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Radiotherapy and immune checkpoints inhibitors for advanced melanoma

      Abstract

      Introduction

      The therapeutic landscape of metastatic melanoma drastically changed after the introduction of targeted therapies and immunotherapy, in particular immune checkpoints inhibitors (ICI). In recent years, positive effects on the immune system associated to radiotherapy (RT) were discovered, and radiation has been tested in combination with ICI in both pre-clinical and clinical studies (many of them still ongoing). We here summarize the rationale and the preliminary clinical results of this approach.

      Materials and methods

      In the first part of this review article, redacted with narrative non-systematic methodology, we describe the clinical results of immune checkpoints blockade in melanoma as well as the biological basis for the combination of ICI with RT; in the second part, we systematically review scientific publications reporting on the clinical results of the combination of ICI and RT for advanced melanoma.

      Results

      The biological and mechanistic rationale behind the combination of ICI and radiation is well supported by several preclinical findings. Retrospective observational series and few prospective trials support the potential synergistic effect between radiation and ICI for metastatic melanoma.

      Conclusion

      RT may potentiate anti-melanoma activity of ICI by enhancing response on both target and non-target lesions. Several prospective trials are ongoing with the aim of further exploring this combination in the clinical setting, hopefully confirming initial observations and opening a new therapeutic window for advanced melanoma patients.

      Keywords

      The main role of the immune system is to restore normal tissues’ homeostasis when altered by pathologic processes, including neoplastic transformation [
      • Vesely M.D.
      • Kershaw M.H.
      • Schreiber R.D.
      • Smyth M.J.
      Natural innate and adaptive immunity to cancer.
      ]. The immune system is often successful in eliminating neoplastic cells. Thus, all established tumours need to overcome immunity to progress and grow, and most of them successfully escape immune control, through different mechanisms [
      • Hanahan D.
      • Weinberg R.A.
      Hallmarks of cancer: the next generation.
      ,
      • Dunn G.P.
      • Old L.J.
      • Schreiber R.D.
      The three Es of cancer immunoediting.
      ]. Until recently, attempts in developing and applying immunotherapeutic strategies aimed to enhance innate and adaptive immune response failed in controlling most of solid tumours, with some exception represented by melanoma and renal cancer, where different forms of immunomodulation have been used for years. The clinical scenario drastically changed after the introduction of immune checkpoints inhibitors (ICI), a new class of targeted drugs able to activate the immune system against cancer cells, and showing efficacy for both solid tumours and haematological malignancies, with striking results leading to unexpected survival gains for advanced/unresectable melanoma [
      • Schadendorf D.
      • Hodi F.S.
      • Robert C.
      • et al.
      Pooled analysis of long-term survival data from phase II and phase III trials of ipilimumab in unresectable or metastatic melanoma.
      ]. Melanoma is actually the first cancer subtype where these immune-activating agents showed an advantage in survival over standard chemotherapy, and data from large clinical trials confirmed a substantial benefit with prolonged survival [
      • Davey R.J.
      • Westhuizen A.V.
      • Bowden N.A.
      Metastatic melanoma treatment: combining old and new therapies.
      ].
      Over the last decade, it was also hypothesized that the combined effects of radiation therapy (RT) and immunotherapy in metastatic tumours might be synergistic, and this research field is currently one of the most stimulating and potentially practice-changing topics in radiation oncology. Several mechanisms have been proposed for explaining the interaction between RT and the immune system. Among them, microenvironment modification, cytokine and danger signals release, pro-inflammatory effect and immunogenic cell death pattern [
      • Golden E.B.
      • Pelliciotta I.
      • Demaria S.
      • Barcellos-Hoff M.H.
      • Formenti S.C.
      The convergence of radiation and immunogenic cell death signaling pathways.
      ,
      • Apetoh L.
      • Ghiringhelli F.
      • Tesniere A.
      • Obeid M.
      • Ortiz C.
      • Criollo A.
      • et al.
      Toll-like receptor 4-dependent contribution of the immune system to anticancer chemotherapy and RT.
      ,
      • Demaria S.
      • Pilones K.A.
      • Vanpouille-Box C.
      • Golden E.B.
      • Formenti S.C.
      The optimal partnership of radiation and immunotherapy: from preclinical studies to clinical translation.
      ]; one of the most attractive experimental hypotheses is that ionizing radiations may act as an “in situ” vaccination in cancer patients, enhancing what has been called the “abscopal” effect after RT [
      • Demaria S.
      • Ng B.
      • Devitt M.L.
      • et al.
      Ionizing radiation inhibition of distant untreated tumours (abscopal effect) is immune mediated.
      ]. Such effect has been occasionally observed in patients undergoing palliative RT (especially in melanoma), and consists in a response of untreated (distant, outside radiation volumes) lesions following a radiation cycle to one lesion [
      • Kingsley D.P.
      An interesting case of possible abscopal effect in malignant melanoma.
      ]. According to preclinical models, the “abscopal” effect is immune-mediated [
      • Demaria S.
      • Ng B.
      • Devitt M.L.
      • et al.
      Ionizing radiation inhibition of distant untreated tumours (abscopal effect) is immune mediated.
      ], and thus may be enhanced by the combined use of ionizing radiation and immune-modulating drugs of different classes, for example the immune checkpoint inhibitor ipilimumab, an anti-CTLA-4 monoclonal antibody which binds to T cells [
      • Demaria S.
      • Kawashima N.
      • Yang A.M.
      • et al.
      Immune-mediated inhibition of metastases after treatment with local radiation and CTLA-4 blockade in a mouse model of breast cancer.
      ].
      Ipilimumab activates T cells by blocking the inhibitory signal mediated by CTLA-4 through the interaction with surface receptors on antigen presenting cells (APC). Retrospective clinical reports showed that the combination of RT and ipilimumab was able to trigger the abscopal effect in a proportion of melanoma patients, and that this effect might prolong survival [
      • Postow M.A.
      • Callahan M.K.
      • Barker C.A.
      • et al.
      Immunologic correlates of the abscopal effect in a patient with melanoma.
      ,
      • Grimaldi A.M.
      • Simeone E.
      • Giannarelli D.
      • et al.
      Abscopal effects of RT on advanced melanoma patients who progressed after ipilimumab immunotherapy.
      ,
      • Barker C.A.
      • Postow M.A.
      • Khan S.A.
      • et al.
      Concurrent RT and ipilimumab immunotherapy for patients with melanoma.
      ]. Additionally, experimental data provided the proof of principle that radiation and immunotherapy may favourably interact to enhance abscopal anti-tumour effects, and radiation may be used as a way to potentiate the effects of immunotherapy [
      • Demaria S.
      • Pilones K.A.
      • Formenti S.
      • Dustin M.
      Exploiting the stress response to radiation to sensitize poorly immunogenic tumours to anti-CLA-4 treatment.
      ]. At the same time, new insights into the mechanisms leading to resistance to the combination of radiation and ICI and possible approaches for overcoming this phenomenon have been discovered [
      • Twyman-Saint Victor C.
      • Rech A.J.
      • Maity A.
      • et al.
      Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer.
      ]. As a consequence of the prolonged survival time achievable with new generation systemic therapies, RT to one or few metastatic sites (either in conventional fractionation or delivering high dose in few fractions) is now widely in use as a local therapy especially for oligo-metastatic disease [
      • Ricardi U.
      • Filippi A.R.
      • Franco P.
      New concepts and insights into the role of radiation therapy in extracranial metastatic disease.
      ].
      Aim of this review is to focus on the potential therapeutic partnership between RT and ICI in advanced melanoma, discussing the most relevant pre-clinical and clinical findings, current research and future challenges.

      Materials and methods

      A narrative methodology was used for selecting and reporting studies on the clinical use of ICI for advanced melanoma, and for describing the biological and mechanistic basis of the combination of radiation and ICI. A systematic review was then performed according to validated guidelines [
      • Moher D.
      • Liberati A.
      • Tetzlaff J.
      • Altman D.G.
      Preferred reporting items for systematic reviews and meta-analyses: the PRISMA statement.
      ,
      • Baumann M.
      • Overgaard J.
      What next?.
      ] for selecting clinical studies reporting on the combination of RT and ICI for advanced melanoma. For this second part, we searched for English-language full length articles published from January 2000 to December 2015 using PubMed, and only studies reporting clinical outcomes following the combination of ICI and RT for metastatic melanoma were included. Studies were excluded if: (a) they were review articles and (b) they were not the most recently published outcomes, in instances of multiple publications from the same study cohort. The search strategy was “metastatic melanoma” OR “advanced melanoma” AND “radiotherapy” OR “radiosurgery” OR “stereotactic body radiotherapy” AND “ipilimumab” OR “pembrolizumab” OR “nivolumab”, which identified 560 articles. Two clinicians reviewed these records to determine which were suitable for inclusion according to the pre-defined criteria, selecting 11 reports. Five more articles were added from reference lists of the selected publications, for a total of 16 reports on the clinical outcomes of the combined treatment.

