An Arthropod Hormone, Ecdysterone, Inhibits the Growth of Breast Cancer Cells via Different Mechanisms
Open accessAbstract
Ecdysterone (Ecdy) is a hormone found in arthropods, which regulates their development. It is also synthesized by a number of plants to combat insect pests. It provides a number of beneficial pharmacological effects including the anabolic and adaptogenic ones. Ecdysterone is widely marketed as food supplement to enhance the physical performance of athletes. In addition to the estrogen receptor beta (ERbeta)-dependent anabolic effect of Ecdy in muscles, the molecular mechanisms of the plethora of other Ecdy-induced pharmacological effects remain unknown. The aim of this study was to investigate the pharmacological effect of ecdysterone on human breast cancer cell lines of different molecular subtypes. Surprisingly, in contrast to the anabolic effect on muscle tissues, we have revealed a tumor suppressive effect of Ecdy on a panel of breast cancer cell lines studied. Using the SeaHorse-based energy profiling, we have demonstrated that Ecdy dampened glycolysis and respiration, as well as greatly reduced the metabolic potential of triple negative breast cancer cell lines. Furthermore, we have revealed that Ecdy strongly induced autophagy. As part of the combined treatment, based on the Combination Index (CI) and Dose Reduction Index (DRI), Ecdy synergized with doxorubicin to induce cell death in several breast cancer cell lines. In contrast, Ecdy had only minor effect on non-transformed human fibroblasts. Collectively, our results indicate that ecdysterone can be considered as a new potential adjuvant for genotoxic therapy in treatment of breast cancer patients.
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Written from the paper’s full text.
TL;DR
Ecdysterone inhibited proliferation and induced death/autophagy in several human breast cancer cell lines, while having little effect on non-transformed fibroblasts. It also synergized with doxorubicin and 2-DG, suggesting potential as an adjuvant in breast cancer therapy.
Key findings
- Ecdysterone reduced growth of MCF7, MDA-MB-231, and MDA-MB-468 cells starting at 250–750 µM; no growth stimulation was observed, and colony number/size were reduced.
- Cell-cycle and death effects varied by line: MCF7 showed increased G1-phase; MDA-MB-468 was insensitive to cell-cycle changes. Cell death increased up to 1.6-fold in MCF7 and MDA-MB-231 and 3.5-fold in MDA-MB-468; Annexin V-positive apoptotic cells increased by 30% in MDA-MB-231 and 48% in MDA-MB-468, but not in MCF7.
- SeaHorse profiling showed metabolic suppression: in MDA-MB-231, 500 and 750 µM Ecdy decreased respiration by ~21% and ~31%; 500 µM reduced stressed OCR by 23% and stressed ECAR by 18%. In MDA-MB-468, 250 µM dampened respiration and glycolysis by 22%, while 750 µM inhibited them by 47% and 28%.
- Ecdy induced autophagy in all three breast cancer lines and synergized with doxorubicin (CI 0.47–0.89; doxorubicin DRI 1.4–17.9 times). Co-treatment with 2-DG increased cell death by 21% in MCF7 and 17% in MDA-MB-231; effects on non-transformed fibroblasts were minimal.
Methods
- Cell lines: MCF7 (luminal ER+PR+HER2−), MDA-MB-231 and MDA-MB-468 (TNBC), plus non-transformed DF2 and WI-38 fibroblasts; MEFs were also used in some assays.
- Assays included MTT, colony formation, proliferation counts, flow cytometry for cell cycle and Annexin V/PI or 7-AAD death, SeaHorse energy profiling, LysoTracker staining, chloroquine-blocked autophagic flux with LC3/p62 western blot and immunofluorescence, cytochrome C immunofluorescence, and Chou-Talalay CI/DRI analysis.
- Statistics: mean ± SD or SEM from at least three biological replicates; Student’s t-test with *p < 0.05 and p < 0.01.
Limitations
- The paper states that additional experiments are required to assess the effectiveness and toxicity of ecdysterone to organs and tissues before its therapeutic potential as an adjuvant can be evaluated, and that its effects on metabolism of different cancer and non-cancer cells should be studied further.
- Evident from the text: all experiments are in vitro cell-culture studies; no in vivo tumor models or clinical data are presented.
- Evident from Table 1: not all tested combinations were synergistic, as some CI values exceeded 1 (e.g., MCF7 with 0.25 µM doxorubicin plus 500 or 750 µM Ecdy, and MDA-MB-231 with 0.15 µM doxorubicin plus 750 µM Ecdy).
Model-written summary. Check it against the paper before relying on it.
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Claims the paper makes
0 of 28 cited sources checked · 0 supported · 0 flagged
- Findingown data
Ecdysterone strongly induced autophagy.
“Furthermore, we have revealed that Ecdy strongly induced autophagy.”
- Also stated in: § Ecdysterone Induces Autophagy · hedged
“Taken together, these data strongly suggest that ecdysterone induces autophagy in all breast cancer cells concomitantly with the increasing ecdysterone concentration.”
Quote in the paperSubstantial claim - Also stated in: § Ecdysterone Induces Autophagy · hedged
- Findingown data
No increase in cell proliferation was detected upon ecdysterone treatment.
“No increase in cell proliferation was detected.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone significantly increased the number of MCF7 cells in the G1-phase.
“It significantly increased the number of MCF7 cells in the G1-phase”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone had no effect on the cell cycle of MDA-MB-468 cells.
