From a Medicinal Mushroom Blend a Direct Anticancer Effect on Triple-Negative Breast Cancer: A Preclinical Study on Lung Metastases
Abstract
:1. Introduction
2. Results
2.1. Raw Materials, Extract Procedure, and Main Active Metabolites of Micotherapy U-Care Blend
2.2. TUNEL Assay
2.3. PARP1, p53, Bax, Bcl2, and PCNA Immunohistochemical Assessment
2.3.1. PARP1
2.3.2. p53
2.3.3. Bax
2.3.4. Bcl2
2.3.5. PCNA
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Animals and Experimental Plan
4.3. Tissue Sampling and Immunohistochemistry
4.3.1. Lung Specimens Preparation
4.3.2. TUNEL Staining
4.3.3. Immunohistochemistry: Apoptotic Pathway Assessment
4.3.4. Immunohistochemical Evaluations
4.4. Statistics
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
- Jemal, A.; Bray, F.; Center, M.M.; Ferlay, J.; Ward, E.; Forman, D. Global cancer statistics. CA Cancer J. Clin. 2011, 61, 69–90. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Cardoso, F.; Harbeck, N.; Fallowfield, L.; Kyriakides, S.; Senkus, E.; ESMO Guidelines Working Group. Locally recurrent or metastatic breast cancer: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up. Ann. Oncol. 2012, 23, vii11–vii19. [Google Scholar] [CrossRef] [PubMed]
- Zeeshan, R.; Mutahir, Z. Cancer metastasis: Tricks of the trade. Bosn. J. Basic Med. Sci. 2017, 17, 172–182. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Schito, L.; Rey, S. Hypoxic pathobiology of breast cancer metastasis. Biochim. Biophys. Acta Rev. Cancer 2017, 1868, 239–245. [Google Scholar] [CrossRef]
- Yadav, B.S.; Chanana, P.; Jhamb, S. Biomarkers in triple negative breast cancer: A review. World J. Clin. Oncol. 2015, 6, 252. [Google Scholar] [CrossRef]
- Yeo, S.K.; Guan, J.L. Breast Cancer: Multiple Subtypes within a Tumor? Trends Cancer 2017, 3, 753–760. [Google Scholar] [CrossRef]
- Yao, Y.; Chu, Y.; Xu, B.; Hu, Q.; Song, Q. Risk factors for distant metastasis of patients with primary triple-negative breast cancer. Biosci. Rep. 2019, 39. [Google Scholar] [CrossRef]
- Iriondo, O.; Liu, Y.; Lee, G.; Elhodaky, M.; Jimenez, C.; Li, L.; Lang, J.; Wang, P.; Yu, M. TAK1 mediates microenvironment-triggered autocrine signals and promotes triple-negative breast cancer lung metastasis. Nat. Commun. 2018, 9, 1994. [Google Scholar] [CrossRef]
- Chitty, J.L.; Filipe, E.C.; Lucas, M.C.; Herrmann, D.; Cox, T.R.; Timpson, P. Recent advances in understanding the complexities of metastasis. F1000Research 2018, 7, 1169. [Google Scholar] [CrossRef]
- Katsuta, E.; Rashid, O.M.; Takabe, K. Clinical relevance of tumor microenvironment: Immune cells, vessels, and mouse models. Hum. Cell 2020, 33, 930–937. [Google Scholar] [CrossRef]
- Buhrmann, C.; Shayan, P.; Banik, K.; Kunnumakkara, A.B.; Kubatka, P.; Koklesova, L.; Shakibaei, M. Targeting NF-κB Signaling by Calebin A, a Compound of Turmeric, in Multicellular Tumor Microenvironment: Potential Role of Apoptosis Induction in CRC Cells. Biomedicines 2020, 8, 236. [Google Scholar] [CrossRef] [PubMed]
- Cacho-Diaz, B.; García-Botello, D.R.; Wegman-Ostrosky, T.; Reyes-Soto, G.; Ortiz-Sánchez, E.; Herrera-Montalvo, L.A. Tumor microenvironment differences between primary tumor and brain metastases. J. Transl. Med. 2020, 18, 1. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Nounou, M.I.; Elamrawy, F.; Ahmed, N.; Abdelraouf, K.; Goda, S.; Syed-Sha-Qhattal, H. Breast Cancer: Conventional Diagnosis and Treatment Modalities and Recent Patents and Technologies. Breast Cancer 2015, 9, 17–34. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Zhao, Y.; Jing, Z.; Li, Y.; Mao, W. Berberine in combination with cisplatin suppresses breast cancer cell growth through induction of DNA breaks and caspase-3-dependent apoptosis. Oncol. Rep. 2016, 36, 567–572. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Czabotar, P.E.; Lessene, G.; Strasser, A.; Adams, J.M. Control of apoptosis by the BCL-2 protein family: Implications for physiology and therapy. Nat. Rev. Mol. Cell Biol. 2014, 15, 49–63. [Google Scholar] [CrossRef]
- Kadam, C.Y.; Abhang, S.A. Apoptosis Markers in Breast Cancer Therapy. Adv. Clin. Chem. 2016, 74, 143–193. [Google Scholar] [CrossRef]
- Elmore, S. Apoptosis: A Review of Programmed Cell Death. Toxicol. Pathol. 2007, 35, 495–516. [Google Scholar] [CrossRef]
- Parton, M.; Dowsett, M.; Smith, I. Studies of apoptosis in breast cancer. BMJ 2001, 322, 1528–1532. [Google Scholar] [CrossRef] [Green Version]
- Hong, S.J.; Dawson, T.M.; Dawson, V.L. PARP and the Release of Apoptosis-Inducing Factor from Mitochondria; Madame Curie Bioscience Database. 2013. Available online: https://www.ncbi.nlm.nih.gov/books/NBK6179/ (accessed on 8 October 2020).
