Therapeutic potential of Catharanthus roseus bioactive compounds as the 4PHO protein inhibitors, a pathogenetic determinative factor of BLV. This work describes the use of computational drug discovery approaches in estimating the interactions conducted between 40 phytocompounds of Catharanthus roseus and the 4PHO protein. Our work has determined the presence of Pamoic acid and Vindolinine hydrochloride as the optimal choice here. Pharmacokinetic screening exhibited drug-like properties to both the highly absorbed compounds in the gastrointestinal tract and Lipinski’s Rule of Five. Molecular docking screening exhibited strong binding of the compounds with 4PHO; pamoic acid exhibited binding energy of –8.7 kcal/mol and vindolinine hydrochloride exhibited –7.8 kcal/mol. VAL62, PHE222, and LEU225 were the most significant residues participating in interaction. Structural validation of 4PHO protein, for instance, through Ramachandran plot analysis confirmed its viability for docking studies. These observations form a platform for the synthesis of plant-derived antiviral drugs against BLV and offer a platform for future in vitro and in vivo studies to assess their therapeutic viability.

Keywords: Molecular docking, Catharanthus roseus, bovine leukemia virus (BLV), cancer, 4PHO protein, drug discovery

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1. Mishra J, Verma N. A brief study on Catharanthus roseus: A review. Int J Res Pharm Pharmaceut Sci. 2017;2(2):20–3.
2. Ajaib M, Khan ZUD, Khan N, Wahab M. Ethnobotanical studies on useful shrubs of District Kotli, Azad Jammu & Kashmir, Pakistan. Pak J Bot. 2010;42:1407–15.
3. Gajalakshmi S, Vijayalakshmi S, Devi V. Pharmacological activities of Catharanthus roseus: A perspective review. Int J Pharma Bio Sci. 2013;4(2):431–9.
4. Aida Y, Murakami H, Takagashi M, Takeshima S-N. Mechanisms of pathogenesis induced by bovine leukemia virus as a model for human T-cell leukemia virus. Front Microbiol. 2013;4:328. doi:10.3389/fmicb.2013.00328.
5. Brujeni GN, Ghorbanpour R, Esmailnejad A. Association of BoLA-DRB3.2 alleles with BLV infection profiles (persistent lymphocytosis/lymphosarcoma) and lymphocyte subsets in Iranian Holstein cattle. Biochem Genet. 2016;54:194–207. doi:10.1007/s10528-016-9712-6.
6. Maclachlan NJ, Dubovi EJ. Fenner’s Veterinary Virology. Cambridge, MA: Academic Press; 2010.
7. Martin F, Vandamme A-M, Mahieux R, Gessain A, Bangham CRM, Watanabe T, et al. Conference highlights of the 15th International Conference on Human Retrovirology: HTLV and related retroviruses, 4–8 June 2011, Leuven, Gembloux, Belgium. Retrovirology. 2011;8:86. doi:10.1186/1742-4690-8-86.
8. Nekouei O, VanLeeuwen JA, Sanchez J, Kelton DF, Tiwari A, Keefe GP. Predicting within-herd prevalence of infection with bovine leukemia virus using bulk-tank milk antibody levels. Prev Vet Med. 2015;122:53–60. doi:10.1016/j.prevetmed.2015.10.009.
9. Moratorio G, Obal G, Trono K, Pritsch O, Esteves P, Cristina J, et al. A detailed molecular analysis of complete bovine leukemia virus genomes isolated from B-cell lymphosarcomas. Vet Res. 2013;44:19. doi:10.1186/1297-9716-44-19.
10. Juliarena MA, Poli M, Ceriani C. Bovine leukemia virus: Current perspectives. Manchester, UK: Dove Press; 2017.
11. Pandey GS, Simulundu E, Mweene AS, Suzuki T, Takada A, Namangala B, et al. Clinical and subclinical bovine leukemia virus infection in a dairy cattle herd in Zambia. Arch Virol. 2017;162:1051–6. doi:10.1007/s00705-016-3205-0.
12. Aida Y, Nosaka T, Nakao M, Nakai M, Masuda T, Ito Y, et al. Further phenotypic characterization of target cells for bovine leukemia virus experimental infection in sheep. Am J Vet Res. 1989;50:1946–51.