“However, MDA-MB-468 cells were insensitive to Ecdy-induced alterations in the cell cycle”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone elevated overall cell death in all three breast cancer cell lines.
“although Ecdy elevated overall cell death in all cell lines”
Quote in the paperSubstantial claim
- Findingown data
Ecdysterone increased the rate of cell death 3.5 times in MDA-MB-468 cells.
“and 3.5 times in the case of MDA-MB-468 cells”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone affected the cell cycle distributions of MCF7 and MDA-MB-231 cells to varying degrees.
“The analysis of the cell cycle has shown that ecdysterone affected to varying degrees the cell cycle distributions of MCF7 and MDA-MB-231 cells, but had no effect on MDA-MB-468 cells”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone increased the population of Annexin V-positive (apoptotic) cells by 30% for MDA-MB-231 cells.
“In contrast, Ecdy increased the population of Annexin V-positive (apoptotic) cells by 30% for MDA-MB-231”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone had only a minor effect on non-transformed human fibroblasts.
“In contrast, Ecdy had only minor effect on non-transformed human fibroblasts.”
- Also stated in: § Introduction · plain
“Importantly, the effect of ecdysterone on non-transformed human fibroblasts was minimal.”
Quote in the paperSubstantial claim - Also stated in: § Introduction · plain
- Findingown data
Ecdysterone synergized with doxorubicin to induce cell death in several breast cancer cell lines.
“As part of the combined treatment, based on the Combination Index (CI) and Dose Reduction Index (DRI), Ecdy synergized with doxorubicin to induce cell death in several breast cancer cell lines.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone increased the rate of cell death up to 1.6 fold for both MCF7 and MDA-MB-231 cells.
“Annexin V/PI staining followed by flow cytometry (Figure 2E) demonstrated that ecdysterone increased the rate of cell death up to 1.6 fold for both MCF7 and MDA-MB-231 cells”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone inhibits the growth of all three breast cancer cell lines in the colony-formation assay.
“Results shown in Figures 1D–F confirm that the treatment with Ecdy inhibits the growth of all three cell lines.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone did not increase the population of Annexin V-positive cells in MCF7 cells.
“it did not increase the population of Annexin V-positive cells”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone treatment down-regulated all three breast cancer cell lines (MCF7, MDA-MB-231, MDA-MB-468) starting at a concentration of 250-750 μM in the MTT assay.
“Results shown in Figures 1A–C clearly demonstrate that in our case the treatment with Ecdy has down-regulated all three cell lines starting with a concentration of 250–750 µM.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone has a tumor suppressive effect on a panel of breast cancer cell lines studied.
“Surprisingly, in contrast to the anabolic effect on muscle tissues, we have revealed a tumor suppressive effect of Ecdy on a panel of breast cancer cell lines studied.”
- Also stated in: § Introduction · plain
“Here, we provide evidence that ecdysterone can inhibit the proliferation of breast cancer cells.”
Quote in the paperSubstantial claim - Also stated in: § Introduction · plain
- Findingown data
Ecdysterone induces autophagy in all breast cancer cells concomitantly with increasing ecdysterone concentration.
“Taken together, these data strongly suggest that ecdysterone induces autophagy in all breast cancer cells concomitantly with the increasing ecdysterone concentration.”
Quote in the paperSubstantial claim - Findingown data
Combined treatment with doxorubicin or 2-DG and 250 μM ecdysterone increased cell death by several times relative to control or individual drug treatment.
“Figures 6A–F and Supplementary Figures S2A–C demonstrate that the combined treatment (either doxorubicin or 2-DG with 250 μM Ecdy) both increased the level of cell death by several times relative to control, or in comparison to the treatment with an individual drug.”
Quote in the paperSubstantial claim - Findingown data
750 μM ecdysterone inhibited respiration and glycolysis further by 47% and 28%, respectively, in MDA-MB-468 cells.
“whereas 750 μM Ecdy inhibited them further by 47 and 28%, respectively”
Quote in the paperSubstantial claim - Findingown data
Co-treatment of Ecdy with 2-DG significantly enhanced both apoptosis and total cell death.
“Co-treatment of Ecdy with 2-DG also significantly enhanced both apoptosis and total cell death (Figures 6C–F).”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone synergizes with doxorubicin to down-regulate the proliferation of breast cancer cells, with a pronounced synergistic interaction (CI 0.47-0.89) across all three Ecdy concentrations.
“CI plots (Figures 5D–F) and Table 1 show that all three Ecdy concentrations have a pronounced synergistic (CI ranges 0.47–0.89) interaction with doxorubicin.”
- Also stated in: § Ecdysterone synergizes With Doxorubicin to Down-Regulate Breast Cancer Cell Lines · boosted
“These data clearly demonstrate the ability of ecdysterone to synergize with doxorubicin to down-regulate the proliferation of breast cancer cells.”
- Also stated in: § Ecdysterone Significantly Enhances Doxorubicin- and 2-DG-Induced Cell Death · hedged
“Taken together, these data suggest that ecdysterone mediates strong synergy with doxorubicin in attenuation of the proliferation of breast cancer cells.”
Quote in the paperSubstantial claim - Also stated in: § Ecdysterone synergizes With Doxorubicin to Down-Regulate Breast Cancer Cell Lines · boosted
- Findingown data
Ecdysterone increased the population of Annexin V-positive (apoptotic) cells by 48% for MDA-MB-468 cells.
