Journal: Cancers

Article Title: Harnessing Induced Essentiality: Targeting Carbonic Anhydrase IX and Angiogenesis Reduces Lung Metastasis of Triple Negative Breast Cancer Xenografts

doi: 10.3390/cancers11071002

Figure Lengend Snippet: Sunitinib induces hypoxia and Carbonic Anhydrase IX (CAIX) expression in primary Triple Negative Breast Cancer (TNBC) tumors. ( a ) Images of MDA-MB-231 LM2-4Luc + primary tumor tissue sections harvested at increasing tumor volumes from animals administered either vehicle or 60 mg/kg sunitinib and immunohistochemically stained for the indicated markers. Scale bars: upper panels, 1 mm; lower panels, 200 μm. ( b to e ) Image-based quantification of ( b ) CD31 + blood vessels, ( c ) CAIX expression, ( d ) pimonidazole and ( e ) proliferation. Data show the mean ± standard error of the mean (SEM). n = 3 animals/group with 10 images/animal. * p < 0.05, *** p < 0.001.

Article Snippet: Formalin-fixed, paraffin-embedded (FFPE) tissues were sectioned (4–5 μm) and stained with primary antibodies against human CAIX (1:50, AF2188; R&D Systems, Minneapolis, MN, USA), CD31 (1:10, DIA-310; Histobiotec, Miami Beach, FL, USA), BrdU (1:10, Roche, Laval, QC, Canada) and pimonidazole (1:100, Hydroxyprobe, Burlington, MA, USA).

Techniques: Expressing, Staining

Journal: Cancers

Article Title: Harnessing Induced Essentiality: Targeting Carbonic Anhydrase IX and Angiogenesis Reduces Lung Metastasis of Triple Negative Breast Cancer Xenografts

doi: 10.3390/cancers11071002

Figure Lengend Snippet: SLC-0111 reduces vessel density and vascular permeability in primary tumors, and decreases lung and liver metastases. ( a ) Representative 3D maximum projections of whole mount primary tumor tissue slices showing CD31 + blood vessels (green) and fluorescently labeled dextran (red). Merged images demonstrate the presence of both intravascular (yellow) and extravasated (arrowheads) dextran. Scale bar = 100 μm. ( b ) Quantification of the number of vessels in whole mount primary tumor slices. Data show the mean ± standard error of the mean (SEM). n = 13–14 images/group. ** p < 0.01, *** p < 0.001. ( c ) Quantification of vascular permeability as assessed by relative area of extravasated dextran in whole mount primary tumor slices. Data show the mean ± SEM. n = 13–16 images/group. * p < 0.05, ** p < 0.01. ( d ) Representative 3D maximum projections of whole mount primary tumor tissue slices showing levels of CAIX expression (green) and the number of CD31 + blood vessels (red). Lower panels, merge. Scale bar = 100 μm. ( e ) Quantification of Carbonic Anhydrase IX (CAIX) expression in whole mount primary tumor tissue slices in panel d. Data show the mean ± SEM. n = 11–15 images/group. * p < 0.05, *** p < 0.001. ( f ) Representative images of lung and liver tissues from animals with MDA-MB-231 LM2-4Luc + orthotopic breast tumors and administered SLC-0111 and sunitinib, either alone or in combination. Visible metastatic nodules (arrows) are indicated. Scale bar = 1 cm. ( g – i ) Analysis of ( g ) lung weight ( n = 9/group), ( h ) liver weight as a function of whole animal body weight ( n = 9/group) and ( i ) number of metastatic nodules present on the liver surface ( n = 7–8/group). For each graph, data show the mean ± SEM. * p < 0.05.

Article Snippet: Formalin-fixed, paraffin-embedded (FFPE) tissues were sectioned (4–5 μm) and stained with primary antibodies against human CAIX (1:50, AF2188; R&D Systems, Minneapolis, MN, USA), CD31 (1:10, DIA-310; Histobiotec, Miami Beach, FL, USA), BrdU (1:10, Roche, Laval, QC, Canada) and pimonidazole (1:100, Hydroxyprobe, Burlington, MA, USA).

Techniques: Permeability, Labeling, Expressing

Journal: Cancers

Article Title: Harnessing Induced Essentiality: Targeting Carbonic Anhydrase IX and Angiogenesis Reduces Lung Metastasis of Triple Negative Breast Cancer Xenografts

doi: 10.3390/cancers11071002

Figure Lengend Snippet: Administration of SLC-0111 reduces lung metastases. ( a ) Representative 3D maximum projections of whole mount lung tissue slices from tumor-bearing mice, showing vimentin-positive metastases (cyan) and CD31 + blood vessels (red). Scale bar = 100 μm. ( b ) Analysis of vimentin positive metastases in whole mount lung tissue sections described in panel a. Data show the mean ± standard error of the mean (SEM). n = 8–10 images/group. * p < 0.05, *** p < 0.001. ( c ) Representative 3D maximum projections of whole mount lung tissue slices from tumor-bearing mice, showing Carbonic Anhydrase IX (CAIX)-positive metastases (green) and CD31 + blood vessels (red). Bar = 100 μm. ( d ) Analysis of CAIX positive metastases in the whole mount lung tissue sections described in panel c. Data show the mean ± SEM. n = 5–8 images/group: No statistically significant differences were observed among the groups.

Article Snippet: Formalin-fixed, paraffin-embedded (FFPE) tissues were sectioned (4–5 μm) and stained with primary antibodies against human CAIX (1:50, AF2188; R&D Systems, Minneapolis, MN, USA), CD31 (1:10, DIA-310; Histobiotec, Miami Beach, FL, USA), BrdU (1:10, Roche, Laval, QC, Canada) and pimonidazole (1:100, Hydroxyprobe, Burlington, MA, USA).

Techniques:

Journal: Cancers

Article Title: Harnessing Induced Essentiality: Targeting Carbonic Anhydrase IX and Angiogenesis Reduces Lung Metastasis of Triple Negative Breast Cancer Xenografts

doi: 10.3390/cancers11071002

Figure Lengend Snippet: Model for targeting angiogenesis and Carbonic Anhydrase IX (CAIX) to reduce metastasis of Triple Negative Breast Cancer (TNBC). Exposure of tumors to anti-angiogenic agents such as sunitinib leads to reduced tumor growth. However, major consequences can include enhanced hypoxia and increased vascular permeability. The exacerbation of hypoxia results in the upregulation of CAIX expression by tumor cells and the potentiation of metastasis. Inhibition of CAIX activity inhibits metastasis through several mechanisms, as shown by previous studies, but may also play a role in reducing vascular permeability and contributing to vascular normalization, potentially reducing the invasion of tumor cells to distant sites.

Article Snippet: Formalin-fixed, paraffin-embedded (FFPE) tissues were sectioned (4–5 μm) and stained with primary antibodies against human CAIX (1:50, AF2188; R&D Systems, Minneapolis, MN, USA), CD31 (1:10, DIA-310; Histobiotec, Miami Beach, FL, USA), BrdU (1:10, Roche, Laval, QC, Canada) and pimonidazole (1:100, Hydroxyprobe, Burlington, MA, USA).

Techniques: Permeability, Expressing, Inhibition, Activity Assay