Assessment of the cytotoxic effects of aporphine prototypes on head and neck cancer cells

  • Dorival Mendes Rodrigues-Junior
  • Nicolie Melanie de Almeida Pontes
  • Gabriela Estrela de Albuquerque
  • Viviane Carlin
  • Givago Prado Perecim
  • Cristiano Raminelli
  • André Luiz VettoreEmail author


Purpose Among alkaloids, abundant secondary metabolites in plants, aporphines constitute a class of compounds with interesting biological activities, including anticancer effects. The present study evaluated the anticancer activities of 14 substances, including four aporphine derivatives acquired through the biomonitoring of (±)-apomorphine hydrochloride total synthesis from 2-phenethylamine and 3,4-dimethoxybenzaldehyde against head and neck squamous cell carcinoma (HNSCC). Methods The cytotoxic effects of compounds against a panel of HNSCC cell lines were determined by PrestoBlue cell viability assay, while the genotoxicity of substances was evaluated by micronucleus test. Cell death was detected by flow cytometry (Annexin V/7AAD) and western blot analysis was used to detect the presence of cleaved Caspase-3 molecules. Results The aporphine and isoquinoline derivatives APO, C1, and A5 significantly reduced HNSCC cell viability and promoted DNA damages in these cells. Further, by activating the Caspase-3 pathway, these substances were able to induce apoptosis. Conclusion Our results revealed that APO, C1, and A5 exhibit cytotoxic effects in HNSCC cells. The mechanisms of action appear to be partly via the generation of DNA damages and apoptosis induction through Caspase-3 pathway activation. This study provides preclinical data that suggest a potential therapeutic role for APO, C1, and A5 against head and neck cancer cells.


Aporphine prototypes Cytotoxic molecules Cancer cells Head and neck cancer 



This study was supported by grants from the Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP (2015/09182–0 and 2017/21990–0). D.M.R.-Jr received a scholarship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, 99999.007922/2014–00) and the São Paulo Research Foundation (FAPESP, 2015/21420–3). G.P.P. also received a scholarship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, 1655331). Both N.M.A.P. and A.L.V. had scholarships from the Conselho Nacional de Desenvolvimento Científico e Tecnológico – CNPq (123641/2017–9 and 300936/2015–0, respectively).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Ethics approval

This study was approved by the institutional ethics committee (CEP-UNIFESP: 7428290317).