      Immune checkpoints inhibitors in melanoma: state of the art and new challenges

      Until recently, the medical management of unresectable metastatic melanoma was based on the use of chemotherapy, either as single agent (dacarbazine, temozolomide or fotemustine) or multi-drug associations, with or without biotherapies such as interferon or interleukin-2. Despite the optimistic results coming from mainly single centre phase II trials suggesting a potential benefit of chemotherapy plus interferon and/or interleukin-2, a series of randomized trials did not demonstrate any advantage [
      • Hamm C.
      • Verma S.
      • Petrella T.
      • Bak K.
      • Charette M.
      Biochemotherapy for the treatment of metastatic malignant melanoma: a systematic review.
      ]. Two large meta-analyses clearly showed that even if bio-chemotherapy may increase the percentage of responses, this does not result in any improvement of survival, while is coupled with a higher toxicity [
      • Ives N.J.
      • Stowe R.L.
      • Lorigan P.
      • Wheatley K.
      Chemotherapy compared with biochemotherapy for the treatment of metastatic melanoma: a meta-analysis of 18 trials involving 2,621 patients.
      ,
      • Sasse A.D.
      • Sasse E.C.
      • Clark L.G.
      • Ulloa L.
      • Clark O.A.
      Chemoimmunotherapy versus chemotherapy for metastatic malignant melanoma.
      ]. The unsatisfactory results obtained by (bio)-chemotherapy were recently summarized in a study analysing the clinical data of more than 2000 patients enrolled onto 42 phase II trials since 1975. An overall 1-year survival of 25.5% and a median survival of 6.2 months were achieved, without any significant improvement over the last 30 years [
      • Korn E.L.
      • Liu P.Y.
      • Lee S.J.
      • et al.
      Meta-analysis of phase II cooperative group trials in metastatic stage IV melanoma to determine progression-free and overall survival benchmarks for future phase II trials.
      ].
      Even if it was well known that T-cell response is regulated through a complex balance of inhibitory and activating signals and that the tumour itself can deregulate these pathways leading therefore to an impairment of the immune system activities [
      • Dunn G.P.
      • Old L.J.
      • Schreiber R.D.
      The three Es of cancer immunoediting.
      ], the relevant new concept which was developed following the failure of cytokine-based immunotherapy was the potential of targeting these inhibitory and activating immunological synapses as a new tool to promote anti-melanoma immune response. Up till now, while the field is rapidly evolving and new drugs are under investigation, two main types of monoclonal antibodies targeting immune checkpoints have been developed and investigated in the treatment of metastatic melanoma. The first targets the cytotoxic T-lymphocyte antigen 4 CTLA-4, the other the Programmed Death 1 (PD-1) pathway. It is of relevance that both compounds physiologically interact with immunological checkpoints leading to inhibitory signals for T cells (priming and effector phases): the blockade of these pathways allows the release of the immune system brake thus fostering, maintaining and stimulating the T-cell response [
      • Pardoll D.E.
      The blockade of immune checkpoints in cancer immunotherapy.
      ].

      Anti-CTLA-4

      Ipilimumab is a fully humanized monoclonal antibody that binds to CTLA-4, a receptor expressed on the T-cell surface that interacts with CD80 (B7-1) and CD86 (B7-2) on the Antigen-Presenting-Cells (APCs) and down regulates T-cell response. CTLA-4 blockade allows CD28 to bind to B7-1 receptors, leading to immune activation, IL-2 secretion, cytotoxic T-cells expansion and proliferation [
      • Korman A.J.
      • Peggs K.S.
      • Allison J.P.
      Checkpoints blockade in cancer immunotherapy.
      ]. The interaction of ipilimumab with the immune system takes place in an early phase of the immune response involving “naive” T lymphocytes and the APCs. This mechanism of action explains the characteristics of the clinical activity as well as the common side effects of this drug, consisting of immune-mediated reactions developing more frequently in the skin, gastro-intestinal tract (mainly diarrhoea), liver and endocrinal glands. Moreover, it gives reason to the delayed occurrence of a relevant clinical response.
      After pre-clinical data and pilot studies showing activity, a randomized phase II study compared different dose regimens in metastatic melanoma (0.3, 3 or 10 mg/kg IV every 3 weeks), showing that both 3 and 10 mg/kg induced optimal response, even if the latter dose was coupled with an increase in immune-related adverse effects (irAEs) [
      • Wolchok J.D.
      • Neyns B.
      • Linette G.
      • et al.
      Ipilimumab monotherapy in patients with pretreated advanced melanoma: a randomised, double-blind, multicentre, phase 2, dose-ranging study.
      ]. In the first phase III trial, ipilimumab ± glycoprotein 100 peptide (gp100) vaccine was compared with gp100 vaccine monotherapy in patients with unresectable stage III or stage IV melanoma. Ipilimumab monotherapy significantly improved median overall survival (OS) compared with gp100 vaccine monotherapy (10.1 months vs. 6.4 months) [
      • Hodi F.S.
      • O’Day S.J.
      • McDermott D.F.
      • et al.
      Improved survival with ipilimumab in patients with metastatic melanoma.
      ]. In a second randomized phase III trial, the combination of ipilimumab (10 mg/kg) and dacarbazine (850 mg/sqm) resulted in significantly superior OS compared to dacarbazine (850 mg/sqm) plus placebo (11.2 months vs. 9.1 months) [
      • Robert C.
      • Thomas L.
      • Bondarenko I.
      • et al.
      Ipilimumab plus dacarbazine for previously untreated metastatic melanoma.
      ].
      Notably, ipilimumab produced a plateau in survival curves: a recent pooled analysis of OS data for 1.861 patients enrolled in 10 prospective and 2 retrospective trials, with up to 10 years follow-up, showed that the survival curve began to plateau around 3 years after treatment. Three-year OS rates were 22%, 26%, and 20% for all, treatment-naive, and previously treated patients, respectively [
      • Schadendorf D.
      • Hodi F.S.
      • Robert C.
      • et al.
      Pooled analysis of long-term survival data from phase II and phase III trials of ipilimumab in unresectable or metastatic melanoma.
      ]. The results of the ipilimumab expanded access programme (EAP) in Italy were consistent with these data, confirming the activity of the drug also in specific patient’s subsets such as the elderly, the mucosal or uveal primaries, and in the presence of brain metastases [
      • Ascierto P.A.
      • Simeone E.
      • Sileni V.C.
      • et al.
      Clinical experience with ipilimumab 3 mg/kg: real-world efficacy and safety data from an expanded access program cohort.
      ].
      Response to ipilimumab can be preceded by an increase in diameter of tumour lesions, reflecting the inflammatory changes and the recall of T-cells to the tumour site. Distinct response patterns have been described: (a) shrinkage in baseline lesions, without new lesions; (b) durable stable disease (in some patients followed by a slow, steady decline in total tumour burden); (c) response after an increase in total tumour burden; and (d) response in the presence of new lesions [
      • Wolchok J.D.
      • Hoos A.
      • O’Day S.
      • et al.
      Guidelines for the evaluation of immune therapy activity in solid tumours: immune-related response criteria.
      ]. This implies the need to adopt specific immune-related criteria to correctly evaluate response, based on the concomitant assessment of both the primary lesions and the appearance of new lesions [
      • Ribas A.
      • Chmielowski B.
      • Glaspy J.A.
      Do we need a different set of response assessment criteria for tumour immunotherapy?.
      ]. Table 1 summarizes the results of phase II/III clinical trials.
      Table 1Summary of the most relevant clinical trials on ipilimumab for advanced melanoma.
      AuthorStudy designDrug(s)No patientsORR (%)DCR (%)Median survival time (months)1-Year OS (%)2-Year OS (%)3-Year OS (%)
      Wolchock et al., 2010
      • Wolchok J.D.
      • Neyns B.
      • Linette G.
      • et al.
      Ipilimumab monotherapy in patients with pretreated advanced melanoma: a randomised, double-blind, multicentre, phase 2, dose-ranging study.
      Randomized, double-blind, phase 2 trialIpilimumab 0.3 mg/kg72013.78.639.618.4NR
      Ipilimumab 3 mg/kg724.226.48.739.324.2
      Ipilimumab10 mg/kg7311.129.211.448.629.8
      Hodi et al., 2010
      • Hodi F.S.
      • O’Day S.J.
      • McDermott D.F.
      • et al.
      Improved survival with ipilimumab in patients with metastatic melanoma.
      Randomized, double-blind phase 3 trialIpilimumab + GP100



      4035.720.11043.621.6NR
      Ipilimumab alone13710.928.510.145.623.5
      GP 100 alone1361.5116.425.313.7
      Robert et al., 2011
      • Robert C.
      • Thomas L.
      • Bondarenko I.
      • et al.
      Ipilimumab plus dacarbazine for previously untreated metastatic melanoma.
      Randomized, double-blind phase 3 trialIpilimumab + Dacarbazine25015.233.211.247.328.520.8
      Dacarbazine + Placebo25210.330.29.136.317.912.2
      Schadendorf et al., 2015
      • Schadendorf D.
      • Hodi F.S.
      • Robert C.
      • et al.
      Pooled analysis of long-term survival data from phase II and phase III trials of ipilimumab in unresectable or metastatic melanoma.
      Pooled analysis (10 prospective
      Including two phase 3 trials (Refs. [24] and [25]).
      and 2 retrospective trials)
      Ipilimumab1861NRNR11.4NRNR22
      Expanded access cohort29859.521
      Abbreviations: ORR = overall response rate; DCR = disease control rate; OS = overall survival; NR = not reported.
      low asterisk Including two phase 3 trials (Refs.
      • Pardoll D.E.
      The blockade of immune checkpoints in cancer immunotherapy.
      and
      • Korman A.J.
      • Peggs K.S.
      • Allison J.P.
      Checkpoints blockade in cancer immunotherapy.
      ).