“and by 48% for MDA-MB-468 cell line”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone allows reduction of the effective doxorubicin dose by 1.4 to 17.9 times depending on the cell line.
“In turn, the use of Ecdy allowed the reduction of the effective dose of doxorubicine from 1.4 to 17.9 times (Table 1), depending on the particular cell line.”
Quote in the paperSubstantial claim - Findingown data
Treatment with Ecdy significantly enhanced doxorubicin-induced release of cytochrome C in all cancer cell lines.
“Taken together, results shown in Figure 7 and Supplementary Figures S2, S3 confirm that treatment with Ecdy significantly enhanced the doxorubicin-induced release of cytochrome C in all cancer cell lines.”
Quote in the paperSubstantial claim - Findingown data
Ecdy down-regulated proliferation of cancer cell lines significantly more robustly compared to normal human fibroblasts at all concentrations tested.
“As shown in Supplementary Figure S4A,B, Ecdy down-regulated the proliferation of cancer cell lines significantly more robustly compared to normal human fibroblasts at all concentrations tested.”
Quote in the paperSubstantial claim - Findingown data
Combined treatment with doxorubicin and ecdysterone has incomparably low inhibitory effect on WI-38 and MEFs in contrast to breast cancer cells.
“Regarding WI-38 and MEFs (Figures 7B,C), the same combined treatment has incomparable low inhibitory effect in contrast to breast cancer cells (Figure 5).”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone strongly induced autophagy in all three breast cancer cell lines, as shown by immunoblotting and immunofluorescence.
“Results of both immunoblotting and immunofluorescence (Figures 4D–G; Supplementary Figure S1) demonstrate that in the case of all three lines, ecdysterone strongly induced autophagy.”
Quote in the paperSubstantial claim - Findingown data
Combined treatment with doxorubicin and ecdysterone displayed a small protection of DF2 cells from doxorubicin.
“In the case of DF2 cells, the combined treatment (doxorubicin + ecdysterone) displayed even a small protection of these cells from doxorubicin (Supplementary Figure S7A).”
Quote in the paperNot a substantial claim - Findingown data
Co-treatment of Ecdy with 2-DG increased the rate of cell death by 21% for MCF7 and 17% for MDA-MB-231 cells.
“It increased the rate of cell death by 21% for MCF7 and 17% for MDA-MB-231 cells.”
Quote in the paperSubstantial claim - Findingown data
Ecdy induced autophagy in fibroblasts to a much lesser extent than in breast cancer cells.
“Results shown in Supplementary Figure S6 demonstrate that Ecdy induced autophagy in fibroblasts to a much lesser extent than in breast cancer cells (Figure 4).”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone inhibited respiration (basic OCR) in MDA-MB-231 cells.
“Energy profiling of MDA-MB-231 cells treated with 0, 250, 500, 750 or 1,000 μM of ecdysterone has shown ecdysterone-mediated inhibition of respiration (basic OCR).”
Quote in the paperSubstantial claim - Findingown data
Flow cytometry analysis showed a strong increase of LysoTracker fluorescence in all three breast cancer cell lines in order of increasing ecdysterone concentration.
“Flow cytometry analysis has shown the strong increase of LysoTracker fluorescence in all three cell lines in order of increase the ecdysterone concentration”
Quote in the paperSubstantial claim - Findingown data
250 μM ecdysterone dampened respiration and glycolysis by 22% in MDA-MB-468 cells.
“Even 250 μM Ecdy dampened respiration and glycolysis by 22%”
Quote in the paperSubstantial claim - Findingown data
In most cases, addition of 250 μM ecdysterone was sufficient to down-regulate growth of breast cancer cells 1.5-2 times more efficiently than corresponding doxorubicin concentrations.
“It is important to notice that in most cases the addition of even 250 μM ecdysterone was sufficient to down-regulate the growth of breast cancer cells 1.5–2 times more efficiently than the corresponding concentrations of doxorubicin (Figures 5A–C).”
Quote in the paperSubstantial claim - Findingown data
500 and 750 μM ecdysterone decreased respiration in MDA-MB-231 cells by approximately 21% and 31%, respectively.
“For instance, 500 and 750 μM Ecdy decreased respiration by approximately 21 and 31%, respectively”
Quote in the paperSubstantial claim - Findingown data
The minimal effective concentration of Ecdy (250 μM) in combination with doxorubicin that induced death of breast cancer cells had almost no effect on DF2 and WI38 fibroblasts.
“Surprisingly, the minimal effective concentration of Ecdy (250 uM) in combination with doxorubicin that induced death of breast cancer cells, had almost no effect on DF2 and WI38 fibroblasts (Figure 6; Supplementary Figure S2).”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone significantly decreased the metabolic potential (both stressed respiration and glycolysis) in MDA-MB-231 cells.
“but significantly decreased the metabolic potential (both stressed respiration (stressed OCR) and glycolysis (stressed ECAR), Figures 3A,B)”
Quote in the paperSubstantial claim - Findingnone given
Ecdysterone mediated the inhibition of growth of all breast cancer cell lines.
“Given that ecdysterone mediated the inhibition of growth of all breast cancer cell lines, we decided to examine its effect on cancer cells a part of the combined treatment with doxorubicin, a genotoxic drug which is widely applied as chemotherapeutics.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone did not alter glycolysis (basic ECAR) in MDA-MB-231 cells.
