Pharmaceutical Research

, Volume 29, Issue 10, pp 2874–2886 | Cite as

Phosphatidylinositol 3-kinase Inhibitor (PIK75) Containing Surface Functionalized Nanoemulsion for Enhanced Drug Delivery, Cytotoxicity and Pro-apoptotic Activity in Ovarian Cancer Cells

  • Meghna Talekar
  • Srinivas Ganta
  • Amit Singh
  • Mansoor Amiji
  • Jackie Kendall
  • William A. Denny
  • Sanjay Garg
Research Paper

ABSTRACT

Purpose

Ovarian cancer is a debilitating disease, which needs multi-pronged approach of targeted drug delivery and enhanced efficacy with the use of combination therapeutics. In this study, we have examined the anticancer activity of PIK75 incorporated in surface functionalized nanoemulsions for targeted delivery to SKOV-3 cells. A pro-apoptotic molecule C6-ceramide was also co-delivered to augment therapeutic efficacy.

Methods

EGFR and FR functionalized nanoemulsions incorporating PIK75 and C6-ceramide were characterized for particle size, surface charge, entrapment efficiency and morphology. Fluorescence and quantitative uptake studies were conducted in SKOV-3 cells to determine intracellular distribution. Cell viability was assessed using MTT assay while mechanism of cytotoxicity was evaluated using capsase-3/7, TUNEL and hROS assay.

Results

Cytotoxicity assay showed 57% decrease in IC50 value of PIK75 following treatment with EGFR targeted nanoemulsion and 40% decrease following treatment with FR targeted nanoemulsion. Combination therapy with PIK75 and ceramide enhanced the cytotoxicity of PIK75 compared to therapy with individual formulations. The increase in cytotoxicity was attributed to increase in cellular apoptosis and hROS activity.

Conclusion

The results of this study showed that the targeted system improved cytotoxicity of PIK75 compared to the non-targeted system. Combination therapy with ceramide augmented PIK75’s therapeutic activity.

KEY WORDS

C6-ceramide EGFR folate nanoemulsion ovarian cancer. phosphatidylinositol 3-kinase inhibitor 

