Radiation Therapy and the In Situ Vaccination Approach

During the past century, from the advent of preclinical modeling to the establishment of clinical trials, the hypothesis that host defenses regulate tumor growth (posited and refined by leaders in the field of cancer immunity) has become accepted as a scientific pillar in oncology. Since the turn of the millennium, a search has been under way for the best therapeutic approach to reprogram the immune system to recognize tumor cells that have undergone “immune escape.” This quest has led some to question conventional scientific views of tumor cell kill, including the role of host immunity in patients treated with radiation therapy. In the last two decades, evidence has accumulated that radiation therapy can effectively convert a potentially lethal cancer into an in situ personalized vaccine. Herein, we review the underlying mechanisms and maneuvers responsible for in situ vaccine production. During the past century, from the advent of preclinical modeling to the establishment of clinical trials, the hypothesis that host defenses regulate tumor growth (posited and refined by leaders in the field of cancer immunity) has become accepted as a scientific pillar in oncology. Since the turn of the millennium, a search has been under way for the best therapeutic approach to reprogram the immune system to recognize tumor cells that have undergone “immune escape.” This quest has led some to question conventional scientific views of tumor cell kill, including the role of host immunity in patients treated with radiation therapy. In the last two decades, evidence has accumulated that radiation therapy can effectively convert a potentially lethal cancer into an in situ personalized vaccine. Herein, we review the underlying mechanisms and maneuvers responsible for in situ vaccine production. A year after his 1908 Nobel prize in recognition for his work on immunity, Paul Ehrlich postulated that host defenses could prevent the development of tumors from neoplastic cells.1Ribatti D. The concept of immune surveillance against tumors. The first theories.Oncotarget. 2017; 8: 7175-7180Crossref PubMed Scopus (115) Google Scholar,2Ehrlich P. Über den jetzigen Stand der Karzinomforschung.Ned Tijdschr Geneeskd. 1909; 5: 273-290Google Scholar However, at the time no experimental models were available to test this hypothesis. It was not until the advent of preclinical models of immunity, when Lewis Thomas and F. Macfarlane Burnet (1960 Nobel laureate for the discovery of acquired immunologic tolerance) further advanced the field of cancer immunity with the provocative hypothesis that tumor cell neoantigens could induce an immunologic response against neoplastic cells. This model proposed that the immune system acts in a coordinated fashion as a homeostatic check to stop the outgrowth of malignant cells. This process was termed “immune surveillance,” whereby circulating and tissue resident immune cells were responsible for the deletion of newly transformed cells.3Burnet F.M. The concept of immunological surveillance.Prog Exp Tumor Res. 1970; 13: 1-27Crossref PubMed Google Scholar The immune surveillance hypothesis has since been regarded as the intellectual underpinning for the field of cancer immunology.3Burnet F.M. The concept of immunological surveillance.Prog Exp Tumor Res. 1970; 13: 1-27Crossref PubMed Google Scholar At the turn of the millennium, Gavin Dunn and Robert Schreiber further refined the immune surveillance hypothesis to explain how transformed cells subvert immune detection and form tumors.4Dunn G.P. Bruce A.T. Ikeda H. et al.Cancer immunoediting: From immunosurveillance to tumor escape.Nat Immunol. 