AU2014200650A1 - Anti-CXCR1 compositions and methods - Google Patents

Abstract

ANTI-CXCR1 COMPOSITIONS AND METHODS Abstract The present invention provides methods of treating cancer by administering an 1L8 CXCR1 pathway inhibitor (e.g., an anti-CXCR1 antibody or Repertaxin) alone or in combination with an additional chemotherapeutic agent such that non-tumorigenic and tumorigenic cancer cells in a subject are killed. The present invention also provides compositions and methods for detecting the presence of and isolating solid tumor stem in a patient (e.g., based on the presence of CXCR1 or FBXO21).

Description

ANTI-CXCRI COMPOSITIONS AND METHODS CROSS-REFERENCE TO RELATED APPLICATIONS The present application claims priority to U.S. Provisional Patent Application No. 5 61/113,458, fi led November 11, 2008, hereby incorporated by reference in its entirety. STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT This invention was made with government support under grant numbers 10 CA66233, CAl 01860, and 5 P30 CA46592 awarded by the NIH. The government has certain rights in the invention. FIELD OF THE INVENTION The present invention provides methods of treating cancer by administering an 15 IL8-CXCRA pathway inhibitor (e.g., an anti-CXCRi antibody or Repertaxin) alone or in combination with an additional chemotherapeutic agent such that non-tumorigenic and tumorigenic cancer cells in a subject are killed. The present invention also provides compositions and methods for detecting the presence of and isolating solid tumor stem cells in a patient (e.g., based on the presence of CXCRi or FBXO21). 20 BACKGROUND Cancer remains the number two cause of mortality in this country, resulting in over 500,000 deaths per year. Despite advances in detection and treatment, cancer mortality remains high. Despite the remarkable progress in understanding the 25 molecular basis of cancer, this knowledge has not yet been translated into effective therapeutic strategies. In particular, breast cancer is the most common cancer in American women, with approximately one in nine women developing breast cancer in their lifetime. Unfortunately, metastatic breast cancer is still an incurable disease. Most wornen with 30 metastatic breast cancer succumb to the disease.

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Traditional modes of therapy (radiation therapy, chemotherapy, and hormonal therapy), while useful, have been limited by the emergence of treatment-resistant cancer cells. Clearly, new approaches are needed to identify targets for treating metastatic breast cancer and cancer generally. 5 SUMMARY OF THE INVENTION The present invention provides methods of treating cancer by administering an IL8-CXCRi pathway inhibitor (e.g., an anti-CXCRI antibody or Repertaxin) alone or in combination with an additional chemotherapeutic agent such that non-tumorigenic and 10 tumorigenic cancer cells in a subject are killed. The present invention also provides compositions and methods for treating and diagnosing the presence of solid tumor stem cells in a patient (e.g., based on the presence of CXCRI or FBXO2 1). In some embodiments, the present invention provides methods of treating cancer comprising: administering an IL8-CXCR1 pathway antagonist and an additional 15 chemotherapeutic agent to a subject. In certain embodiments, the present invention provides methods of reducing or eliminating cancer stem cells and non-tumorigenic cancer cells in a subject comprising: administering Repertaxin or derivative thereof to a subject under conditions such that at least a portion of the cancer stein cells and at least a portion of the non-tumorigenic cancer cells are killed. In other embodiments, the present 20 invention provides methods of reducing or eliminating cancer stem cells and non tumorigenic cancer cells in a subject comprising: administering an IL8-CXCRI pathway antagonist and an additional chemotherapeutic agent to a subject under conditions such that at least a portion of the cancer stein cells and at least a portion of the non tumorigenic cancer cells are killed. In particular embodiments, the present invention 25 provides compositions or kits comprising an IL8-CXCR I pathway antagonist and an additional chemotherapeutic agent. In certain embodiments, the IL8-CXCRI pathway antagonist comprises an agent that specifically blocks the binding of IL8 to CXCRi. In some embodiments, the agent binds to (is specific for) CXCRi, but does not bind to CXCR2. In other embodiments, 30 the agent binds to CXCR 1. In particular embodiments, the agent comprises an anti CXCRI antibody or antibody fragment. In additional embodiments, the agent comprises Repertaxin or a derivative thereof. In further embodiments, the additional chemotherapeutic agent comprises an anti-mitotic compound. In certain embodiments, the anti-mitotic compound is selected from the group consisting of: docetaxel, doxorubicin, paclitaxel, fluorouracil, vincristine, vinblastine, nocodazole, colchicine, 5 podophyllotoxin, steganacin, and combretastatin. In other embodiments, the anti-mitotic compound is a catharalthus alkaloids (e.g., vincristine and vinblastine); or a benzimidazole carbamates such as nocodazole; or colchicine or related compounds such as podophyllotoxin, steganacin or combretastatin; or a taxane such as paclitaxel and docetaxel. In certain embodiments, the additional chemotherapeutic agent comprises 10 docetaxel. In particular embodiments, the subject has a type of cancer that, when treated with a chemotherapeutic, has increased levels of IL-8 production (e.g., which causes an increase in cancer stem cell number of motility). In some embodiments, the subject has a type of cancer selected from the group consisting of: prostate cancer, ovarian cancer, 1s breast cancer, melanoma, non-small cell lung cancer, small-cell lung cancer, and esophageal adenocarcinoma. In other embodiments, the present invention provides methods of detecting solid tumor stem cells comprising; a) providing: i) a sample taken from a tumor of a subject, and ii) an antibody, or antibody fragment (or other binding molecule), specific for the 20 CXCR1 protein or FBXO21 protein (or another protein from Table I); and b) contacting the tissue sample with the antibody, or antibody fragment, under conditions such that the presence or absence of CXCR I+ or FBXO21+ solid tumor stem cells are detected. In particular emb odiments, the antibody, or antibody fragment, is conjugated to a signal molecule. In further embodiments, the signal molecule comprises a fluorescent 25 molecule. In other embodiments, the signal molecule comprises an enzyme that can catalyze a color producing reaction in the presence of a colorimnetric substrate. In certain embodiments, the method further comprises contacting the sample with a secondary antibody, or secondary antibody fragment, specific for the antibody or antibody fragment. In other embodirnents, the secondary antibody, or secondary antibody fragment, 30 comprises a signal molecule. In particular embodiments, no other proteins or nucleic acids are assayed in order to determine the presence or absence of the CXCR1 or 3 FBXO21+ solid tumor stem cells. In additional embodiments, the tumor is selected from the group consisting of: a prostate cancer tumor, an ovarian cancer tumor, a breast cancer tumor, a melanoma, a non-small cell lung cancer tumor, a small-cell lung cancer tumor, and an esophageal adenocarcinoma tumor. 5 In some embodiments, the present invention provides methods of enriching for a population of solid tumor stein cells comprising: a) disassociating a solid tumor to generate disassociated cells; b) contacting the disassociated cells with a reagent that binds CXCR I or FBXO21 (or other protein from Table 1); and c) selecting cells that bind to the reagent under conditions such that an a population enriched for solid tumor stein cells is 10 generated. In certain embodiments, no additional reagents are employed in order to generate the population enriched for solid tumor stem cells. In some embodiments, the tumor is selected from the group consisting of: a prostate cancer tumor, an ovarian cancer tumor, a breast cancer tumor, a melanoma, a non-small cell lung cancer tumor, a small-cell lung 1s cancer tumor, and an esophageal adenocarcinoma tumor. In further embodiments, the reagent is an antibody or antibody fragment (e.g., Fab fragment). In additional embodiments, the reagent is conjugated to a fluorochrome or magnetic particles. In other embodiments, the selecting cells is performed by flow cytometry, fluorescence activated cell sorting, panning, affinity column separation, or magnetic selection. 20 In particular embodiments, the present invention provides an enriched population of solid tumor stem cells isolated by the methods described herein, In some embodiments, the present invention provides isolated populations of cancer stein cells that are: a) tumorigenic; and b) CXCR1+ or FBXO21I+, In certain embodiments, the cancer stem cells are cancer stein cells selected from the group 25 consisting of: prostate cancer stem cells, ovarian cancer stem cells, breast cancer stern cells, skin cancer stem cells, non-small cell lung cancer stem cells, small-cell lung cancer stem cells, and esophageal adenocarcinoma stem cells. In other embodiments, the population comprises at least 60% cancer stein cells and less than 40% non-tumorigenic tumor cells. In further embodiments, the cancer stein cells: are enriched at least two-fold 30 compared to unfractionated non-tumorigenic tumor cells (e.g., 2-fold, 3-fold, 4-fold, 5 fold, ..., 1 0-fold, ... 1 00-fold, .. 1 000-fold). 4 In some embodirents, the present invention provides methods for obtaining from a tumor a cellular composition comprising cancer stem cells and non-tumorigenic tumor cells, wherein at least 60% are tumorigenic stem cells and 40% or less are non tumorigenic tumor cells, the method comprising: a) obtaining a dissociated mixture of 5 tumor cells fr-om a tumor; b) separating the mixture of tumor cells into a first fraction comprising at least 60% cancer stein cells and 40% or less non-tumorigenic tumor cells and a second fraction of tumor cells depleted of cancer stern cells wherein the separating is by contacting the mixture with a reagent against CXCR I or FBXO21; and c) demonstrating the first fraction to be tumorigenic by: i) serial injection into a first host 10 animal and the second fraction to be non-tumorigenic by serial injection into a second host animal. In certain embodiments, the separating is performed by flow cytometry, fluorescence activated cell sorting (FACS), panning, affinity chromatography or magnetic selection. In some embodiments, the separating is performed by fluorescence activated cell sorters (FACS) analysis. 15 In particular embodiments, the present invention provides methods for selecting a treatment for a patient having a solid tumor, comprising: (a) obtaining a sample from the patient; (b) identifying the presence of CXCR-1 or FBXO21+ solid tumor stem cell in the sample; and (c) selecting a treatment for the patient that targets CXCRi+ or FBXO2 1+ solid tumor stem cells (e.g., selecting the use of an anti-CXCRi antibody or 20 antibody fragment). In certain embodiments, the CXCRi+ or FBXO21+- solid tumor stem cells are cancer stem cells selected from the group consisting of: prostate cancer stein cells, ovarian cancer stem cells, breast cancer stem cells, skin cancer stein cells, non-small cell lung cancer stem cells, small-cell lung cancer stem cells, and esophageal adenocarcinoma stem cells. 25 In some embodiments, the present invention provides methods for screening a compound, comprising: a) exposing a sample comprising a CXCR+ ±or FBXO2 1+ cancer stein cell to a candidate anti-neoplastic compound, wherein the candidate anti neoplastic compound comprises a CXCR I or FBXO2 1 antagonist or a IL8-CXCRI signaling pathway antagonist; and b) detecting a change in the cell in response to the 30 compound. 5 In certain embodiments, the sample comprises a non-adherent mammosphere. In further embodiments, the CXCRI or FBXO21 antagonist, or IL8-CXCRi signaling pathway antagonist comprises an antibody or antibody fragment. In some embodiments, the CXCRI antagonist is a derivative of Repartaxin. In other embodiments, the detecting comprises detecting cell death of the tumorigenic breast cell. In further embodiments, the methods further comprise identifying the candidate anti-neoplastic agent as capable of killing tumorigenic cells as well as non-tumorigenic cancer cells. In some embodiments, the present invention provides methods for determining the capability of a test compound to inhibit tumorigenesis of solid tumor stem cells 10 comprising: a) obtaining enriched solid tumor stem cells, wherein the solid tumor stem cells: i) are enriched at least two-fold compared to unfractionated tumor cells; and ii) express CXCRI or FBXO21; b) exposing a first set, but not a second set, of the solid tumor stem cells to a test compound; c) injecting the first set of the solid tumor stein cells into a first host animal and injecting the second set of solid tumor stein cells into a second 1s host animal; and d) comparing a tumor, if present, in the first animal with a tumor formed in the second animal in order to determine if the test compound inhibits tumor formation. In particular embodiments, the test compound is a CXCRI or FBXO21 inhibitor, or a IL8-CXCRi inhibitor pathway inhibitor. In further embodiments, the present invention provides methods for determining 20 the capability of a test compound to inhibit tumorigenesis of solid tumor stem cells comprising: a) obtaining a sample comprising at least 60% solid tumor stein cells, wherein the solid tumor stem cells express CXCR I or FBXO2 1; b) injecting the solid tumor stem cells into first and second host animals; c) treating the first host animal with a test compound, and not treating the second host animal with the test compound; and d) 25 comparing a tumor, if present, in the first animal with a tumor formed in the second animal in order to determine if the test compound inhibits tumor formation. In other embodiments, the test compound is a CXCRI or FBXO21 inhibitor or an IL8-CXCR I pathway inhibitor. 30 6 DESCRIPTION OF FIGURES Figure I shows the ALDEFLUOR-positive cell populations from breast cancer cell lines (MDA-MB-453, SUM 159) have cancer stem cell properties. A-B, G-1. Representative flow cytornetry analysis of ALDH enzymatic activity in MDA-M13-453 5 (A-B) and SUM159 cells (G-HI). The ALDEFLUOR assay was performed as described in Example I below. (C, I) The ALDEFLUOR-positive population was capable of generating tumors in NOD/SCID mice which recapitulated the phenotypic heterogeneity of the initial tumor. (F, L) Tumor growth curves were plotted for different numbers of cells injected (for MDA-MB-453: 50,000 cells, 5,000 cells, and 10 500 cells and for SUM159: 100,000 cells, 10,000 cells, and 1,000 cells) and for each population (ALDEFLUOR-positive, ALDEFL U OR-negative, unseparated). Tumor growth kinetics correlated with the latency and size of tumor formation and the number of ALDEFLUOR-positive cells L). (D, J) H&E staining of ALDEFLUOR-positive cells' injection site, revealing presence of tumor cells (D: MDA-MB-453 15 ALDEFLUOR-positive cells' injection site, and J: SUM59 ALDEFLUOR-positive cells' injection site). (E, K) The ALDEFLUOR-negative ceiis' injection site contained only residual Matrigel, apoptotic cells, and mouse tissue (E: MDA-MB-453 ALDEFLUOR-negative cells' injection site, and K: SUM59 ALDEFLUOR-negative cells' injection site). Data represent mean ± SD. 20 Figure 2 shows classification of the ALDEFLUOR-positive and ALDEFLUOR negative populations isolated from breast cell lines based on the "cancer stem cell signature". Figure 2A. Hierarchical clustering of 16 samples based on a 413-gene expression signature. Each row of the data matrix represents a gene and each column represents a sample. Note the separation between ALDEFLUOR-positive (underlined 25 names) and negative samples (non-underlined names) with the 413 genes for 15 out of the 16 samples. Some genes included in the signature are referenced by their HUGO abbreviation as used in 'Entrez Gene' (Genes down-regulated in the ALDEFLUOR positive populations are labeled in green and genes up-regulated in the ALDEFLUOR positive populations are labeled in red). Fig. 213-C. To confirm the gene expression 30 results, in a set of five breast cancer cell lines sorted for the ALDEFLUOR phenotype, the expression of five discriminator genes overexpressed in ALDEFLUOR-positive populations (CXCRI/IL8RA, FBXO21, NFYA, NOTCH2 and RAD5 LI) were measured by quantitative RT-PCR. The quantitative RT-PCR expression levels of CXCRl and FBXO2 I are presented in this figure. Gene expression levels measured by quantitative RT-PCR confirm the results obtained using DNA microarrays with an 5 increase of CXCR1 and FBXO2 I rRNA level in the ALDEFLUOR-positive population compared to the ALDEFLUOR-negative population (p<0.05). Figure 3 shows the role of the IL8/CXCRi axis in the regulation of breast cancer stem cells. A. Cells expressing CXCRl are contained in the ALDEFLUOR positive population. The ALDEFLUOR-positive and -negative population from four 10 different breast cell lines (HCC1954, SUM 159, MDA-MB-453, BrCa-MZ-01) were isolated by FACS, fixed, and analyzed for the expression of CXCRI protein by imunostaining and FACS analysis. ALDEFLUOR-positive cells were highly enriched in CXCRi-positive cells compared to the ALDEFLUOR-negative population. B. Effect of IL8 treatment on tumorosphere formation of three different cell lines 15 (HCC1954, SUM159, MDA-MB-453). IL8 treatment increased the formation of primary and secondary tumorospheres in a dose-dependent manner. C. Effect of IL8 treatment on the ALDEFLUOR-positive population of four different cell lines cultured in adherent conditions. IL8 increased the ALDEFLUOR-positive population in a dose dependent manner in each of the four cell lines analyzed (* p<0.05/ ** p<0.01, 20 statistically significant differences from the control group). Figure 4 shows ALDEFLUOR-positive cells display increased metastatic potential. A. The IL8/CXCRI axis is involved in cancer stem cell invasion. The role of the IL8/CXCR1 axis in invasion was assessed by a Matrigel invasion assay using serum or IL8 as attractant for three different cell lines (HCC1954, MDA-MB-453, SUM159). 25 ALDEFLUOR-positive cells were 6- to 20-fold more invasive than ALDEFLUOR negative cells (p<0.01). When using 11.8 (100 ng/ml) as attractant, it was observed that a significant increase of ALDEFLUOR-positive cells were invading through Matrigel compared to serum as attiactant (p<0.05). In contrast 118 had -no effect on the invasive capacity of the ALDEFLUOR-negative population. B-M. The AL DE FLUOR -positive 30 population displayed increased metastatic potential. B-D. Quantification of the normalized photon flux measured at weekly intervals following inoculation of 100,000 8 luciferase infected cells from each group (ALDEFLUOR-positive, ALDEFLUOR negative, unseparated). E-J Detection of metastasis utilizing the bioluminescence imaging software (E, G, I: Mice facing down; F, H-, J: Mice facing up). Mice inoculated with ALDEFLUOR-positive cells developed several metastasis localized at 5 different sites (bone, muscle, lung, soft tissue) and displayed a higher photon flux emission than mice inoculated with unseparated cells, which developed no rnore than one metastasis per mouse. In contrast, rice inoculated with ALDEFLUOR-negative cells developed only an occasional small metastasis, which was limited to lymph nodes. K-M. Histologic confirmation, by H&E staining, of metastasis in bone (K), soft tissue 10 (L) and muscle (M) resulting from injection of ALDEFLUOR-positive cells. Figure 5 shows the effect of CXCR1 inhibition on tumor cells viability (Fig. 5A) as well as on cancer stem cell viability (Fig. 5B). Figure 6 shows that Repertaxin treatment induces a bystander effect mediated by the FAS/FAS ligand signaling, and specifically shows that the cell growth inhibition 15 induced by the Repertaxin treatment was partially rescued by the addition of a FAS antagonist and that the cells treated with a FAS agonist displayed a similar cell growth inhibition than the cells treated with Repertaxin. Figure 7 shows the activation of FAK. AKT and FOXOA3 activation without Repertaxin treatment (7A) and in the presence of Repertaxin (7B). 