13. Gansäuer A, Justicia J, Fan C-A, Worgull D, Piestert F. Reductive C—C bond formation after epoxide opening via electron transfer. In: Krische MJ, editor. Metal Catalyzed Reductive C—C Bond Formation: A Departure from Preformed Organometallic Reagents. Springer Science & Business Media; 2007. p. 25–52.
14. Cooper R, Deakin JJ. Africa’s gift to the world. In: Botanical Miracles: Chemistry of Plants That Changed the World. Boca Raton: CRC Press; 2016. p. 46–51.
15. Keglevich P, Hazai L, Kalaus G, Szántay C. Modifications on the basic skeletons of vinblastine and vincristine. Molecules. 2012;17(5):5893–914.
16. Raviña E. Vinca alkaloids. In: The Evolution of Drug Discovery: From Traditional Medicines to Modern Drugs. Weinheim: John Wiley & Sons; 2011. p. 157–9.
17. Morris GM, Huey R, Olson AJ. Using AutoDock for ligand–receptor docking. Methods Mol Biol. 2008;443:365–82.
18. Hirata K, Miyamoto K, Miura Y. Catharanthus roseus L. (Periwinkle): Production of vindoline and catharanthine in multiple shoot cultures. In: Bajaj YPS, editor. Biotechnology in Agriculture and Forestry 26. Medicinal and Aromatic Plants. Berlin: Springer-Verlag; 1994. p. 46–55.
19. Faller BA, Pandi TN. Safety and efficacy of vinorelbine in the treatment of non-small cell lung cancer. Clin Med Insights Oncol. 2011;5:131–44.
20. Ngo QA, Bernadat G, Cavelier F, Baltas M. Synthesis and biological evaluation of Vinca alkaloids and phomopsin hybrids. J Med Chem. 2009;52(1):134–42.
21. Hardouin C, Doris E, Rousseau B, Mioskowski C. Concise synthesis of anhydrovinblastine from leurosine. Org Lett. 2002;4(7):1151–3.
22. Aynilian GH, Hufford CD, Robertson LE, Twohey BE. Catharanthus alkaloids. XXIX. Isolation and structure elucidation of vincoline. J Pharm Sci. 1974;63(4):536–8.
23. Yao XG, Zhu YH, Xu HB, Luo WJ, Zhang JJ, Zhang YL, et al. Natural product vindoline stimulates insulin secretion and efficiently ameliorates glucose homeostasis in diabetic murine models. J Ethnopharmacol. 2013;150(1):285–97.
24. Nishimori A, Maezawa M, Masuda T, Aida Y. Direct polymerase chain reaction from blood and tissue samples for rapid diagnosis of bovine leukemia virus infection. J Vet Med Sci. 2016;78:791–6. doi:10.1292/jvms.15-0577.
25. Manual of Diagnostic Tests and Vaccines for Terrestrial Animals. Paris: World Organisation for Animal Health; 2018. p. 1113–24.
26. Boris-Lawrie K, Kim SJ, Narayan O, Derse D. In vivo study of genetically simplified bovine leukemia virus derivatives that lack tax and rex. J Virol. 1997;71:1514–20. doi:10.1128/jvi.71.2.1514-1520.1997.
27. Pătraşcu I, Enăchescu V, Găină M, Caraba V, Cojocaru C, Alexe C, et al. Specific protection against bovine leukemia virus infection conferred on cattle by the Romanian inactivated vaccine BL-VACC-RO. Virologie. 1980;31:95–102.
28. Ristau E, Morzunow R, Brandt J, Ziebell KL. Protection of sheep against infection with bovine leukemia virus by vaccination with tumour cells or tumour cell preparations from lymph nodes of leukemic cattle. Arch Exp Vet. 1987;41:185–96.
29. Sengupta S, Mitra P, Pal N, Ghosh S. OSADHI – An online structural and analytics based database for herbs of India. J Cheminform. 2022;14(1):129.
30. Kim S, Chen J, Cheng T, Gindulyte A, He J, He S, et al. PubChem in 2021: New data content and improved web interfaces. Nucleic Acids Res. 2021;49(D1):D1388–95.
31. Roderer DJA, Glockshuber R, Ban N. ClyA CC6/264 ox (2-303). Biochemistry. 2014;53:6357–69.
32. Trott O, Olson AJ. AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem. 2010;31(2):455–61.
33. Paul DS, Gautham N. Protein–small molecule docking with receptor flexibility in iMOLSDOCK. arXiv [Preprint]. 2020. arXiv:2010.05475.