      Anti-PD-1

      PD-1 protein is a co-inhibitory receptor expressed on B and T cells, and has been shown to be involved in the negative regulation of T-cell activation [
      • Dunn G.P.
      • Old L.J.
      • Schreiber R.D.
      The three Es of cancer immunoediting.
      ]. PD-1 ligand (PD-L1) is expressed in different tumours, is associated with a worse prognosis and its expression and interaction with T cells is thought to be one of the main mechanisms underlying the immune system escape [
      • Dunn G.P.
      • Old L.J.
      • Schreiber R.D.
      The three Es of cancer immunoediting.
      ]. The discovery that tumour cells were able to activate the PD-1/PD-L1 axis, leading to protection from cytotoxic T cells trough exhaustion, led to the development of specific anti-PD-1 inhibitors [
      • Brahmer J.R.
      • Drake C.G.
      • Wollner I.
      • et al.
      Phase I study of single-agent anti-programmed death-1 (MDX-1106) in refractory solid tumours: safety, clinical activity, pharmacodynamics, and immunologic correlates.
      ]. The anti-PD-1 monoclonal antibodies nivolumab (a fully human anti-PD-1 IgG4) and pembrolizumab (a humanized anti-PD-1 IgG4) were shown to be highly effective for malignant melanoma, and in 2014 they both have been licensed in the United States as second- or third-line treatment for patients with evidence of disease progression after ipilimumab (in BRAF wild-type melanoma) or ipilimumab and BRAF inhibitors (in BRAF V600-mutated melanoma) [
      • Metcalfe W.
      • Anderson J.
      • Trinh V.A.
      • et al.
      Anti-programmed cell death-1 (PD-1) monoclonal antibodies in treating advanced melanoma.
      ]. The toxicity profile of anti-PD-1 agents was reported to be similar to anti-CTLA-4 [
      • Ganghadar T.C.
      • Vonderheide R.H.
      Mitigating the toxic effects of anticancer immunotherapy.
      ]. A randomized phase III study comparing nivolumab vs. dacarbazine in previously untreated melanoma without BRAF mutation demonstrated superior overall response rate (ORR, 40% vs. 13.9%, respectively) and increased 1-year OS (72.9% vs. 42.1%, respectively). Moreover, nivolumab resulted in a better safety profile: treatment-related adverse events occurred in 11.7% of the patients receiving nivolumab and 17.6% of the patients receiving dacarbazine, respectively [
      • Robert C.
      • Long G.V.
      • Brady B.
      • et al.
      Nivolumab in previously untreated melanoma without BRAF mutation.
      ]. In CheckMate 037 phase III trial, patients were randomly assigned 2:1 to receive nivolumab 3 mg/kg every 2 weeks or investigators’ choice chemotherapy (ICC) until progression or unacceptable toxic effects. Primary endpoints were the proportion of patients who had an objective response and overall survival. At first interim analysis on 120 and 47 randomized patients, confirmed objective responses were reported in 31.7% of patients in the nivolumab group vs. 10.6% of patients in the ICC group; no treatment-related deaths occurred [
      • Weber J.
      • D’Angelo S.P.
      • Minor D.
      • et al.
      Nivolumab versus chemotherapy in patients with advanced melanoma who progressed after anti-CTLA-4 treatment (CheckMate 037): a randomised, controlled, open-label, phase 3 trial.
      ].
      The activity of pembrolizumab for advanced melanoma was firstly shown in 2013 by a phase IB study achieving an ORR of 38% in both ipilimumab pre-treated or not pre-treated patients [
      • Hamid O.
      • Robert C.
      • Daud A.
      • et al.
      Safety and tumour responses with lambrolizumab (anti-PD-1) in melanoma.
      ]. Two different doses of pembrolizumab (2 mg/kg and 10 mg/kg) were then investigated and compared with ICC in the Keynote-002 randomized phase II clinical trial. At enrolment, patients had progressive disease after ipilimumab or, if BRAF mutated, after BRAF or MEK inhibitors, or both. Results showed an improvement in progression-free survival (PFS) at 6 months as assessed by independent central review, with HR 0.57 for pembrolizumab 2 mg/kg and 0.50 for 10 mg/kg. Grade 3–4 treatment-related adverse events were more frequent and occurred earlier in patients receiving chemotherapy [
      • Ribas A.
      • Puzanov I.
      • Dummer R.
      • et al.
      Pembrolizumab versus investigator-choice chemotherapy for ipilimumab-refractory melanoma (KEYNOTE-002): a randomised, controlled, phase 2 trial.
      ].
      In a large randomized phase III study, 834 patients with advanced melanoma were treated either with pembrolizumab at a dose of 10 mg/kg every 2 or every 3 weeks or with 4 doses of ipilimumab (3 mg/kg every 3 weeks). The estimated 6-months PFS rates were 47.3% for pembrolizumab every 2 weeks, 46.4% for pembrolizumab every 3 weeks, and 26.5% for ipilimumab, respectively. Estimated 1-year OS rates were 74.1%, 68.4%, and 58.2%, respectively. The response rate was improved when pembrolizumab was administered either every 2 or every 3 weeks, as compared with ipilimumab. Treatment-related adverse events of grade 3–5 severity were lower in the pembrolizumab groups (13.3% and 10.1%) [
      • Robert C.
      • Schachter J.
      • Long G.V.
      • et al.
      Pembrolizumab versus ipilimumab in advanced melanoma.
      ]. Details on phase II-III trials testing anti-PD-1 agents nivolumab or pembrolizumab in advanced melanoma are shown in Table 2.
      Table 2Summary of the most relevant clinical trials on nivolumab or pembrolizumab either alone or in combination with ipilimumab for advanced melanoma.
      AuthorStudy designDrug(s)No patientsORR (%)1-year OS (%)Median PFS (months)
      Robert et al., 2014
      • Robert C.
      • Long G.V.
      • Brady B.
      • et al.
      Nivolumab in previously untreated melanoma without BRAF mutation.
      Randomized, double-blind Phase 3 trialNivolumab

      ICC
      210

      208
      40

      13.9
      72.9

      42.1
      5.1

      2.2
      Weber et al., 2015
      • Weber J.
      • D’Angelo S.P.
      • Minor D.
      • et al.
      Nivolumab versus chemotherapy in patients with advanced melanoma who progressed after anti-CTLA-4 treatment (CheckMate 037): a randomised, controlled, open-label, phase 3 trial.
      Randomize, controlled, open-label, Phase 3 trialNivolumab

      ICC
      120

      47
      Per-protocol interim-analysis.
      31.7

      10.6
      NRNR
      Ribas et al., 2015
      • Ribas A.
      • Puzanov I.
      • Dummer R.
      • et al.
      Pembrolizumab versus investigator-choice chemotherapy for ipilimumab-refractory melanoma (KEYNOTE-002): a randomised, controlled, phase 2 trial.
      Randomized, double-blind Phase 2Pembrolizumab 2 mg/kg

      Pembrolizumab 10 mg/kg

      ICC
      180

      181

      179
      21
      Intention-to-treat, independent-central review, RECIST v1.1.


      25

      4
      NR34% at 6 months
      Intention-to-treat, independent-central review, RECIST v1.1.


      38% at 6 months

      16% at 6 months
      Robert et al., 2015
      • Robert C.
      • Schachter J.
      • Long G.V.
      • et al.
      Pembrolizumab versus ipilimumab in advanced melanoma.
      Randomized, double-blind Phase 2Pembrolizumab Q2W

      Pembrolizumab Q3W

      Ipilimumab
      279

      277

      278
      33.7

      32.9

      11.9
      74.1

      68.4

      58.2
      5.5

      4.1

      2.8
      Postow et al., 2015
      • Postow M.A.
      • Chesney J.
      • Pavlick A.C.
      • et al.
      Nivolumab and ipilimumab versus ipilimumab in untreated melanoma.
      Randomized, double-blind Phase 1Ipilimumab + Nivolumab

      Ipilimumab + Placebo
      Followed by Nivolumab or Placebo.
      72/23

      37/10
      BRAF wild-type/BRAF V600 mutation positive.
      61/52

      11/ 10
      BRAF wild-type/BRAF V600 mutation positive.
      27 (crude rate)

      37 (crude rate)
      Not reached/ 8.5

      4.4/2.7§
      Larkin et al., 2015
      • Larkin J.
      • Chiarion-Sileni V.
      • Gonzalez R.
      • et al.
      Combined nivolumab and ipilimumab or monotherapy in untreated melanoma.
      Randomized, double-blind Phase 3Nivolumab + Ipilimumab

      Ipilimumab

      Nivolumab
      314

      315

      316
      57.6

      19

      43.7
      11.5

      2.9

      6.9
      Hodi et al., 2015
      • Hodi F.S.
      • Postow M.A.
      • Chesney J.A.
      • et al.
      Clinical response, progression-free survival (PFS), and safety in patients (pts) with advanced melanoma (MEL) receiving nivolumab (NIVO) combined with ipilimumab (IPI) vs IPI monotherapy in CheckMate 069 study. Presented at American Society of Clinical Oncology Annual Meeting 2015.
      Randomized, double-blind Phase 2Nivolumab + Ipilimumab

      Ipilimumab
      72

      70
      60

      11
      8.9

      4.7
      Hodi et al., 2015
      • Hodi F.S.
      • Gibney G.
      • Sullivan R.
      • et al.
      An open-label, randomized, phase 2 study of nivolumab given sequentially with ipilimumab in patients with advanced melanoma (CheckMate 064).
      Randomized, open-label Phase 2Nivolumab + Ipilimumab → Nivolumab

      Ipilimumab + Nivolumab → Nivolumab
      68

      70
      41.2

      20
      NRNR
      Abbreviations: ICC = investigator’s choice chemotherapy; ORR = overall response rate; OS = overall survival; PFS = progression-free survival; NR = not reported.
      low asterisk Per-protocol interim-analysis.
      low asterisklow asterisk Intention-to-treat, independent-central review, RECIST v1.1.
      # Followed by Nivolumab or Placebo.
      § BRAF wild-type/BRAF V600 mutation positive.

