murine breast cancer cell line 4t1  (Procell Inc)

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Figure Lengend Snippet: Characterization of mouse tumor model using bioluminescence imaging, flow cytometry and histology. (A) Representative BLI images show an orthotopic 4T1 tumor in mice and its progression overtime. After surgical resection of the primary tumor 14 days after inoculation, metastatic outgrowth and recurrence is observed in the thoracic and abdominal regions. Quantification of the BLI signal from (B) the primary tumor and (C) thoracic region is shown, respectively (n=5). The 4T1 cancer cells (GFP reporter) and CD45+ leukocytes were quantified using flow cytometry in the (D) primary tumor, (E) liver and (F) lungs. (G) Histological evaluation depicts the progression of metastatic disease in the liver and lungs (20x magnification; scale bar = 100 μm; blue: nuclear stain; green:4T1 cancer cells). Mean ± SEM are plotted with statistics by one-/two-way ANOVA with a post hoc Tukey or Sidak’s test. **P<0.01; ***P<0.001; ****P<0.0001.

Article Snippet: The 4T1 model was established in BALB/c mice (Jackson Laboratories).

Techniques: Imaging, Flow Cytometry, Staining

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Figure Lengend Snippet: Quantification of nanoparticle uptake by APCs across different TMEs. (A) Timelines show animal modeling, nanoparticle administration and flow cytometry analysis. (B) Flow cytometry analysis of nanoparticle uptake by 4T1 cancer cells and CD45 + leukocytes in primary tumor. Flow cytometry analysis of nanoparticle uptake by CD11c + dendritic cells and F4/80 + macrophages in (C) primary tumor, (D) lungs at a stage of early metastasis, (E) liver at a stage of early metastasis, (F) lungs at a stage of late metastasis, and (G) liver at a stage of late metastasis (n=5 mice per group). Box and whisker plots (5–95 percentile) with statistics by one-/two-way ANOVA with a post hoc Tukey or Sidak’s test. *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001.

Article Snippet: The 4T1 model was established in BALB/c mice (Jackson Laboratories).

Techniques: Flow Cytometry, Whisker Assay

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Figure Lengend Snippet: Response of good responders to a tumor rechallenge. The treatment scheme of animals bearing 4T1 tumors included surgical resection of the primary tumor, anti-PD1 and untargeted immunostimulatory NP before and after surgery, indicated as mPEG-NP(x2), or only before surgery indicated as mPEG-NP(x1). The good responders from the two groups were rechallenged 42 days after the initial inoculation with 1×10 5 4T1 cells on their right flank. (A) Caliper measurement of the flank tumor size. (B) Kaplan-Meier survival analysis. N=4 mice per condition. Box and whisker plots (5–95 percentile) both with statistics by one-/two-way ANOVA with a post hoc Tukey or Sidak’s test. **P<0.01.

Article Snippet: The 4T1 model was established in BALB/c mice (Jackson Laboratories).

Techniques: Whisker Assay

Journal: Molecular Therapy Oncology

Article Title: Fibulin-4 expressed in metastatic breast cancer is a target of peptide-based imaging probes and experimental therapeutics

doi: 10.1016/j.omton.2026.201207

Figure Lengend Snippet: BLMP6 homes to human cancer metastases (A) Lung paraffin sections from mice without or with 4T1 metastases overlayed with AZDye555-BLMP6 (12.5 μg/mL) reveal binding specificity to metastatic cells. Endomucin IF identifies endothelial cells. Graph: data quantification, representing the average Az555 signal binding across analyzed fields ( n = 3). (B) Lung paraffin section from a mouse with MDA-MB-231 metastases overlayed with AZDye555-BLMP6 (12.5 μg/mL) reveals binding to metastatic cells. N-cadherin IF identifies AZDye555-BLMP6-bound cancer cells that underwent EMT (orange arrows) and stromal cells (green arrows). (C) In a mouse pre-grafted with GFP-expressing MDA-MB-231 metastatic cells, upon i.v. injection of AZDye555-BLMP6 (50 μg), a lung section shows a GFP-positive metastatic lesion with AZDye555-BLMP6 signal, absent in the surrounding non-tumor tissue. (D) Radiolabeled 68 Ga-BLMP6 i.v.-injected into nude mice without and with MDA-MB-231 lung metastases detected by IVIS. At 1 h, PET/CT images are scaled equally (0.4%–1.96% ID/g) to allow cross-reference throughout the study. Arrow, 68 Ga-BLMP6 in metastases; b, bladder; k, kidney. (E) Biodistribution in tissues resected from mice without and with metastases in (D), with gamma counter plotted as percentage of the injected dose per gram tissue (%ID/g tissue). For graphs, shown is mean ± SEM; ∗ p < 0.05, Student t test. Scale bars, 100 μm.

Article Snippet: Spontaneous cancer progression to metastases was modeled by injecting 2 × 10 4 4T1 cells into the mammary fat pad of 10-weeks-old female Balb/c mice (Jackson Laboratory) with a 31-gauge needle.

