BJR © 2020 The Authors.

Published by the British Institute of Radiology

https://​doi.​org/​10.​1259/​bjr.​20190224
Received: Revised: Accepted:
27 February 2019 10 June 2019 14 July 2019

Cite this article as:

Durante M, Formenti S. Harnessing radiation to improve immunotherapy: better with particles?. Br J Radiol 2020; 93: 20190224.

Proton Therapy special feature: Review Article

Harnessing radiation to improve immunotherapy: better

with particles?
1,2
Marco Durante, PhD and 3Silvia Formenti, MD
1
GSI Helmholtzzentrum für Schwerionenforschung, Biophysics Department, Darmstadt, Germany
2
Technische Universität Darmstadt, Institut für Festkörperphysik, Darmstadt, Germany
3
Department of Radiation Oncology, Weill Cornell Medical College, New York, NY, USA

Address correspondence to: Professor Marco Durante

E-mail: ​M.​Durante@​gsi.​de

Abstract
The combination of radiotherapy and immunotherapy is one of the most promising strategies for cancer treatment.
Recent clinical results support the pre-­clinical experiments pointing to a benefit for the combined treatment in meta-
static patients. Charged particle therapy (using protons or heavier ions) is considered one of the most advanced radi-
otherapy techniques, but its cost remains higher than conventional X-­ray therapy. The most important question to be
addressed to justify a more widespread use of particle therapy is whether they can be more effective than X-­rays in
combination with immunotherapy. Protons and heavy ions have physical advantages compared to X-­rays that lead to
a reduced damage to the immune cells, that are required for an effective immune response. Moreover, densely ionizing
radiation may have biological advantages, due to different cell death pathways and release of cytokine mediators of
inflammation. We will discuss results in esophageal cancer patients showing that charged particles can reduce the
damage to blood lymphocytes compared to X-­rays, and preliminary in vitro studies pointing to an increased release
of immune-­stimulating cytokines after heavy ion exposure. Pre-­clinical and clinical studies are ongoing to test these
hypotheses.