      Combination of anti-CTLA-4 and anti PD-1 agents

      The rapid increase of these multiple immunotherapy options opened a new challenging field for metastatic melanoma, with the primary aim being the definition of the best combination strategy in order to improve effectiveness without compromising safety. Recent data demonstrated a substantial improvement in ORR and OS for the combination of ipilimumab and anti-PD-1 agents in different sequencing schedules in which patients are treated with both antibodies. A pilot phase I study tested different approaches: patients received concomitantly intravenous doses of nivolumab and ipilimumab every 3 weeks for 4 doses, followed by nivolumab alone every 3 weeks for 4 doses (concurrent regimen). In a sequenced regimen, patients previously treated with ipilimumab received nivolumab alone every 2 weeks for up to 48 doses. A total of 53 patients received concurrent therapy with nivolumab and ipilimumab, and 33 received sequential treatment. ORR for all patients in the concurrent-regimen group was 40%. Evidence of clinical activity was observed in 65% of patients. At the maximum doses that were associated with an acceptable level of adverse events, 53% of patients had an objective response, all with tumour reduction of 80% or more. Grade 3 or 4 adverse events related to therapy occurred in 53% of patients in the concurrent-regimen group, but were similar to previous experience with monotherapy and were generally reversible [
      • Wolchok J.D.
      • Kluger H.
      • Callahan M.K.
      • et al.
      Nivolumab plus ipilimumab in advanced melanoma.
      ].
      In a double-blind study involving 142 patients with previously untreated metastatic melanoma, patients were assigned 2:1 to receive ipilimumab (3 mg/kg) combined with either nivolumab (1 mg/kg) or placebo once every 3 weeks for four doses, followed by nivolumab (3 mg/kg) or placebo every 2 weeks until disease progression or unacceptable toxic effects. Among patients with BRAF wild-type tumours, ORR was 61% (44 of 72 patients) in the group that received both ipilimumab and nivolumab versus 11% in the group that received ipilimumab and placebo. Median PFS was not reached with the combination therapy and was 4.4 months with ipilimumab monotherapy. Similar results for ORR and PFS were observed in 33 patients with BRAF mutated tumours [
      • Postow M.A.
      • Chesney J.
      • Pavlick A.C.
      • et al.
      Nivolumab and ipilimumab versus ipilimumab in untreated melanoma.
      ]. Another key study showed encouraging data regarding the superiority of the combination over ipilimumab or nivolumab alone. Patients treated with the combination had a PFS of 11.5 months, compared to 2.9 months for those treated with ipilimumab alone and 6 months for those treated with nivolumab alone, respectively; the ORR (57.5%) obtained by the combination was remarkably higher, yet with a higher incidence of grade 3–4 immune-related adverse events (IrAEs), frequently involving more than one organ [
      • Larkin J.
      • Chiarion-Sileni V.
      • Gonzalez R.
      • et al.
      Combined nivolumab and ipilimumab or monotherapy in untreated melanoma.
      ]. The combination was also compared with ipilimumab alone in treatment-naïve patients in a phase II study: ORR was 60% and 11% for the combination treatment and for ipilimumab alone, respectively. Median PFS was 8.9 months for the combination versus 4.7 months for ipilimumab alone. Grade 3–4 drug-related adverse events were reported in 51% of patients receiving nivolumab plus ipilimumab, and in 20% in those receiving ipilimumab alone [
      • Hodi F.S.
      • Postow M.A.
      • Chesney J.A.
      • et al.
      Clinical response, progression-free survival (PFS), and safety in patients (pts) with advanced melanoma (MEL) receiving nivolumab (NIVO) combined with ipilimumab (IPI) vs IPI monotherapy in CheckMate 069 study. Presented at American Society of Clinical Oncology Annual Meeting 2015.
      ]. The recently presented data (in abstract form) of the CheckMate 064 trial, comparing the two sequences of ipilimumab followed by nivolumab vs. nivolumab followed by ipilimumab, suggest a better outcome in terms of ORR for the latter (at 25 weeks 41.2% vs. 20%, respectively) [
      • Hodi F.S.
      • Gibney G.
      • Sullivan R.
      • et al.
      An open-label, randomized, phase 2 study of nivolumab given sequentially with ipilimumab in patients with advanced melanoma (CheckMate 064).
      ]. Details on phase II-III trials testing the combination of nivolumab and ipilimumab are integrated in Table 2.

      Toxicity

      Checkpoint inhibition is associated with a unique spectrum of side effects termed immune-related adverse events (irAEs). IrAEs include dermatologic, gastrointestinal, hepatic, endocrine, and other less common inflammatory events. IrAEs are believed to arise from general immunologic enhancement, and temporary immunosuppression with corticosteroids, tumour necrosis factor-alpha antagonists, mycophenolate mofetil, or other agents can be an effective treatment in most cases. Treatment of moderate or severe irAEs may require a temporary interruption of the drug and the use of corticosteroid immunosuppression, without compromising anti-melanoma activity. Treatment is based upon the severity of the observed toxicity. In Table 3, we report the most common toxic effects associated to the use of anti-CTLA-4, anti-PD-1 or the combination of both for advanced melanoma [
      • Wolchok J.D.
      • Kluger H.
      • Callahan M.K.
      • et al.
      Nivolumab plus ipilimumab in advanced melanoma.
      ,
      • Postow M.A.
      • Chesney J.
      • Pavlick A.C.
      • et al.
      Nivolumab and ipilimumab versus ipilimumab in untreated melanoma.
      ,
      • Larkin J.
      • Chiarion-Sileni V.
      • Gonzalez R.
      • et al.
      Combined nivolumab and ipilimumab or monotherapy in untreated melanoma.
      ,
      • Hodi F.S.
      • Postow M.A.
      • Chesney J.A.
      • et al.
      Clinical response, progression-free survival (PFS), and safety in patients (pts) with advanced melanoma (MEL) receiving nivolumab (NIVO) combined with ipilimumab (IPI) vs IPI monotherapy in CheckMate 069 study. Presented at American Society of Clinical Oncology Annual Meeting 2015.
      ,
      • Hodi F.S.
      • Gibney G.
      • Sullivan R.
      • et al.
      An open-label, randomized, phase 2 study of nivolumab given sequentially with ipilimumab in patients with advanced melanoma (CheckMate 064).
      ]. As described, the combined use of a double ICI is more toxic and should be managed with special attention.
      Table 3Most common side effects of anti-CTLA4, anti-PD1 antibodies and the combination of both for advanced melanoma [Refs.
      • Wolchok J.D.
      • Kluger H.
      • Callahan M.K.
      • et al.
      Nivolumab plus ipilimumab in advanced melanoma.
      ,
      • Postow M.A.
      • Chesney J.
      • Pavlick A.C.
      • et al.
      Nivolumab and ipilimumab versus ipilimumab in untreated melanoma.
      ,
      • Larkin J.
      • Chiarion-Sileni V.
      • Gonzalez R.
      • et al.
      Combined nivolumab and ipilimumab or monotherapy in untreated melanoma.
      ,
      • Hodi F.S.
      • Postow M.A.
      • Chesney J.A.
      • et al.
      Clinical response, progression-free survival (PFS), and safety in patients (pts) with advanced melanoma (MEL) receiving nivolumab (NIVO) combined with ipilimumab (IPI) vs IPI monotherapy in CheckMate 069 study. Presented at American Society of Clinical Oncology Annual Meeting 2015.
      ,
      • Hodi F.S.
      • Gibney G.
      • Sullivan R.
      • et al.
      An open-label, randomized, phase 2 study of nivolumab given sequentially with ipilimumab in patients with advanced melanoma (CheckMate 064).
      ].
      Side effectsIpilimumab (%)Nivolumab (%)Ipilimumab + Nivolumab (%)
      Diarrhoea (any grade)33–371945
      Grade 3–46–1129–11
      Skin rash (any grade)17–332616–40
      Grade 3–40–213–5
      Hepatitis (any grade)4417–21
      Grade 3–40–218–9
      Colitis (any grade)12–13112–23
      Grade 3–47–918–17
      Hypophysitis (any grade)4–718–12
      Grade 3–42–402