  1. 1.
    Cragg GM, Grothaus PG, Newman DJ (2009) Impact of natural products on developing new anti-cancer agents. Chem Rev 109:3012–3043CrossRefGoogle Scholar
  2. 2.
    Hostettmann K, Potterat O, Wolfender JL (1998) The potential of higher plants as a source of new drugs. Chimia 52:10–17Google Scholar
  3. 3.
    Cragg GM, Newman DJ (1999) Discovery and development of antineoplastic agents from natural sources. Cancer Investig 17:153–163CrossRefGoogle Scholar
  4. 4.
    Simmonds MS (2003) Novel drugs from botanical sources. Drug Discov Today 8:721–722CrossRefGoogle Scholar
  5. 5.
    Stévigny C, Bailly C, Quetin-Leclercq J (2005) Cytotoxic and antitumor potentialities of aporphinoid alkaloids. Curr Med Chem Anticancer Agents 5:173–182CrossRefGoogle Scholar
  6. 6.
    Guinaudeau H, Lebœuf M, Cavé A (1994) Aporphinoid Alkaloids, V. J Nat Prod 57:1033–1135CrossRefGoogle Scholar
  7. 7.
    Protais P, Arbaoui J, Bakkali EH, Bermejo A, Cortes D (1995) Effects of various isoquinoline alkaloids on in vitro 3H-dopamine uptake by rat striatal synaptosomes. J Nat Prod 58:1475–1484CrossRefGoogle Scholar
  8. 8.
    Yan R, Wang W, Guo J, Liu H, Zhang J, Yang B (2013) Studies on the alkaloids of the bark of Magnolia officinalis: isolation and on-line analysis by HPLC-ESI-MS(n). Molecules 18:7739–7750CrossRefGoogle Scholar
  9. 9.
    Kashiwada Y, Aoshima A, Ikeshiro Y, Chen YP, Furukawa H, Itoigawa M, Fujioka T, Mihashi K, Cosentino LM, Morris-Natschke SL, Lee KH (2005) Anti-HIV benzylisoquinoline alkaloids and flavonoids from the leaves of Nelumbo nucifera, and structure-activity correlations with related alkaloids. Bioorg Med Chem 13:443–448CrossRefGoogle Scholar
  10. 10.
    Zhang A, Zhang Y, Branfman AR, Baldessarini RJ, Neumeyer JLJ (2007) Advances in development of dopaminergic aporphinoids. Med Chem 50:171–181CrossRefGoogle Scholar
  11. 11.
    Singh IP, Bodiwala HS (2010) Recent advances in anti-HIV natural products. Nat Prod Rep 27:1781–1800CrossRefGoogle Scholar
  12. 12.
    Ponnala S, Chaudhary S, González-Sarrias A, Seeram NP, Harding WW (2011) Cytotoxicity of aporphines in human colon cancer cell lines HCT-116 and Caco-2: an SAR study. Bioorg Med Chem Lett 21:4462–4464CrossRefGoogle Scholar
  13. 13.
    Suresh HM, Shivakumar B, Shivakumar SI (2012) Cytotoxicity of Aporphine alkaloids from the roots of Annona Reticulata on human cancer cell lines. Int J Plant Res 2:57–60CrossRefGoogle Scholar
  14. 14.
    Liu Y, Liu J, Di D, Li M, Fen Y (2013) Structural and mechanistic bases of the anticancer activity of natural Aporphinoid alkaloids. Curr Top Med Chem 13:2116–2126CrossRefGoogle Scholar
  15. 15.
    Ferlay J, Steliarova-Foucher E, Lortet-Tieulent J et al (2012) Cancer incidence and mortality patterns in Europe: estimates for 40 countries in 2012. Eur J Cancer 49:1374–1403CrossRefGoogle Scholar
  16. 16.
    Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D (2011) Global cancer statistics. CA Cancer J Clin 61:69–90CrossRefGoogle Scholar
  17. 17.
    Chin D, Boyle GM, Williams RM, Ferguson K, Pandeya N, Pedley J, Campbell CM, Theile DR, Parsons PG, Coman WB (2005) Novel markers for poor prognosis in head and neck cancer. Int J Cancer 113:789–797CrossRefGoogle Scholar
  18. 18.
    Iyer NG, Tan DS, Tan VK et al (2015) Randomized trial comparing surgery and adjuvant radiotherapy versus concurrent chemoradiotherapy in patients with advanced, nonmetastatic squamous cell carcinoma of the head and neck: 10-year update and subset analysis. Cancer 121:1599–1607CrossRefGoogle Scholar
  19. 19.
    Muraca A, Perecim G, Rodrigues A, Raminelli C (2017) Convergent Total synthesis of (±)-Apomorphine via Benzyne chemistry: insights into the mechanisms involved in the key step. Synthesis 49:3546–3557CrossRefGoogle Scholar
  20. 20.
    Rossini AF, Muraca AC, Casagrande GA, Raminelli C (2015) Total syntheses of Aporphine alkaloids via Benzyne chemistry: an approach to the formation of Aporphine cores. J Organomet Chem 80:10033–10040CrossRefGoogle Scholar
  21. 21.
    Perecim GP, Rodrigues A, Raminelli C (2015) A convenient formation of aporphine core via benzyne chemistry: conformational analysis and synthesis of (R)-aporphine. Tetrahedron Lett 56:6848–6851CrossRefGoogle Scholar
  22. 22.
    Sarto F, Finotto S, Giacomelli L, Mazzotti D, Tomanin R, Levis AG (1987) The micronucleus assay in exfoliated cells of the human buccal. Mutagenesis 2:11–17CrossRefGoogle Scholar
  23. 23.
    Morales-Ramírez P, Vallarino-Kelly T, Cruz-Vallejo VL (2017) The OECD's micronucleus test guideline for single exposure to an agent and the genotox-kinetic alternative. Mutagenesis 32:411–415CrossRefGoogle Scholar
  24. 24.