“At the same time, ecdysterone did not alter glycolysis (basic ECAR, Figure 3B)”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone significantly sensitizes all breast cancer cell lines to doxorubicin treatment.
“The results shown in Figures 5A–C demonstrate that ecdysterone significantly sensitizes all breast cancer cell lines to the treatment with doxorubicin.”
Quote in the paperSubstantial claim - Findingreasoning
Ecdy sensitizes breast cancer cell lines to 2-DG.
“Moreover, we have revealed that Ecdy sensitizes breast cancer cell lines to 2-DG which is in accordance with Ecdy-mediated down-regulation of energy metabolism.”
Quote in the paperSubstantial claim - Findingown data
Ecdy elicited a two-fold increase in cell death of MDA-MB-468 cells relative to MCF7.
“Furthermore, it elicited a two-fold increase in cell death of the MDA-MB-468 cells relative to MCF7.”
Quote in the paperSubstantial claim - Findingown data
Ecdysterone did not stimulate growth of cancer cells.
“Surprisingly, despite the fact that anabolic properties of ecdysterone in muscles have been reported (Parr et al., 2015; Isenmann et al., 2019), we have not observed ecdysterone-mediated growth stimulation of cancer cells.”
Quote in the paperSubstantial claim - Findingown data
Ecdy strongly induced autophagy in breast cancer cells, in contrast to non-transformed human fibroblasts.
“In line with these observations, we have demonstrated that Ecdy strongly induced autophagy in breast cancer cells, in contrast to non-transformed human fibroblasts.”
Quote in the paperSubstantial claim - Findingown data
Administration of ecdysterone caused attenuation of cell growth of breast cancer cells starting from concentration of 250-750 μM.
“Instead, in our MTT experiments administration of ecdysterone caused the attenuation of cell growth of breast cancer cells starting from the concentration of 250–750 μM.”
Quote in the paperSubstantial claim - Findingown data
Ecdy was not able to sensitize non-cancerous fibroblast cells to doxorubicin as observed for breast cancer cells.
“Noteworthy, Ecdy was not able to sensitize non-cancerous (fibroblast) cells to doxorubicin as it was observed for breast cancer cells.”
Quote in the paperSubstantial claim - Findingreasoning
Ecdy strongly enhanced the action of doxorubicin in concentrations (250, 500, and 750 μM), which are sufficient to inhibit energy metabolism and induce autophagy.
“Notably, Ecdy strongly enhanced the action of doxorubicin in concentrations (250, 500, and 750 µM), which are sufficient to inhibit energy metabolism and induce autophagy.”
Quote in the paperSubstantial claim - Findingnone given
When Ecdy was used alone, it failed to significantly down-regulate the proliferation of cancer cells.
“It is important to note that when Ecdy was used alone, it failed to significantly down-regulate the proliferation of cancer cells.”
Quote in the paperSubstantial claim - Interpretationown data
Ecdysterone can be considered as a new potential adjuvant for genotoxic therapy in treatment of breast cancer patients.
“Collectively, our results indicate that ecdysterone can be considered as a new potential adjuvant for genotoxic therapy in treatment of breast cancer patients.”
Quote in the paperSubstantial claim- Rests on an assumption: The concentrations of ecdysterone used in vitro (250–750 µM) are achievable in humans and relevant to breast cancer treatment.
- Rests on an assumption: Findings in breast cancer cell lines translate to breast cancer patients.
- Rests on an assumption: The three cell lines studied (MCF7, MDA-MB-231, MDA-MB-468) are representative of breast cancer molecular subtypes.
- Rests on an assumption: The minor effect of ecdysterone on non-transformed human fibroblasts indicates that it is safe for normal tissues in patients.
- Interpretationown data
Ecdysterone elicited death of MCF7 cells via mechanism(s) other than apoptosis.
“suggesting that Ecdy elicited death of MCF7 cells via mechanism(s) other than apoptosis”
Quote in the paperSubstantial claim - Interpretationown data
Ecdysterone was able to attenuate the level of energy metabolism in TNBC breast cancer cells.
“Taken together, these data suggest Ecdy was able to attenuate the level of energy metabolism in TNBC breast cancer cells.”
Quote in the paperSubstantial claim - Interpretationown data
The ability of ecdysterone to induce cell cycle arrest or elicit cell death of breast cancer cells presumably depends on the genetic background of a particular cell line.
“These data suggest that the ability of ecdysterone to induce cell cycle arrest or elicit cell death of breast cancer cells presumably depends on the genetic background of a particular cell line.”
Quote in the paperSubstantial claim - Interpretationown data
Ecdysterone greatly reduced the capacity of MDA-MB-231 cells for metabolic adaptation.
“Thereby, these results suggest that ecdysterone greatly reduced the capacity of MDA-MB-231 cells for metabolic adaptation.”
Quote in the paperSubstantial claim - Interpretationcitation
The ERbeta-signaling pathway is not the only molecular mechanism utilized by ecdysterone.
“Considering the diversity of ecdysterone-mediated pharmacological activities in the human organism, including different tissues (see Dinan and Lafont, 2006), it seems that the ERbeta-signaling pathway is not the only molecular mechanism which is utilized by ecdysterone.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Dinan and Lafont, 2006
Not checked in the free run.
- Interpretationown data
Ecdy-mediated negative regulation of energy metabolism in TNBC cells can be potentially important for treatment of this most dangerous sub-type of breast malignancy.