      RT and immune checkpoints inhibitors in melanoma: rationale and preclinical data

      Despite the efficacy of ICI, still many patients do not respond or initially respond and then progress, and an anti-tumour immune response powerful enough to control a tumour when it re-emerges by blocking immunosuppressive mechanisms is possible in only a minority of patients [
      • Demaria S.
      • Pilones K.A.
      • Vanpouille-Box C.
      • Golden E.B.
      • Formenti S.C.
      The optimal partnership of radiation and immunotherapy: from preclinical studies to clinical translation.
      ]. This is the result of multiple mechanisms that obstacle the priming and activation of anti-melanoma T cells, their recruitment to the tumour sites as well as their function. These phenomena represent an arduous challenge to effective tumour rejection [
      • Gajewski T.F.
      • Meng Y.
      • Blank C.
      • et al.
      Immune resistance orchestrated by the tumour microenvironment.
      ,
      • Smyth M.J.
      • Godfrey D.I.
      • Trpani A.J.
      A fresh look at the tumour immunosurveillance and immunotherapy.
      ]. Ionizing radiation has been known for a long time to cause pro-inflammatory effects, but the potential benefits of this pro-inflammatory response only recently emerged. The most recent important findings were that radiation is able to convert the tumour in an “in situ” vaccine, altering the microenvironment towards the development of an “immunogenic hub”: radiation is in fact able to promote both the priming and effector phases of anti-tumour immune response [
      • Demaria S.
      • Bhardwaj N.
      • McBride W.H.
      • Formenti S.C.
      Combining RT and immunotherapy: a revived partnership.
      ]. Among the first observations, Lugade et al. evaluated anti-tumour immune responses in mice after treatment of B16 melanoma tumours with single (15 Gy) or fractionated (5 × 3 Gy) doses of localized ionizing radiation. Irradiated mice had cells with greater capability to present tumour antigens and specific T cells that secreted IFN-γ upon peptide stimulation within tumour-draining lymph nodes. Immune activation in tumour-draining lymph nodes correlated with an increase in the number of CD45+ cells infiltrating single dose irradiated tumours compared with non-irradiated mice. Similarly, irradiated mice had increased numbers of tumour-infiltrating lymphocytes that secreted IFN-γ and lysed tumour cell targets [
      • Lugade A.A.
      • Moran J.P.
      • Gerber S.A.
      • Rose R.C.
      • Frelinger J.G.
      • Lord E.M.
      Local radiation therapy of B16 melanoma tumours increases the generation of tumour antigen-specific effector cells that traffic to the tumour.
      ].
      Other crucial observations regarded both the role of radiation in modulating the peptide repertoire, enhancing MHC class I expression and inducing antitumour immunity [
      • Reits E.A.
      • Hodge J.W.
      • Herberts C.A.
      • et al.
      Radiation modulates the peptide repertoire, enhances MHC class I expression, and induces successful antitumour immunotherapy.
      ] and the radiation-induced IFN-gamma production within the tumour microenvironment, that again positively influences antitumour immunity [
      • Lugade A.A.
      • Sorensen E.W.
      • Gerber S.A.
      • Moran J.P.
      • Frelinger J.G.
      • Lord E.M.
      Radiation-induced IFN-gamma production within the tumour microenvironment influences antitumour immunity.
      ].
      Researchers from the University of Chicago then showed, on irradiated mice affected with melanoma and pancreatic cancer, that RT in combination with a specific DNA repair inhibitor was able to trigger immune response by releasing immunomodulatory signals from senescent tumour cells; stimulatory cytokines were discovered to activate T lymphocytes and potentiate immune response [
      • Meng Y.
      • Efimova E.V.
      • Hamzeh K.W.
      • et al.
      Radiation-inducible immunotherapy for cancer: senescent tumour cells as a cancer vaccine.
      ].
      The activation of natural anti-tumour T-cell response requires the uptake and cross-presentation of tumour-derived antigens by dendritic cells (DCs) to T cells; this process depends on type I interferon (IFN-I), which is necessary for DCs recruitment to tumours and their activation. In addition, radiation-induced cell death generates three key molecular signals, that have been shown to promote the uptake and presentation of tumour-derived antigens by DCs: (a) calreticulin translocation from the endoplasmic reticulum to the cell surface acts as a signal for uptake of dying cancer cells by DCs; (b) the release of the nuclear protein high-mobility group box-1 (HMGB1), which binds to toll like receptor (TLR)-4 on DCs, promotes antigen cross-presentation and (c) adenosine triphosphate (ATP) activates the inflammasome via the P2XR7 receptor with downstream release of interleukin (IL)-1β [
      • Golden E.B.
      • Pelliciotta I.
      • Demaria S.
      • Barcellos-Hoff M.H.
      • Formenti S.C.
      The convergence of radiation and immunogenic cell death signaling pathways.
      ]. The generation of these signals collectively defines immunogenic cell death (ICD), which is crucial in promoting the priming phase. Moreover, radiation is able to trigger the effector phase by inducing chemokines and cytokines to recruit effector T-cells at the tumour (CXCL 16), through up-regulation of major histocompatibility complex class I (MHC-I), adhesion molecules, death receptors and NKG2D ligands that enable recognition and elimination of damaged cancer cells [
      • Demaria S.
      • Golden E.B.
      • Formenti S.C.
      Role of local RT in cancer immunotherapy.
      ,
      • Matsumura S.
      • Wang B.
      • Kawashima N.
      • Braunstein S.
      • Badura M.
      • Cameron T.O.
      • et al.
      Radiation-induced CXCL16 release by breast cancer cells attracts effector T cells.
      ]. These discoveries were essential in re-defining the landscape of the interactions between local RT, tumour response and the immune system.
      At the same time, RT may reinforce immunosuppressive pathways that undermine anticancer immune-surveillance by different mechanisms, especially in the priming phase. Radiation can enhance tumour infiltration by myeloid-derived suppressor cells (MDSCs), a subset of bone marrow–derived immature myeloid cells responsible for sustaining chronic immunosuppression [
      • Golden E.B.
      • Pelliciotta I.
      • Demaria S.
      • Barcellos-Hoff M.H.
      • Formenti S.C.
      The convergence of radiation and immunogenic cell death signaling pathways.
      ]. Moreover, regulatory T cells, with a key role in suppression of antitumor immunity, seem more radiation resistant than conventional T cells and show a relative increase after radiation. Also CD47 blockade, whose expression on cancer cells suppress innate immunity, may directly enhance tumour immune-surveillance by CD8 T cells; CD47 expression in tumour microenvironment limits the cooperation between anti-tumour T cell immunity and radiation therapy [
      • Soto-Patoja D.R.
      • Terabe M.
      • Ghosh A.
      • et al.
      CD47 in the tumour microenvironment limits cooperation between anti-tumour T cell immunity and radiation therapy.
      ]. Recently, it was also shown that radiation induces Langerhans’ cells to migrate from the skin to lymph nodes, where they stimulate regulatory T-cells [
      • Price J.G.
      • Idoyaga J.
      • Salmon H.
      • et al.
      CDKN1A regulates Langerhans cell survival and promotes T-reg cell generation upon exposure to ionizing irradiation.
      ,
      • Zitvogel L.
      • Kroemer G.
      Subversion of anticancer immunosurveillance by RT.
      ]. Additionally, counterbalance of the effector phase induced by RT is the increase expression of PD-L1 on cancer cells after radiation, which may de-activate effector T-cells [
      • Golden E.B.
      • Pelliciotta I.
      • Demaria S.
      • Barcellos-Hoff M.H.
      • Formenti S.C.
      The convergence of radiation and immunogenic cell death signaling pathways.
      ].
      As a consequence, the effects of radiation on tumour microenvironment and its interaction with the immune system appear as a complex balance of activating and suppressing signals. The pro-immunogenic “go” signals are countered by immunosuppressive “stop” signals: the balance of stop and go signals determines the development of effective antitumor immune responses. Positive effects of RT should therefore be harnessed to enhance immune activation, especially in combination with immune-activating agents such as immune checkpoint inhibitors [
      • Formenti S.C.
      • Demaria S.
      Combining RT and cancer immunotherapy: a paradigm shift.
      ]. Zegers et al. demonstrated that a radiation-induced antitumor effect can be enhanced by the administration of IL-2 in combination with L19, a selective tumour targeting agent able to improve therapeutic outcome over IL-2 treatment alone. L19 binds to extra domain B, a part of the fibronectin in tumour neovasculature and overexpressed in many solid tumours [
      • Zegers C.M.
      • Rekers N.H.
      • Quaden D.H.
      • et al.
      RT combined with the immunocytokine L19-IL-2 provides long lasting antitumour effects.
      ]. Researchers from the same group lately showed that also in tumour models lacking MHC1 expression and depending on natural killer (NK) immune response, the combination of L19-IL2 plus radiation was able to improve tumour growth delay by an additive effect [
      • Rekers N.H.
      • Zegers C.M.
      • Yaromina A.
      • et al.
      Combination of RT with the immunocytokine L19-IL-2: additive effect in a NK cell dependent tumour model.
      ].
      The combination of RT with ICI has been explored on different fronts across years. Preclinical studies have reported increased loco-regional control when radiation is combined with checkpoint blockade immunotherapy in different cancer subtypes [
      • Sharabi A.B.
      • Lim M.
      • DeWeese T.L.
      • Drake C.G.
      Radiation and checkpoint blockade immunotherapy: radiosensitisation and potential mechanisms of synergy.
      ]. Moreover, increased systemic disease control has been shown by combining radiation with both anti-CTLA-4 and anti-PD-1/PD-L1 inhibitors [
      • Twyman-Saint Victor C.
      • Rech A.J.
      • Maity A.
      • et al.
      Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer.
      ]. A key point in this field was the discovery that the manifestation of the abscopal effect, rarely occurring after RT, is immune-mediated [
      • Demaria S.
      • Ng B.
      • Devitt M.L.
      • et al.
      Ionizing radiation inhibition of distant untreated tumours (abscopal effect) is immune mediated.
      ]. Investigators at Vanderbilt University found that mice that underwent irradiation of a melanoma tumour to 25 Gy in 1 fraction before surgical excision had fewer lung metastases than did mice that underwent excision but no radiation treatment. Greater tumour infiltration by CD8 T cells in mice that had previously received an ex vivo irradiated melanoma vaccine were noted, and the authors suggested that DC-mediated phagocytosis was responsible for the decrease in the frequency of metastases in mice with irradiated tumours [
      • Perez C.A.
      • Fu A.
      • Onishko H.
      • et al.
      Radiation induces an antitumour immune response to mouse melanoma.
      ]. Similarly, investigators at the University of Chicago reported that the efficacy of high-dose ablative radiation therapy in a mouse model of melanoma was immune mediated by CD8 T cells [
      • Lee Y.
      • Auh S.L.
      • Wang Y.
      • et al.
      Therapeutic effects of ablative radiation on local tumour require CD8fl T cells: Changing strategies for cancer treatment.
      ]. Sharabi et al., from the University of California, noted improved tumour control when radiation was combined with anti-PD-1 antibody in a mouse model of breast cancer and melanoma, with enhanced proliferation and activation of antigen-specific T cells and effector memory cells in draining lymph nodes. Stereotactic RT resulted in the development of antigen-specific T cell and B cell-mediated immune responses. These immune-stimulating effects were significantly increased when radiation was combined with either anti-PD-1 therapy or regulatory T cell depletion, resulting in improved local tumour control [
      • Sharabi A.B.
      • Nirschl C.J.
      • Kochel C.M.
      • et al.
      Stereotactic radiation therapy augments antigen-specific PD-1 mediated anti-tumour immune responses via cross-presentation of tumour antigen.
      ]. Park et al. also showed in preclinical melanoma and renal carcinoma models that the combination of stereotactic RT plus PD-1 blockade was able to induce complete regression of the irradiated primary tumour also eliciting a reduction in size of non-irradiated outside the radiation field (abscopal effect). The observed effect was tumour specific and was not dependent on tumour histology or host genetic background, suggesting that RT may induce an abscopal tumour-specific immune response in both the irradiated and non-irradiated tumours, which is potentiated by PD-1 blockade [
      • Park S.S.
      • Dong H.
      • Liu X.
      • et al.
      PD-1 restrains RT-induced abscopal effect.
      ].
      Taken altogether, these data constitute the experimental basis for the combination of ICI and radiation therapy to one or few lesions, with the aim of enhancing the effects of immunotherapy by using radiation as a powerful tool to overcome resistance and to trigger abscopal effect. For the first time, the results of a prospective “proof of principle” trial, smartly testing the combination of local RT and granulocyte–macrophage colony stimulating factor (GM-CSF), as DCs growth factor, successfully showed in various cancer subtypes (not including melanoma) that RT on metastatic sites could allow the activation of the abscopal effect on non-target lesions in a substantial fraction of patients [
      • Golden E.B.
      • Chhabra A.
      • Chachoua A.
      • et al.
      Local RT and granulocyte-macrophage colony-stimulating factor to generate abscopal responses in patients with metastatic solid tumours: a proof-of-principle trial.
      ]. Results of this trial are paramount to further support research in this field, particularly when investigating the combination of RT and ICI. A study investigating on the combination of anti-CTLA-4 in both humans and mouse models of metastatic melanoma showed that the induction of the abscopal effect is limited to a fraction of patients due to an acquired resistance to ipilimumab which is PD-1/PD-L1 mediated. The clinical component of this study was a phase I trial testing the combination of RT on a single lesion (6–8 Gy delivered over two or three fractions) followed by ipilimumab (4 cycles, beginning 3–5 days after the last RT fraction), showing a 36% overall abscopal response rate. Non-responding patients had up regulated PD-L1, and genetic elimination of PD-L1 from therapy-resistant melanoma cells dramatically restored response to ipilimumab plus radiation. This study planted a seed for the sequential combination of radiation and both anti-CTLA-4 and anti-PD1/PD-L1 agents, as a promising strategy to evade immune resistance and trigger the abscopal effect at the highest degree [
      • Barker C.A.
      • Postow M.A.
      • Khan S.A.
      • et al.
      Concurrent RT and ipilimumab immunotherapy for patients with melanoma.
      ]. As well summarized by Ngiow et al., anti-PD-1/PD-L1 antibodies may combat adaptive immune resistance upon localized radiation plus anti-CTLA-4 therapy, and the superior activity of radiation and dual immune checkpoint blockade is mediated by non-redundant immune mechanisms [
      • Ngiow S.F.
      • McArthur G.A.
      • Smyth M.J.
      RT complements immune checkpoint blockade.
      ].
      Fig. 1 illustrates a schematic model of the possible interaction between radiation, tumour microenvironment and the immune system, summarizing the “in situ” vaccination concept and the “abscopal” effect induced by RT. Fig. 2 illustrates the interactions between RT and ICI, with distinctive clinical combinations and sequences, aiming at different possible endpoints (maximizing response, overcoming resistance).
      Figure thumbnail gr1
      Fig. 1The “in situ vaccination” concept: ionizing radiation may increase antigens release from dying cancer cells, activate dendritic cells, expand specific anti-melanoma cytotoxic T cells (CTCs) through cross-priming in draining lymph nodes and increase immune response at both local and distant sites [modified from Ref.
      • Demaria S.
      • Golden E.B.
      • Formenti S.C.
      Role of local RT in cancer immunotherapy.
      ].
      Figure thumbnail gr2
      Fig. 2Different possible therapeutic combinations between RT and ICI for advanced melanoma, with the aim of: (A) maximizing response upfront (concomitant approach, higher toxicity) (B) overcoming resistance in poor responders (sequential approach) and (C) triggering the restoration of immune response and overcoming acquired resistance after initial response (sequential/concomitant approach) [Refs.
      • Demaria S.
      • Golden E.B.
      • Formenti S.C.
      Role of local RT in cancer immunotherapy.
      ,
      • Twyman-Saint Victor C.
      • Rech A.J.
      • Maity A.
      • et al.
      Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer.
      ]. For cells’ shapes and colours see . ICI: immune checkpoint inhibitors.

