Cell Fate Reprogramming in the Era of Cancer Immunotherapy

Abstract

Advances in understanding how cancer cells interact with the immune system allowed the development of immunotherapeutic strategies, harnessing patients' immune system to fight cancer. Dendritic cell-based vaccines are being explored to reactivate anti-tumor adaptive immunity. Immune checkpoint inhibitors and chimeric antigen receptor T-cells (CAR T) were however the main approaches that catapulted the therapeutic success of immunotherapy. Despite their success across a broad range of human cancers, many challenges remain for basic understanding and clinical progress as only a minority of patients benefit from immunotherapy. In addition, cellular immunotherapies face important limitations imposed by the availability and quality of immune cells isolated from donors. Cell fate reprogramming is offering interesting alternatives to meet these challenges. Induced pluripotent stem cell (iPSC) technology not only enables studying immune cell specification but also serves as a platform for the differentiation of a myriad of clinically useful immune cells including T-cells, NK cells, or monocytes at scale. Moreover, the utilization of iPSCs allows introduction of genetic modifications and generation of T/NK cells with enhanced anti-tumor properties. Immune cells, such as macrophages and dendritic cells, can also be generated by direct cellular reprogramming employing lineage-specific master regulators bypassing the pluripotent stage. Thus, the cellular reprogramming toolbox is now providing the means to address the potential of patient-tailored immune cell types for cancer immunotherapy. In parallel, development of viral vectors for gene delivery has opened the door for in vivo reprogramming in regenerative medicine, an elegant strategy circumventing the current limitations of in vitro cell manipulation. An analogous paradigm has been recently developed in cancer immunotherapy by the generation of CAR T-cells in vivo. These new ideas on endogenous reprogramming, cross-fertilized from the fields of regenerative medicine and gene therapy, are opening exciting avenues for direct modulation of immune or tumor cells in situ, widening our strategies to remove cancer immunotherapy roadblocks. Here, we review current strategies for cancer immunotherapy, summarize technologies for generation of immune cells by cell fate reprogramming as well as highlight the future potential of inducing these unique cell identities in vivo, providing new and exciting tools for the fast-paced field of cancer immunotherapy.

Keywords: CAR-T; antigen presentation; cancer immunotherapy; cancer vaccine; cellular reprogramming; dendritic cell; transcription factor; tumor immunology.

Figure 1

Strategies for cancer immunotherapy. Overview of the strategies for immunotherapy translated to clinical practice. Approved strategies include immune checkpoint inhibitors, adoptive T-cell therapy, cancer vaccines, and oncolytic viruses. Checkpoint inhibitors (top left panel) are antibodies targeting key negative regulators of T-cell activation. These include receptors expressed in T-cells targeted with anti-cytotoxic T-lymphocyte associated protein 4 (CTLA-4) (ipilimumab), anti-programmed cell death protein 1 (PD-1) (nivolumab, pembrolizumab) and receptors expressed in tumor cells targeted with anti-programmed death-ligand 1 (PD-L1) (atezolizumab). Adoptive T-cell transfer (top right panel) are autologous cellular therapies currently targeting hematological tumors. Approved Chimeric Antigen Receptor (CAR) T-cells target CD19 (axicabtagene ciloleucel, tisagenlecleucel, brexucabtagene autoleucel, and lisocabtagene maraleucel) or B-cell maturation antigen (BCMA) (idecabtagene vicleucel). Additionally, tumor infiltrated lymphocytes (TILs) expanded ex vivo and natural killer (NK) cells are currently in clinical trials. Cancer vaccines (bottom right panel) exploit the antigen presentation capacity of dendritic cells (DC) to trigger polyclonal T-cell responses to tumor-associated antigens (TAA). These include irradiated tumor cells expressing granulocyte-macrophage colony-stimulating factor (GM-CSF, GVAX), DCs loaded with tumor antigens (sipuleucel-T), or the delivery of mRNA in vivo encoding multiple TAAs. Oncolytic viruses (bottom left panel) target cancer cells own vulnerability to viral infections while inducing immunogenic cell death. Talimogene laherparepvec (T-VEC) is an approved intra-tumoral therapy consisting of a type-1 herpes virus expressing GM-CSF.