“Ecdy-mediated negative regulation of the energy metabolism in TNBC cells can be potentially important for the treatment of this most dangerous sub-type of breast malignancy.”
Quote in the paperSubstantial claim - Premisecitation
Ecdy significantly stimulates the chemotherapeutic effect of cisplatin in mice models.
“Furthermore, Ecdy was shown to significantly stimulate the chemotherapeutic effect of cisplatin in mice models (Konovalova et al., 2002).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Konovalova et al., 2002
Not checked in the free run.
- Premisecitation
Ecdysterone made both drug-resistant and non-drug-resistant cancer cells more susceptible to doxorubicin treatment.
“Our results are in accordance with other studies (Konovalova et al., 2002; Martins et al., 2012; Martins et al., 2015) that have shown that ecdysterone made both drug-resistant and non-drug-resistant cancer cells more susceptible to doxorubicin treatment.”
Quote in the paperSubstantial claim0 of 3 sources checked
- May combine two claims. 21
- Classified as probably not a substantial claim. 13
May combine two claims.
Findingown dataEcdysterone significantly reduced both the number and the size of colonies in all three breast cancer cell lines.
“Moreover, photographs of plates with colonies clearly showed that not only the number of colonies, but also their size, was significantly reduced upon treatment with Ecdy.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataEcdysterone dampened glycolysis and respiration, and greatly reduced the metabolic potential of triple negative breast cancer cell lines.
“Using the SeaHorse-based energy profiling, we have demonstrated that Ecdy dampened glycolysis and respiration, as well as greatly reduced the metabolic potential of triple negative breast cancer cell lines.”
Quote in the paperSubstantial claimMay combine two claims.
Findingnone givenEcdysterone strongly sensitizes breast cancer cells to doxorubicin, resulting in a significant reduction of the effective dose of doxorubicin.
“Moreover, ecdysterone strongly sensitizes breast cancer cells to doxorubicin, resulting in a significant reduction of the effective dose of doxorubicin.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown data500 μM ecdysterone mitigated stressed OCR by 23% and stressed ECAR by 18% in MDA-MB-231 cells.
“Thus, 500 μM Ecdy mitigated stressed OCR by 23% and stressed ECAR by 18%.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataCo-treatment with doxorubicin and ecdysterone elevated apoptosis in MCF7 cells by 23% and in MDA-MB-231 cells by 3.15 times compared to doxorubicin alone.
“Accordingly, co-treatment with doxorubicin and ecdysterone elevated the level of apoptosis in MCF7 cells by 23% and in MDA-MB-231 cells by 3.15 times, respectively, compared to doxorubicin alone.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataIncreased ecdysterone concentration up-regulated survival of WI-38 and MEFs cells, in contrast to breast cancer cell lines.
“Supplementary Figure S8 demonstrates that in contrast to breast cancer cell lines (Figure 5), an increased ecdysterone concentration up-regulated the survival of WI-38 and MEFs cells (Supplementary Figures S7B,C).”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataIn MDA-MB-468 cells, ecdysterone inhibited both baseline respiration and glycolysis.
“In the case of MDA-MB-468 cell line, the same treatment with ecdysterone has led to an inhibition of both the baseline respiration and of glycolysis”
Quote in the paperSubstantial claimMay combine two claims.
FindingreasoningEcdy only weakly inhibits proliferation of fibroblasts in the proliferation assay, but activates fibroblasts in MTT assay.
“Although Ecdy only weakly inhibits proliferation of fibroblasts in the proliferation assay, it does activate fibroblasts in MTT assay which may result from their metabolic activation.”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataSeaHorse energy profiling showed that Ecdy significantly dampened respiration and metabolic potential of MDA-MB-231 cells, and strongly reduced both respiration and glycolysis in MDA-MB-468 cells.
“The SeaHorse energy profiling has shown that Ecdy significantly dampened respiration, as well as the metabolic potential of MDA-MB-231 cells, and strongly reduced both respiration and glycolysis in MD-MB-468 cells.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataEcdy significantly affected the cell cycle distribution of MCF7 cells but had almost no effect on cell cycle of MDA-MB-468 cells.
“While Ecdy significantly affected the cell cycle distribution of MCF7 cells, it had almost no effect on cell cycle of MDA-MB-468 cells.”
Quote in the paperSubstantial claimMay combine two claims.
PremisecitationSeveral studies described ecdysterone-induced sensitization of cancer cells to genotoxic drugs and reduction of tumors in mice.
“It is interesting to note that despite the reported anabolic properties of ecdysterone regarding sport performance, several studies described ecdysterone-induced sensitization of cancer cells to genotoxic drugs and reduction of tumors in mice (Konovalova et al., 2002; Martins et al., 2015).”
Quote in the paperNot a substantial claim0 of 2 sources checked- cites Konovalova et al., 2002
Not checked in the free run.
- cites Martins et al., 2015 source
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May combine two claims.
Interpretationown dataEcdysterone significantly down-regulates cancer cells with no or little effect on fibroblasts.
“Taken together, our experiments show that ecdysterone significantly down-regulates cancer cells with no or little effect on fibroblasts.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationEcdysterone was shown in vitro to stimulate ERbeta, which is involved in skeletal muscle hypertrophy.