      RT and immune checkpoint inhibitors in melanoma: clinical data

      In 1975, Kingsley et al. firstly described a case of abscopal effect in a patient treated with radiation therapy for metastatic melanoma [
      • Kingsley D.P.
      An interesting case of possible abscopal effect in malignant melanoma.
      ]. More recently, two pivotal clinical reports showed how the combination of RT and ipilimumab might be efficient in obtaining disease control on the treated site enhancing abscopal effect on un-irradiated sites. Postow et al. [
      • Postow M.A.
      • Callahan M.K.
      • Barker C.A.
      • et al.
      Immunologic correlates of the abscopal effect in a patient with melanoma.
      ] described the case of a female patient treated with 4 doses of ipilimumab at 10 mg/kg followed by maintenance every 12 weeks. After 1 year she had progressive disease on both a para-spinal mass and spleen/thoracic lymph nodes, and received palliative fractionated RT on the para-spinal mass, while continuing ipilimumab. After 4 months the targeted mass regressed and, remarkably, also a very good partial response was observed on the hilar lymph nodes and spleen lesions, with stable disease at 10 months. The authors performed immunological studies showing an increase in antibody response after RT, consistent with an immune-mediated abscopal effect. Few months later, Hiniker et al. [
      • Hiniker S.M.
      • Chen D.S.
      • Reddy S.
      • et al.
      A systemic complete response of metastatic melanoma to local radiation and immunotherapy.
      ] reported on a case of a male patient who first developed a nodal recurrence after resection of the primary (at 3 years) and then had oligo-recurrent metastatic disease with 2 liver metastases at 4 years. He received 2 doses of ipilimumab followed by RT on 2 out of 7 metastases, followed by 2 more doses of ipilimumab. At 5 months, all liver lesions were in complete response, but the patient relapsed at the site of previous surgery (skin): he was simply observed, and the lesion completely resolved after 2 months. Investigators at Memorial Sloan Kettering Cancer Centre (MSKCC) performed a retrospective analysis of 29 patients who received extra-cranial RT in combination with ipilimumab: no significant increase in adverse effects was observed, and patients receiving RT during maintenance had higher OS than those treated during the induction phase [
      • Barker C.A.
      • Postow M.A.
      • Khan S.A.
      • et al.
      Concurrent RT and ipilimumab immunotherapy for patients with melanoma.
      ]. A retrospective observational series on 23 patients treated with palliative RT after ipilimumab reported the occurrence of abscopal responses in 11/23 (52%); median time between ipilimumab and RT was 5 months, and median OS for patients obtaining an abscopal response was significantly higher than for non-responding patients (22.4 vs. 8.3 months) [
      • Grimaldi A.M.
      • Simeone E.
      • Giannarelli D.
      • et al.
      Abscopal effects of RT on advanced melanoma patients who progressed after ipilimumab immunotherapy.
      ]. Chandra et al., on 47 patients, showed in 68% of cases an improved response on index lesions (outside radiation field) [
      • Chandra R.A.
      • Wilhite T.J.
      • Balboni T.A.
      • et al.
      A systematic evaluation of abscopal responses following RT in metastatic melanoma patients treated with ipilimumab.
      ]. A case of abscopal response after SRS for a brain metastasis and ipilimumab was also reported by Stamell et al. [
      • Stamell E.F.
      • Wolchock J.D.
      • Gnjatic S.
      • et al.
      The abscopal affect associated with a systemic anti-melanoma immune response.
      ]. Barker and Postow recently reviewed the clinical outcomes of the combination of ipilimumab and RT in melanoma, including brain metastases [
      • Barker C.A.
      • Postow M.A.
      Combinations of radiation therapy and immunotherapy for melanoma: a review of clinical outcomes.
      ]. A few reports described successful outcomes in patients treated with ipilimumab and whole-brain radiation therapy (WBRT). Early reports included a 49-year-old patient who received ipilimumab 4 weeks after receiving 30 Gy WBRT, with a significant regression of brain metastases at 12 weeks after the initiation of ipilimumab [
      • Muller-Brenne T.
      • Rudolph B.
      • Schmidberger H.
      Successful therapy of a cerebral metastasized malignant melanoma by whole-brain radiation therapy and ipilimumab.
      ], and a woman with lepto-meningeal disease who received 20 Gy WBRT followed by ipilimumab having a complete radiographic response 2–3 months after completing treatment, without symptoms [
      • Bot I.
      • Blank C.U.
      • Brandsma D.
      Clinical and radiological response of leptomeningeal melanoma after whole brain RT and ipilimumab.
      ]. Gerber et al. reported on 13 patients receiving WBRT and ipilimumab, with a promising overall response rate, yet 10/10 patients with available imaging demonstrated new or increased intralesional bleeding [
      • Gerber N.K.
      • Young R.J.
      • Barker C.A.
      • et al.
      Ipilimumab and whole brain RT for melanoma brain metastases.
      ]. Investigators from Dana-Farber Cancer Institute reported on 16 melanoma patients who received ipilimumab and either WBRT or stereotactic radiosurgery (SRS), 4 concomitantly. Extra-cranial target lesions achieved a response rate of 35% when comparing pre and post RT imaging [
      • Schoenfeld J.D.
      • Mahadevan A.
      • Floyd S.R.
      • et al.
      Ipilimumab and cranial radiation in metastatic melanoma patients: a case series and review.
      ]. Silk et al. from the University of Michigan compared 33 patients with brain metastases receiving either SRS or WBRT and ipilimumab (before or after RT) vs. 37 not receiving ipilimumab, showing improved survival for the combination of SRS and ipilimumab [
      • Silk A.W.
      • Bassetti M.F.
      • West B.T.
      • Tsien C.I.
      • Lao C.D.
      Ipilimumab and radiation therapy for melanoma brain metastases.
      ].
      Knisely et al. reported on 77 patients with brain metastases treated with SRS, with patients who received ipilimumab having a median survival of 21.3 months vs. 4.9 months for those who did not. Survival was not significantly different whether the drug was given before or after SRS [
      • Knisely J.P.
      • Yu J.B.
      • Flanigan J.
      • et al.
      Radiosurgery for melanoma brain metastases in the ipilimumab era and the possibility of longer survival.
      ]. In a similar study from New York University, on 58 patients treated with brain SRS, no difference in local tumour control, survival, or frequency of intracranial haemorrhage was reported for those who did or did not receive ipilimumab, respectively [
      • Mathew M.
      • Tam M.
      • Ott P.A.
      • et al.
      Ipilimumab in melanoma with limited brain metastases treated with stereotactic radiosurgery.
      ]. Tazi et al. reported on the combination of SRS and ipilimumab on 10 patients, showing promising survival results (comparable to those without brain metastases) [
      • Tazi K.
      • Hataway A.
      • Chiuzan C.
      • Shirai K.
      Survival of melanoma patients with brain metastases treated with ipilimumab and stereotactic radiosurgery.
      ].
      Investigators at Memorial Sloan-Kettering Cancer Centre (MSKCC) also reported on 46 patients treated with ipilimumab and brain SRS: on multivariate analysis, prolonged survival was associated with the delivery of SRS during ipilimumab [
      • Kiess A.P.
      • Wolchok J.D.
      • Barker C.A.
      • et al.
      Stereotactic radiosurgery for melanoma brain metastases in patients receiving ipilimumab: safety profile and efficacy of combined treatment.
      ]. An increase in brain metastasis size >150% occurred in 40% of the tumours treated with SRS before or during ipilimumab and, conversely, in 10% of metastases treated with SRS after ipilimumab. Haemorrhage was observed after SRS during ipilimumab in 42% of brain metastases. The potential complications of brain SRS and ipilimumab were studied in a small series of 3 patients: after 20 Gy SRS, radiation necrosis was observed in all, proven radiologically in two and histologically in one patient, respectively [
      • Du-Four S.
      • Wilgenhof S.
      • Duerinck J.
      • et al.
      Radiation necrosis of the brain in melanoma patients successfully treated with ipilimumab: three case studies.
      ]. Radionecrosis was also reported by Knisely et al. (3/27 patients) [
      • Knisely J.P.
      • Yu J.B.
      • Flanigan J.
      • et al.
      Radiosurgery for melanoma brain metastases in the ipilimumab era and the possibility of longer survival.
      ] and Kiess et al. (5/46 patients) [
      • Kiess A.P.
      • Wolchok J.D.
      • Barker C.A.
      • et al.
      Stereotactic radiosurgery for melanoma brain metastases in patients receiving ipilimumab: safety profile and efficacy of combined treatment.
      ]. The higher rate of increasing lesions as well as radio-necrosis features among patients receiving SRS or WBRT in combination with ICI is a matter of debate, but many researchers believe that these findings could be an expression of greater local immune reactions.
      Table 4 summarizes the clinical outcomes reported so far by different series with the combination of RT and ICI in metastatic melanoma.
      Table 4Clinical outcomes following the combination of radiotherapy and ipilimumab for advanced melanoma.
      StudyNo patientsRadiotherapy dose/ fractionation (Type)Ipilimumab schedule (sequence)Targeted siteTarget site response (%)Abscopal response
      Either evaluated with Response Evaluation Criteria in Solid Tumours or with immune-related response criteria.
      (%)
      Postow et al., 2012
      • Postow M.A.
      • Callahan M.K.
      • Barker C.A.
      • et al.
      Immunologic correlates of the abscopal effect in a patient with melanoma.
      128.5 Gy/3 fr (palliative)10 mg/kg (sequential)Para-spinal metastasisPR 100PR 100
      Hiniker et al., 2012
      • Hiniker S.M.
      • Chen D.S.
      • Reddy S.
      • et al.
      A systemic complete response of metastatic melanoma to local radiation and immunotherapy.
      154 Gy/3 fr (SBRT)3 mg/kg (concurrent)Liver metastasisCR 100CR 100
      Barker et al., 2013
      • Barker C.A.
      • Postow M.A.
      • Khan S.A.
      • et al.
      Concurrent RT and ipilimumab immunotherapy for patients with melanoma.
      2930 Gy/10 fr (median) (SBRT/palliative)3–10 mg/kg (concurrent/ maintenance)Non-brain lesionsSymptoms relief 77NR
      Grimaldi et al., 2014
      • Grimaldi A.M.
      • Simeone E.
      • Giannarelli D.
      • et al.
      Abscopal effects of RT on advanced melanoma patients who progressed after ipilimumab immunotherapy.
      2120–24/1; 20 Gy/ 5fr; 30 Gy/10fr; 50 Gy/25 fr (SBRT/SRS/palliative/WBRT)3 mg/kg (sequential)Brain, bone, lymph-node, cutaneous lesionsPR 62PR 43