2002; 3: 991-998Crossref PubMed Scopus (3182) Google Scholar They introduced the concept of “cancer immunoediting,” whereby tumor outgrowth occurs through 3 distinct phases: (1) elimination: tumor cell kill and elimination by natural killer (NK) cells and CD4+ and CD8+ T cells; (2) equilibrium: a state of equilibrium between immune surveillance and tumor outgrowth; and (3) immune escape: the immune system’s failure to recognize and/or eliminate neoplastic cells, leading to tumor outgrowth and clinically detectable tumors. During immune escape, tumor variants either downregulate or lose the expression of tumor antigens (resulting in antigenic drift, a process that occurs through continued deletion of cancer cells expressing T-cell targets), upregulate immune resistance or prosurvival genes, and establish an immunosuppressive microenvironment. Additionally, several peripheral and central immune tolerance mechanisms, such as the proliferation and/or activation of immunosuppressive regulatory T cells (Tregs) and increased expression of immune-inhibitory checkpoints on effector T cells (eg, program death receptor 1 [PD-1], cytotoxic T lymphocyte antigen 4 [CTLA-4], T cell immunoglobulin and mucin domain-containing protein 3, and V-domain Ig suppressor of T cell activation) serve to facilitate immune escape. Through the process of immune surveillance, the immune system unwittingly selects for malignant clones with decreased antigenicity and/or immunogenicity and consequently can no longer suppress their outgrowth. Recently, scientific efforts have focused on reprogramming the immune system to recognize transformed clones that escape immune surveillance. The “cancer immunity cycle” (as described by Daniel Chen and Ira Mellman) serves as an immuno-oncologic framework for the clinical development of cancer immunotherapies.5Chen D.S. Mellman I. Oncology meets immunology: The cancer-immunity cycle.Immunity. 2013; 39: 1-10Abstract Full Text Full Text PDF PubMed Scopus (2572) Google Scholar The proposed framework describes a stepwise progression of immunologic events crucial for the development of an effective anticancer immune response: (1) tumor-cell death with subsequent cancer neoantigen release and uptake by dendritic cells (DCs) for antigen processing; (2) DC neopeptide presentation on major histocompatibility class 1 (MHC-I) and II machinery to naïve T cells, (3) priming, activation, and clonal expansion of T cells recognizing tumor-specific neoantigens, (4) T-cell trafficking into the tumor microenvironment (TME), (5) T-cell tumor infiltration, (6) T-cell tumor recognition (the development of an immune synapse consisting of the T-cell receptor and its cognate antigen bound to MHC-I), and (7) T-cell tumor-cell kill (whereby additional tumor-associated antigens are released resulting in antigen spread/antigen cascade with expansion of effector T cell clones against additional antigenic targets). We and others have shown that radiation therapy is able to perturb each step of the cancer immunity cycle.6Ngwa W. Irabor O.C. Schoenfeld J.D. et al.Using immunotherapy to boost the abscopal effect.Nat Rev Cancer. 2018; 18: 313-322Crossref PubMed Scopus (375) Google Scholar As treatment techniques have evolved to deliver higher doses more safely and precisely, our knowledge of the immunologic effects of radiation has also expanded. Radiation therapy has proven to be a powerful tool to help churn the cancer immunity cycle.7Demaria S. Formenti S.C. The abscopal effect 67 years later: From a side story to center stage.Br J Radiol. 2020; 9320200042Crossref Scopus (22) Google Scholar Herein, we review how an irradiated tumor can be converted effectively from a potentially lethal cancer into an in situ personalized vaccine. Cellular stressors such as intracellular pathogens, conventional chemotherapies, targeted anticancer agents, and physical interventions (including ionizing radiation) expose cryptic tumor-associated antigens within an intact tumor to the adaptive immunologic machinery for subsequent recognition and disposal. As defined by the Nomenclature Committee on Cell Death, immunogenic cell death (ICD) is a form of regulated cell death of tumor cells that drives an adaptive immune response in immunocompetent hosts and is reliant on the antigenicity and adjuvanticity of dying tumor cells. The antigenic determinants of dying tumor cells include neoepitopes (derived from latent endogenous retroviruses, post-translational modifications, translation of cryptic antigenic peptides, or expression of mutated genes or tumor-associated