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We are told in GCSE that mutations and changes to the structure of a nucleotide is so harmful if not dealt with, these various lesion damages being caused by x-rays and whatnot. If so, then this begs the question of why they are so harmful (and how does our body ensure that most, if not all, of these damages are reversed or the effects mitigated,) but more importantly how current developments will help to bring us closer to ‘curing cancer.’ Exploring these three key subtopics will enable one to more clearly understand why this subject is of the utmost importance and thus currently regarded as a vital field of research in the biochemical world.

Considering the necessity for gene transcription and mitosis in body metabolism, the provision of damage-free nucleotides is of the utmost importance. Of course, this is unrealistic, as DNA damage sometimes are unrecognized or irreparable within time constraints (mitosis occurs at such a high rate). Damages either affect the physical structure of the nucleotide (this particularly harmful in slowly or non-dividing cells) or alter and mismatch the bases themselves – this is what bone marrow tissue and other rapidly-dividing cells suffer from. If the latter damage is not recognized before mitosis, it becomes a mutation – repair mechanisms can no longer recognize this damage. This eventually becomes what we know as a tumor, which could then induce various cancers. Currently, various techniques exist to either stimulate the DNA repair mechanisms or the tumor cells themselves, from balanced diets to Boron neutron capture therapy. While each of these mechanisms benefit in some sense, current they all are inefficient, solely mitigate the issue, or cause undesirable side effects.

Lesion Damage:

As replication and transcription occurs at such a high rate, errors and thus DNA lesion damages are inevitable. For example, oxidative damage occurs around 11,500 times a day in humans, this number reaching the 100,000s in rats![1] Fortunately, damage is not always harmful, and rarely fatal to an organism. Yet when it is, DNA damage can lead to mutagenesis, carcinogenesis (formation of tumors) and cell ageing in non-replicating cells – this is where the cell has a progressive decline in resistance to future damages, eventually resulting in cell death (cell-ageing is thus the cause of most age-related diseases)[2] DNA damages include breaks in strands, missing bases in backbones, or nucleotide components being chemically changed – for example the increase of 8-oxo-2’ deoxyguanosine (8-OHdG) in nucleotide sequences.[3] We can classify DNA damage into two main subtopics – endogenous damage and exogenous damage.

Endogenous damage can be defined as naturally-occurring errors, whether during metabolic or hydrolytic (involving breakdown by a reaction with water) processes. These include depurinations, depyrimidinations, cytosine deamination and oxidative damage. Note that deamination of cytosine produces uracil, which isn’t possible in DNA (for various reasons) and is thus cleaved by glycosylases.[4] This would mean at a particular locus one base is missing – which may well be fatal if the sequence is responsible for protein primary structure formation. However, the most well-known and frequent endogenous lesion is oxidative damage, which will be explored further. This is where there is an overall addition of an oxygen atom to a base, changing its structure without changing the base itself. In mitochondria, Oxidative phosphorylationoccurs as a metabolic reaction where Oxygen is reduced to Water by gaining 4 electrons. However, as some electrons are leaked from the transport chain, the reduction is actually partial. These electrons will bind with other Oxygen atoms, forming ROS free radicals which will then oxidize a particular base.[5] As Guanine has the highest oxidation potential out of the 4 bases – this is often the base that is oxidized – it forms 8-hydroxyguanine or C5H5N5O2.[6]

Figure 1 – Diagram of oxidative damage to Guanine. Observe the change of the Hydrogen in the Imidazole ring to a hydroxide as it gains an Oxygen atom. Also note that the structure of 8-hydroxyguanine here is slightly simplified.

On the other hand, external agents can also damage the nucleotide structure and/or composition. Ultraviolet light causes linkage between Cytosine and Thymine (which we know isn’t normal), Ionizing radiation breaks the hydrogen bonds and strands linking consecutive or complementary bases, and industrial chemicals such as vinyl chloride or soot can cause a variety of lesions (damages) including oxidation, crosslinking and formation of ethenobases[7] – the latter a type of DNA adduct, meaning that normal strands of DNA are bounded to human carcinogens. This is what is explored in GCSE, although the particular lesion damage clearly varies with each damaging agent and thus treatment must be specified to combat an overdose or effect.