34. Blaszczyk M, Kurcinski M, Kouza M, Wieteska L, Debinski A, Kolinski A, et al. Modeling of protein–peptide interactions using the CABS-dock web server for binding site search and flexible docking. arXiv [Preprint]. 2015. arXiv:1505.01138.
35. Daina A, Michielin O, Zoete V. SwissADME: A free web tool to evaluate pharmacokinetics, drug-likeness and medicinal chemistry friendliness of small molecules. Sci Rep. 2017;7:42717.
36. Dallakyan S, Olson AJ. AutoDock and AutoDockTools: Automated docking with selective receptor flexibility. Methods Mol Biol. 2015;1115:194–206.
37. DeLano WL. The PyMOL Molecular Graphics System. Schrödinger, LLC; 2020.
38. Trott O, Olson AJ. AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function. J Comput Chem. 2010;31(2):455–61.
39. O’Boyle NM, Banck M, James CA, Morley C, Vandermeersch T, Hutchison GR. Open Babel: An open chemical toolbox. J Cheminform. 2011;3(1):33.
40. Seeliger D, de Groot BL. Ligand docking and binding site analysis with PyMOL and AutoDock/Vina. J Comput Aided Mol Des. 2010;24(5):417–22.
41. Daina A, Michielin O, Zoete V. SwissDock, a protein–small molecule docking web service based on EADock DSS. Nucleic Acids Res. 2013;41(W1):W270–7.
42. Hanwell MD, Curtis DE, Lonie DC, Vandermeersch T, Zurek E, Hutchison GR. Avogadro: An advanced semantic chemical editor, visualization, and analysis platform. J Cheminform. 2012;4(1):17.
43. Li H, Leung T, Wong C. Molecular Docking: A powerful tool for structure-based drug discovery. Curr Comput Aided Drug Des. 2012;8(2):104–13.
44. Dassault Systèmes. BIOVIA Discovery Studio [Internet]. 2021 [cited 2025 Feb 10]. Available from: https://www.3ds.com/products-services/biovia/.
45. Nicholls IA, Pettersson B, Danielson L. Computational approaches to molecular docking and structure-based ligand design. Curr Med Chem. 2008;15(22):2203–12.

Therapeutic potential of Catharanthus roseus bioactive compounds as the 4PHO protein inhibitors, a pathogenetic determinative factor of BLV. This work describes the use of computational drug discovery approaches in estimating the interactions conducted between 40 phytocompounds of Catharanthus roseus and the 4PHO protein. Our work has determined the presence of Pamoic acid and Vindolinine hydrochloride as the optimal choice here. Pharmacokinetic screening exhibited drug-like properties to both the highly absorbed compounds in the gastrointestinal tract and Lipinski’s Rule of Five. Molecular docking screening exhibited strong binding of the compounds with 4PHO; pamoic acid exhibited binding energy of –8.7 kcal/mol and vindolinine hydrochloride exhibited –7.8 kcal/mol. VAL62, PHE222, and LEU225 were the most significant residues participating in interaction. Structural validation of 4PHO protein, for instance, through Ramachandran plot analysis confirmed its viability for docking studies. These observations form a platform for the synthesis of plant-derived antiviral drugs against BLV and offer a platform for future in vitro and in vivo studies to assess their therapeutic viability.

Keywords: Molecular docking, Catharanthus roseus, bovine leukemia virus (BLV), cancer, 4PHO protein, drug discovery

🔒 This is a subscription article

Full text is available to subscribers and institutional members. Please choose an option below to access it.

1. Mishra J, Verma N. A brief study on Catharanthus roseus: A review. Int J Res Pharm Pharmaceut Sci. 2017;2(2):20–3.
2. Ajaib M, Khan ZUD, Khan N, Wahab M. Ethnobotanical studies on useful shrubs of District Kotli, Azad Jammu & Kashmir, Pakistan. Pak J Bot. 2010;42:1407–15.
3. Gajalakshmi S, Vijayalakshmi S, Devi V. Pharmacological activities of Catharanthus roseus: A perspective review. Int J Pharma Bio Sci. 2013;4(2):431–9.
4. Aida Y, Murakami H, Takagashi M, Takeshima S-N. Mechanisms of pathogenesis induced by bovine leukemia virus as a model for human T-cell leukemia virus. Front Microbiol. 2013;4:328. doi:10.3389/fmicb.2013.00328.
