Limits...
Enhanced Specificity and Drug Delivery in Tumors by cRGD - Anchoring Thermosensitive Liposomes.

Dicheva BM, Ten Hagen TL, Seynhaeve AL, Amin M, Eggermont AM, Koning GA - Pharm. Res. (2015)

Bottom Line: Cytotoxic effect of TSL and RGD-TSL was studied on B16Bl6 melanoma, B16F10 melanoma and HUVEC.High resolution intravital microscopy demonstrated specific accumulation of RGD-TSL to the tumor vasculature.Moreover, application of hyperthermia resulted in massive drug release from RGD-TSL.

View Article: PubMed Central - PubMed

Affiliation: Laboratory Experimental Surgical Oncology, Section Surgical Oncology Department of Surgery, Erasmus MC Cancer Center, Rotterdam, The Netherlands. b.dicheva@erasmusmc.nl.

ABSTRACT

Purpose: To develop RGD-targeted thermosensitive liposomes with increased tumor retention, improving drug release efficiency upon mild hyperthermia (HT) in both tumor and angiogenic endothelial cells.

Methods: Standard termosensitive liposomes (TSL) and TSL containing a cyclic Arg-Gly-Asp (cRGD) pentapeptide with the sequence Arg-Cys-D-Phe-Asp-Gly (RGDf[N-Met]C) were synthetized, loaded with Dox and characterized. Temperature- and time-dependent drug release profiles were assessed by fluorometry. Intracellular Dox delivery was studied by flow cytometry and confocal microscopy. Cytotoxic effect of TSL and RGD-TSL was studied on B16Bl6 melanoma, B16F10 melanoma and HUVEC. Intravital microscopy was performed on B16Bl6 tumors implanted in dorsal-skin fold window-bearing mice. Pharmacokinetic and biodistribution of Dox-TSL and Dox-RGD-TSL were followed in B16Bl6 tumor bearing mice upon normothermia or initial hyperthermia conditions.

Results: DLS and cryo-TEM revealed particle homogeneity and size of around 85 nm. Doxorubicin loading efficiency was >95%as assessed by spectrofluorometry. Flow cytometry and confocal microscopy showed a specific uptake of RGD-TSL by melanoma and endothelial cells when compared to TSL and an increased doxorubicin delivery. High resolution intravital microscopy demonstrated specific accumulation of RGD-TSL to the tumor vasculature. Moreover, application of hyperthermia resulted in massive drug release from RGD-TSL. Biodistribution studies showed that initial hyperthermia increases Dox uptake in tumors from TSL and RGD-TSL.

Conclusion: RGD-TSL have potency to increase drug efficacy due to higher uptake by tumor and angiogenic endothelial cells in combination with heat-triggered drug release.

No MeSH data available.


Related in: MedlinePlus

Dox release upon HT treatment from TSL (a) and RGD-TSL (b) in B16Bl6 window chamber bearing mice. Mice were injected with 5 mg/kg Dox in DiD-labelled (purple) TSL or RGD-TSL. After 5 h of liposome circulation, a temperature of 42°C for 1 h was applied to trigger Dox release. Representative images were taken from the beginning of the HT treatment up to 1 h. (c). In vivo quantification of Dox released from RGD-TSL or TSL 1 h after HT treatment, presented as integrated density (IntDen), see materials and methods. (d). Dox uptake in endothelial cells (green) and tumor cells from RGD-TSL after 1 h of HT treatment. Scale bar applies for all images, 50 μm.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4628091&req=5

Fig7: Dox release upon HT treatment from TSL (a) and RGD-TSL (b) in B16Bl6 window chamber bearing mice. Mice were injected with 5 mg/kg Dox in DiD-labelled (purple) TSL or RGD-TSL. After 5 h of liposome circulation, a temperature of 42°C for 1 h was applied to trigger Dox release. Representative images were taken from the beginning of the HT treatment up to 1 h. (c). In vivo quantification of Dox released from RGD-TSL or TSL 1 h after HT treatment, presented as integrated density (IntDen), see materials and methods. (d). Dox uptake in endothelial cells (green) and tumor cells from RGD-TSL after 1 h of HT treatment. Scale bar applies for all images, 50 μm.

Mentions: In order to understand whether RGD-TSL release Dox in vivo upon HT treatment and to follow Dox distribution in the tumor, intravital microscopy was performed. Circulation of TSL and RGD-TSL for 5 h in the blood stream did not cause any premature release of Dox (data not shown). However, when HT at 42°C was applied, immediate Dox release (red) was observed from TSL (Fig. 7a) and RGD-TSL (Fig 7b). Dox from both formulations was released first intravascularly from the circulating DiD-labelled liposomes (purple), after which it was gradually taken up by endothelial cells and tumor cells surrounding the blood vessels (Fig. 7d and supporting information, video 1). The Dox uptake in the both treatment groups increased in time and was maximal after 1 h of HT, when also the lumen of the blood vessels was cleared from Dox. Quantification of the images showed that the amount of delivered Dox to the tumor from RGD-TSL was 1.7 fold higher than from TSL (Fig. 7c). However, this difference was not statistically significant (p-value = 0.8).Fig. 7


Enhanced Specificity and Drug Delivery in Tumors by cRGD - Anchoring Thermosensitive Liposomes.