20 Figure 8 shows the effect of Repertaxin, docetaxel, or the combination thereof on one breast cancer cell line (8A, SUM159) and three human breast cancer xenografts generated from different patients (8B, MCI; 8C. UM2; and 8D, UM3). Figure 9 shows the effect of Repertaxin, docetaxel, or the combination treatment on the cancer stein cell population as assessed by the ALDEFLUOR assay on various 25 cells lines including SUM159 (9A), MCI (9B), UM2 (9C), UM3 (9D). Figure 10 shows the effect of Repertaxin, docetaxel or the combination on serial dilutions of primary tumors (10A. SUM159, 10B. MCI, 10C. UM2, 10D. UM3) that were implanted in the rnammary fat pad of secondary NOD-SCID mice. Figure II shows that Repertaxin treatment reduces the metastatic potential of 30 SUM 159 cell line. Figure 1 IA shows a quantification of the normalized photon flux measured at weekly intervals following inoculation with intracardiac administered SUM 9 159 cells. Metastasis formation was monitored using bioluminescence imaging (] 1B: Mice treated with saline solution; I IC: Mice treated with Repertaxin). Figure 12 shows representations of the overlap between the ALDEFLIJOR positive subpopulation and the CXCR I-positive subpopulation (top) or CXCR2 5 positive subpopulation (bottom) of SUM 159 cells. B-C. SUM159 cells were cultured in adherent conditions and treated with repertaxin (I 00nM) or two specific blocking antibodies for CXCRi (10pg/rnl) or CXCR2 (0 g/ml). After three days, the effect on the cancer stein cell population was analyzed using the ALDEFLUOR assay (B) cell viability was accessed after five days of treatment using the MTT assay (C). A 10 significant reduction of the ALDEFLUOR-positive population and cell viability was observed following treatment with repertaxin or anti-CXCRI antibody. In contrast no significant effect was observed with anti-CXCR2 antibody. D. After 4 days of treatment, the number of apoptotic cells was evaluated utilizing a TUNEL assay. 36% apoptotic cells (stained in green) were detected in repertaxin treated cells compared to 15 the controls where mostly viable cells (stained in blue) were present. E-F. To determine whether cell death was mediated via a bystander effect. CXCRl-positive and CXCRi negative populations were flow sorted and each population treated with various concentrations of repertaxin (D). A decrease in cell viability in CXCRl -positive and unsorted populations were detected whereas no effect was observed in the CXCRI 20 negative population (E). Dialyzed conditioned medium (dCM) from CXCRl -positive cells treated for three days with repertaxin was utilized to treat sorted CXCR1 -positive, CXCRI-negative, or unsorted populations. Serial dilutions of dialyzed conditioned medium were utilized (Control, dCM 1/4, dCM 1/2, dCM 3/4, dCM). After two days of treatment, cell viability was evaluated utilizing the MTT assay. A massive decrease in 25 cell viability was observed in both CXCR I-negative and unseparated populations whereas no effect was observed in the CXCRl-positive population (F). Figure 13 shows tumorigenicity of the ALDEFLUOR-positive/CXCR i-positive and ALDEFLU OR-positive/CXCR1-negative cell populations from SUM1 59 cell line. A. Tumor growth curves were plotted for different numbers of cells injected (50,000 30 cells, 5,000 cells, 1,000 cells, and 500 cells) and for each population (ALDEFLUOR positive/CXCR 1-positive, ALDEFLU OR-positive/CXCR 1-negative). Both cell 10 populations generated tumors. Tumnor growth kinetics correlated with the latency and size of tumor formation and the number of cells injected. B-C. Turnors generated by the ALD EF LUOR-positive/CXCR I -positive population reconstituted the phenotypic heterogeneity of the initial tumor upon serial passages whereas the ALDEFLUOR 5 positive/CXC RI -negative population gave rise to tumors containing only ALDEFLUOR-positive/CXCR I -negative cells. We transplanted both cell population for three passages. Figure 14 shows the effect of CXCR I blockade on tumorsphere formation. SUJM 159 and HCCI 954 cells were cultured in adherent conditions and treated for three 10 days with repertaxin (I 00nM'), an anti-CXCRI blocking antibody (10pg/ml), or an anti CXCR2 blocking antibody (I Ogg/ml). After three days of treatment, cells were detached and cultured in suspension. The number of tumorspheres formed after 5 days of culture were evaluated. Similar results were observed for the both cell lines with a significant decrease in primary and secondary tumorosphere formation in the repertaxin 15 and anti CXCRI-treated conditions compared to controls. In contrast, anti-CXCR2 blocking antibody had no effect on tumorosphere formation. Figure 15 shows the effect of repertaxin treatment on cell viability of SUM 159, HCC1954, and MDA-MB-453 cell lines. Three different cell lines (SUM159, HCC 1954, MDA-MB-453) were cultured in adherent conditions and treated with 20 repertaxin (IO0nM). Cell viability was evaluated after one, three, and five days of treatment using the MTT assay. A decrease in cell viability was observed after 3 days of treatment for STM159 and HCC1954 cell line. However, repertaxin did not effect the viability of MDA-MB 453 cells. Figure 16 shows the effect of CXCR-1 blockade on the ALDEFLJOR-positive 25 population in vitro. A-B. HCC 1954 (A) and MDA-MB-453 (B) cells were cultured in adherent conditions and treated with repertaxin (IOOnM) or two specific blocking antibodies for CXCRi (10 g/m)l) or CXCR2 (10pg/ml). After three days, the effect on the cancer stem cell population was analyzed using the A LDEFLUOR assay. For HCC1954, a significant reduction of the ALDEFLU OR-positive population and cell 30 viability was observed following treatment with repertaxin or anti-CXCRi antibody. In 11 contrast no significant effect was observed with anti-CXCR2 antibody (A). For MDA MB-453, np any effect on the ALDEFLUOR-positive population was observed (B). Figure 17 shows repertaxin treatment induces a bystander effect mediated by FAS/FAS-ligand signaling. A. To determine whether the bystander killing effect 5 induced by the repertaxin treatment was mediated by FA S-ligand, the level of soluble FAS-ligand in the medium was measured utilizing an ELISA assay. After 4 days of treatment, greater than a four-fold increase of soluble FAS-Ligand was detected in the medium of cells treated with repertaxin compared to non-treated controls. B. The level of FAS-ligand mRNA was measured by RT-PCR and confirmed the increase of FAS 10 ligand production after treatment with repertaxin. Similar results were observed after 4 days of treatment with a FAS agonist that activates FAS signaling, with a five-fold increase of the FAS-ligand mRNA compared to the control. C. SUM159 cells were cultured in adherent conditions and treated with repertaxin alone or in combination with an anti- FAS-ligand. Cell growth inhibition induced by the Repertaxin treatment was 15 partially rescued by addition of anti-FAS-Ligand. Cells treated with a FAS agonist displayed similar cell growth inhibition to cells treated with repertaxin alone. D-E. The effect of repertaxin treatment alone or in combination with an anti-FAS-ligand and the treatment of a FAS-agonist on the CXCR1-positve and ALDEFLUOR-positive population was analyzed. The massive decrease in the CXCRI-positive and 20 ALDEFLUOR-positive population induced by repertaxin treatment was not rescued by the anti-FAS-ligand and treatment with FAS-agonist produced a ten-fold and three-fold increase in the percent of the CXCR I -positive and ALDEFLUOR-positive population, respectively. Figure 18 shows the effect of FAS agonist on CXCRI -positive and CXCR-1 25 negative cells. CXCR1-positive and CXCR] -negative populations were flow sorted and each population treated with various concentrations of FAS agonist. A decrease in cell viability in CXCR I-negative and unsorted populations were detected whereas no effect was observed in the CXCRI-positive population. Figure 19 shows analysis of CXCRI protein expression in the normal breast 30 stem/progenitor population and effect of IL-8 treatment on mammosphere formation. A. The ALDEFLUOR-positive and -negative population from normal breast epithelial 12 cells isolated form reduction mammoplasties was isolated by FACS, fixed, and analyzed for the expression of CXCRI protein by inmniostaining and FACS analysis. ALD FLUOR-positive eelIs were highly enriched in CXCR I-positive cells compared to the ALDEFLUOR-negative population. B-C. Effect of IL8 treatment on 5 mammosphere formation. IlL8 treatment increased the formation of primary (B) and secondary mainospheres (C) in a dose-dependent manner. Figure 20 shows the effect of repertaxin treatment on the normal mammary epithelial cells. A. Normal rnammary epithelial cells isolated from reduction mammoplasties were cultured in adherent condition and treated with repertaxin (I OnM 10 or 500nM) or FAS agonist (500ng/ml). After five days of treatment cell viability was evaluated using MTT assay. Repertaxin treatment or the FAS agonist had no effect on the viability of normal mammary epithelial cells cultured in adherent conditions, even when high concentrations of repertaxin (500nM) were utilized. B. The level of soluble FAS-ligand was evaluated by Elisa assay in the medium of normal mannary epithelial 15 cells treated with repertaxin. After 4 days of treatment an increase of soluble FAS ligand was detected in the medium from treated cells. C. Analysis of FAS/CD95 expression in the normal mammary epithelial cells by FACS analysis. No FAS/CD95 expression was detected in the normal mammary epithelial cells cultured in adherent condition. D. Effect of repertaxin treatment on mammosphere formation. Normal 20 mammary epithelial cells were cultured in adherent condition and treated during four, eight, eleven and fifteen days with repertaxin (1 OOnM). After repertaxin treatment cells were detached and cultured in suspension. A significant decrease of mamnmosphere initiating cells was observed in the repertaxin-treated condition. Figure 2I shows the effect of repertaxin treatment on FAK, AKT and FOXO3a 25 activation. To evaluate the effect of repertaxin treatment on CXCR I downstream signaling, two different viral constructs were utilized, one knocking down PTEN expression via a PTEN-siRNA and the other leading to FAK overexpression (Ad FAK). A. SUMI59 control. SUM159 PTEN-siRNA, and SUM159 Ad-FAK cells were cultured in adherent conditions for two days in the absence or presence of 100inM 30 repertaxin and the activation of the FAK/AKT pathway was accessed by western blotting. Repertaxin treatment led to a decrease in FAK Tyr397 and AKT Ser473 13 phosphorylation whereas PTEN deletion and FAK overexpression blocked the effect of repertaxin treatment on FAK and AKT activity. B. Utilizing immunofluorescence training on CXCRI-positve cells, we confirmed that Repertaxin treatment results in a disappearance of phospho-FAK (nembranous staining in red) and phospho-AKT 5 expression (cytoplasmic staining in red). Inmnunofluorescence staining with an anti FOXO3A revealed a cytoplasmic location of FOXO3a (in red) in the untreated cells whereas repertaxin treatment induced a re-localization of FOXO3A to the nucleus. In contrast, cells with PTEN deletion or FAK overexpression display a high level of phospho-FAK, phospho-AKT and cytoplamic FOXO3A expression in both the 10 repertaxin treated and untreated cells. In all samples, nuclei were counterstained with DAPI (in blue). C-D. The effect of Repertaxin on the SUM159 PTEN-siRNA and SU MI 59 Ad-FAK cell viability and on the cancer stem cell population was assessed utilizing the MTT and ALDEFLU OR assays, respectively. After 3 days of treatment, cells with PTEN deletion or FAK overexpression developed resistance to repertaxin 15 (C). Repertaxin treatment did not alter the proportion of ALDEFLU OR-positive SUM159 PTEN knockdown cells. (D). Figure 22 shows the effect of repertaxin treatment on FAK/AKT activation in HCC 1954 and MDA-MB-453 cell lines. To evaluate the effect of repertaxin treatment on CXCRI downstream signaling we utilized a lentiviral construct knocking down 20 PTEN expression via a PTEN-siRNA A. HCC1954 control and HCC1954 PTEN siRNA cells were cultured in adherent conditions for two days in the absence or presence of 1 00nM repertaxin and the activation of the FAK/AKT pathway was accessed by western blotting. Repertaxin treatment led to a decrease in FAK Tyr397 and AKT Ser473 phosphorylation whereas PTEN deletion blocked the effect of 25 repertaxin treatment on FAK and AKT activity. B. Repertaxin treatment did not have any effect on cell viability of MDA-MB-453 cell line wich harbor PTEN mutation. Utilizing western blot analysis we confirmed that FAK/AKT pathway was not perturbated by repertaxin treatment. Figure 23 shows the effect of repertaxin on the HCC 1954 PTEN-siRNA cell 30 viability, assessed utilizing the MTT assay. After 3 days of treatment, cells with PTEN deletion developed resistance to repertaxin. 14 Figure 24 shows expression of FAS-ligand and IL-8 mRNA after docetaxel or repertaxin treatment measured by quantitative RT-PCR. A-B. SUM 159 cells cultured in adherent condition were treated with repertaxin (1 00nM), FAS agonist (50Ong/rnl) or docetaxel (10nrM). After three days of treatment cells were collected and RjNA 5 extracted. Docetaxel, induced both FAS-ligand (A) and IL-8 (B) mRNA in SUMi 59 cells. A 4-fold increase of IL-8 rnRNA level was detected after FAS agonist or docetaxel treatment (B). Figure 25 shows evaluation of PTEN!FAIK/AKT activation in the three different breast cancer xenografts. Western blot analysis revealed that both xenografts presented 10 an expression of PTEN and an activation of FAK/AKT pathway as shovn by FAK Tyr397 and AKT Ser473 phosphorylation. Figure 26 shows Effect of Repertaxin treatment on the breast cancer stem cell population in vivo. A-C. To evaluate the effect of repertaxin treatment on tumor growth and the cancer stem cell population in vivo a breast cancer cell line (SUM 159) 15 and three human breast cancer xenografts generated from different patients (MCI, UM2, UM3) were utilized. A. For each sample. 50,000 cells were injected into the humanized mammary fat pad of NOD/SCID mice and monitored tumor size. When the tumors were about 4 mm, s.c. injection of repertaxin (15mg/Kg) twice/day for 28 days or once/week I.P. injection of docetaxel (IOmg/Kg) or the combination 20 (repertaxin/docetaxel) was initiated. The graph shows the tumor size before and during the course of each indicated treatment (arrow, beginning of the treatment). Similar results were observed for each sample with a statistically significant reduction of the tumor size in docetaxel alone or the combination repertaxin/docetaxel treated groups compared to the control, whereas no difference was observed between the growth of the 25 control tumors and the tumors treated with repertaxin alone. B-C. Evaluation of repertaxin, docetaxel, or the combined treatment on the cancer stem cell population as assessed by the ALDEFLUOR assay (B) and by reimplantation into secondary mice (C). Docetaxel-treated tumor xenografts showed similar or increase percentage of ALDEFLUOR-positive cells compared to the control, whereas repertaxin treatment 30 alone or in combination with docetaxel produced a statistically significant decrease in ALDEFLUOR-positive cells with a 65% to 85% decrease in cancer stem cells 15 compared to the control (p<0.01) (B), Serial dilutions of cells obtained from primary turnors, non treated (control), and treated mice were implanted in the mrammary fat pad of secondary NOD/SCID mice which received no further treatment. Control and docetaxel treated primary tumors formed secondary tumors at all dilutions whereas, 5 only higher numbers of cells obtained from primary turnors treated with repertaxin or in combination with docetaxel were able to form tumors. Furthermore, tumor growth was significantly delayed and resulting tumors were significantly smaller in size than the control or docetaxel treated tumnors (C). D. Xenotransplants from each group were collected and inmunohistochemistry staining was done to detect the expression of 10 phospho-FAK, phospho-AKT, FOXO3A, and ALDI1l. Membranous phospho-FAK expression and cytoplasmic phospho-AKT expression (arrow) was detected in the control and docetaxel-treated tumors whereas no expression was detected in the tumors treated with repertaxin alone or in combination with docetaxel. Nuclear FOXO3A expression (in brown) was detected in the cells treated with docetaxel or repertaxin 15 alone or in combination. A decrease of ALDIH1 expression (arrow) was detected in tumors treated with repertaxin alone or in combination compared to control and the docetaxel-treated tumors, Figure 27 shows the effect of Repertaxin treatment on the breast cancer stem cell population in vivo. A-C. To evaluate the effect of repertaxin treatment on tumor 20 growth and the cancer stem cell population in vivo, a breast cancer cell line (SUM159, A) and three human breast cancer xenografts generated from different patients. For each sample, 50,000 cells were injected into the humanized mammary fat pad of NOD/SCID mice and monitored tumor size. When the tumors were about 4mm, s.c. injection of repertaxin (15mg/Kg) twice/day for 28 days or once/week I.P. injection of 25 docetaxel (10mg/Kg) or the combination (repertaxin/docetaxel) was initiated. The graph shows the tumor size before and during the course of each indicated treatment (arrow, beginning of the treatment). Similar results were observed for each sample with a statistically significant reduction of the turnor size in docetaxel alone or the combination repertaxin/docetaxel treated groups compared to the control whereas no 30 difference was observed between the growth of the control tumors and the tumors treated with repertaxin alone. Evaluation of repertaxin, docetaxel, or the combined 16 treatment on the cancer stem cell population was assessed by the ALDEFL OR assay and by reimplantation into secondary mice. Docetaxel-treated tumor xenografts showed similar or increased percentage of A LDEFLUIOR-positive cells compared to the control, whereas repertaxin treatment alone or in combination with docetaxel produced 5 a statistically significant decrease in ALDEFLUOR-positive cells with a 65% to 85% decrease in cancer stern cells compared to the control (p<0.01). Serial dilutions of cells obtained from primary turnors, non-treated (control), and treated mice were implanted in the mammary fat pad of secondary NOD/SCID mice which received no further treatment. Control and docetaxel treated primary tumors formed secondary tumors at all 10 dilutions whereas, only higher numbers of cells obtained from primary tumors treated with repertaxin or in combination with docetaxel were able to form tumors. Tumor growth was significantly delayed and resulting tumors were significantly smaller in size than the control or docetaxel treated tumors. Figure 28 shows the effect of repertaxin treatment on the breast cancer stem cell 15 population as assessed by the CD44+/CD24- phenotype. A-B. Evaluation of repertaxin, docetaxel, or the combined treatment on the cancer stem cell population was assessed by the presence of CD44-/CD24- cells. In residual tumors treated with docetaxel alone, we consistently observed either an unchanged or increased percent of CD44+/CD24- cells whereas repertaxin treatment alone or in combination with 20 docetaxel resulted in a reduction of the CDI4,/CD24- cell population. A. Flow chart analysis for LTM3 xenograft is presented. B. Sinilar results were observed for MCI, UM2, and UM3. Almost all of SUM159 cells are CD44+/CD24- under all treatment conditions. Figure 29 shows repertaxin treatment reduces the development of systemic 25 metastasis. To evaluate the effect of repertaxin treatment on metastasis formation HCC1954 (A), SUMI159 (B), MDA-MB-453 (C) breast cancer cell lines were infected with a lentivirus expressing luciferase and inoculated 250,000 luciferase infected cells into NOD/SCID mice via intracardial injection. Mice were treated 12 hours after the intracardiac injection either with s.c. injection of saline solution or s.c. injection of 30 repertaxin (15ng/kg), twice a day during 28 days. Metastasis formation was monitored using bioluminescence imaging. Quantification of the normalized photon flux measured 17 at weekly intervals following inoculation revealed a statistically significant decrease in metastasis formation in repertaxin compared to saline controls for mice inoculated with HCC 1954 or SUM 159 cells (A-B). In contrast, repertaxin treatment did not have any effect on metastasis formation for the mice injected with MDA-MB-453 cells. (C). Histologic confirmation, by I&E staining, of metastasis in bone, and soft tissue resulting from mice not treated by repertaxin (D). Figure 30 shows IL-8/CXCRI signalling in cancer stem cells treated with chemotherapy alone or in combination with repertaxin. A. Representation of potential IL-8/CXCRI cell signaling in cancer stem cells. CXCR1 activation following IL-8 10 binding induces phosphorylation of the Focal Adhesion Kinase (FAK). Active FAK phosphorylates AKT and activates the WNT pathway, which regulates stem cell self renewal and FOXO3A that regulates cell survival. Activation of FAK protects cancer stem cells from a FAS-ligand/FAS mediated bystander effect by inhibiting FADD, a downstream effector of FAS signaling. In the presence of chemotherapy, only the bulk 15 tumor cells are sensitive to the treatment and release a high level of IL-8 and FAS ligand proteins during the apoptotic process. Breast cancer stem cells are stimulated via an IL-8 mediated bystander effect and are resistant to the bystander killing effect mediated via FAS-ligand. B. Repertaxin treatment blocks IL-8/CXCRI signaling and inhibits breast cancer stem cell self-renewal and survival. When repertaxin treatment is 20 combined with chemotherapy the cancer stem cells are sensitized to the bystander killing effect mediated by FAS-ligand. DEFINITIONS To facilitate an understanding of the present invention, a number of terms and 25 phrases are defined below: As used herein, the terms "anticancer agent," "conventional anticancer agent," or "cancer therapeutic drug" refer to any therapeutic agents (e.g., chemotherapeutic compounds and/or molecular therapeutic compounds), radiation therapies, or surgical interventions, used in the treatment of cancer (e.g., in manuals). 