Techniques: Binding Assay, Paraffin Section, Expressing, Injection, Positron Emission Tomography-Computed Tomography

Journal: Molecular Therapy Oncology

Article Title: Fibulin-4 expressed in metastatic breast cancer is a target of peptide-based imaging probes and experimental therapeutics

doi: 10.1016/j.omton.2026.201207

Figure Lengend Snippet: MMAE-BLMP6 kills metastatic cells (A) Survival of 4T1 and 4T1-FBLN4-KO cells treated with indicated concentrations of MMAE-BLMP6 for 24 h measured by CellTiter-Blue cell assay. Images: trypan blue staining of 4T1 cells, untreated or treated with 1 μM MMAE-BLMP6. (B) Survival of MDA-MB-231 cells treated with indicated concentrations of MMAE-BLMP6 and MMAE-BLMP5 peptides for 24 h measured by CellTiter-Blue cell assay. Images: trypan blue staining of MDA-MB-231 cells, untreated or treated with 1 μM MMAE-BLMP6. (C) Lung paraffin section from mice with B16F10 metastases injected (sc) with 20 μg of MMAE-BLMP6 and sacrificed 24 h later. TUNEL assay identifies apoptosis in metastatic cells (arrows). (D) Lung paraffin section from mice with MDA-MB-231-luc-GFP metastases injected with MMAE-BLMP6. IF with an antibody against cleaved caspase 3 identifies apoptosis in metastatic cells (green arrows). IB4, isolectin B4 (Cy3-conjugated) binding marking the endothelium. Scale bars, 100 μm.

Article Snippet: Spontaneous cancer progression to metastases was modeled by injecting 2 × 10 4 4T1 cells into the mammary fat pad of 10-weeks-old female Balb/c mice (Jackson Laboratory) with a 31-gauge needle.

Techniques: Staining, Paraffin Section, Injection, TUNEL Assay, Binding Assay

Journal: Molecular Therapy Oncology

Article Title: Fibulin-4 expressed in metastatic breast cancer is a target of peptide-based imaging probes and experimental therapeutics

doi: 10.1016/j.omton.2026.201207

Figure Lengend Snippet: MMAE-BLMP6 suppresses metastasis progression (A) Kaplan-Meier curves showing cumulative survival of C57BL/6 mice i.v.-grafted with B16F10-luc cells and subcutaneously (sc)-injected with MMAE-BLMP6 or PBS as indicated. (B) Bioluminescence analysis of mice from (A) by IVIS revealing metastasis suppression in MMAE-BLMP6-treated mice (days 15 vs. 5). (C) Kaplan-Meier curves showing cumulative survival of Nude-Foxn1nu mice i.v.-grafted with MDA-MB-231-luc-GFP cells and sc-injected with MMAE-BLMP6 or PBS as indicated. (D) Bioluminescence analysis of mice from (D) by IVIS revealing metastasis suppression in MMAE-BLMP6-treated mice (days 21 vs. 12). (E) Mammary tumor growth in BALB/c mice bilaterally grafted with 4T1-luc cells and sc-injected with MMAE-BLMP6 ( N = 8) or scrambled MMAE-BLMP6 ( N = 7) as indicated in (G). Plotted are mean ± SEM; ∗ p < 0.01, Student t test. (F) Bioluminescence analysis of representative mice from (E) by IVIS at week 3 reveals smaller primary tumors (PT) and metastases (M) in mice treated with MMAE-BLMP6. (G) Kaplan-Meier curves showing survival of BALB/c mice orthotopically grafted with 4T1-luc cells and sc-injected with MMAE-BLMP6 or scrambled MMAE-BLMP6 as indicated. (H) Bioluminescence analysis of mice from (F and G) by IVIS revealing metastasis growth suppression in MMAE-BLMP6-treated mice (days 33 vs. 26). (I) Representative lungs from mice that died on the same day showing smaller metastases (arrowheads) in animals treated with MMAE-BLMP6.

Article Snippet: Spontaneous cancer progression to metastases was modeled by injecting 2 × 10 4 4T1 cells into the mammary fat pad of 10-weeks-old female Balb/c mice (Jackson Laboratory) with a 31-gauge needle.

Techniques: Injection

Journal: Molecular Therapy Oncology

Article Title: Fibulin-4 expressed in metastatic breast cancer is a target of peptide-based imaging probes and experimental therapeutics

doi: 10.1016/j.omton.2026.201207

Figure Lengend Snippet: BLMP6 mimics LTBP4 and binds to FBLN4 (A) AlphaFold3 (AF3) modeling of the 3D structure of BLMP6 and mouse LTBP4L (residues 716–733 shown) containing the BLMP6 similarity segment (725–730, highlighted). (B) Left, consensus docking pose predicted by AF3 and CB-Dock shows that BLMP6 interacts with the C terminus of mouse FBLN4 (residues 41–443 shown). Right, a similar binding predicted for a 106 amino acid fragment of LTBP4L (residues 675–780) containing the BLMP6 similarity segment (725–730, highlighted) using AF3 and ClusPro. (C) Biolayer interferometry assay performed with Octet to measure binding affinities. (D) Affinity purification of FBLN4 from 4T1 cell extract with biotinylated BLMP6 immobilized on streptavidin-coated beads. Immunoblotting with anti-FBLN4 antibodies identifies the expected 49 kDa protein, which is not isolated on control beads. (E) Graph: real-time PCR analysis of gene messenger RNA (mRNA) expression, relative to 18S , confirming the knockout of FBLN4 in 4T1 cells. Images: lung paraffin sections from mice that were injected with phage displaying BLMP6. Anti-phage IF demonstrates BLMP6-phage homing to 4T1 metastases but not to 4T1-FBLN4-KO metastases. Scale bars, 100 μm.

Article Snippet: Spontaneous cancer progression to metastases was modeled by injecting 2 × 10 4 4T1 cells into the mammary fat pad of 10-weeks-old female Balb/c mice (Jackson Laboratory) with a 31-gauge needle.

Techniques: Binding Assay, Affinity Purification, Western Blot, Isolation, Control, Real-time Polymerase Chain Reaction, Expressing, Knock-Out, Injection