Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Sep 8:6:8094.
doi: 10.1038/ncomms9094.

In vivo capture and label-free detection of early metastatic cells

Samira M Azarin  1 , Ji Yi  2 , Robert M Gower  3 , Brian A Aguado  2 , Megan E Sullivan  4 , Ashley G Goodman  5 , Eric J Jiang  5 , Shreyas S Rao  5 , Yinying Ren  5 , Susan L Tucker  6 , Vadim Backman  2   7   8 , Jacqueline S Jeruss  9   10 , Lonnie D Shea  5   11   12   13
Affiliations

In vivo capture and label-free detection of early metastatic cells

Samira M Azarin et al. Nat Commun. .

Abstract

Breast cancer is a leading cause of death for women, with mortality resulting from metastasis. Metastases are often detected once tumour cells affect the function of solid organs, with a high disease burden limiting effective treatment. Here we report a method for the early detection of metastasis using an implanted scaffold to recruit and capture metastatic cells in vivo, which achieves high cell densities and reduces the tumour burden within solid organs 10-fold. Recruitment is associated with infiltration of immune cells, which include Gr1(hi)CD11b(+) cells. We identify metastatic cells in the scaffold through a label-free detection system using inverse spectroscopic optical coherence tomography, which identifies changes to nanoscale tissue architecture associated with the presence of tumour cells. For patients at risk of recurrence, scaffold implantation following completion of primary therapy has the potential to identify metastatic disease at the earliest stage, enabling initiation of therapy while the disease burden is low.

PubMed Disclaimer

Conflict of interest statement

Competing financial interests

L.D.S. is an occasional consultant for Pioneer Biosolutions, which licenses some of the technology developed in his lab and used in this research. The remaining authors declare no competing financial interests.

