Enhanced thermo-radiosensitization of tumor cells through suppression of the transcriptional stress rasponse by inhibiting HSF1 activity or expression

Cover Page

Cite item

Full Text

Open Access Open Access
Restricted Access Access granted
Restricted Access Subscription Access

Abstract

Hyperthermia is used in combination with radiation therapy to enhance the radiation response of the target tumor. However, heating of cancer cells activates the HSF1 transcription factor in them and stimulates the HSF1-dependent induction of heat shock proteins (HSPs), which can significantly impair the antitumor effects of hyperthermia and radiation exposure. The aim of this study was to examine the possibility of enhancing the radiosensitizing effect of hyperthermia on cancer cells by suppressing the HSF1-mediated HSP induction in them. The object of the study were HeLa cells derived from a malignant tumor of the human cervix. Before irradiation (2–7 Gy), cells were subjected to heat stress (42°–44°C for 20–60 min) without or in the presence of HSF1 transcriptional activity inhibitors (quercetin, triptolide, KRIBB11). In certain cell samples, HSF1 expression was preliminarily knocked down using small interfering RNAs. Cell death and survival was assessed by the levels of apoptosis/necrosis and clonogenic ability. Expression of HSF1 and HSP was analyzed by immunoblotting. It was found that, compared with the radiosensitizing effects of hyperthermia alone, the combined treatment (HSF1 activity inhibition or HSF1 knockdown + heating) significantly increased the thermo-radiosensitization of cancer cells; this was manifested in the intensification of their post-radiation death (apoptosis + necrosis), as well as in a decrease in clonogenicity. This enhancement of thermo-radiosensitization was observed under the HSP induction blockade. Thus, the combination of hyperthermia with inhibitors of HSF1 activity or expression can effectively sensitize thermoresistant and radioresistant tumors to radiation therapy.

Full Text

Restricted Access

About the authors

Alexander E. Kabakov

Medical Radiological Research Center of the Federal State Budgetary Institution of National Medical Research Center for Radiology of the Ministry of Health of the Russian Federation

Author for correspondence.
Email: aekabakov@hotmail.com
ORCID iD: 0000-0003-1041-1543
Russian Federation, Obninsk

Vera A. Mosina

Medical Radiological Research Center of the Federal State Budgetary Institution of National Medical Research Center for Radiology of the Ministry of Health of the Russian Federation

Email: mva210@rambler.ru
ORCID iD: 0009-0001-7667-6301
Russian Federation, Obninsk

Anna V. Khokhlova

Medical Radiological Research Center of the Federal State Budgetary Institution of National Medical Research Center for Radiology of the Ministry of Health of the Russian Federation

Email: demidkina@yandex.ru
ORCID iD: 0000-0002-4391-6321
Russian Federation, Obninsk

Sergey A. Ivanov

Medical Radiological Research Center of the Federal State Budgetary Institution of National Medical Research Center for Radiology of the Ministry of Health of the Russian Federation

Email: aekabakov@hotmail.com
Russian Federation, Obninsk

Andrey D. Kaprin

Medical Radiological Research Center of the Federal State Budgetary Institution of National Medical Research Center for Radiology of the Ministry of Health of the Russian Federation