    da Silva J, de Freitas TR, Heuser V et al (2000) Effects of chronic exposure to coal in wild rodents (Ctenomys torquatus) evaluated by multiple methods and tissues. Mutat Res 10:39–51CrossRefGoogle Scholar
  25. 25.
    Zurich FM (1913) Apomorphine. The formation of Apomorphine on heating and preserving morphine solutions. Z Physiol Chem 84:363–378CrossRefGoogle Scholar
  26. 26.
    Schwab RS, Amador LV, Lettvin JY (1951) Apomorphine in Parkinson's disease. Trans Am Neurol Assoc 56:251–253Google Scholar
  27. 27.
    Antonini A, Jenner P (2018) Apomorphine infusion in advanced Parkinson disease. Nat Rev Neurol 14:693–694CrossRefGoogle Scholar
  28. 28.
    Kondo Y, Imai Y, Hojo H, Endo T, Nozoe S (1990) Suppression of tumor cell growth and mitogen response by aporphine alkaloids, dicentrine, glaucine, corydine, and apomorphine. J Pharmacobio-Dyn 13:426–431CrossRefGoogle Scholar
  29. 29.
    Lin LZ, Hu SF, Zaw K, Angerhofer CK, Chai H, Pezzuto JM, Cordell GA, Lin J, Zheng DM (1994) Thalifaberidine, a cytotoxic aporphine-benzylisoquinline alkaloid from Thalictrum faberi. J Nat Prod 57:1430–1436CrossRefGoogle Scholar
  30. 30.
    Chen JJ, Ishikawa T, Duh CY, Tsai IL, Chen IS (1996) New dimeric aporphine alkaloids and cytotoxic constituents of Hernandia nymphaeifolia. Planta Med 62:528–533CrossRefGoogle Scholar
  31. 31.
    Chen IS, Chen JJ, Duh CY, Tsai IL, Chang CT (1997) New aporphine alkaloids and cytotoxic constituents of Hernandia nymphaeifolia. Planta Med 63:154–157CrossRefGoogle Scholar
  32. 32.
    Woo SH, Sun NJ, Cassady JM, Snapka RM (1999) Topoisomerase II inhibition by aporphine alkaloids. Biochem Pharmacol 57:1141–1145CrossRefGoogle Scholar
  33. 33.
    Stopper H, Schmitt E, Gregor C, Mueller SO, Fischer WH (2003) Increased cell proliferation is associated with genomic instability: elevated micronuclei frequencies in estradiol-treated human ovarian cancer cells. Mutagen 18:243–247CrossRefGoogle Scholar
  34. 34.
    dos Santos EBR, Daval J, Koziel V, Netter P, Minn A (2001) Toxic effects of apomorphine on rat cultured neurons and glial C6 cells, and protection with antioxidants. Biochem Pharmacol 61:73–85CrossRefGoogle Scholar
  35. 35.
    Gören AC, Zhou BN, Kingston DG (2003) Cytotoxic and DNA damaging activity of some aporphine alkaloids from Stephania dinklagei. Planta Med 69:867–868CrossRefGoogle Scholar
  36. 36.
    Picada JN, Roesler R, Henriques JA (2005) Genotoxic, neurotoxic and neuroprotective activities of apomorphine and its oxidized derivative 8-oxo-apomorphine. Braz J Med Biol Res 38:477–486CrossRefGoogle Scholar
  37. 37.
    Hwang PM, Bunz F, Yu J, Rago C, Chan TA, Murphy MP, Kelso GF, Smith RAJ, Kinzler KW, Vogelstein B (2001) Ferredoxin reductase affects p53-dependent, 5-fluorouracil-induced apoptosis in colorectal cancer cells. Nat Med 7:1111–1117CrossRefGoogle Scholar
  38. 38.
    Kim KK, Kawar NM, Singh RK, Lange TS, Brard L, Moore RG (2011) Tetrathiomolybdate induces doxorubicin sensitivity in resistant tumor cell lines. Gynecol Oncol 122:183–189CrossRefGoogle Scholar
  39. 39.
    Deigner HP, Kinscherf R (1999) Modulating apoptosis: current applications and prospects for future drug development. Curr Med Chem 6:399–414Google Scholar
  40. 40.
    Chen Q, Peng WL, XU AL (2002) Apoptosis of a human non-small cell lung cancer (NSCLC) cell line, PLA-801, induced by acutiaporberine, a novel bisalkaloid derived from Thalictrum acutifolium (Hand.-Mazz) boivin. Biochem Pharmacol 63:1389–1396CrossRefGoogle Scholar
  41. 41.
    Montririttigri K, Moongkarndi P, Joongsomboonkusol S, Chitkul B, Pattanapanyasat K (2008) Apoptotic activity of aporphine from Stephania venosa on human ovarian cancer cells. Mahidol University J Pharm Sci 35:52–56Google Scholar
  42. 42.
    Sun HF, Hou HL, Lu P, Zhang L, Zhao F, Ge C, Wang T, Yao M, Li J (2012) Isocorydine inhibits cell proliferation in hepatocellular carcinoma cell lines by inducing G2/M cell cycle arrest and apoptosis. PLoS One 7:e36808CrossRefGoogle Scholar
  43. 43.
    Slee EA, Adrain C, Martin SJ (2001) Executioner caspase-3, −6, and −7 perform distinct, non-redundant roles during the demolition phase of apoptosis. J Biol Chem 276:7320–7326CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Dorival Mendes Rodrigues-Junior
    • 1
    • 2
  • Nicolie Melanie de Almeida Pontes
    • 2
  • Gabriela Estrela de Albuquerque
    • 1
    • 2
  • Viviane Carlin
    • 2
  • Givago Prado Perecim
    • 3
  • Cristiano Raminelli
    • 3
  • André Luiz Vettore
    • 1
    • 2
    Email author
  1. 1.Departamento de Ciências BiológicasCampus Diadema, Universidade Federal de São PauloDiademaBrazil
  2. 2.Laboratório de Biologia Molecular do CâncerUniversidade Federal de São PauloSão PauloBrazil
  3. 3.Departamento de Química, Campus DiademaUniversidade Federal de São PauloDiademaBrazil

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