Ancient embedded elements in our DNA from generations past can activate a powerful immune response to kill cancer cells like an infection.
The work builds on Princess Margaret Senior Scientist Dr. De Carvalho’s previous ground-breaking discovery known as viral mimicry – the ability to cause cancer cells to behave as though they have been infected, thereby activating the immune system to fight cancer like an infection.
Dr. Daniel De Carvalho and his team have now identified silent ancient DNA elements buried in our genome that when ‘reactivated’ can initiate this immune response.
Importantly, they have also discovered a key enzyme used by cancer cells to prevent this from happening in order to survive.
The enzyme is known as ADAR1, and it acts to prevent the cancer cells from signaling to the immune system. Dr. De Carvalho, Associate Professor, Medical Biophysics, University of Toronto, discovered that by inhibiting this enzyme, cancer cells were more sensitive to new drug therapies that induce viral mimicry.
The research is published online on October 21, 2020 in Nature, under the title, “Epigenetic therapy induces transcription of inverted SINEs and ADAR1 dependency.”
The study first authors are Dr. Parinaz Mehdipour, Dr. Sajid Marhon and Masters’ graduate student Ilias Ettayebi, trainees in Dr. De Carvalho’s laboratory.
“Humans acquired a series of ‘silent’ repetitive elements in our DNA over millions of years of evolution, but it has been unclear why or what purpose they serve,” explains Dr. De Carvalho.
“As ‘genome archeologists’, we set out to identify the function of these ‘DNA relics’ and have found that under the right conditions they can be reactivated and stimulate our immune system.”
Dr. De Carvalho’s discovery of ADAR1 explains how some cancer cells mount a defense against this and protect themselves from our immune system.
“These findings open up a new field of cancer therapies,” says Dr. De Carvalho. “It gives us the opportunity to take advantage of these ancient repetitive DNA elements to fight cancer.”
Studying the potential to modulate the immune response against tumour cells is one of the most rapidly changing and exciting areas in clinical oncology.
While much knowledge has been gained about how the immune system interacts with cancer, leading to the development of novel immunotherapy drugs, there is still a large proportion of cancer patients who do not respond to immunotherapy alone.
In Dr. De Carvalho’s initial discovery, epigenetic drugs were shown to reactivate these repetitive DNA elements and lead to production of double-stranded RNA, a molecular pattern that is also observed following viral infection.
This ‘viral mimicry’ leads to an antiviral response directed specifically against cancer cells.
In this latest research, Dr. De Carvalho’s lab identified the specific ancient repetitive DNA elements as SINEs (Short Interspersed Nuclear Elements). These SINEs usually lie quiet in our genome, having little effect on the host.
However, if activated by new epigenetic drugs, these SINES produce double-stranded RNA – a marker for infection – and can ultimately be used by cells to trigger an innate immune response.
Dr. De Carvalho likens this response “to an ancient dagger that can be used against cancer.”
But cancer cells are wily and have also evolved to evade detection by the immune system even under conditions where the ancient DNA sequences are activated.
Dr. De Carvalho discovered that cancer cells strike back by making more of the ADAR1 enzyme, which functions to disrupts the double-stranded RNA produced by the ancient DNA. In this way ADAR1 prevents the cancer cells from activating the immune system.
Dr. Carvalho and his team went on to demonstrate that deleting ADAR1 from cancer cells makes them exquisitely vulnerable to epigenetic drugs that induce the antiviral response.
“Since the ADAR1 activity is enzymatic, our work provides an exciting new target for drug development efforts for a completely new class of drugs that are able to exploit these ‘ancient weapons’ in our genome,” explains Dr. De Carvalho.
