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. 2016 Mar;19(1):37-44.
doi: 10.3831/KPI.2016.19.005.

Nanopharmaceutical Approach for Enhanced Anti-cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target: -Anti-cancer Potential of Nano-betulinic Acid in Lung Cancer

Jayeeta Das  1 , Asmita Samadder  2 , Sreemanti Das  1 , Avijit Paul  1 , Anisur Rahman Khuda-Bukhsh  1
Affiliations

Nanopharmaceutical Approach for Enhanced Anti-cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target: -Anti-cancer Potential of Nano-betulinic Acid in Lung Cancer

Jayeeta Das et al. J Pharmacopuncture. 2016 Mar.

Abstract

Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA) and its poly (lactide- co-glycolide) (PLGA) nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA) + benzo]undefined[a]pyrene (BaP)]-induced lung cancer in mice in vivo.

Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA) were determined by using transmission electron microscopy (TEM), and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA) as a target were analyzed by using conventional circular dichroism (CD) and melting temperature (Tm) profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA); the ability of NdBA to cross the blood-brain barrier (BBB) was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence-activated cell-sorting (FACS) data analysis.

Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB.

Conclusion: Thus, compared to dBA, NdBA appears to have greater chemoprotective potential against lung cancer.

Keywords: A549 cell line; betulinic acid; drug-DNA interaction; mice; poly (lactide-co-glycolide).

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Conflict of interest statement

Conflict of interest The authors declare that there are no conflict of interest.

Figures

Fig. 1
Fig. 1. TEM images of (A) at low magnification (12000X) and (B) at high magnification (HRTEM, 800000X). (C) CTDNA fragmentation assay: Ln1 = DNA ladder, Ln2 = CTDNA, Ln3 = CT-DNA + dBA, and Ln4 = CT-DNA + NdBA.
Fig. 2
Fig. 2. (A) CD pectral analyses and (B) Tm profiles of CTDNA alone, CT-DNA + dBA and CT-DNA + NdBA.
Fig. 3
Fig. 3. (A) Cellular cytotoxicity assay of A549 lung cancer cells, (B) cellular cytotoxicity assay of normal L6 skeletal muscle cells, and (C) confocal microscopic studies for cellular entry of nanoparticles in A549 cells.
Fig. 4
Fig. 4. (A) Expressions of PARP, Bax and Bcl2 proteins according to indirect ELISA. (B) ROS generation studies of experimental sets of alveolar cells isolated from mice: a = normal control, b = co-mutagenic control, c = co-mutagenic + dBA, d = co-mutagenic + NdBA(I), and e = co-mutagenic + NdBA(II).
Fig. 5
Fig. 5. (A) Chromosomal study of metaphase complements and (B) bone marrow smears showing normal/ micronucleated erythrocytes in different groups of mice: a = normal control, b = co-mutagenic control, c = co-mutagenic + dBA, = co-mutagenic + NdBA(I), and e = co-mutagenic + NdBA(II).
Fig. 6
Fig. 6. (A) Histological analyses of lung tissues before and after treatment with co-mutagens and drugs: a = normal control, b = co-mutagenic control, c = co-mutagenic + dBA, d = co-mutagenic + NdBA(I), and e = co-mutagenic + NdBA(II). (B) Transmission electron microscopy images of brain tissue to detect the capability of NdBA to cross the blood brain barrier: a = normal mouse without any nanoparticle administration, b = mouse brain after administration of NdBA(II). The arrowheads mark the presence of NdBA, and Nu stands for nucleus.
Fig. 7
Fig. 7. Nanopharmaceutical Approach for Enhanced Anti- cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target.
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References

    1. Pershagen G, Nordberg G, Björklund NE. Carcinomas of the respiratory tract in hamsters given arsenic trioxide and/or benzo[a]pyrene by the pulmonary route. Environ Res. 1984;34(2):227–241. doi: 10.1016/0013-9351(84)90091-4. - DOI - PubMed
    1. Deng GE, Rausch SM, Jones LW, Gulati A, Kumar NB, Greenlee H et al. Complementary therapies and integrative medicine in lung cancer: diagnosis and management of lung cancer, 3rd ed: american college of chest physicians evidence based clinical practice guidelines. Chest. 2013;DOI: 10. Chest. 2013 doi: 10.1378/chest.12-2364. - DOI - PubMed
    1. Ellingwood F. The American Materia Medica, Therapeutics and Pharmacognosy. Eclectic Medical Publications; Portland: 1919. 470
    1. Wang L, Bai L, Nagasawa T, Hasegawa T, Yang X, Sakai J et al. Bioactive triterpene saponin from the roots of Phytolacca americana. J Nat Prod. 2008;71(1):35–40. - PubMed
    1. Das J, Das S, Samadder A, Bhadra K, Khuda-Bukhsh AR. Poly (lactide-co-glycolide) encapsulated extract of phytolacca decandra demonstrates better intervention against induced lung adenocarcinoma in mice and on A549 cells. Eur J Pharm Sci. 2012;47(2):313–324. doi: 10.1016/j.ejps.2012.06.018. - DOI - PubMed