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Blood vessel hyperpermeability and pathophysiology in human tumour xenograft models of breast cancer: a comparison of ectopic and orthotopic tumours.

Ho KS, Poon PC, Owen SC, Shoichet MS - BMC Cancer (2012)

Bottom Line: For these results to be meaningful, the hyperpermeable vasculature and reduced lymphatic drainage associated with tumour pathophysiology must be replicated in the model.Dextran accumulation and immunostaining results suggest that small MFP tumours best replicate the vascular permeability required to observe the EPR effect in vivo.A more predictable growth profile and the absence of ulcerated skin lesions further point to the MFP model as a strong choice for long term treatment studies that initiate after a target tumour size has been reached.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Engineering & Applied Chemistry, 200 College Street, Toronto, ON M5S 3E5, Canada.

ABSTRACT

Background: Human tumour xenografts in immune compromised mice are widely used as cancer models because they are easy to reproduce and simple to use in a variety of pre-clinical assessments. Developments in nanomedicine have led to the use of tumour xenografts in testing nanoscale delivery devices, such as nanoparticles and polymer-drug conjugates, for targeting and efficacy via the enhanced permeability and retention (EPR) effect. For these results to be meaningful, the hyperpermeable vasculature and reduced lymphatic drainage associated with tumour pathophysiology must be replicated in the model. In pre-clinical breast cancer xenograft models, cells are commonly introduced via injection either orthotopically (mammary fat pad, MFP) or ectopically (subcutaneous, SC), and the organ environment experienced by the tumour cells has been shown to influence their behaviour.

Methods: To evaluate xenograft models of breast cancer in the context of EPR, both orthotopic MFP and ectopic SC injections of MDA-MB-231-H2N cells were given to NOD scid gamma (NSG) mice. Animals with matched tumours in two size categories were tested by injection of a high molecular weight dextran as a model nanocarrier. Tumours were collected and sectioned to assess dextran accumulation compared to liver tissue as a positive control. To understand the cellular basis of these observations, tumour sections were also immunostained for endothelial cells, basement membranes, pericytes, and lymphatic vessels.

Results: SC tumours required longer development times to become size matched to MFP tumours, and also presented wide size variability and ulcerated skin lesions 6 weeks after cell injection. The 3 week MFP tumour model demonstrated greater dextran accumulation than the size matched 5 week SC tumour model (for P < 0.10). Immunostaining revealed greater vascular density and thinner basement membranes in the MFP tumour model 3 weeks after cell injection. Both the MFP and SC tumours showed evidence of insufficient lymphatic drainage, as many fluid-filled and collagen IV-lined spaces were observed, which likely contain excess interstitial fluid.

Conclusions: Dextran accumulation and immunostaining results suggest that small MFP tumours best replicate the vascular permeability required to observe the EPR effect in vivo. A more predictable growth profile and the absence of ulcerated skin lesions further point to the MFP model as a strong choice for long term treatment studies that initiate after a target tumour size has been reached.

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FITC-Dextran accumulation in tumour tissue normalized to liver tissue control. High molecular weight dextran (2 MDa, ~80 nm) was injected IV into tumour animals as a model nanocarrier and allowed to distribute prior to sacrifice. 3 week old MFP tumours showed higher accumulation of the nanocarrier than 5 week old SC tumours at a 90% confidence interval. All data are shown as the mean of n = 4 animals ± SD. Lines connecting bars denote statistical significance, P < 0.10.
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Figure 2: FITC-Dextran accumulation in tumour tissue normalized to liver tissue control. High molecular weight dextran (2 MDa, ~80 nm) was injected IV into tumour animals as a model nanocarrier and allowed to distribute prior to sacrifice. 3 week old MFP tumours showed higher accumulation of the nanocarrier than 5 week old SC tumours at a 90% confidence interval. All data are shown as the mean of n = 4 animals ± SD. Lines connecting bars denote statistical significance, P < 0.10.

Mentions: Based on fluorescence images of tissue sections, relatively poor dextran uptake was observed in tumour tissue compared to liver tissue across all groups. A threshold was defined to exclude background signal detected in blank tumour and liver tissue and the remaining areas, representing levels above this threshold, were quantified. Less than 1% of the positive signal area observed in the liver control was observed in tumour slices (Figure 2). This can partially be explained by relatively low blood flow through tumour tissue, which has previously been reported to be up to 5-fold lower than in liver [29]. The remaining discrepancy between the dextran accumulation between tumour and liver samples suggests that the model tumour vasculature was less permissive to dextran uptake than the fenestrated liver endothelium, and/or that the lymphatic drainage in the model tumour prevented stable dextran accumulation. Interestingly, dextran uptake in 3 week old MFP tumours was higher than size matched 5 week old SC tumours at 90% confidence (P = 0.08 by one-way ANOVA), suggesting that the orthotopic MFP environment encouraged EPR permissive vasculature and/or lymphovasculature.


