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Antibody-mediated targeting of iron oxide nanoparticles to the folate receptor alpha increases tumor cell association in vitro and in vivo.

Ndong C, Toraya-Brown S, Kekalo K, Baker I, Gerngross TU, Fiering SN, Griswold KE - Int J Nanomedicine (2015)

Bottom Line: Here, we describe molecular targeting of iron oxide nanoparticles (IONPs) to the folate receptor alpha (FOLRα) using an engineered antibody fragment (Ffab).Interestingly, Ffab-IONPs built with custom 120 nm nanoparticles exhibited lower in vitro targeting efficiency when compared to those built with commercially sourced 180 nm nanoparticles.In vivo, however, the two Ffab-IONP platforms achieved equivalent tumor homing, although the smaller 120 nm IONPs were more prone to liver sequestration.

View Article: PubMed Central - PubMed

Affiliation: Thayer School of Engineering, Dartmouth, Hanover, NH, USA.

ABSTRACT
Active molecular targeting has become an important aspect of nanoparticle development for oncology indications. Here, we describe molecular targeting of iron oxide nanoparticles (IONPs) to the folate receptor alpha (FOLRα) using an engineered antibody fragment (Ffab). Compared to control nanoparticles targeting the non-relevant botulinum toxin, the Ffab-IONP constructs selectively accumulated on FOLRα-overexpressing cancer cells in vitro, where they exhibited the capacity to internalize into intracellular vesicles. Similarly, Ffab-IONPs homed to FOLRα-positive tumors upon intraperitoneal administration in an orthotopic murine xenograft model of ovarian cancer, whereas negative control particles showed no detectable tumor accumulation. Interestingly, Ffab-IONPs built with custom 120 nm nanoparticles exhibited lower in vitro targeting efficiency when compared to those built with commercially sourced 180 nm nanoparticles. In vivo, however, the two Ffab-IONP platforms achieved equivalent tumor homing, although the smaller 120 nm IONPs were more prone to liver sequestration. Overall, the results show that Ffab-mediated targeting of IONPs yields specific, high-level accumulation within cancer cells, and this fact suggests that Ffab-IONPs could have future utility in ovarian cancer diagnostics and therapy.

No MeSH data available.


Related in: MedlinePlus

In vitro binding studies of Ffab-IONPs and Bfab-IONPs.Notes: (A) Dose–response binding curves for Ffab-CMD (closed square) and Bfab-CMD (open circles) with rFOLRα protein. (B) Dose–response binding curves for Ffab-BNF (closed square) and Bfab-BNF (open circles) with rFOLRα protein. (C) Binding of FOLRα+ KB cancer cells by Ffab-CMD and Bfab-CMD dosed at 35 μg/mL (0.6 nM). (D) Binding of FOLRα+ KB cancer cells by Ffab-BNF and Bfab-BNF dosed at 35 μg/mL (0.035 nM). Error bars represent standard deviation from technical triplicates. ***P<0.001, two-tailed unpaired t-test.Abbreviations: fab, an engineered monoclonal antibody fragment; Ffab, Farletuzufab, engineered from monoclonal antibody Farletuzumab; Bfab, Botulifab anti-botulinum toxin fab fragment; IONP, iron oxide nanoparticle; CMD, carboxymethyl-dextran; rFOLRα, recombinant folate receptor alpha; BNF, bionized nanoferrite, NP, nanoparticle.
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f4-ijn-10-2595: In vitro binding studies of Ffab-IONPs and Bfab-IONPs.Notes: (A) Dose–response binding curves for Ffab-CMD (closed square) and Bfab-CMD (open circles) with rFOLRα protein. (B) Dose–response binding curves for Ffab-BNF (closed square) and Bfab-BNF (open circles) with rFOLRα protein. (C) Binding of FOLRα+ KB cancer cells by Ffab-CMD and Bfab-CMD dosed at 35 μg/mL (0.6 nM). (D) Binding of FOLRα+ KB cancer cells by Ffab-BNF and Bfab-BNF dosed at 35 μg/mL (0.035 nM). Error bars represent standard deviation from technical triplicates. ***P<0.001, two-tailed unpaired t-test.Abbreviations: fab, an engineered monoclonal antibody fragment; Ffab, Farletuzufab, engineered from monoclonal antibody Farletuzumab; Bfab, Botulifab anti-botulinum toxin fab fragment; IONP, iron oxide nanoparticle; CMD, carboxymethyl-dextran; rFOLRα, recombinant folate receptor alpha; BNF, bionized nanoferrite, NP, nanoparticle.