5. Brujeni GN, Ghorbanpour R, Esmailnejad A. Association of BoLA-DRB3.2 alleles with BLV infection profiles (persistent lymphocytosis/lymphosarcoma) and lymphocyte subsets in Iranian Holstein cattle. Biochem Genet. 2016;54:194–207. doi:10.1007/s10528-016-9712-6.
6. Maclachlan NJ, Dubovi EJ. Fenner’s Veterinary Virology. Cambridge, MA: Academic Press; 2010.
7. Martin F, Vandamme A-M, Mahieux R, Gessain A, Bangham CRM, Watanabe T, et al. Conference highlights of the 15th International Conference on Human Retrovirology: HTLV and related retroviruses, 4–8 June 2011, Leuven, Gembloux, Belgium. Retrovirology. 2011;8:86. doi:10.1186/1742-4690-8-86.
8. Nekouei O, VanLeeuwen JA, Sanchez J, Kelton DF, Tiwari A, Keefe GP. Predicting within-herd prevalence of infection with bovine leukemia virus using bulk-tank milk antibody levels. Prev Vet Med. 2015;122:53–60. doi:10.1016/j.prevetmed.2015.10.009.
9. Moratorio G, Obal G, Trono K, Pritsch O, Esteves P, Cristina J, et al. A detailed molecular analysis of complete bovine leukemia virus genomes isolated from B-cell lymphosarcomas. Vet Res. 2013;44:19. doi:10.1186/1297-9716-44-19.
10. Juliarena MA, Poli M, Ceriani C. Bovine leukemia virus: Current perspectives. Manchester, UK: Dove Press; 2017.
11. Pandey GS, Simulundu E, Mweene AS, Suzuki T, Takada A, Namangala B, et al. Clinical and subclinical bovine leukemia virus infection in a dairy cattle herd in Zambia. Arch Virol. 2017;162:1051–6. doi:10.1007/s00705-016-3205-0.
12. Aida Y, Nosaka T, Nakao M, Nakai M, Masuda T, Ito Y, et al. Further phenotypic characterization of target cells for bovine leukemia virus experimental infection in sheep. Am J Vet Res. 1989;50:1946–51.
13. Gansäuer A, Justicia J, Fan C-A, Worgull D, Piestert F. Reductive C—C bond formation after epoxide opening via electron transfer. In: Krische MJ, editor. Metal Catalyzed Reductive C—C Bond Formation: A Departure from Preformed Organometallic Reagents. Springer Science & Business Media; 2007. p. 25–52.
14. Cooper R, Deakin JJ. Africa’s gift to the world. In: Botanical Miracles: Chemistry of Plants That Changed the World. Boca Raton: CRC Press; 2016. p. 46–51.
15. Keglevich P, Hazai L, Kalaus G, Szántay C. Modifications on the basic skeletons of vinblastine and vincristine. Molecules. 2012;17(5):5893–914.
16. Raviña E. Vinca alkaloids. In: The Evolution of Drug Discovery: From Traditional Medicines to Modern Drugs. Weinheim: John Wiley & Sons; 2011. p. 157–9.
17. Morris GM, Huey R, Olson AJ. Using AutoDock for ligand–receptor docking. Methods Mol Biol. 2008;443:365–82.
18. Hirata K, Miyamoto K, Miura Y. Catharanthus roseus L. (Periwinkle): Production of vindoline and catharanthine in multiple shoot cultures. In: Bajaj YPS, editor. Biotechnology in Agriculture and Forestry 26. Medicinal and Aromatic Plants. Berlin: Springer-Verlag; 1994. p. 46–55.
19. Faller BA, Pandi TN. Safety and efficacy of vinorelbine in the treatment of non-small cell lung cancer. Clin Med Insights Oncol. 2011;5:131–44.
20. Ngo QA, Bernadat G, Cavelier F, Baltas M. Synthesis and biological evaluation of Vinca alkaloids and phomopsin hybrids. J Med Chem. 2009;52(1):134–42.
21. Hardouin C, Doris E, Rousseau B, Mioskowski C. Concise synthesis of anhydrovinblastine from leurosine. Org Lett. 2002;4(7):1151–3.
22. Aynilian GH, Hufford CD, Robertson LE, Twohey BE. Catharanthus alkaloids. XXIX. Isolation and structure elucidation of vincoline. J Pharm Sci. 1974;63(4):536–8.