- Aubrey, B.J.; Kelly, G.L.; Janic, A.; Herold, M.J.; Strasser, A. How does p53 induce apoptosis and how does this relate to p53-mediated tumour suppression? Cell Death Differ. 2018, 25, 104–113. [Google Scholar] [CrossRef] [Green Version]
- Krajewski, S.; Thor, A.D.; Edgerton, S.M.; Moore, D.H.; Krajewska, M.; Reed, J.C. Analysis of Bax and Bcl-2 expression in p53-immunopositive breast cancers. Clin. Cancer Res. 1997, 3, 199–208. [Google Scholar]
- Frenzel, A.; Grespi, F.; Chmelewskij, W.; Villunger, A. Bcl2 family proteins in carcinogenesis and the treatment of cancer. Apoptosis 2009, 14, 584–596. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Slade, D. PARP and PARG inhibitors in cancer treatment. Genes Dev. 2020, 34, 360–394. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Ossovskaya, V.; Koo, I.C.; Kaldjian, E.P.; Alvares, C.; Sherman, B.M. Upregulation of poly (ADP-Ribose) polymerase-1 (PARP1) in triple-negative breast cancer and other primary human tumor types. Genes Cancer 2010, 1, 812–821. [Google Scholar] [CrossRef] [PubMed]
- Yu, S.W.; Andrabi, S.A.; Wang, H.; No, S.K.; Poirier, G.G.; Dawson, T.M.; Dawson, V.L. Apoptosis-inducing factor mediates poly(ADP-ribose) (PAR) polymer-induced cell death. Proc. Nat. Acad. Sci. USA 2006, 103, 18314–18319. [Google Scholar] [CrossRef] [Green Version]
- Domagala, P.; Huzarski, T.; Lubinski, J.; Gugala, K.; Domagala, W. PARP-1 expression in breast cancer including BRCA1-associated, triple negative and basal-like tumors: Possible implications for PARP-1 inhibitor therapy. Breast Cancer Res. Treat. 2011, 127, 861–869. [Google Scholar] [CrossRef] [Green Version]
- Schreiber, V.; Dantzer, F.; Amé, J.C.; De Murcia, G. Poly(ADP-ribose): Novel functions for an old molecule. Nat. Rev. Mol. Cell Biol. 2006, 7, 517–528. [Google Scholar] [CrossRef]
- Bertheau, P.; Lehmann-Che, J.; Varna, M.; Dumay, A.; Poirot, B.; Porcher, R.; Turpin, E.; Plassa, L.F.; De Roquancourt, A.; Bourstyn, E.; et al. P53 in breast cancer subtypes and new insights into response to chemotherapy. Breast 2013, 22, S27–S29. [Google Scholar] [CrossRef]
- Hanahan, D.; Weinberg, R.A. The hallmarks of cancer. Cell 2000, 100, 57–70. [Google Scholar] [CrossRef] [Green Version]
- Wang, S.-C. PCNA: A silent housekeeper or a potential therapeutic target? Trends Pharmacol. Sci. 2014, 35, 178–186. [Google Scholar] [CrossRef]
- Juríková, M.; Danihel, Ľ.; Polák, Š.; Varga, I. Ki67, PCNA, and MCM proteins: Markers of proliferation in the diagnosis of breast cancer. Acta Histochem. 2016, 118, 544–552. [Google Scholar] [CrossRef]
- Varghese, E.; Samuel, S.M.; Sadiq, Z.; Kubatka, P.; Liskova, A.; Benacka, J.; Pazinka, P.; Kruzliak, P.; Büsselberg, D. Anti-Cancer Agents in Proliferation and Cell Death: The Calcium Connection. Int. J. Mol. Sci. 2019, 20, 3017. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Rossi, P.; Difrancia, R.; Quagliariello, V.; Savino, E.; Tralongo, P.; Randazzo, C.L.; Berretta, M. B-glucans from Grifola frondosa and Ganoderma lucidum in breast cancer: An example of complementary and integrative medicine. Oncotarget 2018, 9, 24837–24856. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wasser, S.P. Medicinal mushrooms in human clinical studies. Part I. anticancer, oncoimmunological, and immunomodulatory activities: A review. Int. J. Med. Mush. 2017, 19, 279–317. [Google Scholar] [CrossRef]