30 As used herein, the terms "drug" and "chemotherapeutic agent" refer to pharmacologically active molecules that are used to diagnose, treat, or prevent diseases or 18 pathological conditions in a physiological system (eg., a subject, or in vivo, in vitro, or ex vivo cells, tissues, and organs). Drugs act by altering the physiology of a living organism, tissue, cell, or in vitro systern to which the drug has been administered. It is intended that the terms "drug" and "chemotherapeutic agent" encompass anti 5 hyperproliferative and antineoplastic compounds as well as other biologically therapeutic compounds. An "effective amount" is an amount sufficient to effect beneficial or desired results. An effective arnount can be administered in one or more administrations. As used herein, the term "administration" refers to the act of giving a drug, 10 prodrug, antibody, or other agent, or therapeutic treatment to a physiological systern (e.g. a subject or in vivo, in vitro, or cx vivo cells, tissues, and organs). Exemplary routes of administration to the human body can be through the eyes (ophthalmic), mouth (oral), skin (transdernal), nose (nasal), lungs (inhalant), oral mucosa (buccal), ear, by injection (e.g. intravenously, subcutaneously, intratumorally, intraperitoneally, etc.) and the like. 15 "Coadministration" refers to administration of more than one chemical agent or therapeutic treatment (e.g., radiation therapy) to a physiological system (e.g., a subject or in vivo, in vitro, or ex vivo cells, tissues, and organs). "Coadministration" of the respective chemical agents (e.g. IL8-CXCRI signaling pathway antagonist and additional chemotherapeutic) may be concurrent, or in any temporal order or physical combination. 20 As used herein, the term "regression" refers to the return of a diseased subject, cell, tissue, or organ to a non-pathological, or less pathological state as compared to basal nonpathogenic exemplary subject, cell, tissue, or organ. For example, regression of a tumor includes a reduction of tumor mass as well as complete disappearance of a tumor or tumors. 25 As used herein the term, "in vitro" refers to an artificial environment and to processes or reactions that occur within an artificial environment. In vitro environments can consist of, but are not limited to, test tubes and cell cultures. The term "in vivo" refers to the natural environment (e.g., an animal or a cell) and to processes or reactions that occur within a natural environment. 30 As used herein, the term "cell culture" refers to any in vitro culture of cells. Included within this term are continuous cell lines (e.g., with an immortal phenotype), 19 primary cell cultures, finite cell lines (e.g., non-transformed cells), and any other cell population maintained in vitro, including oocytes and embryos. As used herein, the term "subject" or "patient" refers to organisms to be treated by the methods of the present invention. Such organisms include, but are not limited to, 5 humans and veterinary animals (dogs, cats, horses, pigs, cattle, sheep, goats, and the like). In the context of the invention, the term "subject" or "patient" generally refers to an individual who will receive or who has received treatment. The term "diagnosed," as used herein, refers to the recognition of a disease by its signs and symptoms or genetic analysis, pathological analysis, histological analysis, and 10 the like. As used herein, the term "antisense" is used in reference to nucleic acid sequences (e.g. RNA, p hosphorothioate DNA) that are complementary to a specific RNA sequence (e.g., mRNA). Included within this definition are natural or synthetic antisense RNA molecules, including molecules that regulate gene expression, such as small interfering 15 RNAs or micro RNAs. One type of antisense sequence that may be employed by the present invention is the type that are specific for CXCRI niRNA. The term "test compound" or "candidate compound" refers to any chemical entity, pharmaceutical. drug, and the like, that can be used to treat or prevent a disease, illness, sickness, or disorder of bodily function, or otherwise alter the physiological or cellular 20 status of a sample. Test compounds comprise both known and potential therapeutic compounds. A test compound can be determined to be therapeutic by using the screening methods of the present invention. A "known therapeutic compound" refers to a therapeutic compound that has been shown (e.g., through animal trials or prior experience with administration to humans) to be effective in such treatment or prevention. In 25 preferred embodiments, "test compounds" are anticancer agents. In particularly preferred embodiments, "test compounds" are anticancer agents that induce apoptosis in cells. As used herein, the term "antigen binding protein" refers to proteins which bind to specific antigen. "Antigen binding proteins" include, but are not limited to, immiunoglobulins, including polyclonal, monoclonal, chimeric, single chain, and 30 humanized antibodies, Fab fragments, F(ab')2 fragments, and Fab expression libraries. Various procedures known in the art are used for the production of polyclorial antibodies. 20 For the production of antibodies, various host animals can be immunized by injection with the peptide corresponding to the desired epitope including, but riot limited to, rabbits, mice, rats, sheep, goats, etc. In a preferred embodiment, the peptide is conjugated to an inmunogenic carrier (e.g., diphtheria toxoid, bovine serurn albumin 5 (BSA), or keyhole limpet hemocyanin (KiL-H)). Various adj uvants are used to increase the immunological response, depending on the host species, including, but not limited to, Freund's (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanins, dinitrophenol, and potentially useful human 10 adjuvants such as BCG (Bacille Calmette-Guerin) and Corynebacterium parvum. For preparation of monoclonal antibodies, any technique that provides for the production of antibody molecules by continuous cell lines in culture may be used (See e.g., Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY). These include, but are not limited to, the 15 hybridoma technique originally developed by Kohler and Milstein (K6hler and Milstein, Nature, 256:495-497 (1975)), as well as the trioma technique, the human B-cell hybridoma technique (See e.g., Kozbor c al., Immunol. Today, 4:72 (1983)), and the EBV-hybridoma technique to produce human monoclonal antibodies (Cole et al., in Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96 (1985)). 20 According to the invention, techniques described for the production of single chain antibodies (U.S. 4,946,778; herein incorporated by reference) can be adapted to produce specific single chain antibodies as desired. An additional embodiment of the invention utilizes the techniques known in the art for the construction of Fab expression libraries (Huse et al., Science, 246:1275-1281 (1989)) to allow rapid and easy 25 identification of monoclonal Fab fragments with the desired specificity. Antibody fragments that contain the idiotype (antigen binding region) of the antibody molecule can be generated by known techniques. For example, such fragments include, but are not limited to: the F(ab')2 fragment that can be produced by pepsin digestion of an antibody molecule; the Fab' fragments that can be generated by reducing 30 the disulfide bridges of an F(ab')2 fragment, arid the Fab fragments that can be generated by treating an antibody molecule with papain and a reducing agent. 21 Genes encoding antigen-binding proteins can be isolated by methods known in the art. In the production of antibodies, screening for the desired antibody can be accomplished by techniques known in the art (e.g., radioirnmunoassay, ELISA (enzyme linked imnunosorbant assay), "sandwich" immunoassays, inununoradi metric assays, gel 5 diffusion precipitin reactions, immnunodiffusion assays, in situ immunoassays (using colloidal gold, enzyme or radioisotope labels, for example), Western Blots, precipitation reactions, agglutination assays (e.g., gel agglutination assays, hernaggutination assays, etc. pr utin At assa n etc.), complement fixation assays, inuunofluorescence assays, protein A assays, and immunoelectrophoresis assays, etc.) etc. 10 As used herein, the term "modulate" refers to the activity of a compound to affect (e.g., to promote or retard) an aspect of the cellular function including, but not limited to, cell growth, proliferation, invasion, angiogenesis, apoptosis, and the like. DETAILED DESCRIPTION OF THE INVENTION 15 The present invention provides methods of treating cancer by administering an IL8-CXCRI pathway inhibitor (e.g., an anti-CXCRI antibody or Repertaxin) alone or in combination with an additional chemotherapeutic agent such that non-tumorigenic and tumorigenic cancer cells in a subject are killed, The present invention also provides compositions and methods for treating and diagnosing the presence of solid tumor stem 20 cells in a patient (e.g., based on the presence of CXCRI or FBXO21). L, Tumorigenic Cancer Cells, ALDH, CXCR1, and CXCR1 Inhibition The evolution of a normal cell into a fully transformed one requires the 25 deregulation of multiple cellular processes (1, 2). According to classical models of carcinogenesis, these events can occur in any cell, In contrast, the "cancer stem cell hypothesis" holds that the preferential targets of oncogenic transformation are tissue stem or early progenitor cells that have acquired self-renewal potential (3-6). These "tuior-initiating cells" or "cancer stem cells" (CSC), in turn, are characterized by their 30 ability to undergo self-renewal, a process that drives tumorigenesis and differentiation which contributes to tumor cellular heterogeneity. Recent evidence supporting the 22 cancer stern cell hypothesis has been generated utilizing xenografts of prim ary human tumors. These studies have suggested that tumors are composed of a cellular hierarchy driven by the cancer stem cell component. In addition, recent data suggest that immortalized cell lines derived from both marine and human tissues may also contain a 5 cellular population displaying stem cell properties. Most of these studies have been based on in vitro properties including clonogenic potential, sphere formation and multi lineage differentiation potential (7-10). More limited studies utilizing functional transplantation of immortalized cell lines in xenografts have also suggested the existence of such a hierarchy. These studies have generally utilized Hoechst dye 10 exclusion to identify the so-called "side population" (SP) (7, 9, 11). In addition, cell surface markers defined using primary tumor xenografts such as CD44 and CD 133 have also been utilized to identify similar populations in established cell lines (7, 8). As described in the Examples below, the expression of the stem cell marker Aldehyde dehydrogenase (ALDH) was studied in a series of 33 cell lines derived from 15 human breast cancers and non-transformed breast cells. ALDH is a detoxifying enzyme responsible for the oxidation of intracellular aldehydes and is thought to play a role in stem cell differentiation through metabolism of retinal to retinoic acid (12. 13). ALDH activity as assessed by the fluorescent ALDEFLUOR assay has been successfully utilized to isolate cancer stem cells in multiple myeloma and acute 20 mycloid leukemia (AML) as well as from brain tumors (14-16). It was recently demonstrated that ALDH activity can be utilized to isolate a subpopulation of cells that display stein cell properties from normal human breast tissue and breast carcinomas (17). The ALDEFLUOR-positive population isolated from reduction miammoplasty tissue is able to reconstitute ductal alveolar structures in mammary fat pads of 25 humanized NOD/SCID mice. Furthermore, ALDELFUOR-positive cells isolated from human mammary carcinomas have stem cell properties as demonstrated by their ability to reconstitute tumors on serial passage in NOD/SCID mice as well as to generate the phenotypic heterogeneity of the initial tumors (17). In the Examples below, it is demonstrated that the majority of breast cancer cell lines contain an ALDEFLUOR 30 positive population with a distinct molecular profile that displays cancer stem cell properties. 23 As described in the Examples below, work conducted during the development of embodiments of the present invention identified CXCRI (which is a receptor for the inflammatory chemokine IL8) as a cancer stem cell marker. Only cells within the Aldefluor-positive population expressed CXCRI. Furthermore, it was demonstrated 5 that this receptor plays a functional role in that recombinant IL8 is able to increase the stern cell proportion in cell lines as determined by Aldefluor and sphere formation assays. Although IL8 has been reported to be associated with aggressive breast cancers and is higher in the serum women with metastatic disease, it is believe that the present invention is the first to show a functional link between IL8 and its receptor CXCRI in 10 stem cells. As further described in the Examples below, it was demonstrated that one can selectively target cancer stem cells by blocking the CXCRI receptor in these cells. In one approach described in the Examples, breast cancer cells lines were treated with monoclonal antibodies to CXCR1, but not to the other IL8 receptor CXCR2. Such 1s treatment selectively targeted cancer stem cells as demonstrated by reduced Aldefluor positive populations. Remarkably, it found that although CXCR1 is only expressed in a very small percentage of cells (e.g., less than 1%), that blockade of the CXCRI receptor induced cell death in the majority of other cancer cells despite the fact that they lack the CXCRI receptor. The molecular pathway which mediates the effects of 20 IL on cancer stem cells and accounts for this so-called "bystander effect" of killing other cells has been elucidated. IL8 stimulates stem cell self-renewal by binding to CXCRi, which in turn activates the focal adhesion kinase Fak pathway. This results in activation of Akt which drives stein cell self-renewal, When this pathway is blocked in cancer stem cells, the decrease in Akt signaling causes cytoplasmic sequestration of the 25 Foxo transcription factors resulting in an increased synthesis of Fas ligand. Fas ligand is secreted from cancer stem cells and induces cell death in surrounding cells which contain the Fas receptor. While the present invention is not limited to any particular mechanism, and an understanding of the mechanism is not necessary to practice the present invention, it is 30 believed that CXCRI mediates cancer stem cell self-renewal through a pathway involving Fak and Akt and that blockade of this pathway induces cell death in cancer 24 stein cells as well as surrounding tumor cells. As such, in certain embodiments, the present invention provides compositions and methods for disrupting the IL8-CXCR I pathway (e.g., with anti-CXCRI antibodies, anti-FAK antibodies, or other agents) in order to treat cancer. 5 Since IlL8 is a chemokine involved in tissue inflammation, there has been previous interest in developing inhibitors of IL8 signaling. A small molecule inhibitor, Repartaxin, has been developed as an anti-inflannatory agent to potentially reduce complications of inyocardial infarction and stroke. Repartaxin has been introduced into phase I and phase II clinical trials and has shown little toxicity. As shown in the 10 Examples below, Repartaxin (like anti-CXCR1 antibodies) is able to target cancer stem cells as well as to induce a Fas ligand fas mediated apoptosis by bystander effect in surrounding cells. Importantly, in tumor xenografts, Repartaxin potentiates the effect of chemotherapy. Furthermore, unlike chemotherapy, which preferentially destroys the differentiated cells in tumors sparing the tumor stein cells, Repartaxin is able to target 1s tumor stem cells. As shown in the examples, this was demonstrated by a decrease in the Aldefluor population in Repartaxin treated tumors and by the decrease in ability of these treated tumor cells to form secondary tumors in mice. Also tested was the effects of Repartaxin on the ability to block metastasis. Tumor cells were labeled with luciferase and injected intracardiac in an experimental metastasis model. One day after 20 the tumor cells were introduced, one group of animals was placed on repartaxin alone and the other no treatment. Repartaxin significantly reduced the development of metastasis. The present invention identified the IL8 receptor CXCR I as a target in treating cancer stem cells. The small molecule inhibitor Repartaxin inhibits both CXCRI and 25 CXCR2. The Examples demonstrated that it is CXCR1 that is the most important receptor in cancer stein cells. Furthermore, the Examples indicate that the failure of cytotoxic chemotherapy to affectively treat established cancers may be not only due to the inability of this therapy to target cancer stem cells, but in addition to the documented increase of IL8 secretion upon tumor cytotoxic chemotherapy treatment. 