23. Yao XG, Zhu YH, Xu HB, Luo WJ, Zhang JJ, Zhang YL, et al. Natural product vindoline stimulates insulin secretion and efficiently ameliorates glucose homeostasis in diabetic murine models. J Ethnopharmacol. 2013;150(1):285–97.
24. Nishimori A, Maezawa M, Masuda T, Aida Y. Direct polymerase chain reaction from blood and tissue samples for rapid diagnosis of bovine leukemia virus infection. J Vet Med Sci. 2016;78:791–6. doi:10.1292/jvms.15-0577.
25. Manual of Diagnostic Tests and Vaccines for Terrestrial Animals. Paris: World Organisation for Animal Health; 2018. p. 1113–24.
26. Boris-Lawrie K, Kim SJ, Narayan O, Derse D. In vivo study of genetically simplified bovine leukemia virus derivatives that lack tax and rex. J Virol. 1997;71:1514–20. doi:10.1128/jvi.71.2.1514-1520.1997.
27. Pătraşcu I, Enăchescu V, Găină M, Caraba V, Cojocaru C, Alexe C, et al. Specific protection against bovine leukemia virus infection conferred on cattle by the Romanian inactivated vaccine BL-VACC-RO. Virologie. 1980;31:95–102.
28. Ristau E, Morzunow R, Brandt J, Ziebell KL. Protection of sheep against infection with bovine leukemia virus by vaccination with tumour cells or tumour cell preparations from lymph nodes of leukemic cattle. Arch Exp Vet. 1987;41:185–96.
29. Sengupta S, Mitra P, Pal N, Ghosh S. OSADHI – An online structural and analytics based database for herbs of India. J Cheminform. 2022;14(1):129.
30. Kim S, Chen J, Cheng T, Gindulyte A, He J, He S, et al. PubChem in 2021: New data content and improved web interfaces. Nucleic Acids Res. 2021;49(D1):D1388–95.
31. Roderer DJA, Glockshuber R, Ban N. ClyA CC6/264 ox (2-303). Biochemistry. 2014;53:6357–69.
32. Trott O, Olson AJ. AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem. 2010;31(2):455–61.
33. Paul DS, Gautham N. Protein–small molecule docking with receptor flexibility in iMOLSDOCK. arXiv [Preprint]. 2020. arXiv:2010.05475.
34. Blaszczyk M, Kurcinski M, Kouza M, Wieteska L, Debinski A, Kolinski A, et al. Modeling of protein–peptide interactions using the CABS-dock web server for binding site search and flexible docking. arXiv [Preprint]. 2015. arXiv:1505.01138.
35. Daina A, Michielin O, Zoete V. SwissADME: A free web tool to evaluate pharmacokinetics, drug-likeness and medicinal chemistry friendliness of small molecules. Sci Rep. 2017;7:42717.
36. Dallakyan S, Olson AJ. AutoDock and AutoDockTools: Automated docking with selective receptor flexibility. Methods Mol Biol. 2015;1115:194–206.
37. DeLano WL. The PyMOL Molecular Graphics System. Schrödinger, LLC; 2020.
38. Trott O, Olson AJ. AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function. J Comput Chem. 2010;31(2):455–61.
39. O’Boyle NM, Banck M, James CA, Morley C, Vandermeersch T, Hutchison GR. Open Babel: An open chemical toolbox. J Cheminform. 2011;3(1):33.
40. Seeliger D, de Groot BL. Ligand docking and binding site analysis with PyMOL and AutoDock/Vina. J Comput Aided Mol Des. 2010;24(5):417–22.
41. Daina A, Michielin O, Zoete V. SwissDock, a protein–small molecule docking web service based on EADock DSS. Nucleic Acids Res. 2013;41(W1):W270–7.
42. Hanwell MD, Curtis DE, Lonie DC, Vandermeersch T, Zurek E, Hutchison GR. Avogadro: An advanced semantic chemical editor, visualization, and analysis platform. J Cheminform. 2012;4(1):17.
43. Li H, Leung T, Wong C. Molecular Docking: A powerful tool for structure-based drug discovery. Curr Comput Aided Drug Des. 2012;8(2):104–13.
44. Dassault Systèmes. BIOVIA Discovery Studio [Internet]. 2021 [cited 2025 Feb 10]. Available from: https://www.3ds.com/products-services/biovia/.
45. Nicholls IA, Pettersson B, Danielson L. Computational approaches to molecular docking and structure-based ligand design. Curr Med Chem. 2008;15(22):2203–12.