ABBREVIATIONS

CER

ceramide

EGFR

epidermal growth factor receptor

FR

folate receptor

NE

nanoemulsions

PI3K

Phosphatidylinositol 3-kinase

PIP2

Phosphatidylinositol 4, 5-diphosphate

PIP3

Phosphatidylinositol 3, 4, 5-triphosphate

REFERENCES

  1. 1.
  2. 2.
    Gymnopoulos M, Elsliger MA, Vogt PK. Rare cancer-specific mutations in PIK3CA show gain of function. Proc Natl Acad Sci U S A. 2007;104(13):5569–74.PubMedCrossRefGoogle Scholar
  3. 3.
    Bader AG, Kang S, Vogt PK. Cancer-specific mutations in PIK3CA are oncogenic in vivo. Proc Natl Acad Sci U S A. 2006;103(5):1475–9.PubMedCrossRefGoogle Scholar
  4. 4.
    Frédérick R, Mawson C, Kendall JD, Chaussade C, Rewcastle GW, Shepherd PR, Denny WA. Phosphoinositide-3-kinase (PI3K) inhibitors: Identification of new scaffolds using virtual screening. Bioorg Med Chem Lett. 2009;19(20):5842–7.PubMedCrossRefGoogle Scholar
  5. 5.
    Hu L, Hofmann J, Lu Y, Mills GB, Jaffe RB. Inhibition of phosphatidylinositol 3′-kinase increases efficacy of paclitaxel in in vitro and in vivo ovarian cancer models. Cancer Res. 2002;62(4):1087–92.PubMedGoogle Scholar
  6. 6.
    Karakas B, Bachman KE, Park BH. Mutation of the PIK3CA oncogene in human cancers. Br J Cancer. 2006;94(4):455–9.PubMedCrossRefGoogle Scholar
  7. 7.
    Marone R, Cmiljanovic V, Giese B, Wymann M. Targeting phosphoinositide 3-kinase--Moving towards therapy. Biochim Biophys Acta Protein Proteonomics. 2008;1784(1):159–85.CrossRefGoogle Scholar
  8. 8.
    Hirsch E, Ciraolo E, Ghigo A, Costa C. Taming the PI3K team to hold inflammation and cancer at bay. Pharmacol Ther. 2008;118(2):192–205.PubMedCrossRefGoogle Scholar
  9. 9.
    Maira SM, Voliva C, Garcia-Echeverria C. Class IA phosphatidylinositol 3-kinase: from their biologic implication in human cancers to drug discovery. Expert Opin Ther Targets. 2008;12(2):223–38.PubMedCrossRefGoogle Scholar
  10. 10.
    Knight ZA, Gonzalez B, Feldman ME, Zunder ER, Goldenberg DD, Williams O, Loewith R, Stokoe D, Balla A, Toth B, Balla T, Weiss WA, Williams RL, Shokat KM. A Pharmacological map of the PI3-K family defines a role for p110α in insulin signaling. Cell. 2006;125(4):733–47.PubMedCrossRefGoogle Scholar
  11. 11.
    Chaussade C, Rewcastle GW, Kendall JD, Denny WA, Cho K, Grønning LM, Chong ML, Anagnostou SH, Jackson SP, Daniele N, Shepherd PR. Evidence for functional redundancy of class IA PI3K isoforms in insulin signalling. Biochem J. 2007;404(3):449–58.PubMedCrossRefGoogle Scholar
  12. 12.
    Kendall JD, Rewcastle GW, Frederick R, Mawson C, Denny WA, Marshall ES, Baguley BC, Chaussade C, Jackson SP, Shepherd PR. Synthesis, biological evaluation and molecular modelling of sulfonohydrazides as selective PI3K p110[alpha] inhibitors. Bioorg Med Chem. 2007;15(24):7677–87.PubMedCrossRefGoogle Scholar
  13. 13.
    Hayakawa M, Kawaguchi KI, Kaizawa H, Koizumi T, Ohishi T, Yamano M, Okada M, Ohta M, Tsukamoto SI, Raynaud FI, Parker P, Workman P, Waterfield MD. Synthesis and biological evaluation of sulfonylhydrazone-substituted imidazo[1,2-a]pyridines as novel PI3 kinase p110α inhibitors. Bioorg Med Chem. 2007;15(17):5837–44.PubMedCrossRefGoogle Scholar
  14. 14.
    Dagia NM, Agarwal G, Kamath DV, Chetrapal-Kunwar A, Gupte RD, Jadhav MG, Dadarkar SS, Trivedi J, Kulkarni-Almeida AA, Kharas F, Fonseca LC, Kumar S, Bhonde MR. A preferential p110α/γ PI3K inhibitor attenuates experimental inflammation by suppressing the production of proinflammatory mediators in a NF-κB-dependent manner. Am J Physiol Cell Physiol. 2010;298(4):C929–41.PubMedCrossRefGoogle Scholar