“However, ecdysterone was shown in vitro to stimulate ERbeta, which is involved in skeletal muscle hypertrophy (Parr et al., 2014).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Parr et al., 2014 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTang et al. reported that ecdysterone promotes autophagy in osteoporotic rats.
“Tang and colleagues (Tang et al., 2018) has reported that ecdysterone promotes autophagy in osteoporotic rats.”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Tang et al., 2018 source
Not checked in the free run.
Classified as probably not a substantial claim.
Premisenone givenVery little information is available about the effects of ecdysterone on cancer cells and malignancies.
“Unlike for other hormone-like compounds, very little information is available about the effects of ecdysterone on cancer cells and malignancies.”
Quote in the paperSubstantial claimClassified as probably not a substantial claim.
PremisereasoningOther molecular mechanisms underlying the broad spectrum of ecdysterone-mediated pharmacological effects remain elusive.
“However, other molecular mechanisms underlying the broad spectrum of ecdysterone-mediated pharmacological effects remain elusive.”
Quote in the paperSubstantial claimMay combine two claims.
PremisecitationModulation of cancer-specific metabolic adaptations weakens malignant cells and widens the therapeutic window for effective treatment of TNBC patients.
“The modulation of cancer-specific metabolic adaptations weakens the malignant cells and widens the therapeutic window for effective treatment of TNBC patients (Lanning et al., 2017; Wang et al., 2020).”
Quote in the paperSubstantial claim0 of 2 sources checkedProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
Premisecitation2-DG is a promising inhibitor of glycolysis that decreases energy of cancer cells, making chemotherapy and other treatments more effective, and is most likely useful for treatment of breast cancer including TNBC.
“2-DG is a promising inhibitor of glycolysis, which decreases the energy of the cancer cells thus making chemotherapy and other treatments more effective. It underwent clinical trials and most likely is useful for the treatment of breast cancer including TNBC (Wokoun et al., 2017; Lucantoni et al., 2018; O’Neill et al., 2019).”
Quote in the paperNot a substantial claim0 of 3 sources checkedProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationThis effect is mediated by Ecdy-dependent induction of autophagy, which counteracts the effect of apoptosis.
“This effect is mediated by Ecdy-dependent induction of autophagy, which counteracts the effect of apoptosis.”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Wen et al., 2019 source
Not checked in the free run.
May combine two claims.
Interpretationown dataEcdy can negatively regulate cancer cells through various mechanisms because it inhibited the cell cycle and induced death to a different extent depending on the particular cellular background.
“Apparently, Ecdy can negatively regulate cancer cells through various mechanisms because it inhibited the cell cycle and induced death to a different extend depending on the particular cellular background.”
Quote in the paperSubstantial claimClassified as probably not a substantial claim.
PremisecitationNumerous studies have documented oncogenic properties of male steroid hormones and their derivatives on several human organs, including testis, liver, breast, and others.
“Numerous studies have documented oncogenic properties of male steroid hormones and its derivatives on several human organs, including testis, liver, breast, and others (Sirianni et al., 2012; Salerno et al., 2018).”
Quote in the paperSubstantial claim0 of 2 sources checkedProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationMetabolic reprogramming is a known hallmark of cancer cells, in which they manifest diverse metabolic phenotypes to maintain their proliferation and to combat anticancer therapies.
“Metabolic reprogramming is a known hallmark of cancer cells, in which they manifest diverse metabolic phenotypes to maintain their proliferation and to combat anticancer therapies (Shuvalov et al., 2017; Sun et al., 2020).”
Quote in the paperNot a substantial claim0 of 2 sources checkedProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationEcdy protects from degeneration human nucleus pulposus cells, which form the inner core of the vertebral disc.
“In addition, Ecdy protects from degeneration human nucleus pulposus cells, which form the inner core of the vertebral disc (Wen et al., 2019).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Wen et al., 2019 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationEcdy can promote autophagy upon onset of osteoporosis in rats.
“Ecdy can promote autophagy upon the onset of osteoporosis in rats (Tang et al., 2018).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Tang et al., 2018 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationAutophagy can play dual roles in both tumor promotion and suppression.
“Although autophagy can play dual roles in both tumor promotion and suppression (Yun and Lee, 2018), in terms of chemotherapy autophagy is usually considered as a mechanism of drug-resistance against therapeutics.”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Yun and Lee, 2018 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationEcdysterone possesses a variety of beneficial pharmacological effects on humans, including anabolic and adaptogenic ones.
“However, ecdysterone possesses a variety of beneficial pharmacological effects on humans, including anabolic and adaptogenic ones (Báthori et al., 2008).”
- Also stated in: § Discussion · boosted
“Although positive ecdysterone-mediated pharmacological influence on organisms is well documented (Lafont and Dinan, 2003), we decided to examine possible pharmacological effects of ecdysterone on proliferation of human breast cancer cell lines of different molecular subtypes.”
Quote in the paperNot a substantial claim0 of 2 sources checked- Also stated in: § Discussion · boosted
May combine two claims.
PremisecitationDoxorubicin produces multiple severe side effects including cumulative cardiotoxicity, acute nausea and vomiting, gastrointestinal disturbances, alopecia baldness, and neurologic disturbances.
“The latter produces multiple severe side effects including the cumulative cardiotoxicity, acute nausea and vomiting, gastrointestinal disturbances, alopecia baldness, and neurologic disturbances (Carvalho et al., 2009).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Carvalho et al., 2009 source
Not checked in the free run.