antigens) that are highly immunogenic. The adjuvant components of dying tumor cells provide a spatiotemporal coordinated release and exposure of danger signals that effectively recruit antigen-presenting cells (APCs) (eg, adenosine triphosphate, an activator of the DC P2XR7 purinergic receptor that triggers DC inflammasome activation, secretion of IL-1beta, and subsequent priming of interferon gamma-producing CD8+ T cells) and spatially guide the interaction of APCs and dying cells (eg, ANXA1); APC phagocytosis of dying cells or their corpses (eg, CALR, ERp57, HSP70, HSP90, and F-actin); APC maturation and cross-presentation (eg, adenosine triphosphate, HMGB1 [a DNA-binding protein and TLR4-mediated DC activator], type I interferon [IFN], and mitochondrial transcription factor A); and T cells (eg, CCL2, CXCL1, and CXCL9/10, CXCL16).8Galluzzi L.V. Warren S. Consensus guidelines for the definition, detection and interpretation of immunogenic cell death.J Immunother Cancer. 2020; 8e000337Crossref PubMed Scopus (136) Google Scholar Chemotherapy, radiation therapy, and targeted therapies have been shown to cause tumor cell death in a fashion that affects immunity, including T-cell activity.9Zitvogel L. Galluzzi L. Smyth M.J. et al.Mechanism of action of conventional and targeted anticancer therapies: Reinstating immunosurveillance.Immunity. 2013; 39: 74-88Abstract Full Text Full Text PDF PubMed Scopus (554) Google Scholar,10Yamazaki T. Buqué A. Rybstein M. et al.Methods to detect immunogenic cell death in vivo.Methods Mol Biol. 2020; 2055: 433-452Crossref Scopus (3) Google Scholar Radiation therapy in particular has been shown to induce ICD and promote key components for the development of ICD, an initiating step in the establishment of an in situ vaccine. Although radiation therapy can help convert a tumor into an in situ vaccine, as previously alluded to, it generally requires a partner to efficiently achieve this end. The rare abscopal cases of radiation therapy have demonstrated the capacity of focal radiation to a metastatic site to elicit immune rejections of metastases outside the radiation fields. The exceptionality of these cases is now well explained by the advances in cancer immunology.11Abuodeh Y.P. Venkat Kim S. Systematic review of case reports on the abscopal effect.Curr Probl Cancer. 2016; 40: 25-37Crossref PubMed Scopus (213) Google Scholar By the time that a metastasis has presented clinically, a complex process of immune tolerance and suppression already exists with multiple pathways in place to maintain immune escape.12Vesely M.D. Kershaw M.H. Schreiber R.D. et al.Natural innate and adaptive immunity to cancer.Annu Rev Immunol. 2011; 29: 235-271Crossref PubMed Scopus (1241) Google Scholar Although the abscopal effect of radiation has been of scientific interest, it only represents 1 component of a successful immunotherapy strategy in established tumors.7Demaria S. Formenti S.C. The abscopal effect 67 years later: From a side story to center stage.Br J Radiol. 2020; 9320200042Crossref Scopus (22) Google Scholar A key step in the immune rejection of cancer is the recruitment, activation, and maturation of dendritic cells, a subset of immune cells responsible for cross-presentation of antigens to T-cells. Radiation plays multiple roles in this phase of cancer immune rejection. Radiation releases damage-associated molecular pattern molecules that activate DCs and generate costimulatory signals to activate naïve T-cells.13Apetoh L. Ghiringhelli F. Tesniere A. et al.Toll-like receptor 4-dependent contribution of the immune system to anticancer chemotherapy and radiotherapy.Nat Med. 2007; 13: 1050-1059Crossref PubMed Scopus (2030) Google Scholar Critical to the immunogenicity of radiation therapy is the formation of DNA fragments from the DNA-damage response (DDR) of radiation that originate in the nucleus and transfer to the cytoplasm. In the cytoplasm these DNA fragments lead to activation of cGAS/STING pathways, which constitute the canonical defense against viral infections and result in the production of type I IFNs (IFN-I) and transcription of IFN-stimulated genes including cytokines and chemokines that recruit innate and adaptive immune cells to the tumor.14Kondo T. Kobayashi J. Saitoh T. et al.DNA damage sensor MRE11 recognizes cytosolic double-stranded DNA and induces type I interferon by regulating STING trafficking.Proc Natl Acad Sci U S A. 2013; 110: 2969-2974Crossref PubMed Scopus (211) Google Scholar, 15Vanpouille-Box C. Alard A. Aryankalayil M.J. et al.DNA exonuclease Trex1 regulates radiotherapy-induced tumour immunogenicity.Nat Commun. 