Introduction In recent years the introduction of checkpoint inhibitors

Radiotherapy is an effective and largely used treatment
to achieve local control of solid tumors.1 Conventional
radiotherapy use X-­rays generated by electron accelerators
(linacs), but high-­ energy charged particles produced by
larger cyclotrons or synchrotrons improve precision of dose
delivery and enable to spare more normal tissue compared to
photons.2 Around 170,000 patients have been treated world-
wide with protons, and 25,000 with C-­ions (source: PTCOG,
www.​ptcog.​ch). This remains a small fraction compared to
the millions of patients treated with conventional X-­rays. The
main reason for this difference is the high cost of the particle
accelerators compared to conventional linacs.3 Currently, the
cost effectiveness of particle therapy remains under scrutiny.4
Even with high local control rates achievable with particle
therapy, this advantage fails to translate in improved cancer
survival in most cancers, because of distant metastasis.
Local radiotherapy can trigger an immune response that
can immunize the host, leading to immune destruction
of distant, unirradiated metastasis.5 This phenomenon,
described as an abscopal effect, is rare and the mechanisms
are unclear.6
has proven a breakthrough strategy, capable to induce
powerful tumor rejections and improved survival in meta-
static patients.7 Excellent results have been obtained with
immune checkpoint inhibitors such as anti-­CTLA4 and
anti-­PD1 antibodies in malignant melanoma.8 However,
severe immune-­related side-­effects complicate the use of
immunotherapy and limits its use in cancer patients.9
It is widely acknowledged that the combination of immu-
notherapy with local therapies can further improve the
survival in most solid cancers.10 Because radiation has the
potential to activate an anti tumor immune response,11 it
is an optimal candidate for combinations with immuno-
therapy.12–18 Animal experiments have shown that the
superior activity of radiation and dual immune checkpoint
blockade is mediated by non-­redundant immune mecha-
nisms in cancer.19 Following initial positive results of this
combination,20–22 hundreds of trials have been launched
to test safety and efficacy of radiation and immunotherapy
in different tumors types, including lung cancer,23 the
first cause of cancer-­ related death in United States for
both males and females.24 The PACIFIC trial has shown
 BJR
Figure 1.  Coronal and transverse images of 3D-­CRT plan (left), IMRT plan (middle), and proton plan (right) for esophageal cancer.
The plans clearly show the large tissue sparing using protons. Treatment plans produced at Loma Linda University Medical Center
(CA) details in ref.,32 reproduced with permissions.
significant improvements in disease free survival25 and overall
survival26 in stage III non-­ small-­cell lung cancer (NSCLC)
patients treated with Durvalumab after chemoradiotherapy.
Prospective randomized trials in stage IV patients are still
missing and will be essential to quantify the benefit of combined
radiotherapy and immunotherapy in lung cancer. In the pilot/
feasibility trial NCT02221739 at Weill Cornell Medicine, a
30% disease control was achieved in stage IV NSCLC patients
refractory to anti CTLA-4 alone or in combination with chemo-
therapy by combining Ipilimumab with focal hypofractionated
radiotherapy of a single metastasis.27 Several strategies are under
study to further improve these results, including modifying the
dose per fraction28 and the number of metastasis irradiated.29
Considering the success and promise of the combination of
X-­rays with checkpoint inhibitors, the question is whether
particle therapy can present additional advantages, and result
in better outcomes.30 This question is relevant to the future of
particle therapy, in view of its higher cost. We will discuss here
the potential physical and biological advantages of particle
therapy in combination with immunotherapy.
Physical advantages
Particle therapy is based on the different depth-­dose distribution
of charged particles compared to photons.31 Using protons or
heavier ions, much more normal tissue can be spared in virtu-
ally every tumor site, while preserving dose conformality on the
tumor (see e.g. Figure 1 for esophageal cancer32). Sparing of the
normal tissue, and in particular reducing the exposure of circu-
lating T-­lymphocytes and other immune cells, present advan-
tages to immunotherapy. Non-­ proliferating peripheral blood
lymphocytes are very radiosensitive33 and during radiotherapy
lymphopenia may occur, which is often correlated to a nega-
tive prognosis.34 In fact, both radiotherapy and chemotherapy
damage circulating immune cells,35 compromising the host’s
2 of 7 birpublications.org/bjr
Durante and Formenti
immunity.36 Consistently, blocking immune response in mice
prevents tumor control when radiation therapy is combined with
checkpoint immunotherapy.37
Damage to peripheral blood lymphocytes can be measured during
radiotherapy using the chromosomal aberration assay (Figure 2).
When lymphocytes are irradiated in vitro, charged particles induce
more chromosomal aberrations than X-­rays at the same dose
level, and the relative biological effectiveness (RBE) increases by
increasing the particle linear energy transfer (LET).38 However,
in vivo, the damage mainly depends on the size of the irradiated
volume (or integral dose), and on the presence of lymph nodes in
the field.39 For sites such as the esophagus, an increased field size
leads to a proportional increase of the lymph nodes exposed. In
fact, the chromosomal aberrations detected in circulating lympho-
cytes in esophageal cancer patients are proportional to the radio-
therapy target volume.40 As a result of the reduced normal tissue
volume irradiated with particles (Figure 1), less chromosomal aber-
rations were measured in patients treated with C-­ions than with
X-­rays for esophageal cancer,.41 Similar results were reported for
patients treated with C-­ions or IMRT for prostate cancer.42,43
Damaged lymphocytes eventually die and can lead to lymph-
openia. The absolute lymphocyte count decreases during the
course of radiotherapy. In sites like the esophagus, it can be
expected that the use of protons reduce s the risk of severe lymph-
openia.44 In fact, for the decrease in lymphocyte counts during
the radiotherapy course is less pronounced using protons45 or
C-­ions41 than with X-­rays (Figure 3). Even if the data in Figure 3
