Pharmaceutical Research

, Volume 34, Issue 2, pp 352–364 | Cite as

Design and In Vitro Evaluation of Bispecific Complexes and Drug Conjugates of Anticancer Peptide, LyP-1 in Human Breast Cancer

  • Selin Seda Timur
  • Prashant Bhattarai
  • Reyhan Neslihan Gürsoy
  • İmran Vural
  • Ban-An KhawEmail author
Research Paper



LyP-1, a nine-amino-acid tumor homing peptide, selectively binds to its cognate receptor, p32. Overexpression of p32 in certain tumors should allow use of LyP-1 as a targeting agent for the delivery of therapeutic or diagnostic agents. Peptide conjugates are developed for enhanced pre-targeting of MDA-MB-231 breast cancer cells with peptide-antibody bispecific complexes and targeting with multiple-drug/-fluorophore-conjugated nano-polymers.


LyP-1-anti-DTPA bispecific antibody complexes (LyP-1-bsAbCx) were generated by conjugation of anti-DTPA antibody and LyP-1. LyP-1–doxorubicin (Dox), Dox-DTPA-succinyl-polylysine (Dox-DSPL), Dox-DSPL-LyP-1, DTPA-Dox-poly glutamic acid (D-Dox-PGA) or DTPA-rhodamine conjugated polylysine (DSPL-RITC) were prepared. In vitro therapeutic efficacy and targeting by immunofluorescence in MDA-MB-231 breast cancer cells were assessed with Dox-LyP-1. Immunofluorescence visualization of cancer cells was evaluated after pretargeting with LyP-1-bsAbCx and targeting with DSPL-RITC.


Cytotoxicity of Dox-LyP-1 conjugates was significantly greater than free doxorubicin (p < 0.0001). For fluorescent-labeled LyP-1, internalization occurred in 30 min in tumor cells. Fluorescence intensity of two-step targeted cells showed that pretargeting with LyP-1-bsAbC, followed by targeting with DSPL-RITC was greater than non-pretargeted DSPL-RITC (p < 0.05).


Peptide-conjugates are effective targeting agents for MDA-MB-231 breast cancer cells in culture. LyP-1-bsAbCx and Dox-LyP-1 conjugates may allow development of novel targeted cancer therapy and diagnosis.


peptide-antibody bispecific complexes polymer pro-drug conjugates pre-targeted drug targeting 





Doxorubicin conjugated N-terminal DTPA conjugated PGA




Doxorubicin hydrochloride-




LyP-1 conjugated to Dox-DSPL


LyP-1 conjugated with doxorubicin hydrochloride


N-terminal DTPA conjugated PGA




DTPA conjugated poly-L-lysine conjugated with LyP-1 and Dox


RITC conjugated DPL


LyP-1 conjugated to DSPL


DTPA-succinyl rhodamine conjugated poly-L-lysine


Diethylenetriaminepentaacetic acid


DTPA conjugated bovine serum albumin


Goat anti-mouse IgG antibody conjugated with HRP


9 amino acid peptide ligand specific for mitochondrial membrane receptor p32.


LyP-1 conjugated anti-DTPA bispecific antibody complex


Molecular weight cut-off


5/6-carboxyfluorescein succinimidyl ester


Polymer drug conjugates


Poly-L-glutamic acid




Rhodamine B isothiocyanate


Sodium dodecyl sulfate–polyacrylamide gel electrophoresis


Therapeutic monoclonal antibodies


Tri-nitro benzene sulfonic acid



This study was supported by The Scientific and Technological Research Council of Turkey (TÜBİTAK 2214-International Research Fellowship Programme).


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Selin Seda Timur
    • 1
  • Prashant Bhattarai
    • 2
  • Reyhan Neslihan Gürsoy
    • 1
  • İmran Vural
    • 1
  • Ban-An Khaw
    • 2
    Email author
  1. 1.Pharmaceutical Technology Department, Faculty of PharmacyHacettepe UniversityAnkaraTurkey
  2. 2.Department of Pharmaceutical Sciences, School of Pharmacy, Bouve College of Health SciencesNortheastern UniversityBostonUSA

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