Heterogeneity of stromal fibroblasts in tumors.

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e1 Cancer Biology & Therapy 2007; Vol. 6 Issue 4
Commentary
Heterogeneity of Stromal Fibroblasts in Tumors
Akira Orimo1
Robert A. Weinberg1,2
1Whitehead Institute for Biomedical Research; 2Department of Biology,
Massachusetts Institute of Technology; Cambridge, Massachusetts USA
*Correspondence to: Robert A. Weinberg; Whitehead Institute for Biomedical
Research; Cambridge, Massachusetts USA; Tel.: 617.258.5159; Fax: 617.258.5213;
Email: weinberg@wi.mit.edu
Original manuscript submitted: 04/05/07
Manuscript accepted: 04/10/07
This manuscript has been published online, prior to printing for Cancer Biology &
Therapy, Volume 6, Issue 4. Definitive page numbers have not been assigned. The
current citation is: Cancer Biol Ther 2007; 6(4):
http://www.landesbioscience.com/journals/cc/abstract.php?id=4255
Once the issue is complete and page numbers have been assigned, the citation
will change accordingly.
Key WORds
fibroblast heterogeneity, CAF, cancer,
microenvironment, NG2, SMA, FSP1,
S100A4, PDGFR
Commentary to:
Identification of Fibroblast Heterogeneity in the Tumor
Microenvironment
Hikaru Sugimoto, Thomas M Mundel, Mark W.
Kieran and Raghu Kalluri
Cancer Biol Ther 2006; 5(12):1640 - 6
It is now widely accepted that stromal fibroblasts present within a tumor play signifi-
cant contributions to carcinoma growth.1-4 However, the biological properties of these
fibroblasts are still incompletely understood. In the December, 2006 issue of Cancer
Biology & Therapy, Sugimoto and colleagues5 indicate the existence of a heterogenous cell
population of stromal fibroblasts in tumors based on immunohistological observation
using various fibroblastic makers. We discuss the implications of these findings here.
Fibroblasts are a major stromal cell type that is usually present within human epithelial
carcinomas. The dominant contribution of these fibroblasts to the stroma has encouraged
scientists to study the roles of stromal fibroblasts during tumor progression. For example,
several research groups have determined whether stromal fibroblasts act to promote tumor
growth and progression. Indeed, these studies demonstrated that stromal fibroblasts,
specifically termed carcinoma-associated fibroblasts (CAFs), that have been extracted
from human carcinomas can promote the growth of admixed epithelial carcinoma cells
in immunodeficient mice, doing so far more potently than control fibroblasts extracted
from non-cancerous tissues. Moreover, such CAFs showed an ability to inhibit cancer cell
apoptosis, induce cancer cell proliferation, and stimulate tumor angiogenesis.6,7
Alpha-smooth muscle actin (α-SMA)-positive myofibroblasts have long been recog-
nized as a prominent component of the activated fibroblasts present both in tumors and in
sites of tissue injury.8 Various growth factors, cytokines, and extra cellular matrix (ECM)
proteins produced by myofibroblasts play pivotal roles in tissue repair by stimulating
growth of nearby epithelial cells and facilitating angiogenesis.7,9 Such myofibroblasts
secrete stromal cell-derived factor 1 (SDF-1), which is an angiogenic chemokine, in order
to promote tumor growth and stimulate neoangiogenesis.7,10
In addition to α-SMA, various other markers have been used to detect stromal
fibroblasts in tumors. Included among these are vimentin, fibroblast-specific protein-1
(FSP-1/S100A4),11 NG2 (Neuron-Glial Antigen-2), chondroitin sulfate proteoglycan,5
PDGFR-b,5 fibroblast-activation protein (FAP),12 fibroblast-associated antigen,13 prolyl
4-hydroxylase14 (Fig. 1). Of these, α-SMA and FAP12 are especially useful makers to
resolve myofibroblasts in tumor stroma from stromal fibroblasts, whereas several of the
other markers can be used to reveal both the myofibroblasts and fibroblasts that together
form CAF populations (Fig. 1).
