Despite the natural ability of the immune system to recognize cancer and, in some patients, even to eliminate it, cancer cells have acquired numerous evading mechanisms. With the increasing knowledge and focus shifting from targeting rapidly proliferating cells with chemotherapy to modulating the immune system, there have been recent efforts to integrate (e.g., simultaneously or sequentially) various therapeutic approaches. Combining the oncolytic activity of some chemotherapeutics with immunostimulatory molecules, so-called chemoimmunotherapy, is an attractive strategy. An example of such an immunostimulatory molecule is polyinosinic:polycytidylic acid [Poly(I:C)], a synthetic analogue of double-stranded RNA characterized by rapid nuclease degradation hampering its biological activity. This study investigated the possible interactions of tetracycline and anthracycline chemotherapeutics with different commercial Poly(I:C) molecules and protection against nuclease degradation. Fluorescence spectroscopy and circular dichroism revealed an interaction of all of the selected chemotherapeutics with Poly(I:C)s and the ability of doxycycline and minocycline to prolong the resistance to RNase cleavage, respectively. The partial protection was observed in vitro as well.

BIOCEV 1st Faculty of Medicine Charles University 252 50 Vestec Czech Republic

Department of Paediatrics and Inherited Metabolic Disorders 1st Faculty of Medicine Charles University and General University Hospital 120 00 Prague Czech Republic

Department of Pathological Physiology Faculty of Medicine Masaryk University Kamenice 5 Brno CZ 625 00 Czech Republic

Department of Physiology Faculty of Medicine Masaryk University Kamenice 5 625 00 Brno Czech Republic

Institute of Biotechnology of the Czech Academy of Sciences BIOCEV 252 50 Vestec Czech Republic

Section on Medical Neuroendocrinology Eunice Kennedy Shriver National Institute of Child Health and Human Development National Institutes of Health Building 10 Room 1 3140 10 Center Drive Bethesda Maryland 20892 United States

TumorSHOT Italská 2581 67 Vinohrady Praha 2 Prague 120 00 Czech Republic

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Bray F.; Laversanne M.; Weiderpass E.; Soerjomataram I. The ever-increasing importance of cancer as a leading cause of premature death worldwide. Cancer 2021, 127 (16), 3029–3030. 10.1002/cncr.33587. PubMed DOI

Cui Z.; Willingham M. C.; Hicks A. M.; Alexander-Miller M. A.; Howard T. D.; Hawkins G. A.; Miller M. S.; Weir H. M.; Du W.; DeLong C. J. Spontaneous regression of advanced cancer: identification of a unique genetically determined, age-dependent trait in mice. Proc. Natl. Acad. Sci. U.S.A. 2003, 100 (11), 6682–6687. 10.1073/pnas.1031601100. PubMed DOI PMC

Koch J.; Hau J.; Jensen H. E.; Nielsen C. H.; Rieneck K. The cellular cancer resistance of the SR/CR mouse. Apmis 2012, 120 (12), 974–987. 10.1111/j.1600-0463.2012.02925.x. PubMed DOI

Hanahan D. Hallmarks of Cancer: New Dimensions. Cancer Discov. 2022, 12 (1), 31–46. 10.1158/2159-8290.CD-21-1059. PubMed DOI

Hanahan D.; Weinberg R. A. The hallmarks of cancer. Cell 2000, 100 (1), 57–70. 10.1016/S0092-8674(00)81683-9. PubMed DOI

Hanahan D.; Weinberg R. A. Hallmarks of cancer: the next generation. Cell 2011, 144 (5), 646–674. 10.1016/j.cell.2011.02.013. PubMed DOI

Mokhtari R. B.; Homayouni T. S.; Baluch N.; Morgatskaya E.; Kumar S.; Das B.; Yeger H. Combination therapy in combating cancer. Oncotarget 2017, 8 (23), 38022–38043. 10.18632/oncotarget.16723. PubMed DOI PMC

Apetoh L. Chemoimmunotherapy combinations: translating basic knowledge into clinical successes. Gene Immun. 2024, 25 (2), 99–101. 10.1038/s41435-024-00264-9. PubMed DOI