2017; 8: 15618Crossref PubMed Scopus (645) Google Scholar, 16Fuertes M.B. Kacha A.K. Kline J. et al.Host type I IFN signals are required for antitumor CD8+ T cell responses through CD8{alpha}+ dendritic cells.J Exp Med. 2011; 208: 2005-2016Crossref PubMed Scopus (646) Google Scholar, 17Lugade A.A. Sorensen E.W. Gerber S.A. et al.Radiation-induced IFN-gamma production within the tumor microenvironment influences antitumor immunity.J Immunol. 2008; 180: 3132-3139Crossref PubMed Scopus (297) Google Scholar, 18Burnette B.C. Liang H. Lee Y. et al.The efficacy of radiotherapy relies upon induction of type i interferon-dependent innate and adaptive immunity.Cancer Res. 2011; 71: 2488-2496Crossref PubMed Scopus (480) Google Scholar, 19Deng L. Liang H. Xu M. et al.STING-dependent cytosolic DNA sensing promotes radiation-induced type I interferon-dependent antitumor immunity in immunogenic tumors.Immunity. 2014; 41: 843-852Abstract Full Text Full Text PDF PubMed Scopus (827) Google Scholar, 20Woo S.R. Fuertes M.B. Corrales L. et al.STING-dependent cytosolic DNA sensing mediates innate immune recognition of immunogenic tumors.Immunity. 2014; 41: 830-842Abstract Full Text Full Text PDF PubMed Scopus (726) Google Scholar In addition, radiation-induced DDR also activates innate immune cells present in the TME, including DCs,21Mackenzie K.J. Carroll P. Martin C.-A. et al.cGAS surveillance of micronuclei links genome instability to innate immunity.Nature. 2017; 548: 461-465Crossref PubMed Scopus (523) Google Scholar through mechanisms that remain incompletely characterized.22Gasser S. Raulet D.H. The DNA damage response arouses the immune system.Cancer Res. 2006; 66: 3959-3962Crossref PubMed Scopus (131) Google Scholar, 23Chatzinikolaou G. Karakasilioti I. Garinis G.A. DNA damage and innate immunity: Links and trade-offs.Trends Immunol. 2014; 35: 429-435Abstract Full Text Full Text PDF PubMed Scopus (91) Google Scholar, 24Shiao S.L. Coussens L.M. The tumor-immune microenvironment and response to radiation therapy.J Mammary Gland Biol Neoplasia. 2010; 15: 411-421Crossref PubMed Scopus (92) Google Scholar By inducing the translocation to the cell surface of “eat me signals” such as calreticulin, radiation triggers phagocytosis of cancer cells by DCs and other APCs.25Obeid M. Tesniere A. Ghiringhelli F. et al.Calreticulin exposure dictates the immunogenicity of cancer cell death.Nat Med. 2007; 13: 54-61Crossref PubMed Scopus (1851) Google Scholar Once loaded with tumor antigen and activated by damage-associated molecular patterns in the irradiated TME, cDC1s (classical DC subsets) migrate to the tumor-associated draining lymph node (tDLN) where they activate naïve CD8 T-cells.26Noubade R. Majri-Morrison S. Tarbell K.V. Beyond cDC1: Emerging roles of DC crosstalk in cancer immunity.Front Immunol. 2019; 10: 1014Crossref Scopus (21) Google Scholar Activated CD8-T cells are recruited to the irradiated tumor site by cytokines upregulated by radiation.27Lim J.Y. Gerber S.A. Murphy S.P. et al.Type I interferons induced by radiation therapy mediate recruitment and effector function of CD8(+) T cells.Cancer Immunol Immunother. 2014; 63: 259-271Crossref PubMed Scopus (123) Google Scholar, 28Matsumura S. Wang B. Kawashima N. et al.Radiation-induced CXCL16 release by breast cancer cells attracts effector T cells.J Immunol. 2008; 181: 3099-3107Crossref PubMed Scopus (405) Google Scholar, 29Luo R. Firat E. Gaedicke S. et al.Cisplatin facilitates radiation-induced abscopal effects in conjunction with PD-1 checkpoint blockade through CXCR3/CXCL10-mediated T-cell recruitment.Clin Cancer Res. 2019; 25: 7243-7255Crossref PubMed Scopus (22) Google Scholar In addition, ionizing radiation upregulates MHC-I antigens, death receptors, and NKG2D ligands on cancer cells.30Kim J.