30 The present Examples indicate that the use of CXCRI inhibitors have beneficial effects 25 in being able to specifically target cancer stem cells as well as to block the IL8 stimulation of these cells induced by cytotoxic chemotherapy. Targeting the iL8-CXCRi pathway is not limited to breast cancer, but instead, can be employed in any type of cancer. Preferably, the type of cancer treated is one 5 where there is evidence of increased IL8 production (e.g., in conjunction with chemotherapy). Chernotherapy agents have been shown to directly regulate IL8 transcription in cancer cells. Paclitaxel increases IL8 transcription and secretion in ovarian, breast and lung cancer cell lines (UsIl et al., 2005, Int. J. Gynecol. Cancer, 15:240-245; and Collins et al., 2000, Can. Inm. Immuno., 49:78-84, both of which are 10 herein incorporated by reference). Also, administration of adriamycin and 5-fluoro-2' deoxvuridine to breast cancer cells (DeLarco et al., 2001, Can. Res. 61:2857-2861, herein incorporated by reference), the addition of 5-FU to oral cancer cells (Tamatani et al., 2004, Int., J. Can., 108:912:921, herein incorporated by reference), doxorubicin addition to small cell lung cancer cells (Shibakura et al., 2003, Int. J. Can., 103:380 15 386, herein incorporated by reference) and dacarbazine administration to melanoma cells (Lev et al., 2003, Mol, Can. Ther., 2:753-763. herein incorporated by reference) all result in increased CXCL8 expression. As such, in certain embodiments, the present invention provides agents for targeting the IL-CXCRT, in combination with a chemotherapy agents (e.g., such as those mentioned in the above references) for 20 treating a subject with a type of cancer including, but not limited to, prostate cancer, ovarian cancer, breast cancer, melanoma, non-small cell lung cancer, small-cell lung cancer, and esophageal adenocarcinoma. The present invention is not limited to the type of cancer treated and instead includes, but is not limited to, fibrosarcoma, myxosarcoma, liposarcomna, 25 chondrosarcoma, osteogenic sarcoma, chordomna, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lyniphangioendotheliosarcona, synovioma, mesothelioma, Ewing's tumor, leioniyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squarnous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, 30 papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatorna, bile duct 26 carcinoma, choriocarcinoia, seminoma, embryonal carcinoma, Wilms' tuminor, cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastomna, craniopharyngioma, ependynioma, pinealoma, hernangioblastoma, acoustic neuroma, oligodendroglioma, 5 nieningioma, melanoma, neuroblastoma, and retinoblastoma. IL Detection of Solid Tumor Stein Cell Cancer Markers In some embodiments, the present invention provides methods for detection of expression of stem cell cancer markers (e.g., CXCRi, FBXO21, NFYA, NOTCHI2, 10 RAD5 IL, TBIP, and other proteins from Table 1). In some embodiments, expression is measured directly (e.g., at the RNA or protein level). In son embodiments, expression is detected in tissue samples (e.g., biopsy tissue). In other embodiments, expression is detected in bodily fluids (e.g., including but not limited to, plasma, serum, whole blood, mucus, and urine). 15 The present invention further provides panels and kits for the detection of markers. In some embodiments, the presence of a stem cell cancer marker is used to provide a prognosis to a subject. The information provided is also used to direct the course of treatment. For example, if a subject is found to have a marker indicative of a solid tumor stem cell (e.g., CXCR1, FBXO21, NFYA, NOTCH2, RAD5 ILl, TBP, and 20 other proteins from Table 1), additional therapies (e.g., radiation therapies) can be started at an earlier point when they are more likely to be effective (e.g., before metastasis). In addition, if a subject is found to have a tumor that is not responsive to certain therapy, the expense and inconvenience of such therapies can be avoided. In some embodiments, the present invention provides a panel for the analysis of a 25 plurality of markers (e.g., the combination of CXCR I or FBX2 I and at least one of CD44, CD24, and ESA). The panel allows for the simultaneous analysis of multiple markers correlating with carcinogenesis and/or metastasis. Depending on the subject, panels can be analyzed alone or in combination in order to provide the best possible diagnosis and prognosis. Markers for inclusion on a panel are selected by screening for 30 their predictive value using any suitable method, including but not limited to, those described in the illustrative examples below. 27 1. Detection of R2NA In some embodiments, detection of solid tumor stem cell cancer markers are detected by measuring the expression of corresponding mRNA in a tissue sample. 5 mRNA expression can be measured by any suitable method, including but not limited to, those disclosed below. The accession nurnber for human CXCRI nucleic acid is NM 000634 (herein incorporated by reference) and the accession number for human FBXO2 I is NM 033624 (herein incorporated by reference). These sequences can be used to design primers and probes (as well as siRNA sequences). 10 In some embodiments, RNA is detected by Northern blot analysis. Northern blot analysis involves the separation of RNA and hybridization of a complementary labeled probe. In still further embodiments, RNA (or corresponding cDNA) is detected by hybridization to an oligonucleotide probe). A variety of hybridization assays using a 1s variety of technologies for hybridization and detection are available. For example, in some embodiments, T'aqMan assay (PE Biosystems, Foster City, CA; See e.g., U.S. Patent Nos. 5.962,233 and 5,538,848, each of which is herein incorporated by reference) is utilized. The assay is performed during a PCR reaction. The TaqMan assay exploits the 5'-3' exonuclease activity of the AMPLITAQ GOLD DNA polymerase. A probe 20 consisting of an oligonucleotide with a 5'-reporter dye (eg., a fluorescent dye) and a 3' quencher dye is included in the PCR reaction. During PCR, if the probe is bound to its target, the 5'-3' nucleolytic activity of the AMPLITAQ GOLD polymerase cleaves the probe between the reporter and the quencher dye. The separation of the reporter dye from the quencher dye results in an increase of fluorescence. The signal accumulates 25 with each cycle of PCR and can be monitored with a fluorimeter. In yet other embodiments, reverse-transcriptase PCR (RT-PCR) is used to detect the expression of RNA. In RT-PCR, RNA is enzymatically converted to complementary DNA or "cDNA" using a reverse transcriptase enzyme. The cDNA is then used as a template for a PCR reaction. PCR products can be detected by any suitable method, 30 including but not limited to, gel electrophoresis and staining with a DNA specific stain or hybridization to a labeled probe. In sore embodiments, the quantitative reverse 28 transcriptase PCR with standardized mixtures of competitive templates method described in U.S. Patents 5,639,606, 5,643;765, and 5,876,978 (each of which is herein incorporated by reference) is utilized. 5 2. Detection of Protein In other embodiments, gene expression of stem cell cancer markers is detected by measuring the expression of the corresponding protein or polypeptide (e.g., CXCRI, FBXO21, NFYA, NOTCH2, RAD51LI, TBP, and other proteins from Table 1). Protein expression can be detected by any suitable method. In some embodiments, proteins are 10 detected by immunohistochemistry. In other embodiments, proteins are detected by their binding to an antibody raised against the protein. The accession number for human CXCRI protein is NIP 000625 (herein incorporated by reference) and the accession number for human FBXO2l is NP 296373 (herein incorporated by reference). The generation of antibodies is described below. 15 Antibody binding is detected by techniques known in the art (e.g., radioimmunoassay, ELISA (enzyme linked immunosorbant assay), "sandwich" immunoassays, immunoradiometric assays, gel diffusion precipitation reactions, immunodiffusion assays, in situ immunoassays (e.g., using colloidal gold, enzyme or radioisotope labels, for example), Western blots, precipitation reactions, agglutination 20 assays (e.g., gel agglutination assays, hemagglutination assays, etc.), complement fixation assays, innnunofluorescence assays, protein A assays, and immunoelectrophoresis assays, etc. In one embodiment, antibody binding is detected by detecting a label on the primary antibody. In another embodiment, the primary antibody is detected by detecting 25 binding of a secondary antibody or reagent to the primary antibody. In a further embodiment, the secondary antibody is labeled. Many methods are known in the art for detecting binding in an immunoassay and are within the scope of the present invention. In some embodiments, an automated detection assay is utilized. Methods for the automation of imrnunoassavs include those described in U.S. Patents 5,885,530, 30 4,981,785, 6,159,750, and 5,358,691, each of which is herein incorporated by reference. In some embodiments, the analysis and presentation of results is also automated. For 29 example, in some embodiments, software that generates a prognosis based on the presence or absence of a series of proteins corresponding to cancer markers is utilized. In other embodiments, the immunoassay described in U.S. Patents 5,599,677 and 5,672,480; each of which is herein incorporated by reference. 5 3. cl)A-1 Microarray Technology cDNA inicroarrays are composed of multiple (usually thousands) of different cDNAs spotted (usually using a robotic spotting device) onto known locations on a solid support, such as a glass microscope slide. The cDNAs are typically obtained by PCR 10 amplification of plasmid library inserts using primers complementary to the vector backbone portion of the plasmid or to the gene itself for genes where sequence is known. PCR products suitable for production of microarrays are typically between 0.5 and 2.5 kB in length. Full length cDNAs, expressed sequence tags (ESTs), or randomly chosen cDNAs from any library of interest can be chosen. ESTs are partially sequenced cDN As 15 as described, for example, in Hillier, et al., 1996, 6:807-828. Although some ESTs correspond to known genes, frequently very little or no information regarding any particular EST is available except for a small amount of 3' and/or 5' sequence and, possibly, the tissue of origin of the mRNA from which the EST was derived. As will be appreciated by one of ordinary skill in the art, in general the eDNAs contain sufficient 20 sequence information to uniquely identify a gene within the human genome. Furthermore, in general the cDNAs are of sufficient length to hybridize, selectively, specifically or uniquely, to cDNA obtained from mRNA derived from a single gene under the hybridization conditions of the experiment. In a typical nicroarray experiment, a microarray is hybridized with differentially 25 labeled RNA, DNA, or cDNA populations derived from two different samples. Most commonly RNA (either total RNA or poly A+ RNA) is isolated from cells or tissues of interest and is reverse transcribed to yield cDNA. Labeling is usually performed during reverse transcription by incorporating a labeled nucleotide in the reaction mixture. Although various labels can be used, most commonly the nucleotide is conjugated with 30 the fluorescent dyes Cy3 or Cy5. For example, Cy5-dUTP and Cy3-dUTP can be used. cDNA derived from one sample (representing, for example, a particular cell type, tissue 30 type or growth condition) is labeled with one fluorophore while cDNA derived from a second sample (representing, for example, a different cell type, tissue type, or growth condition) is labeled with the second fluorophore. Similar amounts of labeled material from the two samples are cohybridized to the microarray. In the case of a microarray 5 experiment in which the samples are labeled with Cy5 (which fluoresces red) and Cy3 (which fluoresces green), the primary data (obtained by scanning the microarray using a detector capable of quantitatively detecting fluorescence intensity) are ratios of fluorescence intensity (red/green, RIG). These ratios represent the relative concentrations of cDNA molecules that hybridized to the cDNAs represented on the microarray and thus 10 reflect the relative expression levels of the mRNA corresponding to each cDNA/gene represented on the microarray. Each microarray experiment can provide tens of thousands of data points, each representing the relative expression of a particular gene in the two samples. Appropriate organization and analysis of the data is of key importance, and various computer 1s programs that incorporate standard statistical tools have been developed to facilitate data analysis. One basis for organizing gene expression data is to group genes with similar expression patterns together into clusters, A method for performing hierarchical cluster analysis and display of data derived from microarray experiments is described in Eisen et al., 1998, PNAS 95:14863-14868. As described therein, clustering can be combined with 20 a graphical representation of the primary data in which each data point is represented with a color that quantitatively and qualitatively represents that data point. By converting the data from a large table of numbers into a visual format, this process facilitates an intuitive analysis of the data. Additional information and details regarding the mathematical tools and/or the clustering approach itself can be found, for example, in 25 Sokal & Sneath, Principles of numerical taxonomy, xvi, 359, W. I. Freeman, San Francisco,1963; Hartigan, Clustering algorithms, xiii, 35-1, Wiley, New York, 1975; Paull et al., 1989, J. Nat]. Cancer Inst. 81:1088-92; Weinstein et al. 1992, Science 258:447-51; van Osdol et al., 1994, J. Natl. Cancer Inst. 86:1853-9; and Weinstein et al., 1997, Science, 275:343-9. 30 Further details of the experimental methods used in the present invention are found in the Example below. Additional information describing methods for fabricating 31 and using microarrays is found in U.S. Pat, No. 5,807,522, which is herein incorporated by reference. Instructions for constructing microarray hardware (e.g., arrayers and scanners) using commrercially available parts. Additional discussions of microarray technology and protocols for preparing samples and performing microrarray experiments 5 are found in, for example, DNA arrays for analysis of gene expression, Methods Enzymol, 303:179-205, 1999; Fluorescence-based expression monitoring using microarrays, Methods Enzyrnol, 306: 3-18, 1999; and M. Schena (ed.), DNA Microarrays: A Practical Approach, Oxford University Press, Oxford, UK, 1999. 10 4. Data Analysis In some embodiments, a computer-based analysis program is used to translate the raw data generated by the detection assay (e.g., the presence, absence, or amount of a Civen marker or markers) into data of predictive value for a clinician. The clinician can access the predictive data using any suitable means. Thus, in some embodiments, the 15 present invention provides the further benefit that the clinician, who is not likely to be trained in genetics or molecular biology, need not understand the raw data. The data is presented directly to the clinician in its most useful form. The clinician is then able to immediately utilize the information in order to optimize the care of the subject. The present invention contemplates any method capable of receiving, processing, 20 and transmitting the information to and from laboratories conducting the assays, information provides, medical personal, and subjects. For example, in some embodiments of the present invention, a sample (e.g., a biopsy or a serum or urine sample) is obtained from a subject and submitted to a profiling service (e.g., clinical lab at a medical facility, genomic profiling business, etc.), located in any part of the world 25 (e.g., in a country different than the country where the subject resides or where the information is ultimately used) to generate raw data. Where the sample comprises a tissue or other biological sample, the subject can visit a medical center to have the sample obtained and sent to the profiling center, or subjects can collect the sample themselves and directly send it to a profiling center. Where the sample comprises previously 30 determined biological information, the information can be directly sent to the profiling service by the subject (e.g., an information card containing the information can be 32 scanned by a computer and the data transmitted to a computer of the profiling center using an electronic communication system). Once received by the profiling service, the sample is processed and a profile is produced (e.g., expression data), specific for the diagnostic or prognostic information desired for the subject. 5 The profile data is then prepared in a format suitable for interpretation by a treating clinician. For example, rather than providing raw expression data (e.g. examining a number of the markers), the prepared format can represent a diagnosis or risk assessment for the subject, along with recommendations for particular treatment options. The data can be displayed to the clinician by any suitable method. For example, 10 in some embodiments, the profiling service generates a report that can be printed for the clinician (e.g., at the point of care) or displayed to the clinician on a computer monitor. In some embodiments, the information is first analyzed at the point of care or at a regional facility. The raw data is then sent to a central processing facility for further analysis and/or to convert the raw data to information useful for a clinician or patient. 15 The central processing facility provides the advantage of privacy (all data is stored in a central facility with uniform security protocols), speed, and uniformity of data analysis. The central processing facility can then control the fate of the data following treatment of the subject. For example, using an electronic communication system, the central facility can provide data to the clinician, the subject, or researchers. 20 In some embodiments, the subject is able to directly access the data using the electronic communication system. The subject can chose further intervention or counseling based on the results. In some einbodimenits, the data is used for research use. For example, the data can be used to further optimize the inclusion or elimination of markers as useful indicators of a particular condition or stage of disease. 25 5, Kits In yet other embodiments, the present invention provides kits for the detection and characterization of cancer (e.g. for detecting one or more of the markers, or for modulating the activity of a peptide expressed by one or more of markers). In sore 30 embodiments, the kits contain antibodies specific for a cancer marker, in addition to detection reagents and buffers. In other embodiments, the kits contain reagents specific 33 for the detection of mRNA or cDNA (e.g., oligonucleotide probes or primers). In some embodiments, the kits contain all of the components necessary and/or sufficient to perform a detection assay, including all controls, directions for performing assays, and any necessary software for analysis and presentation of results. 5 Another embodiment of the present invention comprises a kit to test for the presence of the polynucleotides or proteins. The kit can comprise, for example, an antibody for detection of a polypeptide or a probe for detection of a polynucleotide. In addition, the kit can comprise a reference or control sample; instructions for processing samples, performing the test and interpreting the results; and buffers and other reagents 10 necessary for performing the test. In other embodiments the kit comprises pairs of primers for detecting expression of one or more of the genes of the solid tumor stem cell gene signature. In other embodiments the kit comprises a cDNA or oligonucleotide array for detecting expression of one or more of the genes of the solid tumor stem cell gene signature. 