Padrón L. J.; Maurer D. M.; O’Hara M. H.; O’Reilly E. M.; Wolff R. A.; Wainberg Z. A.; Ko A. H.; Fisher G.; Rahma O.; Lyman J. P.; et al. Sotigalimab and/or nivolumab with chemotherapy in first-line metastatic pancreatic cancer: clinical and immunologic analyses from the randomized phase 2 PRINCE trial. Nat. Med. 2022, 28 (6), 1167–1177. 10.1038/s41591-022-01829-9. PubMed DOI PMC

Correale P.; Cusi M. G.; Micheli L.; Nencini C.; Del Vecchio M. T.; Torino F.; Aquino A.; Bonmassar E.; Francini G.; Giorgi G. Chemo-immunotherapy of colorectal carcinoma: preclinical rationale and clinical experience. Invest. New Drugs 2006, 24 (2), 99–110. 10.1007/s10637-006-5932-7. PubMed DOI

Marabelle A.; Tselikas L.; de Baere T.; Houot R. Intratumoral immunotherapy: using the tumor as the remedy. Ann. Oncol. 2017, 28 (suppl_12), xii33–xii43. 10.1093/annonc/mdx683. PubMed DOI

Correia A. S.; Gärtner F.; Vale N. Drug combination and repurposing for cancer therapy: the example of breast cancer. Heliyon 2021, 7 (1), e0594810.1016/j.heliyon.2021.e05948. PubMed DOI PMC

Le Naour J.; Thierry S.; Scuderi S. A.; Boucard-Jourdin M.; Liu P.; Bonnin M.; Pan Y.; Perret C.; Zhao L.; Mao M.; et al. A Chemically Defined TLR3 Agonist with Anticancer Activity. Oncoimmunology 2023, 12 (1), 2227510.10.1080/2162402X.2023.2227510. PubMed DOI PMC

De Waele J.; Verhezen T.; van der Heijden S.; Berneman Z. N.; Peeters M.; Lardon F.; Wouters A.; Smits E. A systematic review on poly(I:C) and poly-ICLC in glioblastoma: adjuvants coordinating the unlocking of immunotherapy. J. Exp. Clin. Cancer Res. 2021, 40 (1), 213.10.1186/s13046-021-02017-2. PubMed DOI PMC

Bianchi F.; Pretto S.; Tagliabue E.; Balsari A.; Sfondrini L. Exploiting poly(I:C) to induce cancer cell apoptosis. Cancer Biol. Ther. 2017, 18 (10), 747–756. 10.1080/15384047.2017.1373220. PubMed DOI PMC

Sales Conniff A.; Encalada G.; Patel S.; Bhandary M.; Al-Takrouri F.; Heller L. Poly(I:C) transfection induces a pro-inflammatory cascade in murine mammary carcinoma and fibrosarcoma cells. RNA Biol. 2022, 19 (1), 841–851. 10.1080/15476286.2022.2084861. PubMed DOI PMC

Kowash H. M.; Potter H. G.; Edye M. E.; Prinssen E. P.; Bandinelli S.; Neill J. C.; Hager R.; Glazier J. D. Poly(I:C) source, molecular weight and endotoxin contamination affect dam and prenatal outcomes, implications for models of maternal immune activation. Brain Behav. Immun. 2019, 82, 160–166. 10.1016/j.bbi.2019.08.006. PubMed DOI

Komal A.; Noreen M.; El-Kott A. F. TLR3 agonists: RGC100, ARNAX, and poly-IC: a comparative review. Immunol. Res. 2021, 69 (4), 312–322. 10.1007/s12026-021-09203-6. PubMed DOI PMC

Sultan H.; Salazar A. M.; Celis E. Poly-ICLC, a multi-functional immune modulator for treating cancer. Semin. Immunol. 2020, 49, 101414.10.1016/j.smim.2020.101414. PubMed DOI

Alvarez M.; Molina C.; Garasa S.; Ochoa M. C.; Rodriguez-Ruiz M. E.; Gomis G.; Cirella A.; Olivera I.; Glez-Vaz J.; Gonzalez-Gomariz J.; et al. Intratumoral neoadjuvant immunotherapy based on the BO-112 viral RNA mimetic. Oncoimmunology 2023, 12 (1), 2197370.10.1080/2162402X.2023.2197370. PubMed DOI PMC

Chukwudi C. U.; Good L. Interaction of the tetracyclines with double-stranded RNAs of random base sequence: new perspectives on the target and mechanism of action. J. Antibiot. 2016, 69 (8), 622–630. 10.1038/ja.2015.145. PubMed DOI