Sugimoto and colleagues5 have now immunostained sections prepared from exper-
imentally generated tumors in mouse and found unique FSP-1-expressing CAF
subpopulations distinct from the α-SMA+ myofibroblasts present in tumor stroma
(Fig. 1). Recently, it was shown that fibroblast-produced FSP-1 promotes tumor metastasis
and that mammary carcinoma cells injected into FSP-1-/- mice showed significant delay
in tumor uptake and decreased tumor incidence15. Interestingly, coinjection of carcinoma
cells with FSP-1+/+ mouse embryonic fibroblasts (MEFs) partially restored the kinetics of
tumor development and the ability to form metastases.15 This suggests that it would be
of interest to determine whether FSP-1-expressing CAF populations can promote tumor
progression compared to those of FSP-1-negative CAF populations.
Another example of the heterogeneity of stromal fibroblasts is provided by a subpopu-
lation of SA-b-gal-positive, senescent fibroblast populations that are present among the
stromal fibroblasts in human ovarian carcinomas; these senescent fibroblasts play pivotal
roles in enhancing carcinogenesis through paracrine signaling mechanisms.16,17 In yet
other studies, a mouse prostate tumor model indicates that carcinoma cells prefentially
induce the clonal expansion of a small fraction of fibroblasts that lack p53, eventually
generating the majority stromal fibroblast population within a tumor;18 this absence of
p53 allowed stromal fibroblasts to generate highly proliferative fibroblasts, resembling
CAFs, that further promote tumor progression. Supporting this line of observation, muta-
tions of the p53 gene were indeed found in stromal regions microdisected from human
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Heterogeneity of Stromal Fibroblasts in Tumor
breast carcinomas.19 Further work will elucidate molecular mecha-
nisms by which loss of p53 in stromal fibroblasts influences nearby
epithelial carcinoma growth.
Stromal fibroblasts within tumors seem to be originally derived
from heterogenous cell types that include bone marrow-derived
progenitors, smooth muscle cells, preadipocytes, fibroblasts, and
myofibroblasts. Sugimoto and colleagues’ observation may reflect
the various distinct cells-of-origin that have been responsible for
generating stromal fibroblasts. Alternatively, a particular fraction
of stromal fibroblasts might be selected or transdifferentiated into
FSP-1-expressing cell populations during tumor progression. Ideally,
future studies should address whether these FSP-1+CAFs exist in
human carcinomas in which they provide a tumor-promoting frac-
tion.
Because stromal fibroblasts are genetically more stable than rapidly
mutating tumor cell populations, tumor immunotherapy specifically
targeting these fibroblasts expressing FAP-1 has been attempted and
has been found to successfully attenuate tumor progression in mouse
models20, 21 In a larger sense, research on stromal fibroblasts will help
us promote understanding of the complex tumor-stroma cell inter-
actions in tumor-prone tissue microenvironments and will provide
clues for the development of new types of anti-tumor therapeutics.
References
1. Bissell MJ, Radisky D. Putting tumours in context. Nat Rev Cancer 2001; 1: 46-54.
2. Mueller MM, Fusenig NE. Friends or foes - bipolar effects of the tumour stroma in cancer.
Nat Rev Cancer 20 4; 4: 839-849.
3. Bhowmick NA, Neilson EG, Moses HL. Stromal fibroblasts in cancer initiation and pro-
gression. Nature 2004; 432: 332-337.
4. Kalluri R, Zeisberg M. Fibroblasts in cancer. Nat Rev Cancer 2006; 6: 392-401.
5. Sugimoto H, Mundel TM, Kieran MW, Kalluri R. Identification of Fibroblast Heterogeneity
in the Tumor Microenvironment. Cancer Biol Ther 2006; 5:1640-6.
6. Olumi AF, Grossfeld GD, Hayward SW, Carroll PR, Tlsty TD, Cunha GR. Carcinoma-
associated fibroblasts direct tumor progression of initiated human prostatic epithelium.
Cancer Res 1999; 59:5002-11.
7. Orimo A, Gupta PB, Sgroi DC, Arenzana-Seisdedos F, Delaunay T, Naeem R, Carey VJ,
Richardson AL, Weinberg RA. Stromal Fibroblasts Present in Invasive Human Breast
Carcinomas Promote Tumor Growth and Angiogenesis through Elevated SDF-1/CXCL12
Secretion. Cell 2005; 121:335-48.
8. Serini G, Gabbiani G. Mechanisms of myofibroblast activity and phenotypic modulation.
Exp Cell Res 1999; 250:273-83.
