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Tumor Growth Need Not Be Driven by Rare Cancer Stem Cells
Priscilla N. Kelly,1,2Aleksandar Dakic,1,2Jerry M. Adams,1*Stephen L. Nutt,1*Andreas Strasser1*
The cancer stem cell hypothesis postulates that tumor growthis driven by a rare subpopulation of tumor cells. Much of thesupporting evidence for this intriguing idea is derived fromxenotransplantation experiments in which human leukemia cellsare grown in immunocompromised mice. We show that, when lymphomasand leukemias of mouse origin are transplanted into histocompatiblemice, a very high frequency (at least 1 in 10) of the tumorcells can seed tumor growth. We suggest that the low frequencyof tumor-sustaining cells observed in xenotransplantation studiesmay reflect the limited ability of human tumor cells to adaptto growth in a foreign (mouse) milieu.
1 Walter and Eliza Hall Institute of Medical Research, Melbourne 3050, Australia. 2 Department of Medical Biology, University of Melbourne, Melbourne 3050, Australia.
* These authors contributed equally to this study.
To whom correspondence should be addressed. E-mail: strasser{at}wehi.edu.au
Cancer biologists are intrigued by the hypothesis that tumorgrowth may be sustained by a rare subpopulation of the cells,termed cancer stem cells. Supporting this concept are the heterogeneouscellular composition of certain tumors and the finding thatonly a minute proportion of the cells (1/106) in some humanacute myeloid leukemia (AML) samples canseed tumor growth whentransplanted in to sublethally irradiated nonobese diabetic(NOD) severe combined immunodeficient (scid) mice (1). The interpretationof such xenotransplantation studies, however, is complicatedby the critical role in tumor growth of interactions with themicroenvironment, which are mediated by both soluble and membrane-boundfactors (2). Notably, many such mouse factors cannot engagethe cognate human receptor and vice versa (3). Thus, the lowfrequency of human AML cells producing tumors in NOD/scid micemight reflect in part the rarity of human tumor cells that canreadily adapt to growth in a foreign (mouse) milieu.
In our view, the frequency of cells that can sustain tumor growth,and thus the generality of the cancer stem cell hypothesis,can best be tested by transfer of titrated numbers of mousetumor cells into nonirradiated histocompatible recipient mice.We isolated primary pre-B/B lymphoma cells from three independentEµ-myc transgenic mice and injected 10 to 105 cells intononirradiated congenic animals. Regardless of the cell numberinjected, all recipients became moribund with disseminated lymphomawithin 35 days (Table 1). Although the number of injected cellsdid not noticeably affect tumor burden, organ infiltration,or disease severity, recipients of 10 or 100 lymphoma cellsusually developed tumors more slowly than those receiving 105cells. Importantly, even transfer of a single cell elicitedfatal lymphoma in three of eight recipients within 33 to 76days (case 2).
Table 1. A large proportion of tumor cells can sustain the growth of murine lymphoid and myeloid malignancies. Cells from primary Eµ-myc pre-B/B lymphomas, Eµ-N-RAS thymic lymphomas, or PU.1–/– AML, all from mice on a C57BL/6 (Ly5.2+) background (>15 backcrosses), were transplanted into nonirradiated congenic C57BL/6 (Ly5.1+) recipient mice. To circumvent problems associated with injection of low cell numbers, we mixed the tumor cells with 106 congenic (C57BL/6-Ly5.1+) spleen cells as carriers. Shown are the fraction of recipients that developed tumors and the average time from transplantation to tumor development. No mice (0/24) injected with carrier spleen cells alone developed any tumor over a 100-day period. ND, not determined.
A small fraction (2 to 5%) of the cells in primary Eµ-myclymphomas displayed the characteristic stem cell markers Sca-1and/or AA4.1. However, when sorted Sca-1+ AA4.1hi or Sca-1+AA4.1lo lymphoma cells were transplanted, as few as 10 cellsof each population elicited fatal lymphoma within 17 to 40 days(Table 1). Similarly, with Eµ-N-RAS thymic lymphomas andfour independent cases of AML caused by PU.1 deficiency, recipientstransplanted with as few as 10 cells developed tumors, althoughonset was delayed in mice receiving only 10 or 100 AML cells(Table 1). For all three malignancies, the cell surface markerphenotype (fig. S1), the gene expression profile (fig. S2),and the invasiveness of the transplanted tumors mirrored thatof the primary tumor.
These observations challenge the concept that growth of AML,and possibly other malignancies, are always sustained by a rarecancer stem cell (1). Although cancer stem cells may well drivethe growth of many cancers, particularly those displaying extensivedifferentiation, our studies of mouse lymphomas and leukemiasindicate that at least certain malignancies (particularly thosewith substantial homogeneity) can be maintained by a relativelylarge proportion (>10%) of tumor cells, perhaps even themajority. Although mouse and human tumors differ in notablerespects, the marked disparity with results from human AML cells(1) suggests that xenotransplantation may underestimate thepercentage of tumor-sustaining cells. With common human solidtumors (for example, brain, colon, and breast), transplantationplaces the tumor growth–sustaining cells within subpopulations(for example, CD133+) that compose up to 20% of the cells (4–6),and most of the remaining cells might be at differentiationstages unsupportable by the mouse microenvironment. The reportedrarity of cancer stem cells in AML (1) and colon cancer (4)might reflect the need to cotransfer an essential human accessorycell (we note that endothelial cell progenitors are also CD133+).
Determining whether the growth of various tumors is sustainedby most of the tumor cells or by a rare subpopulation has importantramifications for the design of novel therapies. Therefore,the cancer stem cell hypothesis merits more rigorous tests.For human tumors, ultimately this will require transfer of tumorcells into mice installed with all the requisite human supportcells. Lastly, because the term "cancer stem cell" also currentlydesignates the normal cell that founded the tumor, we suggestthat the cells sustaining growth of an established tumor bereferred to as "tumor-propagating cells."
Received for publication 15 March 2007. Accepted for publication 25 May 2007.
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In Science Magazine
TECHNICAL COMMENTS
James A. Kennedy, Frédéric Barabé, Armando G. Poeppl, Jean C. Y. Wang, and John E. Dick (14 December 2007) Science318 (5857), 1722c.
[DOI: 10.1126/science.1149590] |Abstract »|Full Text »|PDF »|Supporting Online Material »
TECHNICAL COMMENTS
Jerry M. Adams, Priscilla N. Kelly, Aleksandar Dakic, Stephen L. Nutt, and Andreas Strasser (14 December 2007) Science318 (5857), 1722d.
[DOI: 10.1126/science.1149672] |Abstract »|Full Text »|PDF »
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To whom correspondence should be addressed. E-mail: 
1/106) in some human acute myeloid leukemia (AML) samples canseed tumor growth when transplanted in to sublethally irradiated nonobese diabetic (NOD) severe combined immunodeficient (scid) mice (