Chukwudi C. U.; Good L. Doxycycline inhibits pre-rRNA processing and mature rRNA formation in E. coli. J. Antibiot. 2019, 72 (4), 225–236. 10.1038/s41429-019-0149-0. PubMed DOI

Chukwudi C. U.; Good L. Doxycycline induces Hok toxin killing in host E. coli. PLoS One 2020, 15 (7), e023563310.1371/journal.pone.0235633. PubMed DOI PMC

Chopra I.; Roberts M. Tetracycline antibiotics: mode of action, applications, molecular biology, and epidemiology of bacterial resistance. Microbiol. Mol. Biol. Rev. 2001, 65 (2), 232–260. 10.1128/MMBR.65.2.232-260.2001. PubMed DOI PMC

Warner A. J.; Hathaway-Schrader J. D.; Lubker R.; Davies C.; Novince C. M. Tetracyclines and bone: Unclear actions with potentially lasting effects. Bone 2022, 159, 116377.10.1016/j.bone.2022.116377. PubMed DOI PMC

Nonaka L.; Connell S. R.; Taylor D. E. 16S rRNA mutations that confer tetracycline resistance in Helicobacter pylori decrease drug binding in Escherichia coli ribosomes. J. Bacteriol. 2005, 187 (11), 3708–3712. 10.1128/JB.187.11.3708-3712.2005. PubMed DOI PMC

Brodersen D. E.; Clemons W. M. Jr.; Carter A. P.; Morgan-Warren R. J.; Wimberly B. T.; Ramakrishnan V. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 2000, 103 (7), 1143–1154. 10.1016/S0092-8674(00)00216-6. PubMed DOI

Hori N.; Denesyuk N. A.; Thirumalai D. Shape changes and cooperativity in the folding of the central domain of the 16S ribosomal RNA. Proc. Natl. Acad. Sci. U.S.A. 2021, 118 (10), e202083711810.1073/pnas.2020837118. PubMed DOI PMC

Khan M. A.; Musarrat J. Interactions of tetracycline and its derivatives with DNA in vitro in presence of metal ions. Int. J. Biol. Macromol. 2003, 33 (1–3), 49–56. 10.1016/S0141-8130(03)00066-7. PubMed DOI

Oehler R.; Polacek N.; Steiner G.; Barta A. Interaction of tetracycline with RNA: photoincorporation into ribosomal RNA of Escherichia coli. Nucleic Acids Res. 1997, 25 (6), 1219–1224. 10.1093/nar/25.6.1219. PubMed DOI PMC

Pioletti M.; Schlünzen F.; Harms J.; Zarivach R.; Glühmann M.; Avila H.; Bashan A.; Bartels H.; Auerbach T.; Jacobi C.; et al. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 2001, 20 (8), 1829–1839. 10.1093/emboj/20.8.1829. PubMed DOI PMC

Chen Y. F.; Yang Y. N.; Chu H. R.; Huang T. Y.; Wang S. H.; Chen H. Y.; Li Z. L.; Yang Y. S. H.; Lin H. Y.; Hercbergs A.; et al. Role of Integrin αvβ3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells. Front. Cell Dev. Biol. 2022, 10, 829788.10.3389/fcell.2022.829788. PubMed DOI PMC

Qin Y.; Zhang Q.; Lee S.; Zhong W. L.; Liu Y. R.; Liu H. J.; Zhao D.; Chen S.; Xiao T.; Meng J.; et al. Doxycycline reverses epithelial-to-mesenchymal transition and suppresses the proliferation and metastasis of lung cancer cells. Oncotarget 2015, 6 (38), 40667–40679. 10.18632/oncotarget.5842. PubMed DOI PMC

Sun T.; Zhao N.; Ni C. S.; Zhao X. L.; Zhang W. Z.; Su X.; Zhang D. F.; Gu Q.; Sun B. C. Doxycycline inhibits the adhesion and migration of melanoma cells by inhibiting the expression and phosphorylation of focal adhesion kinase (FAK). Cancer Lett. 2009, 285 (2), 141–150. 10.1016/j.canlet.2009.05.004. PubMed DOI