May combine two claims.
Interpretationown dataIn moderate concentrations, Ecdy sensitizes cancer cells to treatments with chemotherapeutic agents and thus can potentially serve as an adjuvant therapeutic.
“Taken together, these data suggests that in moderate concentrations, Ecdy sensitizes cancer cells to treatments with chemotherapeutic agents and thus can potentially serve as an adjuvant therapeutic.”
Quote in the paperSubstantial claimMay combine two claims.
PremisecitationEcdy enhances the ability to cope with stress and enhances resistance to tiredness.
“Furthermore, since Ecdy enhances the ability to cope with stress and enhances resistance to tiredness (Báthori et al., 2008; Dinan et al., 2009), it seems beneficial to administer it as part of cytotoxic therapy with doxorubicin.”
Quote in the paperSubstantial claim0 of 2 sources checkedProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationDoxorubicin-induced autophagy is involved in development of chemoresistance, and inhibition of autophagy effectively overcomes doxorubicin resistance in a variety of cancers.
“For example, doxorubicin-induced autophagy is involved in the development of chemoresistance, and the inhibition of autophagy effectively overcomes doxorubicin resistance in a variety of cancers (Zhou et al., 2019).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Zhou et al., 2019 source
Not checked in the free run.
The paper relies on this without support.
The concentrations of ecdysterone used in vitro (250–750 µM) are achievable in humans and relevant to breast cancer treatment.
“Then, 0–1,000 μM Ecdy was added for 48 h.”
“Results shown in Figures 1A–C clearly demonstrate that in our case the treatment with Ecdy has down-regulated all three cell lines starting with a concentration of 250–750 µM.”
The paper relies on this without support.
Findings in breast cancer cell lines translate to breast cancer patients.
“Collectively, our results indicate that ecdysterone can be considered as a new potential adjuvant for genotoxic therapy in treatment of breast cancer patients.”
The paper relies on this without support.
The three cell lines studied (MCF7, MDA-MB-231, MDA-MB-468) are representative of breast cancer molecular subtypes.
“We evaluated the effect of this drug on three human breast cancer cell lines with different molecular properties: MCF7 (luminal, ER+Pr+Her2−), MDA-MB-231 (TNBC) and MDA-MB-468 (TNBC).”
The paper relies on this without support.
Ecdysterone does not display hormonal activity in humans because they have no EcRs, nor does it interact with androgen or glucocorticoid receptors.
“However, this compound does not display any hormonal activity in humans because they have no EcRs, nor it interacts with androgen or glucocorticoid receptors.”
The paper relies on this without support.
The observed metabolic down-regulation and autophagy induction are mechanisms by which ecdysterone inhibits breast cancer cell growth.
“An Arthropod Hormone, Ecdysterone, Inhibits the Growth of Breast Cancer Cells via Different Mechanisms”
“Mechanistically, it down-regulates the metabolic potential of cancer cells and induces autophagy.”
The paper relies on this without support.
The minor effect of ecdysterone on non-transformed human fibroblasts indicates that it is safe for normal tissues in patients.
“In contrast, Ecdy had only minor effect on non-transformed human fibroblasts.”
The paper relies on this without support.
Among breast cancers, the TNBC subtype has the worst prognosis with questioned targeted therapies.
“Among breast cancers, the TNBC subtype has the worst prognosis with questioned targeted therapies.”
The paper relies on this without support.
In terms of chemotherapy, autophagy is usually considered as a mechanism of drug-resistance against therapeutics.
“Although autophagy can play dual roles in both tumor promotion and suppression (Yun and Lee, 2018), in terms of chemotherapy autophagy is usually considered as a mechanism of drug-resistance against therapeutics.”
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
In vitro studies associate ecdysterone with endothelial dysfunction prevention, osteoporosis prevention, glucose regulation, cytotoxic effects, antibacterial properties, immunomodulatory effects, and anti-inflammatory activities.
“Moreover, the effects on cell lines are associated with endothelial dysfunction prevention, osteoporosis prevention, glucose regulation, cytotoxic effects, antibacterial properties, immunomodulatory effects, and anti-inflammatory activities [52,53,54,55,56,57,58,60,61,62,63,64,65,66,67,68,69,70,71,72,73].”
From Ecdysterone and Turkesterone—Compounds with Prominent Potential in Sport and Healthy Nutrition (2024) · cites this paper as [61]
The Abstract and Results report that ecdysterone suppressed breast cancer cell growth and enhanced drug-induced cell death, supporting the cytotoxic-effects portion. The Discussion reports reduced glycolysis in breast cancer cell lines, but not general glucose regulation. The Introduction mentions anti-diabetic and immune-stimulating effects without establishing them as in-vitro cell-line findings; our copy does not substantiate the claimed endothelial, osteoporosis-preventive, antibacterial, or anti-inflammatory cell-line effects. (LLM, conf=0.88)
Experimental — accuracy not yet measuredWithin the in vitro studies of ecdysterone, no serious effects were reported.
“Within these studies, no serious effects were reported [52,53,54,55,56,57,58,60,61,62,63,64,65,66,67,68,69,70,71,72,73].”