-Y. Son Y.-O. Park S.-W. et al.Increase of NKG2D ligands and sensitivity to NK cell-mediated cytotoxicity of tumor cells by heat shock and ionizing radiation.Exp Mol Med. 2006; 38: 474-484Crossref PubMed Scopus (116) Google Scholar, 31Ruocco M.G. Pilones K.A. Kawashima N. et al.Suppressing T cell motility induced by anti-CTLA-4 monotherapy improves antitumor effects.J Clin Invest. 2012; 122: 3718-3730Crossref PubMed Scopus (139) Google Scholar, 32Reits E.A. Hodge J.W. Herberts C.A. et al.Radiation modulates the peptide repertoire, enhances MHC class I expression, and induces successful antitumor immunotherapy.J Exp Med. 2006; 203: 1259-1271Crossref PubMed Scopus (954) Google Scholar The DDR induced by radiation therapy triggers transcription and translation of multiple genes that respond to the physical insult of radiation. If genetic mutations of the tumor cell are expressed, this response can originate neoepitopes presented by MHC-I on cancer cells that may circumvent the tolerance associated with established tumors.32Reits E.A. Hodge J.W. Herberts C.A. et al.Radiation modulates the peptide repertoire, enhances MHC class I expression, and induces successful antitumor immunotherapy.J Exp Med. 2006; 203: 1259-1271Crossref PubMed Scopus (954) Google Scholar,33Chakraborty M. Abrams S.I. Camphausen K. et al.Irradiation of tumor cells up-regulates Fas and enhances CTL lytic activity and CTL adoptive immunotherapy.J Immunol. 2003; 170: 6338-6347Crossref PubMed Scopus (326) Google Scholar We have recently provided the first clinical evidence of development of CD8 T cells specific for a mutated neoantigen encoded in a patient with metastatic, treatment-refractory non-small cell lung cancer who had responded to a regimen of ipilimumab and radiation therapy.34Formenti S.C. Rudqvist N.-P. Golden E. et al.Radiotherapy induces responses of lung cancer to CTLA-4 blockade.Nat Med. 2018; 24: 1845-1851Crossref PubMed Scopus (290) Google Scholar This patient carries a mutation in KPNA2, a gene upregulated in expression by radiation.34Formenti S.C. Rudqvist N.-P. Golden E. et al.Radiotherapy induces responses of lung cancer to CTLA-4 blockade.Nat Med. 2018; 24: 1845-1851Crossref PubMed Scopus (290) Google Scholar Tumor-specific T-cell clones appeared in the peripheral blood shortly after completion of radiation to a metastatic site and the first cycle of ipilimumab and remained elevated while the patient achieved a complete response in all of the nonirradiated lesions. A concurrent increase in IFN-I was detectable in the circulation. The findings support the hypothesis that in situ vaccination was achieved in this patient and that the observed abscopal effects were mediated by the neoantigen-specific T cells, directed at least in part against the unique radiation-exposed KPNA2 mutation in the tumor. The mechanisms described suggest a viral mimicry for radiation therapy, whereby with similar mechanisms elicited in response to a viral infection, radiation therapy induces IFN and exposes neoepitopes that may enable the host’s immune system to reject the tumor. A successful cancer vaccine strategy must overcome hurdles including antigen identification (which can vary from patient to patient, cell to cell, and tumor to tumor), mode of vaccine delivery, types of adjuvants, preconceived T-cell responses, and an immunosuppressive microenvironment.35Palucka K. Banchereau J. Dendritic-cell-based therapeutic cancer vaccines.Immunity. 2013; 39: 38-48Abstract Full Text Full Text PDF PubMed Scopus (534) Google Scholar DCs are indispensable for the establishment of effective antitumor immunity given their ability to capture (in situ), process, and present tumor antigens (after maturation [expression of surface MHC class II molecules and costimulatory molecules] and migration [acquisition of chemokine receptors] to tDLN) to T cells for their subsequent clonal expansion (via cytokine secretion). The use of DC-based vaccines to accelerate tumor rejection appears to be a promising approach.36Banchereau J. Steinman R.M. Dendritic cells and the control of immunity.Nature. 