Mattioli R.; Ilari A.; Colotti B.; Mosca L.; Fazi F.; Colotti G. Doxorubicin and other anthracyclines in cancers: Activity, chemoresistance and its overcoming. Mol. Aspects Med. 2023, 93, 101205.10.1016/j.mam.2023.101205. PubMed DOI

Airoldi M.; Barone G.; Gennaro G.; Giuliani A. M.; Giustini M. Interaction of doxorubicin with polynucleotides. A spectroscopic study. Biochemistry 2014, 53 (13), 2197–2207. 10.1021/bi401687v. PubMed DOI

Canzoneri J. C.; Oyelere A. K. Interaction of anthracyclines with iron responsive element mRNAs. Nucleic Acids Res. 2008, 36 (21), 6825–6834. 10.1093/nar/gkn774. PubMed DOI PMC

Ijäs H.; Shen B.; Heuer-Jungemann A.; Keller A.; Kostiainen M. A.; Liedl T.; Ihalainen J. A.; Linko V. Unraveling the interaction between doxorubicin and DNA origami nanostructures for customizable chemotherapeutic drug release. Nucleic Acids Res. 2021, 49 (6), 3048–3062. 10.1093/nar/gkab097. PubMed DOI PMC

Micallef I.; Baron B. Doxorubicin: An overview of the anti-cancer and chemoresistance mechanisms. Ann. Clin. Toxicol. 2020, 3, 1031.

Palù G.; Valisena S.; Ciarrocchi G.; Gatto B.; Palumbo M. Quinolone binding to DNA is mediated by magnesium ions. Proc. Natl. Acad. Sci. U.S.A. 1992, 89 (20), 9671–9675. 10.1073/pnas.89.20.9671. PubMed DOI PMC

Kohn K. W. Mediation of Divalent Metal Ions in the Binding of Tetracycline to Macromolecules. Nature 1961, 191 (4794), 1156–1158. 10.1038/1911156a0. PubMed DOI

Sirajuddin M.; Ali S.; Badshah A. Drug–DNA interactions and their study by UV–Visible, fluorescence spectroscopies and cyclic voltametry. J. Photochem. Photobiol., B 2013, 124, 1–19. 10.1016/j.jphotobiol.2013.03.013. PubMed DOI

Scollo F.; Evci H.; Amaro M.; Jurkiewicz P.; Sykora J.; Hof M. What Does Time-Dependent Fluorescence Shift (TDFS) in Biomembranes (and Proteins) Report on?. Front. Chem. 2021, 9, 738350.10.3389/fchem.2021.738350. PubMed DOI PMC

Lakowicz J. R.; Keating-Nakamoto S. Red-edge excitation of fluorescence and dynamic properties of proteins and membranes. Biochemistry 1984, 23 (13), 3013–3021. 10.1021/bi00308a026. PubMed DOI PMC

Sillén L. G.; Hartiala K. J.; Liem D. H.; Ryhage R.; Stevens R. High-speed Computers as a Supplement to Graphical Methods. III. Twist Matrix Methods for Minimizing the Error-square Sum in Problems with Many Unknown Constants. Acta Chem. Scand. 1964, 18, 1085–1098. 10.3891/acta.chem.scand.18-1085. DOI

Silindir M.; Özer A. Y. Sterilization Methods and the Comparison of E-Beam Sterilization with Gamma Radiation Sterilization. FABAD J. Pharm. Sci. 2009, 34, 43–53.

Lahtz C.; Bates S. E.; Jiang Y.; Li A. X.; Wu X.; Hahn M. A.; Pfeifer G. P. Gamma irradiation does not induce detectable changes in DNA methylation directly following exposure of human cells. PLoS One 2012, 7 (9), e4485810.1371/journal.pone.0044858. PubMed DOI PMC

Tankovskaia S.; Kotb O.; Dommes O.; Paston S. DNA damage induced by gamma-radiation revealed from UV absorption spectroscopy. J. Phys.: Conf. Ser. 2018, 1038, 012027.10.1088/1742-6596/1038/1/012027. PubMed DOI

Husain M. A.; Ishqi H. M.; Rehman S. U.; Sarwar T.; Afrin S.; Rahman Y.; Tabish M. Elucidating the interaction of sulindac with calf thymus DNA: biophysical and in silico molecular modelling approach. New J. Chem. 2017, 41 (24), 14924–14935. 10.1039/C7NJ03698A. DOI