- Blagodatski, A.; Yatsunskaya, M.; Mikhailova, V.; Tiasto, V.; Kagansky, A.; Katanaev, V.L. Medicinal mushrooms as an attractive new source of natural compounds for future cancer therapy. Oncotarget 2018, 9, 29259–29274. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Roda, E.; De Luca, F.; Di Iorio, C.; Ratto, D.; Siciliani, S.; Ferrari, B.; Cobelli, F.; Borsci, G.; Priori, E.C.; Chinosi, S.; et al. Novel medicinal mushroom blend as a promising supplement in integrative oncology: A multi-tiered study using 4t1 triple-negative mouse breast cancer model. Int. J. Mol. Sci. 2020, 21, 3479. [Google Scholar] [CrossRef]
- Zmitrovich, I.V.; Belova, N.V.; Balandaykin, M.E.; Bondartseva, M.A.; Wasser, S.P. Cancer without Pharmacological Illusions and a Niche for Mycotherapy (Review). Int. J. Med. Mush. 2019, 21, 105–119. [Google Scholar] [CrossRef]
- Bechtel, P.E.; Hickey, R.J.; Schnaper, L.; Sekowski, J.W.; Long, B.J.; Freund, R.; Liu, N.; Rodriguez-Valenzuela, C.; Malkas, L.H. A Unique Form of Proliferating Cell Nuclear Antigen Is Present in Malignant Breast Cells. Cancer Res. 1998, 58, 3264–3269. [Google Scholar]
- Malkas, L.H.; Herbert, B.S.; Abdel-Aziz, W.; Dobrolecki, L.E.; Liu, Y.; Agarwal, B.; Hoelz, D.; Badve, S.; Schnaper, L.; Arnold, R.J.; et al. A cancer-associated PCNA expressed in breast cancer has implications as a potential biomarker. Proc. Natl. Acad. Sci. USA 2006, 103, 19472–19477. [Google Scholar] [CrossRef] [Green Version]
- Coccini, T.; Manzo, L.; Roda, E. Safety Evaluation of Engineered Nanomaterials for Health Risk Assessment: An Experimental Tiered Testing Approach Using Pristine and Functionalized Carbon Nanotubes. ISRN Toxicol. 2013, 2013, 825427. [Google Scholar] [CrossRef] [Green Version]
- Baghban, R.; Roshangar, L.; Jahanban-Esfahlan, R.; Seidi, K.; Ebrahimi-Kalan, A.; Jaymand, M.; Kolahian, S.; Javaheri, T.; Zare, P. Tumor microenvironment complexity and therapeutic implications at a glance. Cell Commun. Signal. 2020, 18, 59. [Google Scholar] [CrossRef] [Green Version]
- Liu, Z.; Ding, Y.; Ye, N.; Wild, C.; Chen, H.; Zhou, J. Direct Activation of Bax Protein for Cancer Therapy. Med. Res. Rev. 2016, 36, 313–341. [Google Scholar] [CrossRef] [PubMed]
- Wong, R.S. Apoptosis in cancer: From pathogenesis to treatment. J. Exp. Clin. Cancer Res. 2011, 30, 87. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Yaacoub, K.; Pedeux, R.; Tarte, K.; Guillaudeux, T. Role of the tumor microenvironment in regulating apoptosis and cancer progression. Cancer Lett. 2016, 378, 150–159. [Google Scholar] [CrossRef] [PubMed]
- Oltval, Z.N.; Milliman, C.L.; Korsmeyer, S.J. Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programed cell death. Cell 1993, 74, 609–619. [Google Scholar] [CrossRef]
- Bodrug, S.E.; Aimé-Sempé, C.; Sato, T.; Krajewski, S.; Hanada, M.; Reed, J.C. Biochemical and functional comparisons of Mcl-1 and Bcl-2 proteins: Evidence for a novel mechanism of regulating Bcl-2 family protein function. Cell Death Diff. 1995, 2, 173–182. [Google Scholar]
- Teixeira, C.; Reed, J.C.; Pratt, M.A. Estrogen promotes chemotherapeutic drug resistance by a mechanism involving Bcl-2 proto-oncogene expression in human breast cancer cells. Cancer Res. 1995, 55, 3902–3907. [Google Scholar]
- Qiu, X.; Mei, J.; Yin, J.; Wang, H.; Wang, J.; Xie, M. Correlation analysis between expression of PCNA, Ki-67 and COX-2 and X-ray features in mammography in breast cancer. Oncol. Lett. 2017, 14, 2912–2918. [Google Scholar] [CrossRef] [Green Version]