come from different centers and different chemotherapy drugs,
the tendency to spare more effectively lymphocytes using parti-
cles is supported for esophageal cancer patients. These results
suggest that the physical characteristics of charged particle
lead to sparing of immune cells that can then be engaged in the
immune response triggered by immunotherapy.
Br J Radiol;93:20190224
 Charged particles and immunotherapy
Figure 2.  Damage in peripheral blood lymphocytes of a patient during the course of radiotherapy. The karyotype by mFISH
shows a translocation involving chromosomes 4 and 8.
While a reduced lymphopenia is certainly an advantage for
the patients, whether lymphocyte sparing boosts the immune
response remains to be demonstrated. In esophageal cancer
Figure 3.  Median values of lymphocyte count in esophageal
cancer patients during the course of radiotherapy. Data for
protons (1.8 Gy RBE/fraction) and IMRT (1.8 Gy/fraction) are
from ref.,45 data for 3DCRT (1.6–2.0 Gy/fraction) and C-­ions
(2.7–3.6 Gy RBE/fraction) from ref.41
3 of 7 birpublications.org/bjr
BJR
patients receiving pre-­ operative chemotherapy, a significant
increases in T-­cell receptor diversity was observed in peripheral
T-­cells but not in tumor-­infiltrating lymphocytes.46 In general,
chemotherapy-­ induced depletion of immune cells can be
followed by a ‘rebound overshoot’ effect, leading to transiently
enlarged immune cell numbers, then relaxing to normality.47
Whether this transient excess of immune cells supports immuno-
therapy or not remains to be clarified, and is certainly important
to clarify the potential physical advantages of charged particles in
radioimmunotherapy.
Biological advantages
In addition to the physical advantages, charged particles have
different biological effects compared to X-­rays, caused by the
different kind of DNA lesions induced by densely ionizing radia-
tion.48 High-­LET radiation induce more clustered DNA lesions,
that are difficult to repair,49 and that trigger different DNA
damage repair signals.50–52
DNA repair signaling pathways are strongly related to the
immune response. Recently, it has been directly shown that that
PD-­L1 expression in cancer cells is upregulated in response to
DNA double-­ strand breakage, through the ATM/ATR/Chk1
kinase pathway.53 Similar upregulation of PD-­ L1 has been
Br J Radiol;93:20190224
 BJR
Figure 4.  An illustration of immune-­ mediated effects of
ionizing radiation. The green arrow points to the release of
HGMB1, that interacts with the toll-­like receptor TLR4 activat-
ing the dendritic cell maturation. Cartoon from ref.,60 repro-
duced with permission.
recently shown in melanoma cells exposed to UV radiation.54
Activation of different DNA damage response pathways at
high-­LET, such as resection,55 may have different effects on the
expression of immune receptors. It presently not known whether
the radiation-­induced upregulation of PD-­L1 will actually trans-
late into response to checkpoint inhibitors.56
DNA damage eventually leads to cell death through different
pathways (such as apoptosis, necrosis, mitotic catastrophe or
senescence),57,58 and to the consequent release of small mole-
cules such as ATP, calreticulin, and HMGB159 that can trigger
the immune response (Figure 4).60 In vitro studies with different
human tumor cell lines have shown that proton radiation induces
calreticulin cell-­surface expression, increasing sensitivity to
cytotoxic T-­lymphocyte killing of tumor cells.61 Increased extra-
cellular concentration of HMGB1 has been measured after irra-
diation of human cancer cells with heavy ions,62 and the release
is positively correlated to the particle LET (Figure 5). HMGB1 is
upregulated in the serum of esophageal cancer patients following
chemoradiotherapy,63 and these results are therefore suggestive
of an increased efficiency of densely ionizing radiation compared
to X-­rays.
In addition to the damage to nuclear DNA, recent evidence points
to the sensing of cytoplasmic DNA as a key factor in eliciting
4 of 7 birpublications.org/bjr
Durante and Formenti
Figure 5.  Concentration of HGMB1 in the medium following
irradiation of HeLa cells with sham (0 Gy) radiation (black),
4 Gy C-­ions in the plateau region of the Bragg curve, 13 keV/
μm (red) and 2 Gy C-­ions in the spread-­out-­Bragg-­peak, 70
keV/μm (green). The two doses were chosen to achieve the
same 10% survival in irradiated HeLa cells. Data from ref.62
immune response.64 Double stranded DNA (dsDNA) fragments
generated by exposure to sparsely ionizing radiation are extruded
from the nucleus65,66 and accumulate in the cytosol where
dsDNA sensors, cGAS/STING are activated to transcribe type I
interferon genes and jumpstart an immune response. Interferon
is a powerful promoter of dendritic cells recruitment and activa-
tion, enabling cross-­presentation of radiation induced neoanti-
gens and immune response.67,68 Consistently, metastatic NSCLC
patients responding to focal radiation and CTLA4 blockade
demonstrated an increased serum level of interferon-βcompared
to baseline.27 These results suggest that the cGAS/STING activa-
tion may be an essential pathway for a successful combination
of radiation and immunotherapy.69,70 Increased cytosolic DNA
is observed after irradiation of human cancer cells, but at high
doses per fraction (>10 Gy) cells activate the DNA exonuclease
Trex1, that degrades cytosolic DNA thus blocking the STING
activation.71 Type-­I interferon activation induced by radiation
therefore reflects a balance between sufficient dsDNA induc-
tion to stimulate cGAS/STING and Trex1 activation.72 How this
pathway is affected by charged particles remains unknown, and
experiments are needed to clarify whether the production of
smaller DNA fragments by densely ionizing radiation can lead to
an enhanced response.73
Preliminary animal studies have shown that charged particles
effectively induce abscopal responses for instance by reducing
lung metastasis after irradiation of the primary tumor.74–76
However, these results remain inconclusive and a direct
comparison of X-­rays, protons, and C-­ions is warranted.77
Conclusions
The potential advantages in using protons or heavy ions in combi-
nation with immunotherapy are based on both their physical and
biological properties, warranting experimental verification. Since
many pre-­clinical and clinical trials are ongoing, it is expected that
within a few years the role of charged particles for a combined use
Br J Radiol;93:20190224
 Charged particles and immunotherapy BJR