15 6. In viv Imaging In some embodiments, in vivo imaging techniques are used to visualize the expression of cancer markers in an animal (e.g., a human or non-human mammal). For example, in some embodiments, cancer marker mRNA (e.g., CXCRl or FBXO2 1 20 mRNA) or protein (e.g., CXCRi or FBXO2I protein) is labeled using a labeled antibody specific for the cancer marker. A specifically bound and labeled antibody can be detected in an individual using an in vivo imaging method, including, but not limited to, radionuclide imaging, positron emission tomography, computerized axial tomography, X ray or magnetic resonance imaging method, fluorescence detection, and 25 chemiluminescent detection. Methods for generating antibodies to the cancer markers of the present invention are described below. The in vivo imaging methods of the present invention are useful in the diagnosis of cancers that express the solid tumor stem cell cancer markers of the present invention. In vivo imaging is used to visualize the presence of a marker indicative of the cancer. 30 Such techniques allow for diagnosis without the use of an unpleasant biopsy. The in vivo imaging methods of the present invention are also useful for providing prognoses to 34 cancer patients. For example, the presence of a marker indicative of cancer stein cells can be detected. The in vivo imaging methods of the present invention can further be used to detect metastatic cancers in other parts of the body. In some embodiments, reagents (e.g., antibodies) specific for CXCR I or FBXO21 5 are fluorescently labeled. The labeled antibodies are introduced into a subject (e.g., orally or parenterally). Fluorescently labeled antibodies are detected using any suitable method (e.g., using the apparatus described in U.S. Patent 6,198,107, herein incorporated by reference). In other embodiments, antibodies are radioactively labeled. The use of antibodies 10 for in vivo diagnosis is well known in the art. Sumerdon et al., (Nucl. Med. Biol 17:247 254 [19901 have described an optimized antibody-chelator for the radioimmunoscintographic imaging of tumors using Indium-I II as the label. Griffin et al., (J Clin One 9:631-640 [1991]) have described the use of this agent in detecting tumors in patients suspected of having pancreatic cancer. The use of similar agents with 15 paramagnetic ions as labels for magnetic resonance imaging is known in the art (Lauffer, Magnetic Resonance in Medicine 22:339-342 [1991]). The label used will depend on the imaging modality chosen. Radioactive labels such as Indium-111, Technetium-99m, or Iodine- 131 can be used for planar scans or single photon emission computed tomography (SPECT). Positron emitting labels such as Fluorine-19 can also be used for positron 20 emission tomography (PET). For MR, paramagnetic ions such as Gadolinium (III) or Manganese (II) can be used. Radioactive metals with half-lives ranging from I hour to 3.5 days are available for conjugation to antibodies, such as scandium-47 (3.5 days) gallium-67 (2.8 days), gallium-68 (68 minutes), technetiium-99mn (6 hours), and indium-i 11 (3.2 days), of 25 which gallium-67, technetium-99n, and indium- 1 1 are preferable for gammnna camera imaging, gallium-68 is preferable for positron emission tomography. A useful method of labeling antibodies with such radiomnetals is by means of a bifunctional chelating agent, such as diethylenetriamninepentaacetic acid (IDTPA), as described, for example, by Khaw et al. (Science 209:295 [1980]) for In-i II and Tc-99m, 30 and by Scheinberg et al. (Science 215:1511 [1982]). Other chelating agents can also be used, but the 1 -(p-carboxymethoxybenzyl)EDTA and the carboxycarbonic anhydride of 35 DTPA are advantageous because their use permits conjugation without affecting the antibody's immunoreactivity substantially. Another method for coupling DPTA to proteins is by use of the cyclic anhydride of DTPA, as described by Hnatowich et al. (Int. J. Apple. Radiat. Isot. 33:327 [1982]) for 5 labelino of albumin with In- 111, but which can be adapted for labeling of antibodies. A suitable method of labeling antibodies with Tc-99m which does not use chelation with DPTA is the pretinning method of Crockford et al., (U.S. Pat. No. 4,323,546, herein incorporated by reference). A method of labeling immunoglobulins with Tc-99m is that described by Wong et 10 al. (Int. J. Appl. Radiat. Isot., 29:251 [1978]) for plasma protein, and recently applied successfully by Wong et al. (J. Nucl. Med., 23:229 [1981]) for labeling antibodies. In the case of the radiometals conjugated to the specific antibody, it is likewise desirable to introduce as high a proportion of the radiolabel as possible into the antibody molecule without destroying its immunospecificity. A further improvement can be 15 achieved by effecting radiolabeling in the presence of the specific stem cell cancer marker of the present invention, to insure that the antigen binding site on the antibody will be protected. In still further embodiments, in vivo biophotonic imaging (Xenogen, Almeda, CA) is utilized for in vivo imaging. This real-time in vivo imaging utilizes luciferase, 20 The luciferase gene is incorporated into cells, microorganisms, and animals (e.g., as a fusion protein with a cancer marker of the present invention). When active, it leads to a reaction that emits light. A CCD camera and software is used to capture the image and analyze it. 25 IML Antibodies and Antibody Fragments The present invention provides isolated antibodies and antibody fragments against CXCR1, FBX021, NFYA, NOTCTI2, RAD51L1, TBP, and other proteins from Table 1. The antibody, or antibody fragment, can be any monoclonal or polyclonal antibody that specifically recognizes these proteins. In some embodiments, the present invention 30 provides monoclonal antibodies, or fragments thereof, that specifically bind to CXCRi, FBXO21, NFYA, NOTCH2, RAD51L 1, TBP, and other proteins from Table 1. In some 36 embodiments, the monoclonal antibodies, or fragments thereof, are chimeric or humanized antibodies that specifically bind to these proteins. In other embodiments, the monoclonal antibodies, or fragments thereof, are human antibodies that specifically bind to these proteins. 5 The antibodies against CXCR1, FBXO21, NFYA, NOTCHI2, RAD5 IL1, TBP, and other proteins from Table 1 find use in the experimental, diagnostic and therapeutic methods described herein. In certain embodiments, the antibodies of the present invention are used to detect the expression of a cancer stern cell marker protein in biological samples such as, for example, a patient tissue biopsy, pleural effusion, or blood 10 sample. Tissue biopsies can be sectioned and protein detected using, for example, immunofluorescence or immunohistochemistry. Alternatively, individual cells from a sample are isolated, and protein expression detected on fixed or live cells by FACS analysis. Furthermore, the antibodies can be used on protein arrays to detect expression of a cancer stem cell marker, for example, on tumor cells, in cell lysates, or in other 1s protein samples. In other embodiments, the antibodies of the present invention are used to inhibit the growth of tumor cells by contacting the antibodies with tumor cells either in vitro cell based assays or in vivo animal models. In still other embodiments, the antibodies are used to treat cancer in a human patient by administering a therapeutically effective amount of an antibody against a cancer stem cell marker (e.g., from Table 1). 20 Polyclonal antibodies can be prepared by any known method. Polyclonal antibodies can be raised by immunizing an animal (e.g. a rabbit, rat, mouse, donkey, etc) by multiple subcutaneous or intraperitoneal injections of the relevant antigen (a purified peptide fragment, full-length recombinant protein, fusion protein, etc) optionally conjugated to keyhole limpet hemocyanin (K11), serum albumin, etc. diluted in sterile 25 saline and combined with an adjuvant (e.g. Complete or Incomplete Freund's Adjuvant) to form a stable emulsion. The polyclonal antibody is then recovered from blood, ascites and the like, of an animal so immunized. Collected blood is clotted, and the serum decanted, clarified by centrifugation, and assayed for antibody titer. The polyclonal antibodies can be purified from serum or ascites according to standard methods in the art 30 including affinity chromatography, ion-exchange chromatography, gel electrophoresis, dialysis, etc. 37 Monoclonal antibodies can be prepared using hybridoma methods, such as those described by Kohler and Milstein (1975) Nature 256:495. Using the hybridoma method, a mouse, hamster, or other appropriate host animal, is immunized as described above to elicit the production by lymphocytes of antibodies that will specifically bind to an 5 immunizing antigen. Alternatively, lymphocytes can be immunized in vitro. Following immunization, the lymphocytes are isolated and fused with a suitable myelorna cell line using, for example, polyethylene glycol, to form hybridoma cells that can then be selected away from unfused lymphocytes and myeloma cells. H-ybridomas that produce monoclonal antibodies directed specifically against a chosen antigen as determined by 10 immunoprecipitation, immunoblotting, or by an in vitro binding assay such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA) can then be propagated either in vitro culture using standard methods (Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, 1986) or in vivo as ascites tumors in an animal. The monoclonal antibodies can then be purified from the culture medium 15 or ascites fluid as described for polyclonal antibodies above. Alternatively monoclonal antibodies can also be made using recombinant DNA methods as described in U.S. Patent 4,816,567. The polynucleotides encoding a monoclonal antibody are isolated, such as from mature B-cells or hybridoma cell, such as by RT-PCR using oligonucleotide primers that specifically amplify the genes encoding 20 the heavy and light chains of the antibody, and their sequence is determined using conventional procedures. The isolated polynucleotides encoding the heavy and light chains are then cloned into suitable expression vectors, which when transfected into host cells such as E. coli cells, simian COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells that do not otherwise produce immunoglobulin protein, monoclonal 25 antibodies are generated by the host cells. Also, recombinant monoclonal antibodies or fragments thereof of the desired species can be isolated from phage display libraries as described (McCafferty et al., 1990, Nature, 348:552-554; Clackson et al., 1991, Nature, 352:624-628; and Marks et al., 1991, 1. Mol. Biol., 222:581-597). The polynucleotide(s) encoding a monoclonal antibody can further be modified in 30 a number of different rmanners using recombinant DNA technology to generate alternative antibodies. In one embodiment, the constant domains of the light and heavy 38 chains of, for example, a mouse monoclonal antibody can be substituted 1) for those regions of, for example, a human antibody to generate a chimeric antibody or 2) for a non-immurnoglobulin polypeptide to generate a fusion antibody. In other eibodirnents, the constant regions are truncated or removed to generate the desired antibody fragment 5 of a monoclonal antibody. Furthermore, site-directed or high-density mutagenesis of the variable region can be used to optimize specificity, affinity, etc. of a monoclonal antibody. In some embodiments, of the present invention the monoclonal antibody against a cancer stem cell marker is a humanized antibody. Humanized antibodies are antibodies 10 that contain minimal sequences from non-human (e.g., murine) antibodies within the variable regions. Such antibodies are used therapeutically to reduce antigenicity and HAMA (human anti-mouse antibody) responses when administered to a human subject. In practice, humanized antibodies are typically human antibodies with minimum to no non-human sequences. A human antibody is an antibody produced by a human or an 1s antibody having an amino acid sequence corresponding to an antibody produced by a human. Humanized antibodies can be produced using various techniques known in the art. An antibody can be humanized by substituting the CDR of a human antibody with that of a non-human antibody (e.g. mouse, rat, rabbit, hamster, etc.) having the desired 20 specificity, affinity, and capability (Jones et al., 1986. Nature, 321:522-525; Riechmann et al., 1988, Nature, 332:323-327; Verhoeyen et al., 1988, Science, 239:1534-1536). The humanized antibody can be further modified by the substitution of additional residue either in the Fv framework region and/or within the replaced non-human residues to refine and optimize antibody specificity, affinity, and/or capability. 25 Human antibodies can be directly prepared using various techniques known in the art. Immortalized human B lymphocytes immunized in vitro or isolated from an immunized individual that produce an antibody directed against a target antigen can be generated (See, for example, Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, p. 77 (1985); Boemer et al., 1991, .1. Immunol., 147 (1):86-95; and U.S. 30 Patent 5,750,373). Also, the human antibody can be selected frorn a phage library, where that phage library expresses human antibodies (Vaughan et al., 1996, Nature 39 Biotechnology, 14:309-314; Sheets et al., 1998, PNAS, 95:6157-6162; Hoogenboom and Winter, 1991, J. Mol. Biol., 227:381; Marks et al., 1991, J. Mol. Biol., 222:581). Humanized antibodies can also be made in transgenic mice containing human imrunoglobulin loci that are capable upon immunization of producing the fill repertoire 5 of human antibodies in the absence of endogenous immunoglobulin production. This approach is described in U.S. Patents 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; and 5,661,016. This invention also encompasses bispecific antibodies that specifically recognize cancer stem cell markers. Bispecific antibodies are antibodies that are capable of 10 specifically recognizing and binding at least two different epitopes. Bispecific antibodies can be intact antibodies or antibody fragments. Techniques for making bispecific antibodies are common in the art (Millstein et al., 1983, Nature 305:537-539; Brennan et al., 1985, Science 229:81: Suresh et al, 1986, Methods in Enzymol. 121:120; Traunecker et al., 1991, EMBO J. 10:3655-3659; Shalby et al., 1992. 15 J. Exp. Med. 175:217-225; Kostelny et al., 1992, J. Immunol. 148:1547-1553; Gruber et al., 1994, J. Immunol. 152:5368; and U.S. Patent 5,731,168). In certain embodiments of the invention, it may be desirable to use an antibody fragment, rather than an intact antibody, to increase tumor penetration, for example. Various techniques are known for the production of antibody fragments. Traditionally, 20 these fragments are derived via proteolytic digestion of intact antibodies (for example Morimnoto et al., 1993, Journal of Biochemical and Biophysical Methods 24:107-117 and Brennan ei al., 1985, Science, 229:81). However, these fragments are now typically produced directly by recombinant host cells as described above. Thus Fab, Fv, and scFv antibody fragments can all be expressed in and secreted from E. coli or other host cells, 25 thus allowing the production of large amounts of these fragments. Alternatively, such antibody fragments can be isolated from the antibody phage libraries discussed above. The antibody fragment can also be linear antibodies as described in U.S. Patent 5,641,870, for example, and can be monospecific or bispecific. Other techniques for the production of antibody fragments will be apparent to the skilled practitioner. 30 It may further be desirable, especially in the case of antibody fragments, to modify an antibody in order to increase its serurn half-life. This can be achieved, for 40 example, by incorporation of a salvage receptor binding epitope into the antibody fragment by mutation of the appropriate region in the antibody fragment or by incorporating the epitope into a peptide tag that is then fused to the antibody fragment at either end or in the middle (e.g., by DNA or peptide synthesis). 5 The present invention further embraces variants and equivalents which are substantially homologous to the chimeric, humanized and hurnan antibodies, or antibody fragments thereof, set forth herein. These can contain, for example, conservative substitution mutations, i.e. the substitution of one or more arnino acids by similar amino acids. For example, conservative substitution refers to the substitution of an amino acid 10 with another within the same general class such as, for example, one acidic amino acid with another acidic amino acid, one basic amino acid with another basic amino acid or one neutral amino acid by another neutral amino acid. What is intended by a conservative amino acid substitution is well known in the art. The invention also pertains to inununoconjugates comprising an antibody 15 conjugated to a cytotoxic agent. Cytotoxic agents include chemotherapeutic agents, growth inhibitory agents, toxins (e.g., an enzymatically active toxin of bacterial, fungal, plant, or animal origin, or fragments thereof), radioactive isotopes (i.e., a radioconjugate), etc. Chemotherapeutic agents useful in the generation of such immunoconjugates include, for example, methotrexate, adriamicin, doxorubicin, melphalan, mitomycin C, 20 chlorambucil, daunorubicin or other intercalating agents. Enzymatically active toxins and fragments thereof that can be used include diphtheria A chain, nonbinding active fTagments of diphtheria toxin, exotoxin A chain, ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii proteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAPII, and PAP-S), momordica charantia inhibitor, curcin, crotin, 25 sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenoinycin, enomycin, and the tricothecenes. A variety of radionuclides are available for the production of radioconjugated antibodies including 212Bi, 1311, 13 In, 90Y, and 186Re. Conjugates of the antibody and cytotoxic agent are made using a variety of bifunctional protein coupling agents such as N-succinimidyl-3-(2-pyridyidithiol) propionate (SPDP), 30 iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as 41 glutareldehyde), bis-azido compounds (such as bis(p-azidobenzoyi) hexanediamine), bis diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediarine), diisocyanates (such as tolvene 2,6-diisocyanate), and bis-active fluorine compounds (such as 1,5-difluoro-2,4-dinitrobenzene). Conjugates of an antibody arid one or more 5 small molecule toxins, such as a calicheamicin, naytansinoids, a trichothene, and CCl 065, and the derivatives of these toxins that have toxin activity, can also be used. In some embodiments the antibody of the invention contains human Fe regions that are modified to enhance effector function, for example, antigen-dependent cell mediated cyotoxicity (ADCC) and/or complement dependent cytotoxicity (CDC). This 10 can be achieved by introducing one or more amino acid substitutions in an Fe region of the antibody. For example, cysteine residues) can be introduced in the Fe region to allow interchain disulfide bond formation in this region to improve complement-mediated cell killing and antibody-dependent cellular cytotoxicity (ADCC) (Caron et al., 1992, J. Exp Med. 176:1191-1195; Shopes, 1992, Immunol. 148:2918-2922). Homodimeric 1s antibodies with enhanced anti-tumor activity can also be prepared using heterobifunctional cross-linkers as described in Wolff et al., 1993, Cancer Research 53:2560-2565. Alternatively, an antibody can be engineered which has dual Fe regions (Stevenson et al., 1989, Anti-Cancer Drug Design 3:219-230). 