- Roda, E.; Bottone, M.; Biggiogera, M.; Milanesi, G.; Coccini, T. Pulmonary and hepatic effects after low dose exposure to nanosilver: Early and long-lasting histological and ultrastructural alterations in rat. Toxicol. Rep. 2019, 6, 1047–1060. [Google Scholar] [CrossRef]
Sample Availability: Samples of the compounds are available from the authors. |
Medicinal Mushroom | Fungal Part Used in Micotherapy U-Care | % Contained in Micotherapy U-Care | ID Code |
---|---|---|---|
Agaricus blazei | Fruiting body | 20% | 7700 |
Ophiocordyceps sinensin | Fruiting body and mycelium | 20% | Cm2 |
Ganoderma lucidum | Fruiting body | 20% | Gač |
Grifola frondosa | Fruiting body | 20% | Gf3 |
Lentinula edodes | Fruiting body | 20% | Le.ed.1 |
Antigen | Immunogen | Manufacturer, Species, Mono-Polyclonal, Cat./Lot. No., RRID | Dilution | |
---|---|---|---|---|
Primary Antibodies | Anti-poly (ADP-ribose) polymerase (46D11) | Purified antibody raised against the residues surrounding Gly623 of human PARP-1 | Cell Signaling Technology (Danvers, MA, USA), Rabbit monoclonal IgG, Cat# 9532, RRID:AB_659884 | 1:100 |
Anti-p53 (Ab-5) | Purified antibody raised against the ~53 kDa wild type p53 protein of mouse origin | Sigma-Aldrich (St. Louis, MO, USA), Mouse monoclonal IgG2a, Cat# OP33-100UG, RRID:AB_564977 | 1:100 | |
Anti-Bcl-2-associated X protein (P-19) | Purified antibody Raised against a peptide mapping at the amino terminus of Bax of mouse origin | Santa Cruz Biotechnology (Santa Cruz, CA, USA), Rabbit polyclonal IgG, Cat# sc-526, RRID:AB_2064668 | 1:100 | |
Anti-B-Cell Leukemia/Lymphoma 2 protein (N-19) | Purified antibody raised against a peptide mapping at the N-terminus of Bcl-2 of human origin | Santa Cruz Biotechnology (Santa Cruz, CA, USA), Rabbit polyclonal IgG, Cat# sc-492, RRID:AB_2064290 | 1:100 | |
Anti-Proliferating Cell Nuclear Antigen (Ab-1) | Purified antibody raised against the ~37 kDa PCNA protein of mouse origin | Sigma-Aldrich (St. Louis, MO, USA), Mouse monoclonal IgG2a, Cat# NA03-200UG, RRID:AB_213111 | 2:1000 | |
Secondary Antibodies | Biotinylated horse anti-mouse IgG | Gamma immunoglobulin | Vector Laboratories (Burlingame, CA, USA), Horse, Cat# PK-6102, RRID:AB_2336821 | 1:200 |
Biotinylated goat anti-rabbit IgG | Gamma immunoglobulin | Vector Laboratories (Burlingame, CA, USA), Goat, lot# PK-6101, RRID: AB_2336820 | 1:200 |
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Roda, E.; Luca, F.D.; Locatelli, C.A.; Ratto, D.; Di Iorio, C.; Savino, E.; Bottone, M.G.; Rossi, P. From a Medicinal Mushroom Blend a Direct Anticancer Effect on Triple-Negative Breast Cancer: A Preclinical Study on Lung Metastases. Molecules 2020, 25, 5400. https://doi.org/10.3390/molecules25225400
Roda E, Luca FD, Locatelli CA, Ratto D, Di Iorio C, Savino E, Bottone MG, Rossi P. From a Medicinal Mushroom Blend a Direct Anticancer Effect on Triple-Negative Breast Cancer: A Preclinical Study on Lung Metastases. Molecules. 2020; 25(22):5400. https://doi.org/10.3390/molecules25225400
Chicago/Turabian StyleRoda, Elisa, Fabrizio De Luca, Carlo Alessandro Locatelli, Daniela Ratto, Carmine Di Iorio, Elena Savino, Maria Grazia Bottone, and Paola Rossi. 2020. "From a Medicinal Mushroom Blend a Direct Anticancer Effect on Triple-Negative Breast Cancer: A Preclinical Study on Lung Metastases" Molecules 25, no. 22: 5400. https://doi.org/10.3390/molecules25225400