From Ecdysterone and Turkesterone—Compounds with Prominent Potential in Sport and Healthy Nutrition (2024) · cites this paper as [61]
The Results section reports that ecdysterone had no or little effect on the non-transformed fibroblasts tested, supporting a limited safety observation. It does not establish that no serious effects occurred across in vitro studies; the Discussion says further experiments are needed to assess toxicity to organs and tissues. Page numbers are unavailable in our copy. (LLM, conf=0.84)
Experimental — accuracy not yet measured20E activity observed in TNBC cell lines that lack ER argues that the wide range of pharmacological activities of 20E in tissues of different origins cannot be determined by only MAS1 and ER targets.
“It should be noted, however, that we observed 20E activity also in triple-negative breast cancer (TNBC) cell lines that lack ER [41], arguing that the wide range of pharmacological activities of 20E in tissues of different origins cannot be determined by only these targets.”
From 20-Hydroxyecdysone Boosts Energy Production and Biosynthetic Processes in Non-Transformed Mouse Cells (2024) · cites this paper as [41]
Previous studies revealed 20E-dependent inhibition of glycolysis and respiration in breast carcinoma and non-small cell lung cancer cell lines.
“In contrast to myoblasts and fibroblasts, we have previously revealed the 20E-dependent inhibition of glycolysis and respiration in breast carcinoma and non-small cell lung cancer cell lines [41,42].”
From 20-Hydroxyecdysone Boosts Energy Production and Biosynthetic Processes in Non-Transformed Mouse Cells (2024) · cites this paper as [41]
20E and its derivatives sensitize breast cancer cells to genotoxic stress, induce autophagy, and attenuate multiple drug resistance.
“We and others have shown that 20E and its derivates sensibilize breast cancer cells to genotoxic stress, induce autophagy, and attenuate multiple drug resistance [12,13,14,15,16,17,18,19].”
From 20-Hydroxyecdysone Confers Antioxidant and Antineoplastic Properties in Human Non-Small Cell Lung Cancer Cells (2023) · cites this paper as [12]
20E may possess antineoplastic activity.
“Recent studies have shown that 20E may also possess antineoplastic activity.”
From 20-Hydroxyecdysone Confers Antioxidant and Antineoplastic Properties in Human Non-Small Cell Lung Cancer Cells (2023) · cites this paper as [12]
20E and its derivatives modulate the resistance of cancer cells to genotoxic drugs and mitigate multidrug resistance.
“Furthermore, it has been shown previously that 20E and its derivates modulate the resistance of cancer cells to genotoxic drugs and mitigate multidrug resistance [12,13,14,15,16,17,18,19].”
From 20-Hydroxyecdysone Confers Antioxidant and Antineoplastic Properties in Human Non-Small Cell Lung Cancer Cells (2023) · cites this paper as [12]
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
Each contradiction between two passages listed here was confirmed by an independent check.
Some reported drug combinations are not synergistic
MajorThe paper defines CI < 1 as synergistic, but Table 1 reports CI values above 1 for some combinations. The stated CI range of 0.47-0.89 also excludes those values.
“CI plots (Figures 5D–F) and Table 1 show that all three Ecdy concentrations have a pronounced synergistic (CI ranges 0.47–0.89) interaction with doxorubicin.”
“| MCF7 | 0,25 | 500 | 0,6 | 1,07 | 3,04 | | MCF7 | 0,25 | 750 | 0,57 | 1,13 | 2,17 |”
Colony-formation results are labeled SD and SEM
MinorThe assay methods specify mean ± SD for the colony-formation results, while the Figure 1 caption specifies mean ± SEM for those results.
“Results are represented as the mean ± SD of three biological replicates.”
“Photographs of plates and quantification diagrams are shown. Results are displayed as the mean ± SEM of three experiments.”
Table includes doxorubicin doses outside the stated MTT assay doses
MinorThe described combination MTT assay used 0.15 or 0.25 μM doxorubicin, whereas Table 1 also reports combinations using 0.5 and 0.75 μM.
“To this end, we carried out MTT-assay using ecdysterone (250, 500, or 750 μM) and doxorubicin (0.15 or 0.25 μM) alone or in combination.”
“| MCF7 | 0,75 | 250 | 0,17 | 0,59 | 17,86 |”
The citation lists haven’t been fetched yet
References 36 · Cited by 30
- Authors
- Shuvalov, O., Fedorova, O., Tananykina, E., Gnennaya, Y., Daks, A., Petukhov, A., & Barlev, N. A.
- Affiliations
- Institute of Cytology (RU)
- Federal Almazov North-West Medical Research Centre (RU)
- Venue
- Frontiers in Pharmacology
- Publisher
- Frontiers Media SA
- Published
- 2020-10-30
- Volume
- 11
- Pages
- 561537-561537
- ISSN
- 1663-9812 (ISSN-L)
- PMC
- PMC7663021
- OpenAlex
- W3096417141
- Cited by
- 30
- References
- 36
- Citations by year
- 2026: 3 · 2025: 3 · 2024: 9 · 2023: 6 · 2022: 5 · 2021: 4
- FWCI
- 1.22
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Frontiers in Pharmacology (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Funders
- Russian Science Foundation
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Phytochemical Studies and Bioactivities
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Toxicology › Bioactive Compounds and Antitumor Agents
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Estrogen and related hormone effects
- Keywords
- 20-hydroxyecdysone, breast cancer, tumor suppression, triple-negative breast cancer, glycolysis, cellular respiration, autophagy, doxorubicin synergy, combination index, dose reduction index, cell death, energy profiling, breast cancer molecular subtypes, adjuvant therapy
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