with modern immunotherapy of cancer will be defined. Several
new proton therapy centers are under construction in Europe,
and many of these centers will dedicate “beamtime” to radio-
biology research.78 Often these research efforts are directed to
measurements of proton RBE,79 but one of the key questions to be
answered is whether particles are more effective than X-­rays when
used in combination with immunotherapy.30 Considering the high
cost of the proton or heavy ion therapy facilities and the success
of radioimmunotherapy trials, this research is perhaps the most
important to decide the future of particle therapy.
Acknowledgments
This work was partly supported by EU Horizon2020 grant
73,0983 (INSPIRE). The research in radioimmunotherapy at GSI
is in the frame of FAIR Phase-0 supported by the GSI Helmholtz-
zentrum für Schwerionenforschung in Darmstadt (Germany).

References
1. Baumann M, Krause M, Overgaard J, Debus
J, Bentzen SM, Daartz J, et al. Radiation
oncology in the era of precision medicine.
Nat Rev Cancer 2016; 16: 234–49. doi:
https://​doi.​org/​10.​1038/​nrc.​2016.​18
2. Durante M, Orecchia R, Loeffler JS.
Charged-­Particle therapy in cancer: clinical
uses and future perspectives. Nat Rev Clin
Oncol 2017; 14: 483–95. doi: https://​doi.​org/​
10.​1038/​nrclinonc.​2017.​30
3. Bortfeld TR, Loeffler JS. Three ways to make
proton therapy affordable. Nature 2017; 549:
451–3. doi: https://​doi.​org/​10.​1038/​549451a
4. Mitin T, Zietman AL. Promise and pitfalls
of heavy-­particle therapy. JCO 2014; 32:
2855–63. doi: https://​doi.​org/​10.​1200/​JCO.​
2014.​55.​1945
5. Demaria S, Bhardwaj N, McBride WH,
Formenti SC. Combining radiotherapy and
immunotherapy: a revived partnership. Int J
Radiat Oncol Biol Phys 2005; 63: 655–66. doi:
https://​doi.​org/​10.​1016/​j.​ijrobp.​2005.​06.​032
6. Abuodeh Y, Venkat P, Kim S. Systematic
review of case reports on the abscopal effect.
Curr Probl Cancer 2016; 40: 25–37. doi:
https://​doi.​org/​10.​1016/​j.​currproblcancer.​
2015.​10.​001
7. Kirkwood JM, Butterfield LH, Tarhini
AA, Zarour H, Kalinski P, Ferrone S.
Immunotherapy of cancer in 2012. CA
Cancer J Clin 2012; 62: 309–35. doi: https://​
doi.​org/​10.​3322/​caac.​20132
8. Robert C, Schachter J, Long GV, Arance A,
Grob JJ, Mortier L, et al. Pembrolizumab
versus ipilimumab in advanced melanoma. N
Engl J Med 2015; 372: 2521–32. doi: https://​
doi.​org/​10.​1056/​NEJMoa1503093
9. Postow MA, Sidlow R, Hellmann MD,
Longo DL. Immune-­Related adverse
events associated with immune checkpoint
blockade. N Engl J Med(eds 2018; 378:
158–68. doi: https://​doi.​org/​10.​1056/​
NEJMra1703481
10. Whiteside TL, Demaria S, Rodriguez-­Ruiz
ME, Zarour HM, Melero I. Emerging
opportunities and challenges in cancer
immunotherapy. Clin Cancer Res 2016; 22:
1845–55. doi: https://​doi.​org/​10.​1158/​1078-​
0432.​CCR-​16-​0049
11. Formenti SC, Demaria S. Systemic effects of
local radiotherapy. Lancet Oncol 2009; 10:
718–26. doi: https://​doi.​org/​10.​1016/​S1470-​
2045(​09)​70082-8
12. Ngwa W, Irabor OC, Schoenfeld JD,
Hesser J, Demaria S, Formenti SC. Using
immunotherapy to boost the abscopal
effect. Nat Rev Cancer 2018; 18: 313–22. doi:
https://​doi.​org/​10.​1038/​nrc.​2018.6