Zhou X.; Zhang G.; Pan J. Groove binding interaction between daphnetin and calf thymus DNA. Int. J. Biol. Macromol. 2015, 74, 185–194. 10.1016/j.ijbiomac.2014.12.018. PubMed DOI

Li N.; Hu X.; Pan J.; Zhang Y.; Gong D.; Zhang G. Insights into the mechanism of groove binding between 4–octylphenol and calf thymus DNA. Spectrochim. Acta, Part A 2020, 238, 118454.10.1016/j.saa.2020.118454. PubMed DOI

Genna V.; Iglesias-Fernández J.; Reyes-Fraile L.; Villegas N.; Guckian K.; Seth P.; Wan B.; Cabrero C.; Terrazas M.; Brun-Heath I.; et al. Controlled sulfur-based engineering confers mouldability to phosphorothioate antisense oligonucleotides. Nucleic Acids Res. 2023, 51 (10), 4713–4725. 10.1093/nar/gkad309. PubMed DOI PMC

Thiele D.; Guschlbauer W.; Favre A. Protonated polynucleotide structures. Biochim. Biophys. Acta 1972, 272 (1), 22–26. 10.1016/0005-2787(72)90028-7. PubMed DOI

Kumar G. S.; Das S.; Bhadra K.; Maiti M. Protonated forms of poly[d(G-C)] and poly(dG).poly(dC) and their interaction with berberine. Bioorg. Med. Chem. 2003, 11 (23), 4861–4870. 10.1016/j.bmc.2003.09.028. PubMed DOI

Ucci J. W.; Kobayashi Y.; Choi G.; Alexandrescu A. T.; Cole J. L. Mechanism of interaction of the double-stranded RNA (dsRNA) binding domain of protein kinase R with short dsRNA sequences. Biochemistry 2007, 46 (1), 55–65. 10.1021/bi061531o. PubMed DOI

Douthart R. J.; Burnett J. P.; Beasley F. W.; Frank B. H. Binding of ethidium bromide to double-stranded ribonucleic acid. Biochemistry 1973, 12 (2), 214–220. 10.1021/bi00726a006. PubMed DOI

Lambs L.; Venturim M.; Révérend B. D.-L.; Kozlowski H.; Berthon G. Metal ion-tetracycline interactions in biological fluids: Part 8. Potentiometric and spectroscopic studies on the formation of Ca(II) and Mg(II) complexes with 4-dedimethylamino-tetracycline and 6-desoxy-6-dem. J. Inorg. Biochem. 1988, 33 (3), 193–209. 10.1016/0162-0134(88)80049-7. PubMed DOI

Reuss A.; Vogel M.; Weigand J.; Suess B.; Wachtveitl J. Tetracycline Determines the Conformation of Its Aptamer at Physiological Magnesium Concentrations. Biophys. J. 2014, 107 (12), 2962–2971. 10.1016/j.bpj.2014.11.001. PubMed DOI PMC

Kaiser C.; Vogel M.; Appel B.; Weigand J.; Müller S.; Suess B.; Wachtveitl J. Magnesium Ion-Driven Folding and Conformational Switching Kinetics of Tetracycline Binding Aptamer: Implications for in vivo Riboswitch Engineering. J. Mol. Biol. 2023, 435 (20), 168253.10.1016/j.jmb.2023.168253. PubMed DOI

Jin L.; Amaya-Mazo X.; Apel M. E.; Sankisa S. S.; Johnson E.; Zbyszynska M. A.; Han A. Ca2+ and Mg2+ bind tetracycline with distinct stoichiometries and linked deprotonation. Biophys. Chem. 2007, 128 (2), 185–196. 10.1016/j.bpc.2007.04.005. PubMed DOI

Sun W.; Pertzev A.; Nicholson A. W. Catalytic mechanism of Escherichia coli ribonuclease III: kinetic and inhibitor evidence for the involvement of two magnesium ions in RNA phosphodiester hydrolysis. Nucleic Acids Res. 2005, 33 (3), 807–815. 10.1093/nar/gki197. PubMed DOI PMC

Smole A.; Krajnik A. K.; Oblak A.; Pirher N.; Jerala R. Delivery system for the enhanced efficiency of immunostimulatory nucleic acids. Innate Immun. 2013, 19 (1), 53–65. 10.1177/1753425912450346. PubMed DOI