Self-induced DNA repair mechanisms as a form of treatment occur throughout the cell’s life cycle, but also right after replication. Whilst after replication, proofreading and mismatch repair mechanisms serve to detect errors in the ‘nicks’ of the newly synthesized DNA (for eukaryotes) and thus correct the nucleotide[8], the most interesting and debatably important mechanisms combat damage accumulation during the lifetime of a cell. The three main types are Direct reversal, Single-stranded repair and Double-stranded repair. Direct reversal is most energy efficient as it only involves reversing the lesion-causing reaction as supposed to introducing any new nucleotides (see below), yet it only works for very specific types of lesion damage. For example, a photoreactivation reaction can reverse some pyrimidine dimerization damages – each reaction is specific to a lesion[9]. As this is a very intuitive repair mechanism, we now look at the other two methods, which are much more versatile and can deal with a variety of lesion damages.

a) Single-stranded repair:

As per the name, this is where only one strand of the double helix is defected. Repair mechanisms are reliant on the complementary strand being used as a template for correction (much like mRNA in protein synthesis). For errors in only one base, the BER (Base excision repair) mechanism is used. This is useful in damages like deamination or oxidation, which are some of the most frequent lesions (see above). Glycosylase enzymes remove the damaged base by cleaving the N-glycosidic bond, leaving an AP site (no purine or pyrimidine base at the locus, but adjacent bases unaffected). Then, the AP endonuclease enzyme removes a certain number of bases, depending on whether a short-patch or long-patch repair is desired. (choice mechanism currently unknown)[10] Leaving a 5’ phosphate end and a 3’ hydroxyl end, the corrected base can now be inserted with the help of Pol β (A type of DNA polymerase).[11] Depending on the patch length, DNA ligase or Flap endonucleases are used to attach 1 or 2-10 nucleotides respectively.

Figure 2 – A diagram representing BER in a strand of DNA. Ignore the role of dRpase.
Source: Departamento de Bioquímica e Imunologia, Universidade Federal de Minas Gerais. Escherichia coli as a model system to study DNA repair genes of eukaryotic organisms. Published March 31st 2003.

However, for damages to more than one base (note that still only one strand is damaged), there exists another excision method known as NER (Nucleotide excision repair) which cleaves 12 to 24 nucleotides using different proteins, yet the mechanism is almost identical. This is necessary in helix-distorting damages (i.e. that disrupts the helix structure), including pyrimidine dimerization from UV light, where adjacent bases on one of the strands join. For example, adjacent thymine bases will bind, leaving the adenine bases on the complementary strand isolated and thus NER must be utilized to create a new strand in which the thymine bases can bind with the isolated adenine bases. [12]

b) Double-stranded repair:

We first note that lesions affecting both strands are potentially much more harmful and difficult to repair, as no template exists at the specific locus. This would eventually lead to large segments of chromosomes either ‘dying’ in the next mitosis or worse still, mutating. Lesions of the sort are generally caused by high-energy radiation, thus explaining why the Chernobyl tragedy was much more severe than accidents seen previously. Two mechanisms exist for self-induced repair, yet they are both reliant on various factors or are not as effective as single-stranded repair mechanisms.

Homologous recombination is the favored method of repair, as there is a much lower risk of new mutations forming. This is where an identical nucleotide sequence found at another locus is used as a template – for example from a sister chromatid after meiosis 2. It is crucial during this process that genome integrity is kept, and tumor suppressors including Rad51 are necessary for the new strand to safely ‘invade’ the damaged nucleotide region without being damaged.[13]

On the other hand, if no identical strand can be found as a template (for example if the cell isn’t a gamete and is very specialized), one must resort to Non-homologous end joining (NHEJ). Perhaps rather crude, this mechanism involves ‘gluing’ the two adjacent correct regions back together. DNA ligase IV, found in chromosome 13, catalyzes the formation of an ester bond between the phosphate backbone and nucleotides of both chromosomes on both helixes (remember these lesions affects both strands). This mechanism would inevitably lead to further mutation formation as nucleotides cleaved aren’t replaced, yet in many nucleotides this is still preferred to the loss of a complete chromosome arm.[14]

Rarely, a lesion is so damaging that no repair mechanism above can reverse or even mitigate the effects. With the potential risk of tumor formation, the cell would resort to its last repair mechanism – cell apoptosis. This is where the cell programs a ‘suicide’ in a variety of ways, may it be cell shrinkage, nuclear fragmentation or chromatin condensation.[15] However, due to the rapid rate of mitosis, apoptosis is not always performed and thus unable to completely prevent tumor formation – this is why cancer is still such a large problem to date.

Figure 4 – A table to summarize different types of Lesions and the repair processes utilized. As you can see, each process requires many specific proteins. Source: Wikipedia

Shortfalls of self-induced repair:

As explained above, apoptosis doesn’t always solve the problem of lesion damage. Even before that, each repair mechanism is not always effective or even efficient – this is what we are striving to solve. As said before, with the frequency of DNA replications and thus lesion damages, repair mechanisms are often unable to recognize the damage in time (many observe a change in the base, but if it has already replicated then there is no observed difference.) To combat this, it is possible to delay the cell replication cycle, but this is problematic in other ways – think of the importance in quickly forming platelets to construct eschars. (scabs)

In addition, many lesions do not alter the base sequence, but still influences which genes are expressed. These so called epimutations are also unrecognizable by repair mechanisms. Worse still, lesion damage can occur in repair genes themselves, and especially for those involved in Homologous recombination or mismatch repair, mutation rates and tumor formation rates increase rapidly (shown in mice.)[16] Is there anything we can do about this?