Email: aekabakov@hotmail.com
Russian Federation, Obninsk

References

  1. Мкртчян Л.С., Замулаева И.А., Киселева В.И., Титова В.А., Крикунова Л.И. Рак шейки матки: химиолучевая терапия и прогностическая роль вируса папилломы человека. Под ред. С.А. Иванова и А.Д. Каприна. М.: ГЕОС, 2022. 190 с. [Mkrtchjan L.S., Zamulaeva I.A., Kiseleva V.I., Titova V.A., Krikunova L.I. Rak shejki matki: himioluchevaja terapija i prognosticheskaja rol’ virusa papillomy cheloveka. Рod red. S.A. Ivanova i A.D. Kaprina). Moskva: GEOS, 2022. 190 p. (In Russ)].
  2. Bhatla N., Aoki D., Sharma D.N., Sankaranarayanan R. FIGO Cancer Report 2018 Cancer of the cervix uteri. Int. J. Gynecol. Obstet. 2018;143(2):22–36.
  3. Datta N.R., Ordonez S.G., Gaipl U.S. et al. Local hyperthermia combined with radiotherapy and-/or chemotherapy: recent advances and promises for the future. Cancer Treat. Rev. 2015;41(9):742–753.
  4. Kokura S., Yoshikawa T., Ohnishi T. Hyperthermic oncology from bench to bedside. Springer, 2016. 444 p.
  5. Кудрявцев В.А., Макарова Ю.М., Кабаков А.Е. Термосенсибилизация опухолевых клеток ингибиторами активности и экспрессии шаперонов. Биомед. химия. 2012;58(6):662-672. [Kudryavtsev V.A., Makarova Yu.M., Kabakov A.E. Thermosensitization of tumor cells with inhibitors of chaperone activity and expression. Biochemistry (Moscow), Supplement Series B: Biomedical Chemistry. 2012;6(1): 61–67. (In Russ)]
  6. Кабаков А.Е., Анохин Ю.Н., Лебедева Т.В. Реакции нормальных и опухолевых клеток и тканей на гипертермию в сочетании с ионизирующей радиацией. Обзор. Радиация и риск. 2018;27(4):141-154. [Kabakov A.E., Anokhin Yu.N., Lebedeva T.V. Reactions of normal and tumor cells and tissues to hyperthermia in combination with ionizing radiation. review. Radiation and risk. 2018;27(4):141–154. (In Russ)]. doi: 10.21870/0131-3878-2018-27-4-141-154.
  7. Кабаков А.Е., Кудрявцев В.А., Хохлова А.В. и др. Апоптоз в опухолевых клетках, подвергнутых сочетанному действию гипертермии и облучения: исследование молекулярных механизмов и мишеней. Радиация и риск. 2018;27(2):62-75. [Kabakov A.E., Kudryavtsev V.A., Khokhlova A.V. et al. Apoptosis in tumor cells subjected to the combined action of hyperthermia and irradiation: a study of the molecular mechanisms and targets. Radiation and risk. 2018;27(2):62–75. (In Russ)]. doi: 10.21870/0131-3878-2018-27-2-62-75.
  8. Rossi A., Ciafre S., Balsamo M. et al. Targeting the heat shock factor 1 by RNA interference: a potent tool to enhance hyperthermochemotherapy efficacy in cervical cancer. Cancer Res. 2006;66(15):7678-7685. doi: 10.1158/0008-5472.CAN-05-4282.
  9. Hosokawa N., Hirayoshi K., Kudo H. et al. Inhibition of the activation of heat shock factor in vivo and in vitro by flavonoids. Mol. Cell. Biol. 1992;12(8):3490–3498.
  10. Westerheide S.D., Kawahara T.L.A., Orton K. et al. Triptolide, an inhibitor of the human heat shock response that enhances stress-induced cell death. J. Biol. Chem. 2006;281(14):9616–9622.
  11. Yoon Y.J., Kim J.A., Shin K.D. et al. KRIBB11 inhibits HSP70 synthesis through inhibition of heat shock factor 1 function by impairing the recruitment of positive transcription elongation factor b to the hsp70 promoter. J. Biol. Chem. 2011;286(3):1737–1747.
  12. Zaarur N., Gabai V.L., Porco Jr. J.A. et al. Targeting heat shock response to sensitize cancer cells to proteasome and Hsp90 inhibitors. Cancer Res. 2006;66(3): 1783–1791. doi: 10.1158/0008-5472.CAN-05-3692.
  13. Kudryavtsev V.A., Khokhlova A.V., Mosina V.A. et al. Induction of Hsp70 in tumor cells treated with inhibitors of the Hsp90 activity: A predictive marker and promising target for radiosensitization. PLoS One. 2017;2(3):e0173640. doi: 10.1371/journal.pone.0173640. eCollection 2017.
  14. Kabakov A.E., Gabai V.L. Cell death and survival assays. Methods Mol. Biol. 2018;1709:107–127.
  15. Kabakov A.E., Malyutina Ya.V., Latchman D.S. Hsf1-mediated stress response can transiently enhance cellular radioresistance. Radiat. Res. 2006;165(4):410–423. doi: 10.1667/rr3514.1.
  16. Малютина Я.В., Кабаков А.Е. Предрадиационная индукция белков теплового шока повышает клеточную радиорезистентность. Радиац. биология. Радиоэкология. 2007;47(3): 273-279. [Malyutina Ja.V., Kabakov A.E. Predradiatsionnaja induktsija belkov teplovogo shoka povyshaet kletochnuju radiorezistentnost. Radiats. Biol. Radioecol. 2007;47(3): 273–279. (In Russ)].
  17. Якимова А.О., Кабаков А.Е. Высокая термочувствительность клеток MDA-MB-231 как предпосылка для терморадиосенсибилизации трижды негативного рака молочной железы в клинической практике. Радиац. биология. Радиоэкология. 2023;63(1):273-279. [Yakimova A.O., Kabakov A.E. High thermosensitivity of MDA-MB-231 cells as a basis for thermoradiosensitization of triple negative breast cancer in clinical practice. Radiats. Biol. Radioecol.2023;63(1):273–279. (In Russ)]. doi: 10.31857/S0869803123010113