Blood vessel hyperpermeability and pathophysiology in human tumour xenograft models of breast cancer: a comparison of ectopic and orthotopic tumours.

Ho KS, Poon PC, Owen SC, Shoichet MS - BMC Cancer (2012)

FITC-Dextran accumulation in tumour tissue normalized to liver tissue control. High molecular weight dextran (2 MDa, ~80 nm) was injected IV into tumour animals as a model nanocarrier and allowed to distribute prior to sacrifice. 3 week old MFP tumours showed higher accumulation of the nanocarrier than 5 week old SC tumours at a 90% confidence interval. All data are shown as the mean of n = 4 animals ± SD. Lines connecting bars denote statistical significance, P < 0.10.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3539979&req=5

Figure 2: FITC-Dextran accumulation in tumour tissue normalized to liver tissue control. High molecular weight dextran (2 MDa, ~80 nm) was injected IV into tumour animals as a model nanocarrier and allowed to distribute prior to sacrifice. 3 week old MFP tumours showed higher accumulation of the nanocarrier than 5 week old SC tumours at a 90% confidence interval. All data are shown as the mean of n = 4 animals ± SD. Lines connecting bars denote statistical significance, P < 0.10.
Mentions: Based on fluorescence images of tissue sections, relatively poor dextran uptake was observed in tumour tissue compared to liver tissue across all groups. A threshold was defined to exclude background signal detected in blank tumour and liver tissue and the remaining areas, representing levels above this threshold, were quantified. Less than 1% of the positive signal area observed in the liver control was observed in tumour slices (Figure 2). This can partially be explained by relatively low blood flow through tumour tissue, which has previously been reported to be up to 5-fold lower than in liver [29]. The remaining discrepancy between the dextran accumulation between tumour and liver samples suggests that the model tumour vasculature was less permissive to dextran uptake than the fenestrated liver endothelium, and/or that the lymphatic drainage in the model tumour prevented stable dextran accumulation. Interestingly, dextran uptake in 3 week old MFP tumours was higher than size matched 5 week old SC tumours at 90% confidence (P = 0.08 by one-way ANOVA), suggesting that the orthotopic MFP environment encouraged EPR permissive vasculature and/or lymphovasculature.

Bottom Line: For these results to be meaningful, the hyperpermeable vasculature and reduced lymphatic drainage associated with tumour pathophysiology must be replicated in the model.Dextran accumulation and immunostaining results suggest that small MFP tumours best replicate the vascular permeability required to observe the EPR effect in vivo.A more predictable growth profile and the absence of ulcerated skin lesions further point to the MFP model as a strong choice for long term treatment studies that initiate after a target tumour size has been reached.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Engineering & Applied Chemistry, 200 College Street, Toronto, ON M5S 3E5, Canada.

ABSTRACT

Background: Human tumour xenografts in immune compromised mice are widely used as cancer models because they are easy to reproduce and simple to use in a variety of pre-clinical assessments. Developments in nanomedicine have led to the use of tumour xenografts in testing nanoscale delivery devices, such as nanoparticles and polymer-drug conjugates, for targeting and efficacy via the enhanced permeability and retention (EPR) effect. For these results to be meaningful, the hyperpermeable vasculature and reduced lymphatic drainage associated with tumour pathophysiology must be replicated in the model. In pre-clinical breast cancer xenograft models, cells are commonly introduced via injection either orthotopically (mammary fat pad, MFP) or ectopically (subcutaneous, SC), and the organ environment experienced by the tumour cells has been shown to influence their behaviour.

Methods: To evaluate xenograft models of breast cancer in the context of EPR, both orthotopic MFP and ectopic SC injections of MDA-MB-231-H2N cells were given to NOD scid gamma (NSG) mice. Animals with matched tumours in two size categories were tested by injection of a high molecular weight dextran as a model nanocarrier. Tumours were collected and sectioned to assess dextran accumulation compared to liver tissue as a positive control. To understand the cellular basis of these observations, tumour sections were also immunostained for endothelial cells, basement membranes, pericytes, and lymphatic vessels.

Results: SC tumours required longer development times to become size matched to MFP tumours, and also presented wide size variability and ulcerated skin lesions 6 weeks after cell injection. The 3 week MFP tumour model demonstrated greater dextran accumulation than the size matched 5 week SC tumour model (for P < 0.10). Immunostaining revealed greater vascular density and thinner basement membranes in the MFP tumour model 3 weeks after cell injection. Both the MFP and SC tumours showed evidence of insufficient lymphatic drainage, as many fluid-filled and collagen IV-lined spaces were observed, which likely contain excess interstitial fluid.

Conclusions: Dextran accumulation and immunostaining results suggest that small MFP tumours best replicate the vascular permeability required to observe the EPR effect in vivo. A more predictable growth profile and the absence of ulcerated skin lesions further point to the MFP model as a strong choice for long term treatment studies that initiate after a target tumour size has been reached.

Show MeSH
Related in: MedlinePlus