Human efforts and developing technologies:

Human efforts to aid the process of DNA repair or prevent DNA damage from causing harm to an extent have had varied results, with new techniques constantly being discovered, each with their own benefits but also shortfalls and potential risks. While it may seem that increasing the rate of DNA transcription is the best way to combat lesion damage (by ensuring that damaged cells can be replaced), and thus resorting to commercial production of various transcriptional activators as they stabilize the RNA polymerase holoenzyme responsible for this increase[17], this is not always the case. The method above does not address the damaged cells themselves nor the risk of tumor formation – i.e. this is merely dodging the problem at hand. Thus, this essay will explore two areas researched as alternative repair mechanisms – protein inhibition and controlled tumor destruction.

a) Protein Inhibition:

DNA synthesis unsurprisingly relies on a complex mixture of enzymes and other proteins. By inhibiting particular proteins, insufficient genetic information or oxidative stress will ‘encourage’ cell apoptosis[18]. However, it is important to mention that to prevent normal cell apoptosis (we only target cancerous cells), methods must address a difference between normal and cancerous cells. Three such inhibitors are PARP inhibitors, F06 protein inhibitors and RPA protein inhibitors.

PARPi or Poly-ADP ribose polymerase inhibitors disrupt the ability of these protein to catalyze mismatch repair processes – they usually help repair the ‘nicks’ in DNA. Inhibition will lead to formation of multi-stranded breaks. However, the neat trick lies in the fact that while these lesions are usually reparable in normal cells, the high mitotic rate in cancerous cells means that many lesions will form and eventually lead to cell death or apoptosis. However, currently this method is only effective in Ovarian and some Prostate cancers, as tumorous cells are PTEN-defective, this being important to allow integration of PARPi[19] – I will not get into this as it is much more complicated.

The latter two mechanisms disrupt the NER pathways rather than that of mismatch repairs. The F06 protein is responsible for inhibiting the protein-protein binding between ERCC1 and XPF in lung cancer cells, this significant in two aspects. While when bonded both proteins are essential to NER repair, as monomers their toxicity is also harmful to the tumor cells, which will also lack a repair mechanism for the toxins (no bonded ERCC1-XPF proteins).[20]

On the other hand, one could inhibit RPA proteins. While RPA is not directly involved in the survival of cancerous damaged cells, their fold structure contains oligosaccharide to saccharide bindings (OB bindings), which stimulates ATR and CHK1 (ataxia-telangiectasia-mutated-and-Rad3-related kinase and checkpoint-kinase 1 respectively[21].) These two chemicals are responsible for limiting the efficacy of radiation and chemical therapy as methods for killing cancerous cells. Thus, by inhibiting RPA proteins, one creates a snowball effect which would eventually aid current therapeutic methods. Side effects or shortfalls are to these 3 protein inhibition mechanisms are currently unknown, but considering their recent discovery, (especially the latter mechanism) problems are inevitable.[22]

b) Controlled tumor destruction:

We all have heard of chemotherapy and radiotherapy as methods of ‘curing’ different cancers. However, we explore a recently developed alternative mechanism – Boron neutron capture therapy or BNCT. With the successes of proton therapy and the current development of cyclotrons to be distributed to various countries (for example by Hitachi)[23], this method is often missed. A reaction between Boron-10 and thermal neutrons produces alpha particles, a lithium-7 nucleus and high-LET (linear energy transfer) radiation. As B-10 is more readily accepted by cancerous cells, these harmful cells are destroyed, with as few as possible normal cells also undergoing cell death.[24] In tumors like Malignant pleural mesothelioma (MPM), the proximity of the lungs means dosage or alternative radiation treatments are harmful – and thus BNCT is utilized in places like Kyoto University’s reactor institute.

Figure 5- Drawn representation of BNCT. Note that thermal neutrons ignore the normal cells, and thus can collide with the cancerous cells with high efficiency.

Of course, treatment of the sort requires highly-skilled surgeons for targeting, cost large amounts due to the requirements of a dosimetry system and CT scan, and clearly have unknown side effects (only 50% of patients in Japan survive for unknown reasons[25].) The mechanism is still not ideal – and we can strive for better.

A brighter future?

Clearly, we have yet to reach a definitive ‘cure’ to cancer without any risks or chance of failure. While it may seem that with every new mechanism new issues are uncovered, we are slowly finding a way to incorporate various strategies in such a way that they complement the strengths of each other and alleviate the further issues caused. Whether it be utilizing the developing CRISPR/Cas9 technology, or creating molecules to stimulate self-repair, scientists are discovering new methods every day, each hailed as the possible cure and many raking in 100s of thousands of dollars for further research.