Mentions: FOLRα binding of the Ffab-CMD and Ffab-BNF constructs, and their respective Bfab-IONP negative control counterparts was evaluated by ELISA (Figure 4A, B). Compared to the monomeric Ffab antibody, both Ffab-IONP constructs exhibited orders of magnitude higher apparent affinities (Ffab-BNF EC50 =0.013 nM; Ffab-CMD EC50 =0.16 nM), which reflects avidity effects derived from the nanoparticles’ polyvalent nature. Conversely, no substantial binding was observed with the Bfab-BNF or Bfab-CMD negative control nanoparticles. These results demonstrated that Ffab binding of rFOLRα was not compromised during IONP conjugation, and that the Bfab-IONP controls had no inherent affinity for the recombinant receptor. It is noteworthy that the larger Ffab-BNF particles exhibited 10-fold higher apparent affinity compared to the Ffab-CMD particles. This effect likely results from their 10-fold difference in number of conjugated antibodies (500 vs 50 antibodies for BNF and CMD, respectively; Table 1). The higher maximum signal of the Ffab-BNF particles is likely due, in part, to their higher apparent affinity, and in part, due to the fact that BNF nanoparticles possess greater iron content than CMD nanoparticles (0.6 mg iron/mg of particles vs 0.5 mg iron/mg of particles, respectively).


Antibody-mediated targeting of iron oxide nanoparticles to the folate receptor alpha increases tumor cell association in vitro and in vivo.

Ndong C, Toraya-Brown S, Kekalo K, Baker I, Gerngross TU, Fiering SN, Griswold KE - Int J Nanomedicine (2015)

In vitro binding studies of Ffab-IONPs and Bfab-IONPs.Notes: (A) Dose–response binding curves for Ffab-CMD (closed square) and Bfab-CMD (open circles) with rFOLRα protein. (B) Dose–response binding curves for Ffab-BNF (closed square) and Bfab-BNF (open circles) with rFOLRα protein. (C) Binding of FOLRα+ KB cancer cells by Ffab-CMD and Bfab-CMD dosed at 35 μg/mL (0.6 nM). (D) Binding of FOLRα+ KB cancer cells by Ffab-BNF and Bfab-BNF dosed at 35 μg/mL (0.035 nM). Error bars represent standard deviation from technical triplicates. ***P<0.001, two-tailed unpaired t-test.Abbreviations: fab, an engineered monoclonal antibody fragment; Ffab, Farletuzufab, engineered from monoclonal antibody Farletuzumab; Bfab, Botulifab anti-botulinum toxin fab fragment; IONP, iron oxide nanoparticle; CMD, carboxymethyl-dextran; rFOLRα, recombinant folate receptor alpha; BNF, bionized nanoferrite, NP, nanoparticle.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4388088&req=5