A large amount of data has been accumulated about the composition of circulating cell-free DNA (cfDNA) in the blood of higher organisms, but the biological function of cfDNAs remains to be discussed. Nowadays, several measurements of cfDNA characteristics are used in cancer patient liquid biopsies: SNP analysis of proto-oncogenes and onco-suppressors, changes in the pattern of DNA methylation, and the abundance of oncogene fragments and other specific fragments of tumor-derived DNA microsatellites, tandem repeats, and mobile genetic elements.1, 2, 3
Although attempts have been made to use cfDNAs as indicators of cancer development, based on the content of specific DNA sequences in the blood, the low sensitivity of cfDNA measurement methods, and failure to apply developed markers for early-stage tumor development, has significantly limited the clinical application of this analysis.4,5
On the other hand, increases in microsatellite content, tandem repeats, and mobile genetic elements are commonly detected in the early stages of cancer, both in experimental tumor models and patients, causing researchers to consider these sequences as potential markers for prognosis, as well as factors in disease progression and cure.6,7
Retrotransposons, a type of mobile genetic element (MGE), are able to self-reproduce in the genome through RNA intermediates. These elements can be located in a tandem manner (satellite heterochromatin, telomeres, etc.) or scattered throughout the genome.8
Dispersed short-interspersed nuclear elements (SINEs) are short DNA sequences (less than 500 base pairs) formed by reverse transcription of short RNA molecules: 5S rRNA, tRNA, and various microRNAs (miRNAs).9, 10, 11
SINEs do not encode proteins and their transposition in the genome depends on other mobile elements.12
The most famous and well-investigated SINEs are human Alu-repeats and their murine B-family homologs.13 Long-interspersed nuclear elements (LINEs) are longer retrotransposons (several thousand base pairs in length) containing 3′ end poly(A)-tracts, adenine-rich sequences, or tandem-repeating sequences.14 LINEs are independent elements that encode their own reverse transcriptase (RT) and endonuclease, allowing for genome migration.10
The high abundance of SINEs (particularly Alu) in the blood of cancer patients was first reported in 1977.15 The much higher abundance of LINEs in the cfDNA of cancer patients, compared with healthy donors, was found later, but the described results were controversial.16,17
Nevertheless, researchers have accumulated extensive data on the high levels of SINEs and LINEs, as well as the multiple mutations found in these elements, in the circulating cfDNA of cancer patients and murine models.18, 19, 20, 21
Nowadays, the level of Alu repeats is commonly used to measure the concentration of specific tumor-derived cfDNA in blood serum.22
Despite the extensive data, the role of SINEs and LINEs is still not fully understood, though there is evidence that the elements are regulators that activate onco-genes and suppress onco-suppressors.
For example, suppression of the LINE-encoded RT in tumor cells leads to a reduction in tumor progression, both in vitro and in vivo.23 Insertions or deletions of various SINEs and LINEss also lead to the development or suppression of various forms of cancer and other diseases.24, 25, 26, 27
However, the role of the repetitive part of the LINE-element remains unclear, and a reasonable explanation for the role of these elements in carcinogenesis is still unknown. The fact that metastases are often accompanied by a change in the number of SINEs and LINEs in tumor cells, metastatic cells, and the pool of cfDNA requires special attention and additional research.28,29
Previously, DNase I was demonstrated to display high anti-metastatic activity in several tumor models.30, 31, 32, 33 The significant reduction (up to 90%) in the number and area of metastases in Lewis lung carcinoma-bearing mice after treatment with DNase I correlated with a decrease in blood cfDNA concentration and with the restoration of total deoxyribonuclease activity in blood serum to the level of healthy animals.34
Later, we revealed that the molecular targets of DNase I are oncogenes and tandem repeats, including the SINEs and LINEs overrepresented during tumor progression.35
The main goal of our study was to reveal the effect of DNase I on SINEs and LINEs in the pool of circulating cfDNA under the tumor progression in mice and to find the relationship between the level of these tandem repeats and metastasis/tumor development. For these purposes we used three murine models: Lewis lung carcinoma, melanoma B16, and multiple drug resistant (MDR)-positive lymphosarcoma RLS40 (hereafter LLC, B16, and RLS40, respectively).
The ability of the SINEs and LINEs derived from the blood serum of tumor-bearing mice to penetrate into tumor cells of another origin was studied to clarify the possibility of horizontal transfer.