CRISPR/Cas9 technology is a relatively new mechanism that enables site-specific genomic targeting in any organism.[26] By binding to the protospacer adjacent motif in DNA sequence, a desired mutation can be introduced. Recently, the ability to transport the genetic material into the nucleus where it can then be introduced has been sped up through the use of electrical stimulation or electroporation.[27] Due to the fact that this technology can be used as a complement to protein inhibition (as mutations introduced can help to cause this), the efficiency shortfalls of methods explored above can hopefully be reduced. However, this doesn’t directly address the problem of tumor formation, instead increasing the rate of repair – a mitigation method rather than a full-blown ‘cure.’ Moreover, there is a risk that the technology will ‘snip’ too much as the region affected cannot currently be calculated.[28]

On the other hand, one could look to relatively unknown methods recently brought to light. An article published in Nature 2016 showed how one could protect the TERT or telomerase reverse transcriptase gene which is a promoter site, thus definitively reducing the rate of tumor formation[29], while multiple studies on RAD51 (eukaryotic homologue of E.coli recombinase) have shown that if overexpression can be encouraged in noncancerous cells, increased resistance to radiation damage is possible – the stimulatory factor has now been found as C20H16Br2N2O3S or the RS-1 stimulatory compound.[30][31]

Figure 6- The cover of Time magazine presenting CRISPR as a potential cure for cancer. Yet, while CRISPR works, it only helps very specific cases – and thus cannot be called a definitive ‘cure’ to cancer.

 Source: Time magazine, Volume 188 no.1. Published July 4th2016


So, how optimistic can we be about eventually solving this worldwide issue and giving a Nobel Prize to the individual who discovers a breakthrough? Technology is only set to improve, further enabling those in the field to utilize or discover new precise techniques, which may prove to be the missing part in current repair mechanisms. Whatever the case, even though the battle in one’s cell between lesion damage and repair mechanisms is ever present, our involvement in the complex workings of genetics is set to soon uncover something which will unequivocally cure what is the second highest cause of death[32], yet stems from just one small mishap.

[1] Helbock, H.J, Beckman, K.B, Shigenaga, M.K. DNA oxidation matters: the HPLC-electrochemical detection assay of 8-deoxyguanosine and 8-oxo-guanine. 1998.

[2] Université de Montreal. The study of the molecular mechanisms involved in cellular ageing.

[3] Kaneko, T, Tahara, S, Taguchi, T, Kondo, H. Accumulation of oxidative DNA damage, 8-oxo-2’deoxyguanosine, and change of repair systems during in vitro cellular aging of cultured human skin fibroblasts. PubMed Nov 2001.


[5] DNA Oxidation – DNA damage by ROS. Youtube video published Sep 2017.



[8] Dexheimer, T, S. DNA repair pathways and mechanisms, 2013



[11] Braithwaite, E,K et al. DNA polymerase lambda mediates a back-up base excision repair activity in extracts of mouse embryonic fibroblasts. J. Biol, May 2005.


[13] Image from Kanehisa Laboratories, September 2016. Taken from

[14] Alberts, B. Johnson, A. DNA repair (2002).

[15] Elmore, S. Apoptosis: A review of Programmed Cell Death. Published in NCBI December 2007.

[16] Hegan, D,C et al. Elevated levels of mutation in multiple tissues of mice deficient in the DNA mismatch repair gene. National academy of Sciences of the USA, December 2006


[18] Long, J.S, Ryan, K.M. New frontiers in promoting tumor cell death: targeting apoptosis, necroptosis and autophagy. Oncogen, published December 2012.


[20] Gavande, N S et al. DNA repair targeted therapy: The past or future of cancer treatment? ScienceDirect February 2016.

[21] Qiu, Z. Oleinick, N. Zhang, J. ATR/CHK1 inhibitors and cancer therapy. Science direct, published March 2018.

[22] Glanxer, J,G. Liu, S. RPA Inhibition increases Replication Stress and Suppresses Tumor Growth. NCBI, Published July 2014.


[24] Nedunchezhian, K et al. Boron Neutron Capture Therapy – A literature review. NCBI, Published December 2016

[25] – Taken from Barth et al, 2005.




[29] Wong, J.W.H. Differential DNA repair underlies mutation hotspots at active promoters in cancer genomes. Nature, 2016

[30] Raderschall, E et al. Elevated Levels of Rad51 Recombination Protein in Tumor Cells. Cancer research journal, published 2002.

[31] Mason, J.M, Logan, H.L, Budke, B. The RAD51-stimulatpory compound RS-1 can exploit the RAD51 overexpression that exists in cancer cells and tumors. PubMed, July 2014

[32] Ma, X. Yu, H. Global Burden of Cancer. Yale Journal, Published October 2007.

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