Dicheva BM, Ten Hagen TL, Seynhaeve AL, Amin M, Eggermont AM, Koning GA - Pharm. Res. (2015)

Dox release upon HT treatment from TSL (a) and RGD-TSL (b) in B16Bl6 window chamber bearing mice. Mice were injected with 5 mg/kg Dox in DiD-labelled (purple) TSL or RGD-TSL. After 5 h of liposome circulation, a temperature of 42°C for 1 h was applied to trigger Dox release. Representative images were taken from the beginning of the HT treatment up to 1 h. (c). In vivo quantification of Dox released from RGD-TSL or TSL 1 h after HT treatment, presented as integrated density (IntDen), see materials and methods. (d). Dox uptake in endothelial cells (green) and tumor cells from RGD-TSL after 1 h of HT treatment. Scale bar applies for all images, 50 μm.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4628091&req=5

Fig7: Dox release upon HT treatment from TSL (a) and RGD-TSL (b) in B16Bl6 window chamber bearing mice. Mice were injected with 5 mg/kg Dox in DiD-labelled (purple) TSL or RGD-TSL. After 5 h of liposome circulation, a temperature of 42°C for 1 h was applied to trigger Dox release. Representative images were taken from the beginning of the HT treatment up to 1 h. (c). In vivo quantification of Dox released from RGD-TSL or TSL 1 h after HT treatment, presented as integrated density (IntDen), see materials and methods. (d). Dox uptake in endothelial cells (green) and tumor cells from RGD-TSL after 1 h of HT treatment. Scale bar applies for all images, 50 μm.
Mentions: In order to understand whether RGD-TSL release Dox in vivo upon HT treatment and to follow Dox distribution in the tumor, intravital microscopy was performed. Circulation of TSL and RGD-TSL for 5 h in the blood stream did not cause any premature release of Dox (data not shown). However, when HT at 42°C was applied, immediate Dox release (red) was observed from TSL (Fig. 7a) and RGD-TSL (Fig 7b). Dox from both formulations was released first intravascularly from the circulating DiD-labelled liposomes (purple), after which it was gradually taken up by endothelial cells and tumor cells surrounding the blood vessels (Fig. 7d and supporting information, video 1). The Dox uptake in the both treatment groups increased in time and was maximal after 1 h of HT, when also the lumen of the blood vessels was cleared from Dox. Quantification of the images showed that the amount of delivered Dox to the tumor from RGD-TSL was 1.7 fold higher than from TSL (Fig. 7c). However, this difference was not statistically significant (p-value = 0.8).Fig. 7

Bottom Line: Cytotoxic effect of TSL and RGD-TSL was studied on B16Bl6 melanoma, B16F10 melanoma and HUVEC.High resolution intravital microscopy demonstrated specific accumulation of RGD-TSL to the tumor vasculature.Moreover, application of hyperthermia resulted in massive drug release from RGD-TSL.

View Article: PubMed Central - PubMed

Affiliation: Laboratory Experimental Surgical Oncology, Section Surgical Oncology Department of Surgery, Erasmus MC Cancer Center, Rotterdam, The Netherlands. b.dicheva@erasmusmc.nl.

ABSTRACT

Purpose: To develop RGD-targeted thermosensitive liposomes with increased tumor retention, improving drug release efficiency upon mild hyperthermia (HT) in both tumor and angiogenic endothelial cells.

Methods: Standard termosensitive liposomes (TSL) and TSL containing a cyclic Arg-Gly-Asp (cRGD) pentapeptide with the sequence Arg-Cys-D-Phe-Asp-Gly (RGDf[N-Met]C) were synthetized, loaded with Dox and characterized. Temperature- and time-dependent drug release profiles were assessed by fluorometry. Intracellular Dox delivery was studied by flow cytometry and confocal microscopy. Cytotoxic effect of TSL and RGD-TSL was studied on B16Bl6 melanoma, B16F10 melanoma and HUVEC. Intravital microscopy was performed on B16Bl6 tumors implanted in dorsal-skin fold window-bearing mice. Pharmacokinetic and biodistribution of Dox-TSL and Dox-RGD-TSL were followed in B16Bl6 tumor bearing mice upon normothermia or initial hyperthermia conditions.

Results: DLS and cryo-TEM revealed particle homogeneity and size of around 85 nm. Doxorubicin loading efficiency was >95%as assessed by spectrofluorometry. Flow cytometry and confocal microscopy showed a specific uptake of RGD-TSL by melanoma and endothelial cells when compared to TSL and an increased doxorubicin delivery. High resolution intravital microscopy demonstrated specific accumulation of RGD-TSL to the tumor vasculature. Moreover, application of hyperthermia resulted in massive drug release from RGD-TSL. Biodistribution studies showed that initial hyperthermia increases Dox uptake in tumors from TSL and RGD-TSL.

Conclusion: RGD-TSL have potency to increase drug efficacy due to higher uptake by tumor and angiogenic endothelial cells in combination with heat-triggered drug release.

No MeSH data available.


Related in: MedlinePlus