References
1. Soliman S.E., Alhanafy A.M., Habib M.S.E., Hagag M., Ibrahem R.A.L. Serum circulating cell free DNA as potential diagnostic and prognostic biomarker in non small cell lung cancer. Biochem. Biophys. Rep. 2018;15:45–51. [PMC free article] [PubMed] [Google Scholar]
2. Rouvinov K., Mermershtain W., Dresler H., Ariad S., Riff R., Shani-Shrem N., Keizman D., Neulander E.Z., Douvdevani A. Circulating cell-free DNA levels in patients with metastatic renal cell carcinoma. Oncol. Res. Treat. 2017;40:707–710. [PubMed] [Google Scholar]
3. Tang Z., Li L., Shen L., Shen X., Ju S., Cong H. Diagnostic value of serum concentration and integrity of circulating cell-free DNA in breast cancer: a comparative study with CEA and CA15-3. Lab. Med. 2018;49:323–328. [PubMed] [Google Scholar]
4. Spindler K.G. Methodological, biological and clinical aspects of circulating free DNA in metastatic colorectal cancer. Acta Oncol. 2017;56:7–16. [PubMed] [Google Scholar]
5. Haber D.A., Velculescu V.E. Blood-based analyses of cancer: circulating tumor cells and circulating tumor DNA. Cancer Discov. 2014;4:650–661. [PMC free article] [PubMed] [Google Scholar]
6. Zhang R., Pu W., Zhang S., Chen L., Zhu W., Xiao L., Xing C., Li K. Clinical value of ALU concentration and integrity index for the early diagnosis of ovarian cancer: A retrospective cohort trial. PLoS ONE. 2018;13:e0191756. [PMC free article] [PubMed] [Google Scholar]
7. Sobhani N., Generali D., Zanconati F., Bortul M., Scaggiante B. Cell-free DNA integrity for the monitoring of breast cancer: Future perspectives? World J. Clin. Oncol. 2018;9:26–32. [PMC free article] [PubMed] [Google Scholar]
8. Leng S., Zheng J., Jin Y., Zhang H., Zhu Y., Wu J., Xu Y., Zhang P. Plasma cell-free DNA level and its integrity as biomarkers to distinguish non-small cell lung cancer from tuberculosis. Clin. Chim. Acta. 2018;477:160–165. [PubMed] [Google Scholar]
9. Bhargava P.M., Shanmugam G. Uptake of nonviral nucleic acids by mammalian cells. Prog. Nucleic Acid Res. Mol. Biol. 1971;11:103–192. [PubMed] [Google Scholar]
10. Batzer M.A., Deininger P.L. Alu repeats and human genomic diversity. Nat. Rev. Genet. 2002;3:370–379. [PubMed] [Google Scholar]
11. Jelinek W.R., Schmid C.W. Repetitive sequences in eukaryotic DNA and their expression. Annu. Rev. Biochem. 1982;51:813–844. [PubMed] [Google Scholar]
12. López-Flores I., Garrido-Ramos M.A. The repetitive DNA content of eukaryotic genomes. Genome Dyn. 2012;7:1–28. [PubMed] [Google Scholar]
13. Umylny B., Presting G., Efird J.T., Klimovitsky B.I., Ward W.S. Most human Alu and murine B1 repeats are unique. J. Cell. Biochem. 2007;102:110–121. [PubMed] [Google Scholar]
14. Adams J.W., Kaufman R.E., Kretschmer P.J., Harrison M., Nienhuis A.W. A family of long reiterated DNA sequences, one copy of which is next to the human beta globin gene. Nucleic Acids Res. 1980;8:6113–6128. [PMC free article] [PubMed] [Google Scholar]
15. Stroun M., Anker P., Maurice P., Gahan P.B. Circulating nucleic acids in higher organisms. Int. Rev. Cytol. 1977;51:1–48. [PubMed] [Google Scholar]
16. Cheng J., Cuk K., Heil J., Golatta M., Schott S., Sohn C., Schneeweiss A., Burwinkel B., Surowy H. Cell-free circulating DNA integrity is an independent predictor of impending breast cancer recurrence. Oncotarget. 2017;8:54537–54547. [PMC free article] [PubMed] [Google Scholar]
17. Servomaa K., Rytömaa T. UV light and ionizing radiations cause programmed death of rat chloroleukaemia cells by inducing retropositions of a mobile DNA element (L1Rn) Int. J. Radiat. Biol. 1990;57:331–343. [PubMed] [Google Scholar]
18. Iskow R.C., McCabe M.T., Mills R.E., Torene S., Pittard W.S., Neuwald A.F., Van Meir E.G., Vertino P.M., Devine S.E. Natural mutagenesis of human genomes by endogenous retrotransposons. Cell. 2010;141:1253–1261. [PMC free article] [PubMed] [Google Scholar]
19. Lee E., Iskow R., Yang L., Gokcumen O., Haseley P., Luquette L.J., 3rd, Lohr J.G., Harris C.C., Ding L., Wilson R.K., Cancer Genome Atlas Research Network Landscape of somatic retrotransposition in human cancers. Science. 2012;337:967–971. [PMC free article] [PubMed] [Google Scholar]