13. Demaria S, Golden EB, Formenti SC.
Role of local radiation therapy in cancer
immunotherapy. JAMA Oncol 2015; 1: 1325–
32. doi: https://​doi.​org/​10.​1001/​jamaoncol.​
2015.​2756
14. Formenti SC, Demaria S. Combining
radiotherapy and cancer immunotherapy: a
paradigm shift. J Natl Cancer Inst 2013; 105:
256–65. doi: https://​doi.​org/​10.​1093/​jnci/​
djs629
15. Weichselbaum RR, Liang H, Deng L, Fu
Y-­X. Radiotherapy and immunotherapy: a
beneficial liaison? Nat Rev Clin Oncol 2017;
14: 365–79. doi: https://​doi.​org/​10.​1038/​
nrclinonc.​2016.​211
16. Durante M, Reppingen N, Held KD.
Immunologically augmented cancer
treatment using modern radiotherapy. Trends
Mol Med 2013; 19: 565–82. doi: https://​doi.​
org/​10.​1016/​j.​molmed.​2013.​05.​007
17. Van Limbergen EJ, De Ruysscher DK,
Olivo Pimentel V, Marcus D, Berbee M,
Hoeben A, et al. Combining radiotherapy
with immunotherapy: the past, the present
and the future. Br J Radiol 2017; 90:
20170157. doi: https://​doi.​org/​10.​1259/​bjr.​
20170157
18. Herrera FG, Bourhis J, Coukos G.
Radiotherapy combination opportunities
Leveraging immunity for the next oncology
practice. CA Cancer J Clin 2017; 67: 65–85.
doi: https://​doi.​org/​10.​3322/​caac.​21358
19. Twyman-­Saint Victor C, Rech AJ, Maity
A, Rengan R, Pauken KE, Stelekati E,
et al. Radiation and dual checkpoint
blockade activate non-­redundant immune
mechanisms in cancer. Nature 2015;
520: 373–7. doi: https://​doi.​org/​10.​1038/​
nature14292
20. Postow MA, Callahan MK, Barker
CA, Yamada Y, Yuan J, Kitano S, et al.
Immunologic correlates of the abscopal effect
in a patient with melanoma. N Engl J Med
2012; 366: 925–31. doi: https://​doi.​org/​10.​
1056/​NEJMoa1112824
21. Hiniker SM, Chen DS, Knox SJ. Abscopal
effect in a patient with melanoma. N Engl J
Med 2012; 366: 2035–6. doi: https://​doi.​org/​
10.​1056/​NEJMc1203984
22. Golden EB, Chhabra A, Chachoua A,
Adams S, Donach M, Fenton-­Kerimian M,
et al. Local radiotherapy and granulocyte-­
macrophage colony-­stimulating factor to
generate abscopal responses in patients
with metastatic solid tumours: a proof-­
of-­principle trial. Lancet Oncol 2015; 16:
795–803. doi: https://​doi.​org/​10.​1016/​S1470-​
2045(​15)​00054-6
23. Ko EC, Raben D, Formenti SC. The
integration of radiotherapy with
immunotherapy for the treatment of non-­
small cell lung cancer. Clin Cancer Res 2018;
24: 5792–806. doi: https://​doi.​org/​10.​1158/​
1078-​0432.​CCR-​17-​3620
24. Siegel RL, Miller KD, Jemal A. Cancer
statistics, 2019. CA Cancer J Clin 2019; 69:
7–34. doi: https://​doi.​org/​10.​3322/​caac.​21551
25. Antonia SJ, Villegas A, Daniel D, Vicente
D, Murakami S, Hui R, et al. Durvalumab
after chemoradiotherapy in stage III non-­
small-­cell lung cancer. N Engl J Med 2017;
377: 1919–29. doi: https://​doi.​org/​10.​1056/​
NEJMoa1709937
26. Antonia SJ, Villegas A, Daniel D, Vicente D,
Murakami S, Hui R, et al. Overall survival
with Durvalumab after chemoradiotherapy
in stage III NSCLC. N Engl J Med 2018;
379: 2342–50. doi: https://​doi.​org/​10.​1056/​
NEJMoa1809697
27. Formenti SC, Rudqvist N-­P, Golden E,
Cooper B, Wennerberg E, Lhuillier C, et al.
Radiotherapy induces responses of lung
cancer to CTLA-4 blockade. Nat Med 2018;

