant. This suggested that antigen drainage from the tumor might be available to APCs in the tdLN and we further hypothesized that therapies that lead to immunogenic tumor cell death might synergize with a tdLN-targeted adjuvant approach. While cytotoxic modalities, such as chemotherapy and radiotherapy, have traditionally been used for their ability to kill tumor cells, tumor cells shed antigens and debris upon cell death that drain to the tdLN. Consistent with our hypothesis, targeting an adjuvant to the tdLN enhanced therapeutic benefits of chemo- and radio- therapy, while targeting a non-tdLN did not. We hypothesized that targeting the tdLN with NP vaccines might switch the tdLN microenvironment from a suppressive to an immunogenic one. This suggested that direct targeting of active suppressive mechanisms in the tumor might further improve immunotherapy. We thus engineered a nano-sized micellar carrier loaded with 6-thioguanine (MC-6TG), a cytotoxic drug used in leukemia, and explored its use in targeting T cell- impairing myeloid-derived suppressor cells (MDSCs) in order to enhance the efficacy of therapeutic vaccines. MC-6TG entirely eradicated both Ly6c+ monocytic and Ly6g+ granulocytic MDSCs for up to 7 d in the blood, tumor and tdLN of tumor-bearing mice. Since MC-6TG also targeted certain subsets of macrophages and DCs, depleting MDSCs did not have an impact on the efficacy of a NP vaccine, which relies on APCs for efficacy. However, MC-6TG synergized with adoptively transferred CD8+ T cells, significantly delaying tumor growth and enhancing survival in a murine model of implantable melanoma. Overall, this thesis describes the design and implementation of a novel approach to immunotherapy: we engineered NP-based therapeutic vaccines, optimized their delivery route to enhance efficacy, and simultaneously tackled immune suppressive cells in the tumor microenvironment to allow optimal therapeutic outcomes. The technologies and methods developed in this work are particularly relevant to clinical oncology and have the potential to benefit cancer patients by improving cancer therapy efficacy.

EPFL2013

Development of a nano-adjuvant cancer vaccine and its evaluation for melanoma in combination with radiotherapy

Sylvie Hauert

Cancer is the second leading cause of death in the world and melanoma is the deadliest type of skin cancer. Although surgery, radiotherapy and chemotherapy are still standard treatments, the discovery of the role played by the immune system in cancer has allowed the development of new treatments, called immunotherapies. The goal of these treatments is to help the immune system eradicate cancerous cells, for example by increasing the anti-tumor response of the patient. In this particular case, the treatment has to activate cytotoxic CD8+ and CD4+ helper T cells against the cancer cells. This can be done by using subunit vaccines composed of tumor antigens, adjuvants and a delivery system, which will be internalized by dendritic cells (DCs) and induce its activation followed by the migration to the lymph nodes where it will educate T cells against specific antigens. Our laboratory has developed two different delivery systems, nanoparticles (NPs) and polymersomes (PSs) that can enhance the vaccineâs immune response by efficiently entering the lymphatic system, due to their size, and drain to the lymph node. Once they reach that location, they will be internalized by DCs and induce an anti-tumor response. In this thesis, we used these two nanocarriers to create a vaccine composed of different nano-adjuvants capable of activating several toll-like receptors simultaneously and test their efficacy at inducing an anti-tumor response in melanoma. We have developed and characterized the different nano-adjuvants, PS-MPLA, PS-CL075 and NP-CpG-B, and demonstrated, in vitro, synergistic effects on DC activation when PS-MPLA + PS-CL075 and PS-MPLA + NP-CpG-B were combined. In vivo, we showed the enhanced immune response with the different nano-adjuvant combinations, but only PS-MPLA + NP-CpG-B was able to delay tumor growth in melanoma. We then tested this vaccine in combination with radiotherapy, mimicking a clinical situation. We first demonstrated an enhanced immune response when combining a high irradiation dose, 15 Gy, with the nano-adjuvant vaccine draining the tumor-draining lymph node. Moreover, the vaccination of NP-CpG-B + PS-MPLA and radiotherapy increased mice survival by 17 days compared to mice only receiving radiotherapy. â Overall, we have demonstrated that combining different nano-adjuvants, targeting the tumor-draining lymph node, with radiotherapy enhanced significantly the survival in a melanoma model. This approach is particularly promising since ionizing radiation is a standard treatment in cancer and our vaccine can be applied to a multitude of cancer since it does not contain any antigen and its composition can easily be modified.

EPFL2018

Targeting and Modulating Tumor-Associated Sites with Novel Nanoparticle-Based Immunotherapies

Laura Jeanbart

EPFL2013

Development of a nano-adjuvant cancer vaccine and its evaluation for melanoma in combination with radiotherapy

Sylvie Hauert

EPFL2018