20 IV. Drug Screening In some embodiments, the present invention provides drug screening assays (e.g., to screen for anticancer drugs). The screening methods of the present invention utilize stem cell cancer markers (e.g., CXCR1, FBXO2I1, NFYA, NOTCHI2, RAD51L1, TBP, and other proteins from Table 1) identified using the methods of the present invention. 25 For example, in some embodiments, the present invention provides methods of screening for compounds that alter (e.g., increase or decrease) the expression of, or activity of, CXCR I or FBX02 1. In some embodiments, candidate compounds are antisense agents or siRNA agents (e.g., oligonucleotides) directed against cancer markers. In other embodiments, candidate compounds are antibodies that specifically bind to a stem cell 30 cancer marker of the present invention. In certain embodiments, libraries of compounds of small molecules are screened using the methods described herein. 42 In one screening method, candidate compounds are evaluated for their ability to alter stein cell cancer marker expression by contacting a compound with a cell expressing a stem cell cancer marker and then assaying for the effect of the candidate compounds on expression. In some embodiments, the effect of candidate compounds on expression of a 5 cancer marker gene is assayed by detecting the level of cancer marker mRNA expressed by the cell. mRNA expression can be detected by any suitable rnethod. In other embodiments, the effect of candidate compounds on expression of cancer marker genes is assayed by measuring the level of polypeptide encoded by the cancer markers. The level of polypeptide expressed can be measured using any suitable method, including but not 10 limited to, those disclosed herein. In some embodiments, other changes in cell biology (e.g., apoptosis) are detected. Specifically, the present invention provides screening methods for identifying modulators, i.e., candidate or test compounds or agents (e.g., proteins, peptides, peptidomimetics, peptoids, small molecules or other drugs) which bind to, or alter the 15 signaling or function associated with the cancer markers of the present invention, have an inhibitory (or stimulatory) effect on, for example, stem cell cancer marker expression or cancer markers activity, or have a stimulatory or inhibitory effect on, for example, the expression or activity of a cancer marker substrate. Compounds thus identified can be used to modulate the activity of target gene products (e.g., stem cell cancer marker genes, 20 such as CXCRI or FBXO21) either directly or indirectly in a therapeutic protocol, to elaborate the biological function of the target gene product, or to identify compounds that disrupt normal target gene interactions. Compounds which inhibit the activity or expression of cancer markers are useful in the treatment of proliferative disorders, e.g., cancer, particularly mnetastatic cancer or eliminating or controlling tumor stem cells to 25 prevent or reduce the risk of cancer. In one embodiment, the invention provides assays for screening candidate or test compounds that are substrates of a cancer markers protein or polypeptide or a biologically active portion thereof. In another embodiment, the invention provides assays for screening candidate or test compounds that bind to or modulate the activity of a 30 cancer marker protein or polypeptide or a biologically active portion thereof. 43 The test compounds of the present invention can be obtained using any of the numerous approaches in combinatorial library methods known in the art, including biological libraries; peptoid libraries (libraries of molecules having the functionalities of peptides, but with a novel, non-peptide backbone, which are resistant to enzymatic 5 degradation but which nevertheless remain bioactive; see, e.g., Zuckennann et al., J. Med. Chern. 37: 2678-85 [1994]); spatially addressable parallel solid phase or solution phase libraries; synthetic library rnethods requiring deconvolution; the 'one-bead one compound' library method; and synthetic library rnethods using affinity chromatography selection. The biological library and peptoid library approaches are preferred for use 10 with peptide libraries, while the other four approaches are applicable to peptide, non peptide oligomer or small molecule libraries of compounds (Lam (1997) Anticancer Drug Des. 12:145). Examples of methods for the synthesis of molecular libraries can be found in the art, for example in: DeWitt et al., Proc. Natl. Acad. Sci. U.S.A. 90:6909 [1993]; Erb et 15 al., Proc. Nad. Acad. Sci. USA 91:11422 [1994]; Zuckermann et al., J. Med. Chem. 37:2678 [1994]; Cho et al,, Science 261:1303 [1993]; Carrell et al., Angew. Chem. Int. Ed. Engl. 33.2059 [1994]; Carell et al., Angew. Chem, Iit. Ed. Engl. 33:2061 [1994]; and Gallop et al., J. Med. Chem. 37:1233 [1994]. Libraries of compounds can be presented in solution (e.g., Houghten, 20 Biotechniques 13:412-421 [1992]), or on beads (Lam, Nature 354:82-84 [1991]), chips (Fodor, Nature 364:555-556 [19931), bacteria or spores (U.S. Patent No. 5,223,409; herein incorporated by reference), plasmids (Cull et al., Proc. Nad. Acad. Sci. USA 89:18651869 [1992]) or on phage (Scott and Smith, Science 249:386-390 [1990]; Devlin Science 249:404-406 [1990]; Cwirla et al., Proc. Nati. Acad. Sci. 87:6378-6382 [1990]; 25 Felici, J. Mol. Biol. 222:301 [991]) In one embodiment, an assay is a cell-based assay in which a cell that expresses a stem cell cancer marker protein or biologically active portion thereof is contacted with a test compound, and the ability of the test compound to the modulate cancer marker's activity is determined. Determining the ability of the test compound to modulate sten 30 cell cancer marker activity can be accomplished by monitoring, for example, changes in enzymatic activity. The cell, for example, can be of mammalian origin. 44 The ability of the test compound to modulate cancer marker binding to a compound, e.g., a stem cell cancer marker substrate, can also be evaluated. This can be accomplished, for example, by coupling the compound, e.g., the substrate, with a radioisotope or enzymatic label such that binding of the compound, e.g., the substrate, to 5 a cancer marker can be determined by detecting the labeled compound, e.g., substrate, in a complex. Alternatively, the stem cell cancer marker is coupled with a radioisotope or enzymatic label to monitor the ability of a test compound to modulate cancer marker binding to a cancer markers substrate in a complex. For example, compounds (e.g., 10 substrates) can be labeled with 1251, 35S 14C or 311, either directly or indirectly, and the radioisotope detected by direct counting of radioemmission or by scintillation counting. Alternatively, compounds can be enzymatically labeled with, for example, horseradish peroxidase, alkaline phosphatase, or luciferase, and the enzymatic label detected by determination of conversion of an appropriate substrate to product. 15 The ability of a compound (e.g., a stein cell cancer marker substrate) to interact with a stem cell cancer marker with or without the labeling of any of the interactants can be evaluated. For example, a microphysiorneter can be used to detect the interaction of a compound with a cancer marker without the labeling of either the compound or the cancer marker (McConnell et al. Science 257:1906-1912 [1992]). As used herein, a 20 "microphysiometer" (e.g., Cytosensor) is an analytical instrument that measures the rate at which a cell acidifies its environment using a light-addressable potentioinetric sensor (LAPS). Changes in this acidification rate can be used as an indicator of the interaction between a compound and cancer markers. In yet another embodiment, a cell-free assay is provided in which a cancer marker 25 protein or biologically active portion thereof is contacted with a test compound and the ability of the test compound to bind to the stem cell cancer marker protein or biologically active portion thereof is evaluated. Biologically active portions of the cancer markers proteins to be used in assays of the present invention include fragments that participate in interactions with substrates or other proteins, e.g., fragments with high surface 30 probability scores. 45 Cell-free assays involve preparing a reaction mixture of the target gene protein and the test compound under conditions and for a time sufficient to allow the two components to interact and bind, thus forming a complex that can be removed and/or detected. 5 The interaction between two molecules can also be detected, e.g., using fluorescence energy transfer (FRET) (see, for example, Lakowicz et al., U.S. Patent No. 5,631,169; Stavrianopoulos et al., U.S. Patent No. 4,968,103; each of which is herein incorporated by reference). A fluorophore label is selected such that a first donor molecule's emitted fluorescent energy will be absorbed by a fluorescent label on a 10 second, 'acceptor' molecule, which in turn is able to fluoresce due to the absorbed energy. Alternately, the 'donor' protein molecule can simply utilize the natural fluorescent energy of tryptophan residues. Labels are chosen that emit different wavelengths of light, such that the 'acceptor' molecule label can be differentiated from that of the 'donor'. Since the efficiency of energy transfer between the labels is related to the distance separating 15 the molecules, the spatial relationship between the molecules can be assessed. In a situation in which binding occurs between the molecules, the fluorescent emission of the 'acceptor' molecule label in 1 5 the assay should be maximal. An FRET binding event can be conveniently measured through standard fluorometric detection means well known in the art (e.g., using a fluorimeter). 20 In another embodiment, determining the ability of the stem cell cancer markers protein to bind to a target molecule can be accomplished using real-time Biomolecular Interaction Analysis (BIA) (see, e.g., Sjolander and Urbaniczky, Anal. Chem. 63:2338 2345 [1991] and Szabo et al. Curr. Opin. Struct. Biol. 5:699-705 [1995]). "Surface plasmon resonance" or "BIA" detects biospecific interactions in real time, without 25 labeling any of the interactants (e.g., BlAcore). Changes in the mass at the binding surface (indicative of a binding event) result in alterations of the refractive index of light near the surface (the optical phenomenon of surface plasmuon resonance (SPR)), resulting in a detectable signal that can be used as an indication of real-time reactions between biological molecules. 30 In one embodiment, the target gene product or the test substance is anchored onto a solid phase. The target gene product/test compound complexes anchored on the solid 46 phase can be detected at the end of the reaction. The target gene product can be anchored onto a solid surface, and the test compound, (which is not anchored), can be labeled, either directly or indirectly, with detectable labels discussed herein. It may be desirable to immobilize stem cell cancer markers, an anti-cancer marker 5 antibody or its target molecule to facilitate separation of complexed from non-cornplexed forms of one or both of the proteins, as well as to accommodate automation of the assay. Binding of a test compound to a stem cell cancer marker protein, or interaction of a cancer marker protein with a target molecule in the presence and absence of a candidate compound, can be accomplished in any vessel suitable for containing the reactants. 10 Examples of such vessels include microtiter plates, test tubes, and micro-centriftige tubes. In one embodiment, a fusion protein can be provided which adds a domain that allows one or both of the proteins to be bound to a matrix. For example, glutathione-S transferase-cancer marker fusion proteins or glutathione-S-transferase/target fusion proteins can be adsorbed onto glutathione Sepharose beads (Sigma Chemical, St. Louis, 15 MO) or glutathione-derivatized microtiter plates, which are then combined with the test compound or the test compound and either the non-adsorbed target protein or cancer marker protein, and the mixture incubated under conditions conducive for complex formation (e.g., at physiological conditions for salt and pH). Following incubation, the beads or microtiter plate wells are washed to remove any unbound components, the 20 matrix immobilized in the case of beads, complex determined either directly or indirectly, for example, as described above. Alternatively, the complexes can be dissociated from the matrix, and the level of cancer markers binding or activity determined using standard techniques. Other techniques for immobilizing either cancer markers protein or a target molecule on 25 matrices include using conjugation of biotin and streptavidin, Biotinylated cancer marker protein or target molecules can be prepared from biotin-NIS (N-hydroxy-succinimide) using techniques known in the art (e.g., biotinylation kit, Pierce Chemicals, Rockford, IL), and immobilized in the wells of streptavi din-coated 96 well plates (Pierce Chernical). In order to conduct the assay, the non-imrnmobilized component is added to the 30 coated surface containing the anchored component. After the reaction is complete, unreacted components are removed (e.g., by washing) under conditions such that any 47 complexes formed will remain immobilized on the solid surface. The detection of complexes anchored on the solid surface can be accomplished in a number of ways. Where the previously non-immobilized component is pre-labeled, the detection of label immobilized on the surface indicates that complexes were formed. Where the previously 5 non-imnobilized component is not pre-labeled, an indirect label can be used to detect complexes anchored on the surface; e.g., using a labeled antibody specific for the immobilized component (the antibody, in turn, can be directly labeled or indirectly labeled with, e.g., a labeled anti-lIgG antibody). This assay is performed utilizing antibodies reactive with stem cell cancer marker 10 protein or target molecules but which do not interfere with binding of the stem cell cancer markers protein to its target molecule. Such antibodies can be derivatized to the wells of the plate, and unbound target or cancer markers protein trapped in the wells by antibody conjugation. Methods for detecting such complexes, in addition to those described above for the GST-immobilized complexes, include immunodetection of complexes using 15 antibodies reactive with the cancer marker protein or target molecule, as well as enzyme linked assays which rely on detecting an enzymatic activity associated with the cancer marker protein or target molecule. Alternatively, cell free assays can be conducted in a liquid phase. In such an assay, the reaction products are separated from unreacted components, by any of a 20 number of standard techniques, including, but not limited to: differential centrifugation (see, for example, Rivas and Minton, Trends Biochem Sci 18:284-7 [19931); chromatography (gel filtration chromatography, ion-exchange chromatography); electrophoresis (see, e.g., Ausubel et al., eds. Current Protocols in Molecular Biology 1999, J. Wiley: New York.); and immunoprecipitation (see, for example, Ausubel et al., 25 eds. Current Protocols in Molecular Biology 1999, J. Wiley: New York). Such resins and chromatographic techniques are known to one skilled in the art (See e.g., Heegaard J. Mol. Recognit 11:141-8 [1998]; lageand Tweed J. Chromnatogr. Biomned. Sci. AppI 699:499-525 [1997]). Further, fluorescence energy transfer can also be conveniently utilized, as described herein, to detect binding without further purification of the complex 30 from solution. 48 The assay can include contacting the stein cell cancer markers protein or biologically active portion thereof with a known compound that binds the cancer marker to form an assay mixture, contacting the assay mixture with a test compound, and determining the ability of the test compound to interact with a cancer marker protein, 5 wherein determining the ability of the test compound to interact with a cancer marker protein includes determining the ability of the test compound to preferentially bind to cancer markers or biologically active portion thereof, or to modulate the activity of a target molecule, as compared to the known compound. To the extent that stein cell cancer markers can, in vivo, interact with one or more 10 cellular or extracellular macromolecules, such as proteins, inhibitors of such an interaction are useful. A homogeneous assay can be used can be used to identify inhibitors. For example, a preformed complex of the target gene product and the interactive cellular or extracellular binding partner product is prepared such that either the target 1s gene products or their binding partners are labeled, but the signal generated by the label is quenched due to complex formation (see, e.g., U.S. Patent No. 4,109,496, herein incorporated by reference, that utilizes this approach for immunoassays). The addition of a test substance that competes with and displaces one of the species from the preformed complex will result in the generation of a signal above background. In this way, test 20 substances that disrupt target gene product-binding partner interaction can be identified. Alternatively, cancer markers protein can be used as a "bait protein" in a two-hybrid assay or three-hybrid assay (see, eg., T.S. Patent No. 5,283,317; Zervos et al. Cell 72:223-232 [1993]; Madura et al., J. Biol. Chem. 268,12046-12054 [19931; Bartel et al. Biotechniques 14:920-924 [1993]; iwabuchi et al., Oncogene 8:1693-1696 [1993]; and 25 Brent WO 94/10300; each of which is herein incorporated by reference), to identify other proteins, that bind to or interact with cancer markers ("cancer marker-binding proteins" or "cancer marker-bp") and are involved in cancer marker activity. Such cancer marker-bps can be activators or inhibitors of signals by the cancer marker proteins or targets as, for example, downstream elements of a cancer markers-mediated signaling pathway. 30 Modulators of cancer markers expression can also be identified. For example, a cell or cell free mixture is contacted with a candidate compound and the expression of 49 cancer marker nRNA or protein evaluated relative to the level of expression of stem cell cancer marker mRNA or protein in the absence of the candidate compound. When expression of cancer marker mRNA or protein is greater in the presence of the candidate compound than in its absence, the candidate compound is identified as a stimulator of 5 cancer marker niRNA or protein expression. Alternatively, when expression of cancer marker mRNA or protein is less (i.e., statistically significantly less) in the presence of the candidate compound than in its absence, the candidate compound is identified as an inhibitor of cancer marker mRNA or protein expression. The level of cancer markers mRNA or protein expression can be determined by methods described herein for 10 detecting cancer markers mRNA or protein. This invention further pertains to novel agents identified by the above-described screening assays (See e.g., below description of cancer therapies). Accordingly, it is within the scope of this invention to further use an agent identified as described herein (e.g., a cancer marker modulating agent, an antisense cancer marker nucleic acid 15 molecule, a siRNA molecule, a cancer marker specific antibody, or a cancer marker binding partner) in an appropriate animal model (such as those described herein) to determine the efficacy, toxicity, side effects, or mechanism of action, of treatment with such an agent. Furthermore, novel agents identified by the above-described screening assays can be, e.g.. used for treatments as described herein (e.g. to treat a human patient 20 who has cancer). In certain embodiments, the present invention employs non-adherent mammospheres for various screening procedures, including methods for screening CXCR I or FBX021 signaling pathway antagonists. Non-adherent mammospheres are an in vitro culture system that allows for the propagation of primary human mammary 25 epithelial stem and progenitor cells in an undifferentiated state, based on their ability to proliferate in suspension as spherical structures. Non-adherent mammospheres have previously been described in Dontu et al Genes Dev. 2003 May 15;17(10):1253-70, and Dontu et al., Breast Cancer Res. 2004;6(6):R605-15, both of which are herein incorporated by reference. These references are incorporated by reference in their 30 entireties and specifically for teaching the construction and use of non-adherent mammospheres. As described in Dontu et al., niammospheres have been characterized as 50 being composed of sten and progenitor cells capable of self-renewal and multi-lineage differentiation. Dontu et al. also describes that mamnmospheres contain cells capable of clonally generating complex functional ductal-alveolar structures in reconstituted 3-D culture systems in Matrigel. 