5 of 7 birpublications.org/bjr Br J Radiol;93:20190224
 BJR
24: 1845–51. doi: https://​doi.​org/​10.​1038/​
s41591-​018-​0232-2
28. Formenti SC. Optimizing dose per fraction:
a new chapter in the story of the Abscopal
effect? Int J Radiat Oncol Biol Phys 2017; 99:
677–9. doi: https://​doi.​org/​10.​1016/​j.​ijrobp.​
2017.​07.​028
29. Brooks ED, Chang JY. Time to abandon
single-­site irradiation for inducing abscopal
effects. Nat Rev Clin Oncol 2019; 16: 123–35.
doi: https://​doi.​org/​10.​1038/​s41571-​018-​
0119-7
30. Durante M, Brenner DJ, Formenti SC. Does
heavy ion therapy work through the immune
system? Int J Radiat Oncol Biol Phys 2016; 96:
934–6. doi: https://​doi.​org/​10.​1016/​j.​ijrobp.​
2016.​08.​037
31. Durante M, Paganetti H. Nuclear physics
in particle therapy: a review. Rep Prog Phys
2016; 79: 096702. doi: https://​doi.​org/​10.​
1088/​0034-​4885/​79/​9/​096702
32. Ling TC, Slater JM, Nookala P, Mifflin R,
Grove R, Ly AMGrove R, Patyal B, Yang
G, et al. Analysis of intensity-­modulated
radiation therapy (IMRT), proton and 3D
conformal radiotherapy (3D-­CRT) for
reducing perioperative cardiopulmonary
complications in esophageal cancer patients.
Cancers 2014; 6: 2356–68. doi: https://​doi.​
org/​10.​3390/​cancers6042356
33. Heylmann D, Badura J, Becker H, Fahrer J,
Kaina B. Sensitivity of CD3/CD28-­stimulated
versus non-­stimulated lymphocytes to
ionizing radiation and genotoxic anticancer
drugs: key role of ATM in the differential
radiation response. Cell Death Dis 2018; 9:
1053. doi: https://​doi.​org/​10.​1038/​s41419-​
018-​1095-7
34. Venkatesulu BP, Mallick S, Lin SH, Krishnan
S. A systematic review of the influence of
radiation-­induced lymphopenia on survival
outcomes in solid tumors. Crit Rev Oncol
Hematol 2018; 123(January): 42–51. doi:
https://​doi.​org/​10.​1016/​j.​critrevonc.​2018.​01.​
003
35. Yovino S, Kleinberg L, Grossman SA,
Narayanan M, Ford E. The etiology of
treatment-­related lymphopenia in patients
with malignant gliomas: modeling radiation
dose to circulating lymphocytes explains
clinical observations and suggests methods
of modifying the impact of radiation on
immune cells. Cancer Invest 2013; 31: 140–4.
doi: https://​doi.​org/​10.​3109/​07357907.​2012.​
762780
36. Lobachevsky PN, Ventura J,
Giannakandropoulou L, Forrester H,
Palazzolo JS, Haynes NM, et al. A functional
immune system is required for the systemic
genotoxic effects of localized irradiation. Int
J Radiat Oncol Biol Phys 2019; 103: 1184–93.
6 of 7 birpublications.org/bjr
doi: https://​doi.​org/​10.​1016/​j.​ijrobp.​2018.​11.​
066
37. Crittenden MR, Zebertavage L, Kramer
G, Bambina S, Friedman D, Troesch V,
et al. Tumor cure by radiation therapy and
checkpoint inhibitors depends on pre-­
existing immunity . Sci Rep 2018; 8: 7012.
doi: https://​doi.​org/​10.​1038/​s41598-​018-​
25482-w
38. Ritter S, Durante M. Heavy-­Ion induced
chromosomal aberrations: a review. Mutat
Res 2010; 701: 38–46. doi: https://​doi.​org/​10.​
1016/​j.​mrgentox.​2010.​04.​007
39. d'Alesio V, Pacelli R, Durante M,
Canale Cama G, Cella L, Gialanella G, et al.
Lymph nodes in the irradiated field influence
the yield of radiation-­induced chromosomal
aberrations in lymphocytes from breast
cancer patients. Int J Radiat Oncol Biol Phys
2003; 57: 732–8. doi: https://​doi.​org/​10.​1016/​
S0360-​3016(​03)​00664-3
40. Durante M, Yamada S, Ando K, Furusawa
Y, Kawata T, Majima H, et al. Measurements
of the equivalent whole-­body dose during
radiation therapy by cytogenetic methods.
Phys Med Biol 1999; 44: 1289–98. doi: https://​
doi.​org/​10.​1088/​0031-​9155/​44/​5/​314
41. Durante M, Yamada S, Ando K, Furusawa
Y, Kawata T, Majima H, et al. X-­Rays vs.
carbon-­ion tumor therapy: cytogenetic
damage in lymphocytes. Int J Radiat Oncol
Biol Phys 2000; 47: 793–8. doi: https://​doi.​
org/​10.​1016/​S0360-​3016(​00)​00455-7
42. Hartel C, Nikoghosyan A, Durante M,
Sommer S, Nasonova E, Fournier C, et al.
Chromosomal aberrations in peripheral
blood lymphocytes of prostate cancer
patients treated with IMRT and carbon ions.
Radiother Oncol 2010; 95: 73–8. doi: https://​
doi.​org/​10.​1016/​j.​radonc.​2009.​08.​031
43. Pignalosa D, Lee R, Hartel C, Sommer S,
Nikoghosyan A, Debus J, et al. Chromosome
inversions in lymphocytes of prostate cancer
patients treated with x-­rays and carbon ions.