f4-ijn-10-2595: In vitro binding studies of Ffab-IONPs and Bfab-IONPs.Notes: (A) Dose–response binding curves for Ffab-CMD (closed square) and Bfab-CMD (open circles) with rFOLRα protein. (B) Dose–response binding curves for Ffab-BNF (closed square) and Bfab-BNF (open circles) with rFOLRα protein. (C) Binding of FOLRα+ KB cancer cells by Ffab-CMD and Bfab-CMD dosed at 35 μg/mL (0.6 nM). (D) Binding of FOLRα+ KB cancer cells by Ffab-BNF and Bfab-BNF dosed at 35 μg/mL (0.035 nM). Error bars represent standard deviation from technical triplicates. ***P<0.001, two-tailed unpaired t-test.Abbreviations: fab, an engineered monoclonal antibody fragment; Ffab, Farletuzufab, engineered from monoclonal antibody Farletuzumab; Bfab, Botulifab anti-botulinum toxin fab fragment; IONP, iron oxide nanoparticle; CMD, carboxymethyl-dextran; rFOLRα, recombinant folate receptor alpha; BNF, bionized nanoferrite, NP, nanoparticle.
Mentions: FOLRα binding of the Ffab-CMD and Ffab-BNF constructs, and their respective Bfab-IONP negative control counterparts was evaluated by ELISA (Figure 4A, B). Compared to the monomeric Ffab antibody, both Ffab-IONP constructs exhibited orders of magnitude higher apparent affinities (Ffab-BNF EC50 =0.013 nM; Ffab-CMD EC50 =0.16 nM), which reflects avidity effects derived from the nanoparticles’ polyvalent nature. Conversely, no substantial binding was observed with the Bfab-BNF or Bfab-CMD negative control nanoparticles. These results demonstrated that Ffab binding of rFOLRα was not compromised during IONP conjugation, and that the Bfab-IONP controls had no inherent affinity for the recombinant receptor. It is noteworthy that the larger Ffab-BNF particles exhibited 10-fold higher apparent affinity compared to the Ffab-CMD particles. This effect likely results from their 10-fold difference in number of conjugated antibodies (500 vs 50 antibodies for BNF and CMD, respectively; Table 1). The higher maximum signal of the Ffab-BNF particles is likely due, in part, to their higher apparent affinity, and in part, due to the fact that BNF nanoparticles possess greater iron content than CMD nanoparticles (0.6 mg iron/mg of particles vs 0.5 mg iron/mg of particles, respectively).

Bottom Line: Here, we describe molecular targeting of iron oxide nanoparticles (IONPs) to the folate receptor alpha (FOLRα) using an engineered antibody fragment (Ffab).Interestingly, Ffab-IONPs built with custom 120 nm nanoparticles exhibited lower in vitro targeting efficiency when compared to those built with commercially sourced 180 nm nanoparticles.In vivo, however, the two Ffab-IONP platforms achieved equivalent tumor homing, although the smaller 120 nm IONPs were more prone to liver sequestration.

View Article: PubMed Central - PubMed

Affiliation: Thayer School of Engineering, Dartmouth, Hanover, NH, USA.

ABSTRACT
Active molecular targeting has become an important aspect of nanoparticle development for oncology indications. Here, we describe molecular targeting of iron oxide nanoparticles (IONPs) to the folate receptor alpha (FOLRα) using an engineered antibody fragment (Ffab). Compared to control nanoparticles targeting the non-relevant botulinum toxin, the Ffab-IONP constructs selectively accumulated on FOLRα-overexpressing cancer cells in vitro, where they exhibited the capacity to internalize into intracellular vesicles. Similarly, Ffab-IONPs homed to FOLRα-positive tumors upon intraperitoneal administration in an orthotopic murine xenograft model of ovarian cancer, whereas negative control particles showed no detectable tumor accumulation. Interestingly, Ffab-IONPs built with custom 120 nm nanoparticles exhibited lower in vitro targeting efficiency when compared to those built with commercially sourced 180 nm nanoparticles. In vivo, however, the two Ffab-IONP platforms achieved equivalent tumor homing, although the smaller 120 nm IONPs were more prone to liver sequestration. Overall, the results show that Ffab-mediated targeting of IONPs yields specific, high-level accumulation within cancer cells, and this fact suggests that Ffab-IONPs could have future utility in ovarian cancer diagnostics and therapy.

No MeSH data available.


Related in: MedlinePlus