Supplementary files

Supplementary Files
Action
1. JATS XML
Download
2. Fig. 1. Segments of blots, showing how HSF1 knockdown with siRNA or inhibition of the HSF1 activity with quercetin prevents accumulation of the inducible form of HSP70 in heat-stressed HeLa cells: a – immunoblotting with anti-HSF1 antibodies to confirm the suppressed expression of HSF1 and anti-actin antibodies to ensure equal loading of cellular protein matter; b – immunoblotting with anti-HSP70 antibodies detecting the HSP70 level in cell lysates prepared at different time points after heating (43°C, 60 min) cells without HSF1 knockdown (upper panel) or with HSF1 knockdown (siRNA); c – immunoblotting with anti-HSP70 antibodies detecting the HSP70 level in cell lysates prepared at different time points after heating (43°C, 60 min) without the HSF1 inhibition (upper panel) or in the presence of 30 µmol/l quercetin (+querc). Time points following heating are indicated (in hours) at the upper edges of blots.

Download (281KB)
Indexing metadata
3. Fig. 2. The decrease in clonogenicity of HeLa cells subjected to hyperthermia alone (A) or hyperthermia followed by irradiation (B) after suppression in them of the expression or activity of HSF1. Colony survival in the untreated cells (Contr.) was considered as 100%. Before irradiation (3–7 Gy), the cells were incubated under hyperthermic conditions (Hyp.: 43°C, 60 min) without additional treatments or after knockdown of HSF1 (siRNA), or in the presence of inhibitors of HSF1 activity: 30 μmol/l quercetin (Querc), 10 n mol/l triptolide (Tript) or 15 µ mol/l KRIBB11. A: * Significant difference from control, p < 0.05; ** – significant difference from the control and also from the effect of monotreatment with hyperthermia marked *, p < 0.01. B: Аll values obtained for every dose of radiation after the combined treatments hyperthermia + HSF1 inhibitor (4 lower curves) are significantly different from the corresponding values in the control and also from the curve obtained for monotreatment with hyperthermia, p < 0.05.

Download (152KB)
Indexing metadata
4. Fig. 3. The enhanced cell death (apoptosis and necrosis) in HeLa cells subjected to hyperthermia alone or hyperthermia followed by irradiation after suppression of HSF1 expression or activity in them. Before irradiation (4 or 7 Gy), cells were heat-stressed (43°C, 60 min) without additional treatments or after knockdown of HSF1 (siRNA), or in the presence of inhibitors of HSF1 activity: 30 μmol/l quercetin (Querc), 10 n μmol/l M triptolide (Tript) or 15 µ μmol/l KRIBB11. * Significant difference from control, p < 0.05; ** significant difference from the control and from the value marked *, p < 0.05.

Download (286KB)
Indexing metadata

Copyright (c) 2024 Russian Academy of Sciences