1998; 392: 245-252Crossref PubMed Scopus (11831) Google Scholar,37Trombetta E.S. Mellman I. Cell biology of antigen processing in vitro and in vivo.Annu Rev Immunol. 2005; 23: 975-1028Crossref PubMed Scopus (825) Google Scholar To date, limited clinical success of tumor rejection from the use of DC-based tumor vaccines has been achieved, thus necessitating further refinement. Thus far, DC-based vaccines have included peptide- and nucleic acid–based vaccines, antigen-coupled DC-targeting antibodies, and ex vivo–generated DCs preloaded with tumor antigens. An alternative approach is to convert the tumor into an immunogenic hub with locally disruptive technologies for the establishment of a personalized tumor vaccine that could be applied across various hosts and tumor types. Radiation therapy is one such tool that can expose DCs to the determinants required for the development of a tumor vaccine. Various strategies have been proposed, including locally disruptive technologies (eg, radiation therapy, cryoablation, microwave ablation), to fragment intact tumors and to release their contents into the adjacent milieu for subsequent APC uptake. Unfortunately, locally disruptive technologies alone are poorly immunogenic as they result in the release of factors that are both proinflammatory and immune suppressive. Thus, these local treatments require the addition of an adjuvant to unleash their immunogenic potential. In a screen of a combination of immunotherapies, the intratumoral injection of the toll-like receptor 9 (TLR9) agonist CpG oligonucleotide (which engages TLR9, a receptor expressed in APCs that recognizes pathogen-associated molecular patterns and subsequently initiates cytokine production responsible for innate and adaptive immunity) and anti-OX40 agonistic antibodies (OX40, a secondary costimulatory immune checkpoint molecule) provided impressive results in murine models. The combination cured multiple types of cancer and prevented the development of spontaneous genetically driven tumors. Additional TLR agonists with similar local APC maturation effects have been shown to promote T-cell clonal expansion, including poly I:C (TLR3 agonist), monophosphoryl lipid A (TLR4), flagellin (TLR5), and imiquimod (TLR7).38Dubensky Jr., T.W. Reed S.G. Adjuvants for cancer vaccines.Semin Immunol. 2010; 22: 155-161Crossref PubMed Scopus (179) Google Scholar Because of the systemic effects of the local administration of these adjuvants, the additional benefit of combining locally disruptive therapies to increase the availability to cryptic tumor antigens has been pursued. Locally disruptive technologies have been shown to augment responses of TLR agonists. In a preclinical lymphoma model, intratumoral injection of a TLR9 agonist (CpG oligonucleotides) induced systemic antitumor immunity and cured large, disseminated tumors. In a subsequent study of patients with low-grade B-cell lymphoma, radiation therapy plus TLR9 agonists to a single tumor site resulted in a 25% response rate in distant untreated tumors. This maneuver was shown to induce systemic antilymphoma clinical responses without the required production of a customized vaccine product.39Brody J.D. Ai W.Z. Czerwinski D.K. et al.In situ vaccination with a TLR9 agonist induces systemic lymphoma regression: A phase I/II study.J Clin Oncol. 2010; 28: 4324-4332Crossref PubMed Scopus (345) Google Scholar A similar maneuver combining Fms-like tyrosine kinas receptor 3 ligand, radiation therapy, and a TLR3 agonist was shown to induce antitumor effector T-cell responses and systemic (abscopal) cancer remission in patients with advanced stage low-grade lymphoma, substantiating the view that recruiting and activating intratumoral, crosspriming DCs is achievable with a combined approach.40Hammerich L. Marron T.U. Upadhyay R. et al.Systemic clinical tumor regressions and potentiation of PD1 blockade with in situ vaccination.Nat Med. 2019; 25: 814-824Crossref PubMed Scopus (139) Google Scholar Likewise, in a TS/A murine model of breast cancer with cutaneous