      SD 10
      Chandra et al., 2015
      • Chandra R.A.
      • Wilhite T.J.
      • Balboni T.A.
      • et al.
      A systematic evaluation of abscopal responses following RT in metastatic melanoma patients treated with ipilimumab.
      4726 Gy (median) (SBRT/SRS/palliative/WBRT)3–10 mg/kg (sequential)Brain, soft tissue, bone, intrathoracic, abdominovisceralNRPR 36
      Stamell et al., 2013
      • Stamell E.F.
      • Wolchock J.D.
      • Gnjatic S.
      • et al.
      The abscopal affect associated with a systemic anti-melanoma immune response.
      1NR (SRS)NRBrainNRCR 100
      Muller-Brenne et al., 2003
      • Muller-Brenne T.
      • Rudolph B.
      • Schmidberger H.
      Successful therapy of a cerebral metastasized malignant melanoma by whole-brain radiation therapy and ipilimumab.
      130 Gy/10 fr WBRT3 mg/kg (sequential)BrainCR 100NR
      Bot et al., 2012
      • Bot I.
      • Blank C.U.
      • Brandsma D.
      Clinical and radiological response of leptomeningeal melanoma after whole brain RT and ipilimumab.
      120 Gy/5fr WBRT3 mg/kg (sequential)BrainCR 100NR
      Gerber et al.
      • Gerber N.K.
      • Young R.J.
      • Barker C.A.
      • et al.
      Ipilimumab and whole brain RT for melanoma brain metastases.
      1330 Gy/10 fr (median) WBRT3–10 mg/kg (concurrent)BrainPR/ SD 56%NR
      Schoenfeld et al., 2015
      • Schoenfeld J.D.
      • Mahadevan A.
      • Floyd S.R.
      • et al.
      Ipilimumab and cranial radiation in metastatic melanoma patients: a case series and review.
      1622 Gy; 36 Gy (SRS/WBRT)3–10 mg/kg (sequential/concurrent)BrainNRPR 35
      Silk et al., 2013
      • Silk A.W.
      • Bassetti M.F.
      • West B.T.
      • Tsien C.I.
      • Lao C.D.
      Ipilimumab and radiation therapy for melanoma brain metastases.
      3314–24 Gy/1–5 fr; 30–37.5 Gy/10–13 fr (SRS/WBRT)3 mg/kg (sequential)BrainPR 56.7NR
      Knisely et al., 2012
      • Knisely J.P.
      • Yu J.B.
      • Flanigan J.
      • et al.
      Radiosurgery for melanoma brain metastases in the ipilimumab era and the possibility of longer survival.
      27NR (SRS)NR (sequential)BrainNRNR
      Mathew et al., 2013
      • Mathew M.
      • Tam M.
      • Ott P.A.
      • et al.
      Ipilimumab in melanoma with limited brain metastases treated with stereotactic radiosurgery.
      2520 Gy/1 fr (median) (SRS)3 mg/kg (sequential/concurrent)Brain6-months LC 63%NR
      Tazi et al., 2015
      • Tazi K.
      • Hataway A.
      • Chiuzan C.
      • Shirai K.
      Survival of melanoma patients with brain metastases treated with ipilimumab and stereotactic radiosurgery.
      10NR (SRS)3 mg/kg (sequential/concurrent)BrainNRNR
      Kiess et al., 2015
      • Kiess A.P.
      • Wolchok J.D.
      • Barker C.A.
      • et al.
      Stereotactic radiosurgery for melanoma brain metastases in patients receiving ipilimumab: safety profile and efficacy of combined treatment.
      4615–24 Gy/1 fr (SRS)3–10 mg/kg (sequential/concurrent)BrainNRNR
      Du-Four et al., 2012
      • Du-Four S.
      • Wilgenhof S.
      • Duerinck J.
      • et al.
      Radiation necrosis of the brain in melanoma patients successfully treated with ipilimumab: three case studies.
      320 Gy/1 fr (SRS)3 mg/kg (sequential)BrainCR 100NR
      Abbreviations: Gy: Gray; fr: fractions; WBRT: whole-brain radiotherapy; CR: complete response; PR: partial response; SD: stable disease; LC: local control, NR: not reported¸ SRS: stereotactic radiosurgery; SBRT: stereotactic body radiation therapy.
      low asterisk Either evaluated with Response Evaluation Criteria in Solid Tumours or with immune-related response criteria.