20. Solyom S., Ewing A.D., Rahrmann E.P., Doucet T., Nelson H.H., Burns M.B., Harris R.S., Sigmon D.F., Casella A., Erlanger B. Extensive somatic L1 retrotransposition in colorectal tumors. Genome Res. 2012;22:2328–2338. [PMC free article] [PubMed] [Google Scholar]
21. Gualtieri A., Andreola F., Sciamanna I., Sinibaldi-Vallebona P., Serafino A., Spadafora C. Increased expression and copy number amplification of LINE-1 and SINE B1 retrotransposable elements in murine mammary carcinoma progression. Oncotarget. 2013;4:1882–1893. [PMC free article] [PubMed] [Google Scholar]
22. Lehner J., Stötzer O.J., Fersching D.M., Nagel D., Holdenrieder S. Plasma DNA integrity indicates response to neoadjuvant chemotherapy in patients with locally confined breast cancer. Int. J. Clin. Pharmacol. Ther. 2013;51:59–62. [PubMed] [Google Scholar]
23. Oricchio E., Sciamanna I., Beraldi R., Tolstonog G.V., Schumann G.G., Spadafora C. Distinct roles for LINE-1 and HERV-K retroelements in cell proliferation, differentiation and tumor progression. Oncogene. 2007;26:4226–4233. [PubMed] [Google Scholar]
24. Helman E., Lawrence M.S., Stewart C., Sougnez C., Getz G., Meyerson M. Somatic retrotransposition in human cancer revealed by whole-genome and exome sequencing. Genome Res. 2014;24:1053–1063. [PMC free article] [PubMed] [Google Scholar]
25. Rodríguez-Martín C., Cidre F., Fernández-Teijeiro A., Gómez-Mariano G., de la Vega L., Ramos P., Zaballos Á., Monzón S., Alonso J. Familial retinoblastoma due to intronic LINE-1 insertion causes aberrant and noncanonical mRNA splicing of the RB1 gene. J. Hum. Genet. 2016;61:463–466. [PubMed] [Google Scholar]
26. Roberts S.A., Lawrence M.S., Klimczak L.J., Grimm S.A., Fargo D., Stojanov P., Kiezun A., Kryukov G.V., Carter S.L., Saksena G. An APOBEC cytidine deaminase mutagenesis pattern is widespread in human cancers. Nat. Genet. 2013;45:970–976. [PMC free article] [PubMed] [Google Scholar]
27. Stacey S.N., Kehr B., Gudmundsson J., Zink F., Jonasdottir A., Gudjonsson S.A., Sigurdsson A., Halldorsson B.V., Agnarsson B.A., Benediktsdottir K.R. Insertion of an SVA-E retrotransposon into the CASP8 gene is associated with protection against prostate cancer. Hum. Mol. Genet. 2016;25:1008–1018. [PMC free article] [PubMed] [Google Scholar]
28. Hancks D.C., Kazazian H.H., Jr. SVA retrotransposons: Evolution and genetic instability. Semin. Cancer Biol. 2010;20:234–245. [PMC free article] [PubMed] [Google Scholar]
29. Belancio V.P., Roy-Engel A.M., Deininger P.L. All y’all need to know ‘bout retroelements in cancer. Semin. Cancer Biol. 2010;20:200–210. [PMC free article] [PubMed] [Google Scholar]
30. Salganik R.I., Martynova R.P., Matienko N.A., Ronichevskaya G.M. Effect of deoxyribonuclease on the course of lymphatic leukaemia in AKR mice. Nature. 1967;214:100–102. [PubMed] [Google Scholar]
31. Sugihara S., Yamamoto T., Tanaka H., Kambara T., Hiraoka T., Miyauchi Y. Deoxyribonuclease treatment prevents blood-borne liver metastasis of cutaneously transplanted tumour cells in mice. Br. J. Cancer. 1993;67:66–70. [PMC free article] [PubMed] [Google Scholar]
32. Wen F., Shen A., Choi A., Gerner E.W., Shi J. Extracellular DNA in pancreatic cancer promotes cell invasion and metastasis. Cancer Res. 2013;73:4256–4266. [PMC free article] [PubMed] [Google Scholar]
33. Alexeeva L.A., Patutina O.A., Sen’kova A.V., Zenkova M.A., Mironova N.L. Inhibition of invasive properties of murine melanoma by bovine pancreatic DNase I in vitro and in vivo. Mol. Biol. (Mosk.) 2017;51:637–646. [PubMed] [Google Scholar]
34. Patutina O., Mironova N., Ryabchikova E., Popova N., Nikolin V., Kaledin V., Vlassov V., Zenkova M. Inhibition of metastasis development by daily administration of ultralow doses of RNase A and DNase I. Biochimie. 2011;93:689–696. [PubMed] [Google Scholar]
35. Alekseeva L.A., Mironova N.L., Brenner E.V., Kurilshikov A.M., Patutina O.A., Zenkova M.A. Alteration of the exDNA profile in blood serum of LLC-bearing mice under the decrease of tumour invasion potential by bovine pancreatic DNase I treatment. PLoS ONE. 2017;12:e0171988. [PMC free article] [PubMed] [Google Scholar]
More information: Parinaz Mehdipour et al, Epigenetic therapy induces transcription of inverted SINEs and ADAR1 dependency, Nature (2020). DOI: 10.1038/s41586-020-2844-1