involvement, the combination of topical imiquimod (TLR7 agonist) and local radiation therapy induced complete regression of treated lesions and improved systemic tumor control, leading to enhanced survival.41Dewan M.Z. Vanpouille-Box C. Kawashima N. et al.Synergy of topical toll-like receptor 7 agonist with radiation and low-dose cyclophosphamide in a mouse model of cutaneous breast cancer.Clin Cancer Res. 2012; 18: 6668-6678Crossref PubMed Scopus (107) Google Scholar To realize the potential of radiation-induced in situ vaccination there are several factors pertaining to the technique, delivery, targeting, and quality of radiation therapy that must be considered and may affect the treatment-related vaccinal effect. Technological advances including the advent of image guided precision radiation therapy and widespread clinical uptake of stereotactic body radiation therapy (stereotactic body radiation therapy [SBRT] or SABR) have permitted dose escalation of the tumor and sparing of adjacent normal tissues in hypofractionated regimens. However, there is limited understanding of how alternative dose-fractionation schedules or target selection modify the underlying radiobiological and immunologic mechanisms of radiation therapy. Preclinical work has highlighted a complex interplay between dose per fraction and Trex1-regulated immunogenicity as well as a dose-dependent induction of ICD,15Vanpouille-Box C. Alard A. Aryankalayil M.J. et al.DNA exonuclease Trex1 regulates radiotherapy-induced tumour immunogenicity.Nat Commun. 2017; 8: 15618Crossref PubMed Scopus (645) Google Scholar,42Golden E.B. Frances D. Pellicciotta I. et al.Radiation fosters dose-dependent and chemotherapy-induced immunogenic cell death.Oncoimmunology. 2014; 3e28518Crossref PubMed Scopus (262) Google Scholar,43Gameiro S.R. Jammeh M.L. Wattenberg M.M. et al.Radiation-induced immunogenic modulation of tumor enhances antigen processing and calreticulin exposure, resulting in enhanced T-cell killing.Oncotarget. 2014; 5: 403-416Crossref PubMed Scopus (230) Google Scholar thus providing rationale to explore this phenomenon in the clinic and suggesting that additional radiation-related factors are likely to affect the ability to generate an effective in situ vaccine. The majority of clinical studies exploring the interaction of radiation and immunotherapy have generally targeted a single metastatic site and examined local control of irradiated lesions and out-of-field responses in nonirradiated lesions. Although anecdotal clinical vignettes of abscopal responses exist in the literature and several nascent prospective studies have reported out-of-field responses, the observed responses with the addition of radiation therapy to immunotherapy have been modest and challenging to consistently reproduce in the clinic.44Xie C. Duffy A.G. Brar G. et al.Immune checkpoint blockade in combination with stereotactic body radiotherapy in patients with metastatic pancreatic ductal adenocarcinoma.Clin Cancer Res. 2020; 26: 2318-2326Crossref Scopus (14) Google Scholar, 45Postow M.A. Knox S.J. Goldman D.A. et al.A prospective, phase 1 trial of nivolumab, ipilimumab, and radiotherapy in patients with advanced melanoma.Clin Cancer Res. 2020; 26: 3193-3201Crossref Scopus (9) Google Scholar, 46Kwon E.D. Drake C.G. 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The rationale for targeting multiple metastatic sites with SBRT is multifaceted, including the recognition of cancer heterogeneity at metastatic stages,49De Mattos-Arruda L. Sammut S.-J. Ross E.M. et al.The genomic and immune landscapes of lethal metastatic breast cancer.Cell Rep. 2019; 27: 2690-2708.e10Abstract Full Text Full Text PDF PubMed Scopus (44) Google Scholar as well as the potential immunologic advantages of a reduction in systemic disease burden. Importantly, several lines of clinical evidence have recently suggested that local consolidation with radiation therapy in select subsets of patients with low metastatic disease burden can meaningfully extend survival.50Gomez D.R. Tang C. Zhang J. et al.Local consolidative therapy vs. maintenance therapy or observation for patients w