      Dose/fractionation issues and the role of stereotactic ablative RT

      The technological advances in the field of Radiation Oncology now allow for rapid non-invasive delivery of very high radiation doses to various metastatic sites. Stereotactic Body Radiation Therapy (SBRT) or Stereotactic Ablative RT (SABR) has been extensively investigated in recent years in metastatic patients [
      • Ricardi U.
      • Filippi A.R.
      • Franco P.
      New concepts and insights into the role of radiation therapy in extracranial metastatic disease.
      ,
      • Ricardi U.
      • Badellino S.
      • Filippi A.R.
      Clinical applications of stereotactic radiation therapy for oligometastatic cancer patients: a disease-oriented approach.
      ], showing high local control rates and promising progression-free survival estimates, at the price of a very limited toxicity. With the term SBRT/SABR, we now refer to a “philosophy” of cancer treatment with high focused doses in 1 or few sessions (<10), administered with ablative intent. Very few melanoma patients have been included in studies on SABR for extra-cranial metastases, mostly for lung and liver [
      • Ricardi U.
      • Filippi A.R.
      • Franco P.
      New concepts and insights into the role of radiation therapy in extracranial metastatic disease.
      ]: efficacy and toxicity, when reported, appear similar to any other primary histology.
      The best combination strategy, as well as the best dose/fractionation regimen for RT is still under debate. When combined with ipilimumab, preclinical comparisons of different dose fractionation schedules (24 Gy in 3 fractions, 30 Gy in 5 fractions, or 20 Gy in single fraction) in breast cancer models suggest that multi-fraction approach is superior to single fraction regimens, in terms of abscopal effect induction [
      • Dewan M.Z.
      • Galloway A.E.
      • Kawashima N.
      • et al.
      Fractionated but not single-dose RT induces an immune-mediated abscopal effect when combined with anti-CTLA-4 antibody.
      ]. Golden et al. reported on the combination of ipilimumab and 30 Gy in 5 fractions on liver lesions in a patient with refractory lung cancer, with complete response of treated and untreated lesions and the patient alive at one year [
      • Golden E.B.
      • Demaria S.
      • Schiff P.B.
      • et al.
      An abscopal response to radiation and ipilimumab in a patient with metastatic non-small cell lung cancer.
      ].
      The clinical data reported so far on melanoma patients appear consistent with these findings, as shown in Table 4. In some reports, ablative fractionation has been used to target visceral lesions, with 18 Gy × 3, in others a slight hypo-fractionation was preferred, delivering 9.5 Gy × 3. However, abscopal effect was also observed after ipilimumab when standard fractionation was used. The only prospective trial testing the combination of radiation therapy and dendritic cell stimulation with GM-CSF in various cancer subtypes employed a 35 Gy/10 fractions approach on 2 index lesions treated subsequently [
      • Golden E.B.
      • Chhabra A.
      • Chachoua A.
      • et al.
      Local RT and granulocyte-macrophage colony-stimulating factor to generate abscopal responses in patients with metastatic solid tumours: a proof-of-principle trial.
      ]. The biological mechanisms of radiation induced cell death at fraction sizes greater than 810 Gy might be different from the classical radiobiology paradigm of conventionally fractionated RT. In addition to DNA damaging events, experimental models suggest that endothelial membrane alterations including sphingomyelin mediated endothelial apoptosis lead to microvasculature dysfunction [
      • Fuks Z.
      • Kolesnik R.
      Engaging the vascular component of the tumour response.
      ]. A marked increase in MHC-1 molecules expression was observed after high doses (25 Gy) when irradiating melanoma cells at different dose levels [
      • Reits E.A.
      • Hodge J.W.
      • Herberts C.A.
      • et al.
      Radiation modulates the peptide repertoire, enhances MHC class I expression, and induces successful antitumour immunotherapy.
      ]. Lee et al. demonstrated that the reduction of tumour burden after ablative RT depends largely on T-cell responses: ablative radiation may dramatically increase T-cell priming in draining lymphoid tissues, leading to reduction/eradication of the primary tumour or distant metastasis in a cytotoxic T cell-dependent fashion [
      • Lee Y.
      • Auh S.L.
      • Wang Y.
      • et al.
      Therapeutic effects of ablative radiation on local tumour require CD8fl T cells: Changing strategies for cancer treatment.
      ]. Smilowitz et al. showed that increasing radiation dose may improve the outcome of immunotherapy when irradiating an advanced intracerebral melanoma model on mice at different dose levels in single fraction (15, 18.75 or 22.5 Gy) [
      • Smilowitz H.M.
      • Micca P.L.
      • Sasso D.
      • et al.
      Increasing radiation dose improves immunotherapy outcome and prolongation of tumor dormancy in a subgroup of mice treated for advanced intracerebral melanoma.
      ]. The use of ablative radiation doses on small targets in combination with immunotherapy seems a good strategy also because of draining lymph nodes sparing, allowing for APC migration, T-cell activation, cross priming and in situ effector T cells migration. However, we need to wait for prospective ongoing trials to fully answer to this question.
      Likewise, the optimal timing of the combination between RT and immune checkpoints inhibitors is a matter of debate: in melanoma, the establishment of anti-tumour immunity seems to be enhanced when anti-CTLA-4 antibody precedes RT, or when given concomitantly [
      • Demaria S.
      • Bhardwaj N.
      • McBride W.H.
      • Formenti S.C.
      Combining RT and immunotherapy: a revived partnership.
      ]. In a recent Editorial, it has been suggested that radiation should be regarded as a complex “drug”, and its combination with immunotherapy warrants a systematic approach to optimize the efficacy of the radiation component [
      • Golden E.B.
      • Formenti S.C.
      Radiation therapy and immunotherapy: growing pains.
      ]. An alternative approach to the concomitant use of ablative doses and ICI is a strategy where an intervention aimed to increase priming (i.e. GM-CSF or toll-like receptor agonists) is followed by concomitant radiation and anti-CTLA-4, and next by anti-PD1/anti-PD-L1 agents to prevent T cell exhaustion. From experimental data, this approach seems the most promising in advanced melanoma [
      • Barker C.A.
      • Postow M.A.
      • Khan S.A.
      • et al.
      Concurrent RT and ipilimumab immunotherapy for patients with melanoma.
      ], however many combinations might be efficient and ready to be explored in the clinical setting (Fig. 2).
      A summary of ongoing clinical trials testing the combination of RT and immune checkpoints inhibitors in melanoma is provided in Table 5.
      Table 5Prospective clinical trials combining either anti-CTLA-4 or anti-PD-1 agents and radiotherapy for advanced melanoma (from www.clinicaltrials.gov, December 2015, in order of estimated completion date).
      Registration numberStudy designEligibility criteriaInterventionPrimary endpointEstimated enrolmentEstimated study completion date
      NCT01689974Phase IILocally unresectable, metastatic melanoma, with at least 2 distinct measurable metastatic sites, one of at least 1 cm or largerArm A: IPI alone

      Arm B: IPI and RT
      Response rate10Completed in March 2015
      NCT01497808Phase I/IIMetastatic melanomaIPI and SBRTDose- limiting toxicity40June 2015 (ongoing, not recruiting)
      NCT01557114Phase IUnresectable locally advanced or metastatic melanoma with at least one melanoma metastasis accessible to radiation therapyInduction IPI (4 courses), →RT→

      Maintenance IPI
      Maximum Tolerated Dose of RT in combination with IPI30March 2016
      NCT01449279Single institution, open-label, pilot studyStage IV melanomaIPI and palliative radiation therapyPercentage of patients experiencing serious adverse events in the first 4 months of treatment20June 2016
      NCT01996202Phase IResected patients at high risk of recurrence/ Neoadjuvant- definitive approach for locally advanced patientsRT and IPIIncidence of immune related adverse events associated with IPI¸ acute and late radiation toxicities24June 2016
      NCT01970527Phase IIRecurrent/stage IV Melanoma

      Index lesion between 1 and 5 cm
      SBRT (3 fractions) between days 1 and 13 → IPI every 3 weeks (4 courses)Late toxicity, immune- related clinical response, immune-related PFS, OS40September 2016
      NCT02115139Phase IIMelanoma brain metastasesWhole brain RT with concurrent IPI1-year OS66October 2016
      NCT02097732Phase IIMelanoma brain metastasesStandard arm: SRS → IPI (4 cycles)

      Experimental arm: IPI (2 cycles) → SRS → IPI (2 cycles)
      Local control rate40May 2017
      NCT02107755Phase IIOligo-metastatic melanomaSBRT with concurrent IPIPFS32June 2017
      NCT02406183Phase IMetastatic melanoma with at least 3 extra-cranial measurable lesionsSBRT with concurrent IPIMaximum Tolerated dose, with dose-limiting toxicity in 25% of patient21July 2017
      NCT01565837Phase IIOligo-metastatic but unresectable melanomaSBRT with concurrent IPIOS, safety and tolerability (acute and subacute toxicity)50November 2017
      NCT02407171Phase IIa (expansion cohort)Metastatic melanoma (with at least one site of measurable disease suitable for SBRT)SBRT (at maximum tolerated dose discovered in phase I) and Pembro (200 mg every 2 weeks)Overall response rate60December 2018
      NCT02562625Phase IIUnresectable or stage IV melanoma with 1-3 lesions targets for high dose radiotherapy and at least one other lesion which will not be irradiated to assess the abscopal effect of the treatment

      Arm 1: Pembro alone

      Arm 2: Pembro and RT (24 Gy/ 3 fr)
      Abscopal effect234October 2019
      NCT01703507Phase IMelanoma brain metastasesArm A: IPI and WBRT

      Arm B: IPI and SRS
      Maximum tolerated dose of IPI24November 2019
      NCT02318771Phase IMetastatic melanoma (among other tumour types)
      • RT (8 Gy/1 fr–20 Gy/5 fr) → re-biopsy → Pembro
      • Pembro → RT → Pembro
      Change in PD-LI levels40January 2020
      Abbreviations: SRS: stereotactic radiosurgery; SBRT: stereotactic body radiation therapy; PFS: progression-free survival; OS: overall survival; IPI: ipilimumab; Pembro: pembrolizumab; NA: not applicable; NR: not reported.

      Conclusions

      Both preclinical models and clinical data showed that RT to one or few melanoma metastases might trigger and/or enhance the so-called abscopal effect. This effect is amplified in melanoma when combining RT with ICI such as anti-CTLA-4 or anti-PD-1/anti-PD-L1 antibodies, or the concomitant/sequential combination of both. Recent discoveries led to a better understanding of the mechanisms underlying this effect, and clinical data from retrospective observational studies and few prospective studies confirmed this hypothesis, suggesting prolonged response and survival. Currently, several prospective trials are ongoing with the aim of defining which is the best combination strategy, as well the best RT dose/fractionation regimen. Results of these studies will give answers to very important questions, hopefully creating a new window of therapeutic opportunities for metastatic melanoma patients, especially for those who still do not respond to ICI, where radiation to one or few lesions could play a major role in enhancing immune-mediated anti-tumour effects.

      Conflict of interest statement

      The Authors declare no conflict of interest with the material included in this review article.

      Acknowledgements

      The Authors would like to thank Pierluigi Fresia for his precious assistance in preparing the Figures.

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