Radiother Oncol 2013; 109: 256–61. doi:
https://​doi.​org/​10.​1016/​j.​radonc.​2013.​09.​021
44. Fang P, Shiraishi Y, Verma V, Jiang W, Song
J, Hobbs BP, et al. Lymphocyte-­Sparing
effect of proton therapy in patients with
esophageal cancer treated with definitive
chemoradiation. Int J Part Ther 2017; 4:
23–32. doi: https://​doi.​org/​10.​14338/​IJPT-​17-​
00033.1
45. Davuluri R, Jiang W, Fang P, Xu C, Komaki
R, Gomez DR, et al. Lymphocyte nadir and
esophageal cancer survival outcomes after
chemoradiation therapy. Int J Radiat Oncol
Biol Phys 2017; 99: 128–35. doi: https://​doi.​
org/​10.​1016/​j.​ijrobp.​2017.​05.​037
46. Zhang C, Palashati H, Tan Q, Ku W, Miao
Y, Xiong H, et al. Immediate and substantial
Durante and Formenti
evolution of T-­cell repertoire in peripheral
blood and tumor microenvironment of
patients with esophageal squamous cell
carcinoma treated with preoperative
chemotherapy. Carcinogenesis 2018; 39:
1389–98. doi: https://​doi.​org/​10.​1093/​carcin/​
bgy116
47. Proietti E, Moschella F, Capone I, Belardelli
F. Exploitation of the propulsive force of
chemotherapy for improving the response to
cancer immunotherapy. Mol Oncol 2012; 6:
1–14. doi: https://​doi.​org/​10.​1016/​j.​molonc.​
2011.​11.​005
48. Durante M. New challenges in high-­energy
particle radiobiology. Br J Radiol 2014; 87:
20130626. doi: https://​doi.​org/​10.​1259/​bjr.​
20130626
49. Asaithamby A, Hu B, Chen DJ. Unrepaired
clustered DNA lesions induce chromosome
breakage in human cells. Proc Natl Acad Sci
U S A 2011; 108: 8293–8. doi: https://​doi.​org/​
10.​1073/​pnas.​1016045108
50. Löbrich M, Jeggo PA. The impact of a
negligent G2/M checkpoint on genomic
instability and cancer induction. Nat Rev
Cancer 2007; 7: 861–9. doi: https://​doi.​org/​
10.​1038/​nrc2248
51. Sage E, Harrison L. Clustered DNA
lesion repair in eukaryotes: relevance to
mutagenesis and cell survival. Mutat Res
2011; 711(1–2): 123–33. doi: https://​doi.​org/​
10.​1016/​j.​mrfmmm.​2010.​12.​010
52. Bukowska B, Karwowski BT. The Clustered
DNA Lesions - Types, Pathways of Repair
and Relevance to Human Health. Curr Med
Chem 2018; 25: 2722–35. doi: https://​doi.​org/​
10.​2174/​0929​8673​2566​6180​2261​10502
53. Sato H, Niimi A, Yasuhara T, Permata TBM,
Hagiwara Y, Isono M, et al. Dna double-­
strand break repair pathway regulates PD-­L1
expression in cancer cells. Nat Commun
2017; 8. doi: https://​doi.​org/​10.​1038/​s41467-​
017-​01883-9
54. Wang W, Chapman NM, Zhang B, Li M,
Fan M, Laribee RN, et al. Upregulation of
PD-­L1 via HMGB1-­Activated IRF3 and
NF-κB contributes to UV radiation-­induced
immune suppression. Cancer Res 2019; 79:
2909–22. doi: https://​doi.​org/​10.​1158/​0008-​
5472.​CAN-​18-​3134
55. Averbeck NB, Ringel O, Herrlitz M, Jakob
B, Durante M, Taucher-­Scholz G. Dna end
resection is needed for the repair of complex
lesions in G1-­phase human cells. Cell Cycle
2014; 13: 2509–16. doi: https://​doi.​org/​10.​
4161/​15384101.​2015.​941743
56. Shevtsov M, Sato H, Multhoff G, Shibata A.
Novel approaches to improve the efficacy of
Immuno-­Radiotherapy. Front Oncol 2019; 9:
156. doi: https://​doi.​org/​10.​3389/​fonc.​2019.​
00156
Br J Radiol;93:20190224
 Charged particles and immunotherapy
57. Paludan SR, Reinert LS, Hornung V. Dna-­
Stimulated cell death: implications for host
defence, inflammatory diseases and cancer.
Nat Rev Immunol 2019; 19: 141–53. doi:
https://​doi.​org/​10.​1038/​s41577-​018-​0117-0
58. Vitale I, Galluzzi L, Castedo M, Kroemer
G. Mitotic catastrophe: a mechanism for
avoiding genomic instability. Nat Rev Mol
Cell Biol 2011; 12: 385–92. doi: https://​doi.​
org/​10.​1038/​nrm3115
59. Lotze MT, Tracey KJ. High-­Mobility group
box 1 protein (HMGB1): nuclear weapon
in the immune arsenal. Nat Rev Immunol
2005; 5: 331–42. doi: https://​doi.​org/​10.​1038/​
nri1594
60. Ebner DK, Tinganelli W, Helm A, Bisio
A, Yamada S, Kamada T, et al. The
immunoregulatory potential of particle
radiation in cancer therapy. Front Immunol
2017; 8: 99. doi: https://​doi.​org/​10.​3389/​
fimmu.​2017.​00099
61. Gameiro SR, Malamas AS, Bernstein MB,
Tsang KY, Vassantachart A, Sahoo N, et al.
Tumor cells surviving exposure to proton
or photon radiation share a common
immunogenic modulation signature,
rendering them more sensitive to T cell-­