Figure 2

Cell reprogramming as a source of immune cells. Cellular reprogramming provides an opportunity to generate tailored immune cells for immunotherapy. Somatic cells can be reprogrammed to induced pluripotent stem cells (iPSCs) with the transcription factors OCT4, SOX2, KLF4, and c-MYC. iPSCs are then differentiated to hematopoietic progenitor cells (HPCs) with the cytokines stem cell factor (SCF), vascular endothelial growth factor (VEGF), and bone morphogenetic protein 4 (BMP4). Protocols for the differentiation of HPCs into myeloid and lymphoid lineages involve exposing cells to complex cytokine cocktails and/or feeder layers including MS-5, OP9-DL1, or artificial thymic organoids (ATO) supporting lymphopoiesis. Due to the self-renewal capacity of pluripotent stem cells, iPSC reprogramming (top panel) can generate immune cells on a large scale and is suitable for ex vivo genome editing. Cells undergo rejuvenation and erasure of epigenetic marks associated with aging or exhaustion. However, iPSC differentiation into mature immune cells is challenging (dashed lines) and leads to the generation of heterogeneous populations. Direct reprogramming (also known as transdifferentiation, bottom panel) refers to a change in cell fate mediated by lineage transcription factors that, unlike in iPSC reprogramming, does not involve pluripotent intermediates. The direct reprogramming of fibroblasts to macrophages (MØ) or conventional dendritic cells type 1 (cDC1) induce exclusively the target immune cell fate. Direct reprogramming is a faster and more efficient process. It can be elicited both in vitro and within the target tissue, thus having potential for in vivo reprogramming. It also reduces tumorigenic risks from contaminating stem cells. Moreover, direct reprogramming may retain epigenetic aging hallmarks, making it more suitable for modeling aging-related disease but more difficult to achieve complete epigenetic reprogramming. Transcription factors can also be applied to accelerate and direct the differentiation of iPSCs (also known as forward reprogramming, top panel). FLT3L, Fms-like tyrosine kinase 3 ligand; TPO, Thrombopoietin; IL, interleukin; G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; cDC2, conventional dendritic cells type 2 DCs; pDCs, plasmacytoid DCs; NK, natural killer; ILCs, innate lymphoid cells.

Figure 3

Disrupting cancer cell tumorigenicity with cellular reprogramming. Several tumor cell types have been described to be amenable to cell reprogramming towards induced pluripotent stem cells (iPSCs). Melanoma, glioblastoma, sarcoma, gastrointestinal (GI), as well as breast, bladder, and liver cancers, acute myeloid leukemia (AML), and chronic myelogenous leukemia (CML) have all been successfully reprogrammed towards iPSC with OCT4, SOX2, KLF4, and c-MYC. Following reprogramming to iPSCs, cancer cells can be differentiated into benignity by either differentiation to their original cell type or by converting them into a different lineage (highlighted by the colored lines). Similarly, in vitro direct reprogramming has been demonstrated in glioma, B-cell acute leukemia (B-ALL), Burkitt lymphoma (BL), squamous cell carcinoma (SCC), and hepatic cellular carcinoma (HCC). Lineage-specific factors rewrite the cancerous epigenetic and transcriptional program into mature terminally differentiated cells (glioma to neuron; SCC to melanocyte; HCC to hepatocyte), or by modifying cell fate and therefore disrupting the tumorigenic drive [B-leukemias to macrophage (MØ)].

Figure 4

In vivo reprogramming strategies for regenerative medicine and immunotherapy. Overview of parallel in vivo approaches for cell fate reprogramming and immune cell engineering. In vivo cell fate reprogramming offers an approach to restore damaged cell types and tissue function (top panel). The concept has been demonstrated in multiple tissues and cellular conversions. Examples include the reprogramming of Müller glia cells to photoreceptors leading to restoration of vision in congenitally blind mice; reprogramming of astrocytes and glial cells to neurons; the conversion of fibrotic tissue after myocardial infarction to functional cardiomyocytes and the reprogramming of pancreatic acinar/exocrine cells into insulin-producing β-cells. These strategies mainly rely on the delivery of lineage-specific factors through viral vectors. In parallel, in vivo engineering of immune cells has been developed allowing the generation of genetically modified immune cells, often designated immune reprogramming (bottom panel). Recent examples in cancer immunotherapy include the conversion from macrophage type 2 (M2) to macrophage type 1 (M1), manipulation of cancer cells to express immunogenic circuitry and cytokines, and the in vivo delivery of chimeric antigen receptors (CAR) to circulating T-cells. Delivery strategies are diverse including lipid nanoparticles coupled with DNA or replicative RNA, and viral vectors (lentivirus, AAV, retrovirus, adenovirus, Sendai virus). In vivo approaches in regenerative medicine and immunotherapy converge towards the creation of off-the-shelf, immunocompatible therapies that harbor low tumorigenic risks. Simplified clinical translation and manufacturing are balanced by challenges in delivery and the control of cell-type specificity and numbers. AAV, adeno-associated virus.