  15. 15.
    Tiwari SB, Amiji MM. Improved oral delivery of paclitaxel following administration in nanoemulsion formulations. J Nanosci Nanotechnol. 2006;6(9–10):3215–21.PubMedCrossRefGoogle Scholar
  16. 16.
    Lo Prete AC, Maria DA, Rodrigues DG, Valduga CJ, Ibañez OCM, Maranhão RC. Evaluation in melanoma-bearing mice of an etoposide derivative associated to a cholesterol-rich nanoemulsion. J Pharm Pharmacol. 2006;58(6):801–8.PubMedCrossRefGoogle Scholar
  17. 17.
    Ohguchi Y, Kawano K, Hattori Y, Maitani Y. Selective delivery of folate-PEG-linked, nanoemulsion-loaded aclacinomycin A to KB nasopharyngeal cells and xenograft: Effect of chain length and amount of folate-PEG linker. J Drug Target. 2008;16(9):660–7.PubMedCrossRefGoogle Scholar
  18. 18.
    Tagne JB, Kakumanu S, Nicolosi RJ. Nanoemulsion preparations of the anticancer drug dacarbazine significantly increase its efficacy in a xenograft mouse melanoma model. Mol Pharm. 2008;5(6):1055–63.PubMedCrossRefGoogle Scholar
  19. 19.
    Tagne JB, Kakumanu S, Ortiz D, Shea T, Nicolosi RJ. A nanoemulsion formulation of tamoxifen increases its efficacy in a breast cancer cell line. Mol Pharm. 2008;5(2):280–6.PubMedCrossRefGoogle Scholar
  20. 20.
    Kawakami S, Yamashita F, Hashida M. Disposition characteristics of emulsions and incorporated drugs after systemic or local injection. Adv Drug Deliv Rev. 2000;45(1):77–88.PubMedCrossRefGoogle Scholar
  21. 21.
    Milane L, Duan Z, Amiji M. Development of EGFR-targeted polymer blend nanocarriers for combination paclitaxel/lonidamine delivery to treat multi-drug resistance in human breast and ovarian tumor cells. Mol Pharm. 2011;8(1):185–203.PubMedCrossRefGoogle Scholar
  22. 22.
    Magadala P, Amiji M. Epidermal growth factor receptor-targeted gelatin-based engineered nanocarriers for DNA delivery and transfection in human pancreatic cancer cells. AAPS J. 2008;10(4):565–76.PubMedCrossRefGoogle Scholar
  23. 23.
    Liu Y, Sun J, Cao W, Yang J, Lian H, Li X, Sun Y, Wang Y, Wang S, He Z. Dual targeting folate-conjugated hyaluronic acid polymeric micelles for paclitaxel delivery. Int J Pharm. 2011;421(1):160–9.PubMedCrossRefGoogle Scholar
  24. 24.
    Kang C, Yuan X, Zhong Y, Pu P, Guo Y, Albadany A, Yu S, Zhang Z, Li Y, Chang J, Sheng J. Evaluation of folate-PAMAM for the delivery of antisense oligonucleotides to rat C6 glioma cells in vitro and in vivo. J Biomed Mater Re. 2010;93(2):585–94.Google Scholar
  25. 25.
    Leamon CP, Low PS. Selective targeting of malignant cells with cytotoxin-folate conjugates. J Drug Target. 1994;2(2):101–12.PubMedCrossRefGoogle Scholar
  26. 26.
    Asadishad B, Vossoughi M, Alemzadeh I. Folate-receptor-targeted delivery of doxorubicin using polyethylene glycol-functionalized gold nanoparticles. Ind Eng Chem Res. 2010;49(4):1958–63.CrossRefGoogle Scholar
  27. 27.
    Huang W, Chen, C., Lin, Y, Lin, C. Apoptotic sphingolipid ceramide in cancer therapy. Journal of Lipids. 2011;2011.Google Scholar
  28. 28.
    Van Vlerken LE, Duan Z, Little SR, Seiden MV, Amiji MM. Modulation of intracellular ceramide using polymeric nanoparticles to overcome multidrug resistance in cancer. Cancer Res. 2007;67(10):4843–50.PubMedCrossRefGoogle Scholar
  29. 29.
    Devalapally H, Duan Z, Seiden MV, Amiji MM. Paclitaxel and ceramide co-administration in biodegradable polymeric nanoparticulate delivery system to overcome drug resistance in ovarian cancer. Int J Cancer. 2007;121(8):1830–8.PubMedCrossRefGoogle Scholar
  30. 30.
