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Nearly all DNA lesions block DNA replication, although some can be bypassed by error-prone translesion
[18]
DNA polymerases . The ability of cells to manage this remarkable daily lesion load is a reflection of
the high efficiency of DNA repair systems. That said, DNA damage can cause mutations, chromosome
structural alterations, cell cycle arrest, senescence, and cell death. Among the hundreds of types of DNA
lesions, DSBs are among the most cytotoxic, and the cytotoxicity of genotoxic chemicals and ionizing
radiation is largely due to DSBs [19,20] . Other double-strand lesions, such as inter-strand crosslinks, are also
[21]
highly cytotoxic .
Cells respond to DNA damage by activating checkpoint signaling and DNA repair pathways, collectively
termed the DDR. DDR promotes cell survival and suppresses cancer by promoting genome stability,
but it also triggers programmed cell death when damage is excessive. Altered expression or mutation of
DDR proteins predispose to cancer, determine tumor response to chemo- and radiotherapy, and underlie
several congenital conditions including multiple types of Seckel syndrome, primordial dwarfism, and
premature aging syndromes [22-24] . The DDR is a major determinant of cancer cell responses to chemo- and
radiotherapy, and is thus an enticing target to augment cancer therapy [25-30] . DDR components are often
defective in cancer, but because the DDR is a complex network of interacting/cross-talking pathways, cells
can respond to alterations in one pathway with compensatory changes in other pathways. Compensatory
pathways within the DDR network represent formidable obstacles to successful cancer treatment. A better
understanding of DDR pathways can reveal synthetic lethal relationships that can be exploited to augment
cancer therapy in general, and to develop personalized therapies [31-35] .

The DDR includes two checkpoint signaling pathways, one centered on ataxia telangiectasia mutated
(ATM), a kinase that responds to DSBs and one centered on ataxia telangiectasia and Rad3 related (ATR)
kinase that is triggered by single-stranded DNA (ssDNA) generated by 5’-3’ resection of DSB ends and
by decoupling of the replication machinery from MCM helicase at stalled replication forks [36-39] . ATM and
ATR, along with DNA-PKcs, are PI3 kinase-like kinases (PIKKs) that are “early responders” to DSBs and
replication stress. PIKKs phosphorylate large networks of proteins [40-42] including the downstream effector
kinases Chk1 and Chk2 that phosphorylate p53 and other targets to arrest the cell cycle in response to
damage, promote DNA repair, and promote programmed cell death pathways when damage exceeds a
threshold [43-46] [Figure 1]. The DDR thus presents two broad targets to manipulate for therapeutic gain:
inhibiting DNA repair sensitizes cells to damage and inhibiting checkpoint signaling prevents cell cycle
arrest in response to damage, increasing replication stress, fork collapse to DSBs, genome instability, and
cell death [20,47-50] .

DSBs are repaired by error-prone non-homologous end-joining (NHEJ) or by homologous recombination
(HR) repair [Figure 2] [51,52] , templated from sister chromatids (restricted to S/G2 phases), homologous
chromosomes, or short sequence repeats if the double-strand damage occurs within or nearby repeated
sequences - not uncommon given the human genome comprises > 50% repetitive elements (Alu, MIRs,
[53]
SINEs, LINEs, etc.) . HR is generally accurate, but it does pose risks of genome rearrangements
including large-scale loss of heterozygosity and translocations that can initiate tumorigenesis and drive
tumor progression [27,54,55] . When the primary NHEJ or HR pathways fail, even more error-prone DSB
repair pathways serve as back-up, including alternative (microhomology-mediated) NHEJ, single-strand
annealing, and break-induced replication [56-62] .

RADIOBIOLOGICAL PROPERTIES OF THERAPEUTIC IONIZING RADIATION
Three types of external beam radiation are used to treat cancer. X-rays and protons are low linear energy
transfer (LET) radiation, although proton LET varies (see below). LET is a measure of ionization density,
thus low LET X-rays (and protons for the most part) are sparsely ionizing. This means that most X-ray
lesions, including DSBs, are widely dispersed. X-rays are massless photons that interact weakly with