5 In certain embodiments, the following exemplary screening methods are employed. For in vitro studies, one could treat cells with either adenoviral constructs expressing control or CXCRi or FBXO2I candidate siRNA (m.o.i. 10 to 100) for 3 days or a small molecule candidate (e.g., PI 1A665752 derivative) (0.1 -0.5 uM) for 3 days and compare the ability of CXCR1+ or FBXO21 + cells to form tumor spheres compared in 10 untreated vs. treated cells. For in vivo studies, one could infect human breast cancer cells with a lentivirus expressing luciferase to monitor tumor growth. Luciferase-expressing cancer cells could be injected into breast tissue and tumors of approximately 0.5-0.7 cm in size could be established, with 5 animals per group. Animals with established tumors could then be treated with either a candidate CXCR1 or FBXO21 inhibitor (daily iv. 15 30mg/kg/day for 7 days), or vehicle control. Parallel studies could be performed using infection with adenovirus expressing control or candidate CXCRi or FBXO21 siRNA (m.o.i. 100 or 500 for 7 days). Animals could be imaged at day 7, 14, 21, and 28 to assess tumor size and then be sacrificed. Tumor size could be further assessed at autopsy and a portion of the tumor stained to assess tumor histology. The remaining tumor could 20 be harvested and sorted to assess the percentage of CXCRi or FBXO2 1 positive and CXCR 1 or FBXO2I negative cells. To verify that administration of candidate CXCR 1 or FBXO21 inhibitor and candidate CXCRI or FBXO21 siRNA adenovirus infection is inhibiting CXCR I or FBXO2 1 signaling function, phosphorylation of downstream mediators such as Gab-i and ERK could be examined (see, Chistensen et al., Cancer 25 Res., 2003; 63:7345-7355, herein incorporated by reference). V. Cancer Therapies In some embodiments, the present invention provides therapies for cancer. In some embodiments, therapies target cancer markers (e.g., including but not limited to, 30 CXCR1 or FBXO21 and proteins in the CXCRi or FBXO21 signaling pathway). In some embodiments, any known or later developed cancer stem cell therapy may be used. 51 For example, cancer stein cell therapeutic agents are described in U.S. Pats, 6,984,522 and 7,115,360 and applications W003/050502, W005/074633, and W005/005601, herein incorporated by reference in their entireties. 5 Antibody Therapy In some embodiments, the present invention provides antibodies that target tumors that express a stem cell cancer marker of the present invention. Any suitable antibody (e.g., monoclonal, polyclonal, or synthetic) can be utilized in the therapeutic methods disclosed herein. In some embodiments, the antibodies used for cancer therapy 10 are humanized antibodies. Methods for humanizing antibodies are well known in the art (See e.g., U.S. Patents 6,180,370, 5,585,089, 6,054,297, and 5,565,332; each of which is herein incorporated by reference). In some embodiments, the therapeutic antibodies comprise an antibody generated against a stem cell cancer marker of the present invention, wherein the antibody is 15 conjugated to a cytotoxic agent. In such embodiments, a tumor specific therapeutic agent is generated that does not target normal cells, thus reducing many of the detrimental side effects of traditional chemotherapy. For certain applications, it is envisioned that the therapeutic agents will be pharmacologic agents that will serve as useful agents for attachment to antibodies, particularly cytotoxic or otherwise anticellular agents having 20 the ability to kill or suppress the growth or cell division of endothelial cells, The present invention contemplates the use of any pharmacologic agent that can be conjugated to an antibody, and delivered in active form. Exemplary anticellular agents include chemotherapeutic agents, radioisotopes, and cytotoxins. The therapeutic antibodies of the present invention can include a variety of cytotoxic moieties, including but not 25 limited to, radioactive isotopes (e.g., iodine- 131, iodine-123, technicium-99in, indium I 11, rhenium-188, rhenium-I 86, galliun-67, copper-67, yttrium-90, iodine-125 or astatine-21 1), hormones such as a steroid, antimetabolites such as cytosines (e.g., arabinoside, fluorouracil, methotrexate or arninopterin; an anthracycline;r mitomycin C), vinca alkaloids (e.g., demecolcine; etoposide; mithramycin), and antitumor alkylating 30 agent such as chlorambucil or melphalan. Other embodiments can include agents such as a coagulant, a cytokine, growth factor, bacterial endotoxin or the lipid A moiety of 52 bacterial endotoxin, For example, in some embodiments, therapeutic agents will include plant-, fungus- or bacteria-derived toxin, such as an A chain toxins, a ribosome inactivating protein, u-sarcin, aspergillin, restrictocin, a ribonuclease, diphtheria toxin or pseudomonas exotoxin, to mention just a few examples. In some embodiments, 5 deglycosylated ricin A chain is utilized. In any event, it is proposed that agents such as these can, if desired, be successfully conjugated to an antibody, in a manner that will allow their targeting, internalization, release or presentation to blood components at the site of the targeted tumor cells as required using known conjugation technology (See, e.g., Ghose et al., 10 Methods Enzymol., 93:280 [1983]). For example, in some embodiments the present invention provides immunotoxins targeted at stem cell cancer marker of the present invention. Immunotoxins are conjugates of a specific targeting agent typically a tumor-directed antibody or fragment, with a cytotoxic agent, such as a toxin moiety. The targeting agent directs the toxin to, 15 and thereby selectively kills, cells canying the targeted antigen. In some embodiments, therapeutic antibodies employ crosslinkers that provide high in vivo stability (Thorpe et al., Cancer Res., 48:6396 [19881). In other embodiments, particularly those involving treatment of solid tumors, antibodies are designed to have a cytotoxic or otherwise anticellular effect against the 20 tumor vasculature, by suppressing the growth or cell division of the vascular endothelial cells. This attack is intended to lead to a tumor-localized vascular collapse, depriving the tumor cells, particularly those tumor cells distal of the vasculature, of oxygen and nutrients, ultimately leading to cell death and tumor necrosis. In some embodiments, antibody based therapeutics are formulated as 25 pharmaceutical compositions as described below. In some embodiments, administration of an antibody composition of the present invention results in a measurable decrease in cancer (e.g., decrease or elimination of tumor). VL Therapeutic Compositions and Administration 30 A pharmaceutical composition containing a regulator of turnorigenesis according the present invention can be administered by any effective method. For 53 example, an IL8-CXCR I signaling pathway antagonist, or other therapeutic agent that acts as an antagonist of proteins in the IL8-CXCRI signal transduction/response pathway can be administered by any effective method. In certain embodiments of the present invention, the therapeutic agent comprises Repertaxin or a derivative thereof. 5 In certain embodiments, a physiologically appropriate solution containing an effective concentration of an IL8-CXCRI signaling pathway antagonist can be administered topically, intraocularly, parenterally, orally, intranasally, intravenously, intramuscularly, subcutaneously or by any other effective means. In particular, the 118 CXCRI signaling pathway antagonist agent may be directly injected into a target cancer 10 or tumor (e.g., into breast tissue) by a needle in amounts effective to treat the tumor cells of the target tissue. Alternatively, a cancer or tumor present in a body cavity such as in the eye, gastrointestinal tract, genitourinary tract (e.g., the urinary bladder), pulmonary and bronchial system and the like can receive a physiologically appropriate composition (e.g., a solution such as a saline or phosphate buffer, a suspension, or an enulsion, which 15 is sterile) containing an effective concentration of an IL8-CXCR1 signaling pathway antagonist via direct injection with a needle or via a catheter or other delivery tube placed into the cancer or tumor afflicted hollow organ. Any effective imaging device such as X ray, sonogram, or fiber-optic visualization system may be used to locate the target tissue and guide the needle or catheter tube. In another alternative, a physiologically 20 appropriate solution containing an effective concentration of an IL8-CXCRi signaling pathway antagonist can be administered systemically into the blood circulation to treat a cancer or tumor that cannot be directly reached or anatomically isolated. Such manipulations have in common the goal of placing the IL8-CXCRl signaling pathway antagonist in sufficient contact with the target tunor to permit the 25 antagonist to contact, transduce or transfect the tumor cells (depending on the nature of the agent). In one embodiment, solid tumors present in the epithelial linings of hollow organs may be treated by infusing the suspension into a hollow fluid filled organ, or by spraying or misting into a hollow air filled organ. Thus, the tumor cells (such as a solid tumor stern cells) may be present in or among the epithelial tissue in the lining of 30 pulmonary bronchial tree, the lining of the gastrointestinal tract, the lining of the female reproductive tract, genitourinary tract, bladder, the gall bladder and any other organ 54 tissue accessible to contact with the 1L8-CXCRI signaling pathway antagonist. In another embodiment, the solid tumor may be located in or on the lining of the central nervous system, such as, for example, the spinal cord, spinal roots or brain, so that the IL8-CXCRI signaling pathway antagonist infused in the cerebrospinal fluid contacts 5 and transduces the cells of the solid tumor in that space. One skilled in the art of oncology can appreciate that the antagonist can be administered to the solid tumor by direct injection into the tumor so that the antagonist contacts and affects the tumor cells inside the tumor. The tumorigenic cells identified by the present invention can also be used to 10 raise anti-cancer cell antibodies. In one embodiment, the method involves obtaining an enriched population of tumorigenic cells or isolated tumorigenic cells; treating the population to prevent cell replication (for example, by irradiation); and administering the treated cell to a human or animal subject in an amount effective for inducing- an immune response to solid tumor stem cells. For guidance as to an effective dose of 1s cells to be injected or orally administered; see,. U.S. Pat. Nos. 6,218,166, 6,207,147, and 6,156,305, incorporated herein by reference. In another embodiment, the method involves obtaining an enriched population of solid tumor stem cells or isolated solid tumor stem cells: mixing the tumor stem cells in an in vitro culture with immune effector cells (according to immunuological methods known in the art) from a human 20 subject or host animal in which the antibody is to be raised; removing the immune effector cells from the culture; and transplanting the immune effector cells into a host animal in a dose that is effective to stimulate an immune response in the animal. In some em bodiments of the present invention, the anti-tumorigenic therapeutic agents (e.g. IL8-CXCRi signaling pathway antagonists) of the present invention are co 25 administered with other anti-neoplastic therapies. A wide range of therapeutic agents find use with the present invention. Any therapeutic agent that can be co-administered with the agents of the present invention, or associated with the agents of the present invention is suitable for use in the methods of the present invention. Various classes of antineoplastic (e.g., anticancer) agents are contemplated for use 30 in certain embodiments of the present invention. Anticancer agents suitable for use with the present invention include, but are not limited to, agents that induce apoptosis, agents 5 that inhibit adenosine deaminase function, inhibit pyrimidine biosynthesis, inhibit purine ring biosynthesis, inhibit nucleotide interconversions, inhibit ribonucleotide reductase, inhibit thymidine monophosphate (TMP) synthesis, inhibit dihydrofolate reduction, inhibit DNA synthesis, form adducts with DNA, damage DNA, inhibit DNA repair, 5 intercalate with DNA, deaminate asparagines, inhibit RNA synthesis, inhibit protein synthesis or stability, inhibit microtubule synthesis or function, and the like. In sorne embodiments, exemplary anticancer agents suitable for use in compositions and methods of the present invention include, but are not limited to: 1) alkaloids, including microtubule inhibitors (e.g., vincristine, vinblastine, and vindesine, 10 etc.), microtubule stabilizers (e.g., paclitaxel (TAXOL), and docetaxel, etc.), and chromatin function inhibitors, including topoisomerase inhibitors, such as epipodophyllotoxins (e.g., etoposide (VP-16), and teniposide (VM-26), etc.), and agents that target topoisomerase I (e.g., camptothecin and isirinotecan (CPT- 11), etc.); 2) covalent DNA-binding agents (alkylating agents), including nitrogen mustards (e.g., 15 mechlorethamine, chlorambtucil, cyclophosphamide, ifosphamide, and busulfan (MYLERAN), etc.), nitrosoureas (e.g., carmustine, lomustine, and semustine, etc.), and other alkylating agents (e.g., dacarbazine, hydroxymethylmelamine, thiotepa, and mitomycin, etc.); 3) noncovalent DNA-binding agents (antitumor antibiotics), including nucleic acid inhibitors (e.g., dactinomycin (actinomycin D), etc.), anthracyclines (e.g., 20 daunorubicin (daunomycin, and cerubidine), doxorubicin (adriamycin), and idarubicin (idamycin), etc.), anthracenediones (e.g., anthracycline analogues, such as mitoxantrone, etc.), bleomycins (BLENOXANE), etc., and plicamycin (mithramycin), etc.; 4) antimetabolites, including antifolates (e.g., methotrexate, FOLEX, and MEXATE, etc.), purine antimetabolites (e.g., 6-mercaptopurine (6-MP, PURINETHOL), 6-thioguanine (6 25 TG), azathioprine, acyclovir, ganciclovir, chlorodeoxyadenosine,2 chlorodeoxyadenosine (CdA), and 2'-deoxycoformycin (pentostatin), etc.), pyrimidine antagonists (e.g., fluoropyrimi dines (e.g., 5-fluorouracil (ADRUCIL), 5 fluorodeoxyuridine (Fd.Trd) (floxuridine)) etc.), and cytosine arabinosides (e.g., CYTOSAR (ara-C) and fludarabine, etc.); 5) enzymes, including L-asparaginase, and 30 hydroxyurea, etc.; 6) hormones, including glucocorticoids, antiestrogens (e.g., tamoxifen, etc.), nonsteroidal antiandrogens (e.g., flutamide, etc.), and aromatase 56 inhibitors (e.g., anastrozole (AR IMIDEX), etc.); 7) platinum compounds (g cisplatin and carboplatin, etc.); 8) monoclonal antibodies con jugated with anticancer drugs, toxins, and/or radionuclides, etc.; 9) biological response modifiers (e.g., interferons (e.g., IFN-a, etc.) and interleukins (e.g., IL-2, etc.), etc.); 10) adoptive inununotherapy; 11) 5 hematopoietic growth factors; 12) agents that induce tumor cell differentiation (e.g., all trans-retinoic acid, etc.); 13) gene therapy techniques; 14) antisense therapy techniques; 15) tumor vaccines; 16) therapies directed against tumor metastases (e.g., batimastat, etc.); 17) angiogenesis inhibitors; 18) proteosome inhibitors (e.g., VELCADE); 19) inhibitors of acetylation and/or methylation (e.g. HDAC inhibitors); 20) modulators of 10 NF kappa B; 21) inhibitors of cell cycle regulation (e.g., CDK inhibitors); 22) modulators of p53 protein function; and 23) radiation. Any oncolytic agent that is routinely used in a cancer therapy context finds use in the compositions and methods of the present invention. For example, the U.S. Food and Drug Administration maintains a formulary of oncolytic agents approved for use in the 15 United States. International counterpart agencies to the U.S.F.D.A. maintain similar formularies. Table 3 provides a list of exemplary antineoplastic agents approved for use in the U.S. Those skilled in the art will appreciate that the "product labels" required on all U.S. approved chemotherapeutics describe approved indications, dosing information, toxicity data, and the like, for the exemplary agents. 20 TABLE 3 Aldesleukin Proleukin Chiron Corp., (des-alanyl-1, serine-1 25 human interleukin-2) Emeryville, CA Alemtuzumab Campath Millennium and (Ig(IGl K anti CD52 antibody) ILEX Partners, LT Cambridge, MA Alitretinoin Panretin Ligand (9-cis-retinoic acid) Pharmaceuticals, Inc., San Diego CA Allopurinol Zvloorim GlaxoSmithKline, (1 ,5-dihydro-4 H -pyrazolo[3,4-d]pyrimidin-4-or Research Triangie monosodium salt) Park, NC 57 Altretamine Hexalen US Bioscience, West (N,N,N',N ,NN,- hexamethyl- 1,3,5-triazine-2, 4, Conshohocken, PA 6-triamine) Amifostine Ethyol US Bioscience (ethanethiol, 2-[(3 -aminoplropyl)amino]-, dihydrogen phosphate (ester)) Anastrozole Arimidex AstraZeneca ( ,3-Benzenediacetonitrile, a, a, a', at'-etramethyl- Pharmaceuticals, LP 5-(1-1I,2,4-triazoi-1-ylmnethyl)) Wilmington, DE Arsenic trioxide Trisenox Cell Therapeutic, Inc., Seattle, WA Asparaginase Elspar Merck & Co., Inc., (L-asparagine amidohydrolase type EC-2) Whitehouse Station, NJ BCG Live TICE BCG Organon Teknika, lyophilizedd preparation of an attenuated strain of Corp., Durham, NC Mvcobacterium bovis (Bacillus Calmette-Gukin [BCG], substrain Montreal) bexarotene capsules Targretin Ligand (4-[ 1-(5,6,7-tetrahydro-3,5,5,8,8-pentametl-2- Pharmaceuticals napthalenyl) ethenyl] benzoic acid) bexarotene gel Targretin Ligand Pharmaceuticals Bleomycin Blenoxane Bristol-Myers Squibb (cytotoxic glycopeptide antibiotics produced by Co., NY, NY Strepbomyces verticillus; bleomycin A and bleomycin B2) Capecitabine Xeloda Roche (5'-deoxy-5-liuoro-N-[ (pentyloxy)carboonyl] cytidine) Carboplatin Paraplatin Bristol-Myers Squibb (platinum, diammine [1,1 cyclobutanedicarboxylato(2 -)-O, 0'] -,(SP-4-2)) Carmustine BCNU, BiCNU Bristol-Myers Squibb (1, 3-bis(2-chloroethyl)-1-nitrosourea) 58 Carmustine with Polifeprosan 20 Implant Gliadel Wafer Guilford Pharmaceuticals, Inc., Baltimore, MD Celecoxib Celebrex Searle (as 4-[5-(4-methylphenyi)-3- (trifluoromethyl)- Pharmaceuticals, I1-pyrazol- 1 -VI] England benzenesulfonamide) Chlorambucil Leukeran Glaxo SmithKline (4- bis(2chlorethyl)amino]benzenebutanoic acid) Cisplatin Platmiol Bristol-Myers Squibb (PtCl2HN 2 ) Cladribine Leustatin, 2-CdA R.W. Johnson (2-chloro-2-deoxy-b-D-adenosinei Pharmaceutical Research Institute, Raritan, NJ Cyclophosphamide Cytoxan, Neosar Bristol-Myers Squibb (2-[bis(2-chioroethyl)amino] tetrahydro-2H-13.2 oxazaphosphorine 2-oxide monohydrate) Cytarabine Cvtosar-U Pharmacia & Upjohn (1 -h-D-Arabinofuranosylcytosin e, C 9 1HI-NN 3 0) Company cytarabine liposomal DepoCyt Skye Pharmaceuticals, Inc., San Diego, CA Dacarbazinc DTIC-Dome Bayer AG, (5-(3,3-dimethyi-1-triazeno)-imidazole-4- Leverkusen, carboxamide (DTIC)) Germany Dactinomycin, actinomycin ) Cosmegen Merck (actmomycin produced by Streptomyces parvuilus, Darbepoetin al fa Arariesp Amgen, Inc (recombinant peptide) Thousand Oaks, CA dautnorub icin lposotnal DanuoXome Nexstar ((8S-cis)-8-acetyi-10-[(3-aniino-2 .3,6-trideoxy-a- Pharnaceuticals Inc., L-iyxo-hexopyranosyi)oxy]-7,8,9,1 0-tetrahydro- Boulder, CO 59 6,8,11 -trihydroxy-l1-rnethoxy-5 1 2 Daimnrubicin HCI daimiomycin Cerubidine Wyveth Averst, ((I S ,3 S )-3 -Ace-tyl- 1,2,3,4,6111 -hexafivdro- Madison, Nj 3,5,1 2-trihyvdroxy- IlO-meth oxy-6,i I1-dioxo- I maplitimcenyi -mn-, 6lrdoy4lh) lyxIo -hexopyranoside hydrochloride) Denileukia diftitox ( 1 )ntak Seragen, I n., (recomibinanmt peptide) I-iopkirmtori MA Dexra-zoxanie Zintecard Phammcta & Ult ((S) -4,4'-(i11-methyl- 2 -etharnediyi)bi s-2, 6 - (iomp'ry pilperazinectonoI Docetaxel Taxotere Aventis ((R, S-Ncabov3 phnvissein.N-tent- Pharmaeuticals, Inc., butyl ester, 13-ester with 5b-20-epoxy- Bridgewater, -NJ 12a,,7bi~b13ahex hydoxyax11 -en-9-on0 4 aceate 2-benzoate. tiihydrate) Doxcorubicirn 1IC Adniamycin, Piafroacia & ITpjohln (8S, 1OS)-104[(3-amn~ino-2,3.6-trideoxy-ai-L-lyxo- RUbex Companmy hexopyran osylboxy] -8-glycoiyi-7,8,9,1O tetalydro.