    Li Z, Zhao R, Wu X, Sun Y, Yao M, Li J, Xu Y, Gu J. Identification and characterization of a novel peptide ligand of epidermal growth factor receptor for targeted delivery of therapeutics. FASEB J. 2005;19(14):1978–85.PubMedCrossRefGoogle Scholar
  31. 31.
    Song S, Liu D, Peng J, Sun Y, Li Z, Gu JR, Xu Y. Peptide ligand-mediated liposome distribution and targeting to EGFR expressing tumor in vivo. Int J Pharm. 2008;363(1-2):155–61.PubMedCrossRefGoogle Scholar
  32. 32.
    van Vlerken LE, Duan Z, Little SR, Seiden MV, Amiji MM. Biodistribution and pharmacokinetic analysis of paclitaxel and ceramide administered in multifunctional polymer-blend nanoparticles in drug resistant breast cancer model. Mol Pharm. 2008;5(4):516–26.PubMedCrossRefGoogle Scholar
  33. 33.
    Ganta S, Amiji M. Coadministration of paclitaxel and curcumin in nanoemulsion formulations to overcome multidrug resistance in tumor cells. Mol Pharm. 2009;6(3):928–39.PubMedCrossRefGoogle Scholar
  34. 34.
    Ganta S, Devalapally H, Amiji M. Curcumin enhances oral bioavailability and anti-tumor therapeutic efficacy of paclitaxel upon administration in nanoemulsion formulation. J Pharm Sci. 2010;99(11):4630–41.PubMedCrossRefGoogle Scholar
  35. 35.
    Ganta S, Paxton JW, Baguley BC, Garg S. Pharmacokinetics and pharmacodynamics of chlorambucil delivered in parenteral emulsion. Int J Pharm. 2008;360(1–2):115–21.PubMedCrossRefGoogle Scholar
  36. 36.
    Talekar M, Kendall J, Denny W, Garg S. Targeting of nanoparticles in cancer: drug delivery and diagnostics. Anti Canc Drugs. 2011;22(10):949–62.CrossRefGoogle Scholar
  37. 37.
    Werner ME, Karve S, Sukumar R, Cummings ND, Copp JA, Chen RC, Zhang T, Wang AZ. Folate-targeted nanoparticle delivery of chemo- and radiotherapeutics for the treatment of ovarian cancer peritoneal metastasis. Biomaterials. 2011;32(33):8548–54.PubMedCrossRefGoogle Scholar
  38. 38.
    Shayesteh L, Lu Y, Kuo WL, Baldocchi R, Godfrey T, Collins C, Pinkel D, Powell B, Mills GB, Gray JW. PlK3CA is implicated as an oncogene in ovarian cancer. Nat Genet. 1999;21(1):99–102.PubMedCrossRefGoogle Scholar
  39. 39.
    Esmaeili F, Ghahremani MH, Ostad SN, Atyabi F, Seyedabadi M, Malekshahi MR, Amini M, Dinarvand R. Folate-receptor-targeted delivery of docetaxel nanoparticles prepared by PLGA-PEG-folate conjugate. J Drug Target. 2008;16(5):415–23.PubMedCrossRefGoogle Scholar
  40. 40.
    Orth JD, Krueger EW, Weller SG, McNiven MA. A novel endocytic mechanism of epidermal growth factor receptor sequestration and internalization. Cancer Res. 2006;66(7):3603–10.PubMedCrossRefGoogle Scholar
  41. 41.
    Thevissen K, François IEJA, Winderickx J, Pannecouque C, Cammue BPA. Ceramide involvement in apoptosis and apoptotic diseases. Mini-Reviews in Medicinal Chem. 2006;6(6):699–709.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Meghna Talekar
    • 1
  • Srinivas Ganta
    • 2
  • Amit Singh
    • 3
  • Mansoor Amiji
    • 3
  • Jackie Kendall
    • 4
  • William A. Denny
    • 4
  • Sanjay Garg
    • 1
    • 5
  1. 1.School of Pharmacy, Faculty of Medical & Health SciencesThe University of AucklandAucklandNew Zealand
  2. 2.Nemucore Medical Innovations, Inc.WorcesterUSA
  3. 3.Department of Pharmaceutical Sciences, School of PharmacyNortheastern UniversityBostonUSA
  4. 4.Auckland Cancer Society Research Centre Faculty of Medical & Health SciencesThe University of AucklandAucklandNew Zealand
  5. 5.School of Pharmacy & Medical SciencesUniversity of South Australia (UniSa)AdelaideAustralia

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