mediated killing. Int J Radiat Oncol Biol Phys
2016; 95: 120–30. doi: https://​doi.​org/​10.​
1016/​j.​ijrobp.​2016.​02.​022
62. Onishi M, Okonogi N, Oike T, Yoshimoto
Y, Sato H, Suzuki Y, et al. High linear
energy transfer carbon-­ion irradiation
increases the release of the immune
mediator high mobility group box 1 from
human cancer cells. J Radiat Res 2018; 59:
541–6. doi: https://​doi.​org/​10.​1093/​jrr/​
rry049
63. Suzuki Y, Mimura K, Yoshimoto Y,
Watanabe M, Ohkubo Y, Izawa S, et al.
Immunogenic tumor cell death induced
by chemoradiotherapy in patients with
esophageal squamous cell carcinoma. Cancer
Res 2012; 72: 3967–76. doi: https://​doi.​org/​
10.​1158/​0008-​5472.​CAN-​12-​0851
7 of 7 birpublications.org/bjr
64. Corrales L, Matson V, Flood B, Spranger S,
Gajewski TF. Innate immune signaling and
regulation in cancer immunotherapy. Cell
Res 2017; 27: 96–108. doi: https://​doi.​org/​10.​
1038/​cr.​2016.​149
65. Mackenzie KJ, Carroll P, Martin C-­A, Murina
O, Fluteau A, Simpson DJ, et al. cGAS
surveillance of micronuclei links genome
instability to innate immunity. Nature 2017;
548: 461–5. doi: https://​doi.​org/​10.​1038/​
nature23449
66. Harding SM, Benci JL, Irianto J, Discher DE,
Minn AJ, Greenberg RA. Mitotic progression
following DNA damage enables pattern
recognition within micronuclei. Nature 2017;
548: 466–70. doi: https://​doi.​org/​10.​1038/​
nature23470
67. Chen Q, Sun L, Chen ZJ. Regulation and
function of the cGAS-­STING pathway of
cytosolic DNA sensing. Nat Immunol 2016;
17: 1142–9. doi: https://​doi.​org/​10.​1038/​ni.​
3558
68. Zitvogel L, Galluzzi L, Kepp O, Smyth MJ,
Kroemer G. Type I interferons in anticancer
immunity. Nat Rev Immunol 2015; 15:
405–14. doi: https://​doi.​org/​10.​1038/​nri3845
69. Galluzzi L, Vanpouille-­Box C, Bakhoum
SF, Demaria S. Snapshot: cGAS-­STING
signaling. Cell 2018; 173: 276–276.e1. doi:
https://​doi.​org/​10.​1016/​j.​cell.​2018.​03.​015
70. Vanpouille-­Box C, Demaria S, Formenti
SC, Galluzzi L. Cytosolic DNA sensing in
organismal tumor control. Cancer Cell 2018;
34: 361–78. doi: https://​doi.​org/​10.​1016/​j.​
ccell.​2018.​05.​013
71. Vanpouille-­Box C, Alard A, Aryankalayil
MJ, Sarfraz Y, Diamond JM, Schneider
RJ, et al. Dna exonuclease TREX1
regulates radiotherapy-­induced tumour
immunogenicity. Nat Commun 2017;
8: 15618. doi: https://​doi.​org/​10.​1038/​
ncomms15618
72. Vanpouille-­Box C, Formenti SC, Demaria S.
Trex1 dictates the immune fate of irradiated
cancer cells. Oncoimmunology 2017; 6:
BJR
e1339857. doi: https://​doi.​org/​10.​1080/​
2162402X.​2017.​1339857
73. Durante M, Formenti SC. Radiation-­
Induced chromosomal aberrations and
immunotherapy: micronuclei, cytosolic
DNA, and Interferon-­Production pathway.
Front Oncol 2018; 8: 192. doi: https://​doi.​org/​
10.​3389/​fonc.​2018.​00192
74. Shimokawa T, Ma L, Ando K, Sato K, Imai
T. The future of combining Carbon-­Ion
radiotherapy with immunotherapy: evidence
and progress in mouse models. Int J Part
Ther 2016; 3: 61–70. doi: https://​doi.​org/​10.​
14338/​IJPT-​15-​00023.1
75. Ando K, Fujita H, Hosoi A, Ma L,
Wakatsuki M, Seino K-­I, et al. Intravenous
dendritic cell administration enhances
suppression of lung metastasis induced by
carbon-­ion irradiation. J Radiat Res 2017;
58: 446–55. doi: https://​doi.​org/​10.​1093/​jrr/​
rrx005
76. Sato K, Nitta N, Aoki I, Imai T, Shimokawa
T. Repeated photon and C-­ion irradiations
in vivo have different impact on alteration of
tumor characteristics. Sci Rep 2018; 8: 1458.
doi: https://​doi.​org/​10.​1038/​s41598-​018-​
19422-x
77. Ebner DK, Tinganelli W, Helm A, Bisio A,
Simoniello P, Natale F, et al. Generating and
grading the abscopal effect: proposal for
comprehensive evaluation of combination
immunoradiotherapy in mouse models.
Transl Cancer Res 2017; 6(S5): S892–S899.
doi: https://​doi.​org/​10.​21037/​tcr.​2017.​06.​01
78. Durante M. Proton beam therapy in Europe:
more centres need more research. Br J Cancer
2019; ; 120: 777–8. doi: https://​doi.​org/​10.​
1038/​s41416-​018-​0329-x
79. Paganetti H. Relative biological effectiveness
(RBE) values for proton beam therapy.
variations as a function of biological
endpoint, dose, and linear energy transfer.
Phys Med Biol 2014; 59: R419–R472. doi:
https://​doi.​org/​10.​1088/​0031-​9155/​59/​22/​
R419
Br J Radiol;93:20190224