Figures

Figure 1
Figure 1. PLG scaffolds recruit metastatic tumor cells
Tissues were isolated at day 28 post-tumor inoculation (21 days after scaffold implantation or mock surgery). (a,b) Bioluminescence imaging (BLI) of peritoneal fat pads receiving scaffold implants (a) or mock surgeries (b). (c–e) Hematoxylin and eosin (H&E) staining of the primary tumor (c), a fat pad containing a scaffold (white circle indicates metastatic cluster) (d), and a fat pad without a scaffold (e). Scale bars indicate 100 μm.
Figure 2
Figure 2. Recruitment of tumor cells to scaffolds reduces tumor burden in lung
(a) Flow cytometric analysis of the percentage of tdTomato-positive tumor cells in cells isolated from lungs at day 28 post-tumor inoculation. Data shown as mean ± SEM (n = 8, 2 independent replicates). * P < 0.01 compared to mock surgery (Mann-Whitney test). (b) H&E staining of lung section (which circles indicate metastatic clusters). Scale bar indicates 200 μm. (c) Histological analysis of H&E-stained lung sections to determine the number of tumor clusters per section. Data shown as mean ± SEM (n = 12, 2 independent replicates). * P < 0.05 compared to mock surgery (Mann-Whitney test).
Figure 3
Figure 3. Early detection of tumor cells in scaffolds
Flow cytometric analysis of tdTomato-positive tumor cells in tissues isolated from mice at day 14 and 28 post-tumor inoculation. (a) Number of mice with tumor cells detectable in each tissue in a group of 8 mice at day 14 or 28 post-tumor inoculation. Each mouse received two intraperitoneal (IP) scaffolds. P values from Fisher’s exact test. (b) Percentage of tdTomato-positive cells in the total cell population isolated from IP scaffolds and subcutaneous (SQ) scaffolds at day 14 post-tumor inoculation. Data shown as mean ± SEM (n = 16 for IP scaffold, n = 10 for SQ scaffold, 2 independent replicates). * P < 0.05 compared to IP scaffold (Mann-Whitney test).
Figure 4
Figure 4. Detection of tumor cells in scaffold using IS-OCT
(a,b) Representative 3-D maps of D generated from in situ IS-OCT analysis of subcutaneous scaffolds implanted in tumor-free (a) and tumor-bearing (b) mice at day 14 post-tumor inoculation. Scale bars indicate 200 μm. (c) Average D value for subcutaneous scaffolds in tumor-free (“No Tumor”) and tumor-bearing (“Tumor”) mice. Data shown as mean ± SEM (n = 6, 2 independent replicated). * P < 0.05 compared to tumor-free mice (Mann-Whitney test).
Figure 5
Figure 5. Evaluation of the immune environment within scaffolds
(a,b) CD-45 immunolabeling (green) at day 28 post-tumor inoculation of fat pads receiving mock surgery (a) or scaffold implant (b). Nuclei are blue. Scale bar indicates 100 μm. (c,d) Number of migrating 231BR (c) or 4T1 (d) cells in the presence of splenocyte-conditioned media (SCM) or non-conditioned media (non-SCM). Data shown as mean ± SEM (n = 6, 2 independent replicates). * P < 0.05 and ** P < 0.005 compared to non-conditioned media (Mann-Whitney test). (e–h) Flow cytometric analysis of cells removed from lungs (e,f) and scaffolds (g,h) of tumor-free and tumor-bearing (Day 14 and Day 28) NSG (e,g) or BALB/c (f,h) mice. The model used for mice with tumors involved the inoculation of cells at day 0, with scaffolds implanted at day 7. The evaluation of scaffolds in tumor free mice was performed following day 7 post-implantation. Cell populations are reported as percentage of total CD45-positive leukocytes. Data shown as mean ± SEM (n = 5 for lungs, n = 10 for scaffolds, 2 independent replicates). * P < 0.05 and ** P < 0.005 compared to Day 0 (e,f) or Day 14 (g,h) (Mann-Whitney test).
Figure 6
Figure 6. Immunomodulation of the scaffold microenvironment influences recruitment of tumor cells
Flow cytometric analysis at day 14 post-tumor inoculation of cells removed from scaffolds containing a CCL22 or β-galactosidase (control) vector implanted in NSG (a,b) or BALB/c (c,d) mice. Cell populations are reported as percentage of total CD45-positive leukocytes (a,c) or total number of tumor cells in the scaffold (b,d). Data shown as mean ± SEM (n = 10, 2 independent replicates). * P < 0.05, ** P < 0.005 and *** P < 0.001 compared to β-galactosidase scaffolds (Mann-Whitney test). (e) Number of migrating 231BR cells in the presence of Gr1hiCD11b+ cell-conditioned media (Gr1hiCD11b+-CM) or non-conditioned media (non-SCM Data shown as mean ± SEM (n = 6, 2 independent replicates). *** P < 0.001 compared to non-conditioned media (Mann-Whitney test). (f,g) Flow cytometric analysis at day 14 post-tumor inoculation of cells removed from blank scaffolds or scaffolds seeded with Gr1hiCD11b+ cells. Cell populations are reported as the percentage of Gr1hiCD11b+ cells in the total leukocyte population (f) or total number of tumor cells in the scaffold (g). Data shown as mean ± SEM (n = 10, 2 independent replicates). * P < 0.05 compared to blank scaffold (Mann-Whitney test).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Griffith OL, Gray JW. ‘Omic approaches to preventing or managing metastatic breast cancer. Breast Cancer Res. 2011;13:230. - PMC - PubMed
    1. Nagrath S, et al. Isolation of rare circulating tumour cells in cancer patients by microchip technology. Nature. 2007;450:1235–1239. - PMC - PubMed
    1. Stott SL, et al. Isolation of circulating tumor cells using a microvortex-generating herringbone-chip. Proc Natl Acad Sci USA. 2010;107:18392–18397. - PMC - PubMed
    1. Yoon HJ, et al. Sensitive capture of circulating tumour cells by functionalized graphene oxide nanosheets. Nat Nanotechnol. 2013;8:735–741. - PMC - PubMed
    1. Bednarz-Knoll N, Alix-Panabieres C, Pantel K. Clinical relevance and biology of circulating tumor cells. Breast Cancer Res. 2011;13:228. - PMC - PubMed

Publication types

MeSH terms

Substances