9. Ronnov-Jessen L, Van Deurs B, Nielsen M, Petersen OW. Identification, paracrine genera-
tion, and possible function of human breast carcinoma myofibroblasts in culture. In Vitro
Cell Dev Biol 1992; 28A:273-83.
10. Allinen M, Beroukhim R, Cai L, Brennan C, Lahti-Domenici J, Huang H, Porter D, Hu
M, Chin L, Richardson A, Schnitt S, Sellers WR, Polyak K. Molecular characterization of
the tumor microenvironment in breast cancer. Cancer Cell 2004; 6:17-32.
11. Strutz F, Okada H, Lo CW, Danoff T, Carone RL, Tomaszewski JE, Neilson EG.
Identification and characterization of a fibroblast marker: FSP1. J Cell Biol 1995; 130:393-
405.
12. Park JE, Lenter MC, Zimmermann RN, Garin-Chesa P, Old LJ, Rettig WJ. Fibroblast acti-
vation protein, a dual specificity serine protease expressed in reactive human tumor stromal
fibroblasts. J Biol Chem 1999; 274:36505-12.
13. Ronnov-Jessen L, Celis JE, Van Deurs B, Petersen OW. A fibroblast-associated antigen:
characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stro-
mal cells. J Histochem Cytochem 1992; 40:475-86.
14. Konttinen YT, Nykanen P, Nordstrom D, Saari H, Sandelin J, Santavirta S, Kouri T. DNA
synthesis in prolyl 4-hydroxylase positive fibroblasts in situ in synovial tissue. An autoradi-
ography-immunoperoxidase double labeling study. J Rheumatol 1989; 16:339-45.
15. Grum-Schwensen B, Klingelhofer J, Berg CH, El-Naaman C, Grigorian M, Lukanidin E.
& Ambartsumian, N. Suppression of tumor development and metastasis formation in mice
lacking the S100A4(mts1) gene. Cancer Res 2005; 65: 3772-3780.
16. Krtolica A, Parrinello S, Lockett S, Desprez PY, Campisi J. Senescent fibroblasts promote
epithelial cell growth and tumorigenesis: a link between cancer and aging. Proc Natl Acad
Sci USA 2001; 98:12072-7.
17. Yang G, Rosen DG, Zhang Z, Bast RC, Jr., Mills GB, Colacino JA, Mercado-Uribe I, Liu,
J. The chemokine growth-regulated oncogene 1 (Gro-1) links RAS signaling to the senes-
cence of stromal fibroblasts and ovarian tumorigenesis. Proc Natl Acad Sci USA 2006; 103:
16472-7.
18. Hill R, Song Y, Cardiff RD, Van Dyke T. Selective evolution of stromal mesenchyme with
p53 loss in response to epithelial tumorigenesis. Cell 2005; 123:1001-11.
19. Kurose K, Gilley K, Matsumoto S, Watson PH, Zhou XP, Eng C. Frequent somatic muta-
tions in PTEN and TP53 are mutually exclusive in the stroma of breast carcinomas. Nat
Genet 2002; 32:355-7.
20. Loeffler M, Kruger JA, Niethammer AG, Reisfeld RA. Targeting tumor-associated fibro-
blasts improves cancer chemotherapy by increasing intratumoral drug uptake. J Clin Invest
2006; 116:1955-62.
21. Lee J, Fassnacht M, Nair S, Boczkowski D, Gilboa, E. Tumor immunotherapy targeting
fibroblast activation protein, a product expressed in tumor-associated fibroblasts. Cancer
Res 2005; 65:11156-63.
Figure 1. Immunohistological markers of stromal fibroblasts in tumor.
Carcinoma-associated fibroblasts (CAFs) are stromal fibroblast populations
present within tumor and these CAFs include populations of both myofibro-
blasts and fibroblasts. Various fibroblastic markers are used to detect stromal
fibroblasts in tumor: α-SMA, fibroblast-activation protein (FAP), vimentin,
NG2 (Neuron-Glial Antigen-2) chondroitin sulfate proteoglycan, PDGFR-b,
fibroblast-associated antigen, prolyl 4-hydroxylase, and fibroblast-specific
protein-1 (FSP-1/S100A4). Of these, α-SMA and FAP seems to be specific
markers for myofibroblasts.