-6,8,1-thdtu--teoy-' riaphthacenedione hydrochloride) dOOxnubiin Adlrimiycit PF'S PhajrnmaCia & UJpiohnr Intr avenlous Compan~y doxorubicin liposomal Doxil Sequn-s !Pharmnaceuticals,In, Menlo nark, CA dromtostan olot e propionat e ![romostanoionte Eli 'Lil ly & Company, (1 '7b-Hydro_-xy-2a-mothvi-5 a-aridrostan-3 -one Indianapolis, IN propionate) dromostanolone propionate Masteronle Syntex, Corp., Palo tnpjection Alto, CA Elliott's B Solution Eiott's B Orphan Medical, Inc SoIlution1 60 Epirubicin Ellence Pharnacia & Upjohn ((8S-cis)-10-[(3-amino-2,3,6-trideox-a-L- Company arabino- hexopyranosyl)oxy]-78,9,10-tetrahydro 6,8,1 1--trihydroxy-8- (hydroxvacetyl)-I--methoxy 5,12-naphthacenedione hydrochloride) Epoetin alfa Epogen Amgen Inc (recombinant peptide) Estrainustine Emcyt Pharmacia & Upjohn (estra-1 3 5(10)-triene-3,1 7-diol(i 7(beta))-, 3- Company [bis(2-chloroethl)carbamate] I 7-(dihydrogen phosphate), disodium salt, monohydrate. or estradiol 3-[bis(2-chioroethyl)carbamate] 17 (dihydrogen phosphate), disodium salt, monohydrate) Etoposide phosphate Etopophos Bristol-Myers Squibb (4'-Demethylepi odophyllotoxin 9-[4,6-O-(R) ethylidene-(beta)-D-glucopyranoside], 4 (dihydrogen phosphate)) etoposide, VP-6 Vepesid Bristol-Myers Squibb (4'-demethlepipodoplhyllotoxin 9-[4,6-0-(R) ethylidene-(beta)-D)-glucopyranoside]) Exemestane Aromasin Pharmacia & Upjohn (6-methylenandrosta-1,4-diene-3, 17-dione) Company Filgrastim Neupogen Amgen, Inc (r-metHuG-CSF) floxuridine (intraarterial) FUDR Roche (2'-deoxy-5 -fluorouridine) Fludaralbine Fludara Berlex Laboratories, (fluorinated nucleotide analog of the antiviral Inc., Cedar Knolls, agent vidaralbine, 9-b -D-arabinoftranosyladenine NJ (ara-A)) Fluorouracil, 5-FU Adrucil ICN Pharmaceuticals, (5-fluoro-2,4(11,3H)-pyrinidinedione) Inc., Iumacao, Puerto Rico Fulvestrant Faslod ex I IPR Pharmaceuticals, 61 (7-aipha-[9-(4,4,5,5,5 -penta fluoropentylsuiphinyl) Guayarna, Puerto 3-o3-yi]estra-1 ,3,5-(iO)- tri ene-3,1 Y7- beta-d iol1) Rico Gomcitabino Gemizar Eli Lilly (2.'-deoxy--2, -d ill;)orocyfidinie monohydrochioride (b-isomer)) Gemiuzumnab Ozogamicii Mlyiotarg Wvyeth Ayerst (anti-CD33 hP67.6) Gosreln cetteZoladex Implant AsrtaZetieca (acetate salt of [D-Ser(But)jAzgiyI' ]LHRH; pyro- Pharmaceuticals (Iiiu-HIis-Ttp- Set.IIyr. D Ser(But)- I ki.-rg- Pro Az-ly-NH'-2 acetate [C 5 91,HsN X0 14 (C!241402), Hydroxymrca Hvdrea Bristol-Myers Squiiblb Ibribumomab 1 iuxetan Zevalin Bioger IDEC, Inc, (immiunocoiijugale resulfintig frm a tI iot ca Canfbridge MA'\ covalemtbond between the. monocIonail antibody lIbritUrnomab) and thlinkec~ hoator nuiUXILn [2-bis(carboxymethyl)amimo]-3 -(p isoth iocyanatophenlyl)- PropYi]-[N-[2 bis(cairboxymethyl)aimino]-2--(iqe~hyi) ethyil-iyc me) idarubicin Idamycin Pharmacia & Upjohin (5, 12-Naplithacenedionie, 9-acetyi-7-V3 -amino- Company hexopyranosylIoxy]-7,8,9.] O i-tetrahvdro-6 ,9,l -1 Inhycoxyhyrochlrde,('7S.- cis ) If'osfamnido IFEX Bristol-Myers Squibb (3-(2-chloroethyfi2-r(2 chioroethiyi)amiino':tetrahv-,dro-2H- 1,3,2 oxazalpi)ostphorme 2--oxide) Imatinib Mesilate Gleevec N'ovartis AG, Basel, (4- (4-ethl- -pperzinv~meI~y] -- [4me~yi-SwitZerland 3-:4(-yiiv'-_- mdn ai~] phenl~bnzaidemethanesuif-onate) Interferon aia-2a Rofecron-A I-ioffiriann-La Roche, (recombinant peptide) Inc., Nutley, NJ 62 Interferon alfa-2b Intron A Schering AG, Berlin, (recombinant peptide) (Lyophilized Germany Betaseron) Irinotecan H]Cl Camptosar Pharmacia & Upjohn ((4S)-4,1 I -diethyl-4-hydroxy-9-[(4- piperi- Company dinopipeirido)carbonyioxy]-]l-pyrano[3 4': 6.7] indolizino[1,2-b] quinoline-3,14(41H, 12H) dione hydrochloride trihydrate) Letrozole Fernara Novartis (4, 4'-(IH-i,2,4 -Triazol-i-yhnethylene) dibenzonitrile) Leucovorin Wellcovorin, Immunex, Corp., (L-Glutamic acid. N[4[[(2amino-5-formyl- Leucovorin Seattle. WA 1,4,5,6, 7,S hexahydro4oxo6 pteridinyl)methyl]amino]benzoyl], calcium salt (1:1)) Levamisole HC] Ergarmisol Janssen Research ((( S.)-2.3,5, 6-tetrahydro-6-phenylimidazo [2, - Foundation, b] thiazole monohydrochloride C] H 2

N

2 S-HCl) Titusville, NJ Lomustine CeeNU Bristol-Myers Squibb (1 -(2-chloro-ethyi)-3-cyciohexyl- 1-nitrosourea) Meclorethamine,nitrogen mustard Mustargen Merck (2-chloro-N-(2-chloroethyl)-N-methylethanamine hydrochloride) Megestrol acetate Megace Bristol-Myers Squibb 17ai( acetyloxy)- 6- methylpregna.- 4,6- diene 3,20- dione Melphalan, L-PAM Alkeran GlaxoSmilthK line (4- bis(2-chloroethyl) amino] -L-phenylalan ine) lercaptopurine, 6-IP Purinetho GlaxoSmilthK line (1,7-dihydro-6 H -purine-6-thione monohydrate) Mesa Mesnex Asta Medica (sodium 2-mercaptoethane sulfonate) M ethotrexate Methotrexate Lederle Laboratories 63 (N-[4-[[(2,4-diamino-6 pteridinyiirnethyl]methylamnino] benzoyl-L glutaric acid) Methoxsalen Uvadex Therakos, Inc., Way (9-methoxy-7H-furo[3 2-g][ ]-benzopyran-7-one) Exton, Pa Mlilomycin C Mutamycin Brislol-Myers Squibb mitomycin C Mitozytrex SuperGen, Inc., Dublin, CA Mitotane Lysodren Bristol-Myers Squibb (1, 1-dihoro -2-(o- chiorophenyl)-2-(p chlorophenyl) ethane) Iitoxantron e Novantrone Immunex (1.4-dihydroxy-5,8-bis[[2- [(2- Corporation hydroxyethyl )amino]ethyi]amino]-9,10 anthracenedione dihydrochloride) Nandrolone phenpropion ate Durabol in -50 Organon, Inc., West Orange, NJ Nofetumomab Verluma Boehringer Ingelhfeim Pharma KG, Germany Oprelvekin Neumega Genetics Institute, (IL- I1) Inc., Alexandria, VA Oxaliplatin Eloxatin Sanofi Syvnthelabo, (cis-[(]R,2R)-1,2-eyclohexanediamine-N,N'] Inc., NY, NY [oxalato(2-)-O,O'] platinum) Pacliaxel TALXOL Bristol-Myers Squibb (5B, 20-Epoxy-1,2a, 4,7, 10B, 13a hexahtydroxytax- I1 -en-9-one 4,10-diacetate 2 benzoate 13-ester with (2R, 3 S)- N-benzoyl-3 phenyisoserine) Pamidronate Aredia Novartis (phosphonic acid (3-amino-1-hydroxypropylidene) bis-, disodium salt, pentahydrate, (APD)) 64 Pegademase Adagen Enzon ((monomethoxzypolyeIhylene g lycol succinimidyl) (Pegademase Pharmaceuticals, Inc., i1 - 17 -adenosine deaminase) Bovine) Bridgewater, NJ Pegaspargase Oncaspar Enzon (monomethoxypolyethylene glycol succinimidyl L-asparaginase) Pegfilgrastim Neulasta Amgen, Inc (covalent conjugate of recombinant methionyl human G-CSF (Filgrastim) and monomethoxypolyethyleeti glycol) Pentostatin Nipent Parke-Davis Pharmaceutical Co., Rockville, MD Pipobroman Vercyte Abbott Laboratories, Abbott Park. IL Plicarmycin, Mithramycin Mithracin Pfizer, Inc., NY, NY (antibiotic produced by Streptomyces plicatus) Porfirmer sodium Photofrin QLT Phototherapeutics, Inc., Vancouver, Canada Procarbazine Matulane Sigma Tau (N-isopropVl-p-(2-3ethylhydrazmno -p -tol uamid e Pharmaceuticals, Inc., monohydrochloride) Gaithersburg,MD Quinacrine Alabrine Abbott Labs (6-chloro-9-( 1 methyl-4-diethyl-amine) butylammno -2-ruethoxyacridin e) Rasburicase Elitek Sanofi-Synthelabo, (recotmbinant peptide) Inc, Rituximab Rituxan Genentech, Inc., (recotbitnt anti-CD20 antibody) South San Francisco, CA Sargramostimt Prokine Immunex Corp (recombinant peptide) 65 Streptozocin Zanosar Pharnacia & Upjohn (streptozocin 2 -- deoxy - 2 - Company [[(methyinitrosoamino)carbonyi]amino] - a(and b - 1) glucopyrantose and 220 mg citric acid anhydrous) Talc Sclerosol Bryan, Corp. (MgISi4mOm (011)2) Wobum, MA Tamoxi fen Nolvadex AstraZeneca ((Z)2-[4-(1,2-diphenyl-1-butenyl) phenoxy] -N, N- Pharmaccuicals dimethylethanamne 2-hydroxy-1,2,3 propanetricarboxylate (1:1)) Temozolomide Temodar Schering (3.4-dihydro-3-methyl-4-oxoimidazo[5, 1 -d]-as tetrazine-8-carboxamide) teniposide, VM-26 Vumon Bristol-Myers Squibb (4'-demethyleperipodoph~yllotoxin 9-[4,6-0-(R)-2 thenylidene-(beta)-D-glucopyranoside]) Testolactone Teslac Bristol-Myers Squibb (13-hydroxy-3-oxo-13.17-secoandrosta-1,4-dien I7-oic acid [der ]-lactone) Thioguanine., 6-TG Thioguanine GlaxoSmithKline (2-amino-I1,7-dihvdro-6 H - purine-6-thione) Thiotepa Thioplex Immunex (Aziridine, 1, ,1 "-phosphinothioylidynetris-, or Corporation Tris (1 -aziridinvl) phosphine sulfide) Topotecan HCl Hycamtin GlaxoSmithKline ((S)-10-[(dimethylamino) methyl]-4-ethyl-4,9 dihydroxy-1H-pyrano[3', 4': 6,7] indolizino [1,2-b] quinoline-3,14-(411 12-dione monohydrochloride) Toremifene Fareston Roberts (2-(p-[(Z)-4-chloro- 1 ,2-diphenyl-1 -butenyl Pharmaceutical phenoxy)-N,N-dimethyiethylamine ciltate (1:1)) Corp., Eatontown, NJ Tositumomab, I 131 Tositumomab Bexxar Corixa Corp., Seattle 66 (recombinant murine immunotherapeutic WA monoclonal IgG2n lambda anti-CD20 antibody (I 131 is a radioimmunotherapeutic antibody)) Trastuzumab Herceplin Genentech, Inc (recombinant monoclonal IgG 1 kappa anti-HER2 antibody) Tretinoin, ATRA Vesanoid Roche (all-trans retinoic acid) Uracil Mustard tJracil Mustard Roberts Labs Capsules Valrubicin, N-trifluoroacetyladrianycin-14- Valstar Anthra -- > Medeva valerate ((2S-cis)-2- [1,2,3,4,6,11-hexahydr-2,5,12 trihydroxy-7 methoxy-6, i -dioxo-[[4 2,3,6 trideoxy-3- [trifluoroacetyI)-amino-a-L-lyxo hexopyranosvl]oxyl] -2-naphthacenyi] -2-oxoethyl pentanoate) Vinblastine, Leurocristine Velban Eli Lilly (C46HscN4010-H2SO4) Vincristine Oncovin Eli Lilly (C46TlIcN4010'o-lSO4) Vinorelbine Navelbine GlaxoSmithlK line (3' ,4'-didehydro-4'-deoxy-C' norvincaleukoblastine [R-(R *,R*)-2,3 dihydroxybutanedioate (1:2)(salt)]) Zoledronate, Zoledronic acid Zoneta Novartis ((1 -Hydroxy-2-imidazol- 1 -yl-phosphonoethyl) phosphonic acid monohydrate) Antimicrobial therapeutic agents may also be used as therapeutic agents in the present invention. Any agent that can kill, inhibit, or otherwise attenuate the function of 5 microbial organisms may be used, as well as any agent contemplated to have such 67 activities. Antimicrobial agents include, but are not limited to, natural and synthetic antibiotics, antibodies, inhibitory proteins (e.g., defensins), antisense nucleic acids, membrane disruptive agents and the like, used alone or in combination. Indeed, any type of antibiotic may be used including, but not limited to, antibacterial agents, antiviral 5 agents, antifungal agents, and the like. In still further embodiments, the present invention provides compounds of the present invention (and any other chemotherapeutic agents) associated with targeting agents that are able to specifically target particular cell types (e.g., tumor cells). Generally, the therapeutic compound that is associated with a targeting agent, targets 10 neoplastic cells through interaction of the targeting agent with a cell surface moiety that is taken into the cell through receptor mediated endocytosis. Any moiety known to be located on the surface of target cells (e.g., tumor cells) finds use with the present invention. For example, an antibody directed against such a moiety targets the compositions of the present invention to cell surfaces containing the 15 moiety. Alternatively, the targeting moiety may be a ligand directed to a receptor present on the cell surface or vice versa. Similarly, vitamins also may be used to target the therapeutics of the present invention to a particular cell. As used herein, the term "targeting molecules" refers to chemical moieties, and portions thereof useful for targeting therapeutic compounds to cells, tissues, and organs 20 of interest. Various types of targeting molecules are contemplated for use with the present invention including, but not limited to, signal peptides, antibodies, nucleic acids, toxins and the like. Targeting moieties may additionally promote the binding of the associated chemical compounds (e.g., small molecules) or the entry of the compounds into the targeted cells, tissues, and organs. Preferably, targeting moieties are selected 25 according to their specificity, affinity, and efficacy in selectively delivering attached compounds to targeted sites within a subject, tissue, or a cell, including specific subeellular locations and organelles. Various efficiency issues affect the administration of all drugs--and of highly cytotoxic drugs (e.g., anticancer drugs) in particular. One issue of particular importance 30 is ensuring that the administered agents affect only targeted cells (e.g., cancer cells), 68 tissues, or organs. The nonspecific or unintended delivery of highly cytotoxic agents to nontargeted cells can cause serious toxicity issues. Numerous attempts have been made to devise drug-targeting schemes to address the problems associated with nonspecific drug delivery. (See e.g., K.N. Syrigos and A.A. 5 Epenetos Anticancer Res., 19:606-614 (1999); Y.J. Park et al, J. Controlled Release, 78:67-79 (2002); R.V.J. Chari, Adv. Drug Deliv. Rev., 31:89-104 (1998); and D. Putnam and J. Kopecek, Adv. Polymer Sci.. 122:55-123 (1995)). Conjugating targeting moieties such as antibodies and ligand peptides (e.g., RDG for endothelium cells) to drug molecules has been used to alleviate some collateral toxicity issues associated with 10 particular drugs. The compounds and anticancer agents may be administered in any sterile, biocompatible pharmaceutical carrier, including, but not limited to, saline, buffered saline, dextrose, and water. In some embodiments, the pharmaceutical compositions of the present invention may contain one agent (e.g., an antibody). In other embodiments, 15 the pharmaceutical compositions contain a mixture of at least two agents (e.g., an antibody and one or more conventional anticancer agents). In still further embodiments, the pharmaceutical compositions of the present invention contain at least two agents that are administered to a patient under one or more of the following conditions: at different periodicities, at different durations, at different concentrations, by different 20 administration routes, etc. In some embodiments, the IL8-CXCRi signaling pathway antagonist is administered prior to the second anticancer agent, e.g., 0.5,1,23, 4,5, 10 12, or 18 hours, 1, 2, 3, 4, 5, or 6 days, 1, 2, 3, or 4 weeks prior to the administration of the anticancer agent. In some embodiments, the iL8-CXCRI signaling pathway antagonist is administered after the second anticancer agent, e.g., 0.5, 1, 2 3, 4, 5, 10, 12, 25 or 18 hours, 1, 2, 3, 4, 5, or 6 days, 1, 2, 3, or 4 weeks after the administration of the anticancer agent. In some embodiments, the IL.8-CXCR I signaling pathway antagonist and the second anticancer agent are administered concurrently but on different schedules, e.g., the IL8-CXCR 1 signaling pathway antagonist compound is administered daily while the second anticancer agent is administered once a week, once every two weeks, once 30 every three weeks, or once every four weeks. In other embodiments, the IL8-CXCRI signaling pathway antagonist is administered once a week while the second anticancer 69 agent is administered daily, once a week, once every two weeks, once every three weeks, or once every four weeks. Depending on the condition being treated, preferred embodiments of the present pharmaceutical compositions are formulated and administered systemically or locally. 5 Techniques for formulation and administration can be found in the latest edition of "Remington's Pharmaceutical Sciences" (Mack Publishing Co, Easton Pa.). Suitable routes may, for example, include oral or transmucosal administration as well as parenteral delivery (e.g., intramuscular, subcutaneous, intramedullary, intrathecal, intraventricular, intravenous, intraperitoneal, or intranasal administration). 10 The present invention contemplates administering therapeutic compounds and, in some embodiments, one or more conventional anticancer agents, in accordance with acceptable pharmaceutical delivery methods and preparation techniques. For example, therapeutic compounds and suitable anticancer agents can be administered to a subject intravenously in a pharmaceutically acceptable carrier such as physiological saline. 15 Standard methods for intracellular delivery of pharmaceutical agents are contemplated (e.g., delivery via liposome). Such methods are well known to those of ordinary skill in the art. In some embodiments, the formulations of the present invention are useful for parenteral administration (e.g., intravenous, subcutaneous, intramuscular, intramedullary, 20 and intraperitoneal). Therapeutic co-administration of some contemplated anticancer agents (eg., therapeutic polypeptides) can also be accomplished using gene therapy reagents and techniques. In some embodiments of the present invention, therapeutic compounds are administered to a subject alone, or in combination with one or more conventional 25 anticancer agents (e.g., nucleotide sequences, drugs, hormones, etc.) or in pharmaceutical compositions where the components are optionally mixed with excipient(s) or other pharmaceutically acceptable carriers, In preferred embodiments of the present invention, pharmaceutically acceptable carriers are biologically inert. In preferred embodiments, the pharmaceutical compositions of the present invention are formulated using 30 pharmaceutically acceptable carriers well known in the art in dosages suitable for oral administration. Such carriers enable the pharmaceutical compositions to be formulated as 70 tablets, pills, capsules, dragees, liquids, gels, syrups, slurries, solutions, suspensions and the like, for respective oral or nasal ingestion by a subject. Pharmaceutical preparations for oral use can be obtained by combining the active compounds with solid excipients, optionally grinding the resulting mixture, and 5 processing the mixture into granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are carbohydrate or protein fillers such as sugars, including lactose, sucrose, mrannitol, or sorbitol; starch from corn, wheat, rice, potato, etc.; cellulose such as methyl cellulose, hvdroxypropy Imethyl-cel lulose, or sodium carboxymethylcellulose; gums including arabic and tragacanth; and proteins such 10 as gelatin and collagen. If desired, disintegrating or solubilizing agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, alginic acid or a salt thereof such as sodium alginate. In preferred embodiments, dosing and administration regimes are tailored by the clinician, or others skilled in the pharmacological arts, based upon well known 1s pharmacological and therapeutic considerations including, but not limited to, the desired level of therapeutic effect, and the practical level of therapeutic effect obtainable. Generally, it is advisable to follow well-known pharmacological principles for administrating chemotherapeutic agents (e.g.. it is generally advisable to not change dosages by more than 50% at time and no more than every 3-4 agent half-lives). For 20 compositions that have relatively little or no dose-related toxicity considerations, and where maximum efficacy (e.g., destruction of cancer cells) is desired, doses in excess of the average required dose are not uncommon. This approach to dosing is commonly referred to as the "maximal dose" strategy. In certain embodiments, the IL8-CXCR 1 signaling pathway antagonist is administered to a subject at a dose of 1-40 mg per day 25 (e.g. for 4-6 weeks). In certain embodiments, subject is administered a loading dose of between 15-70 mg of the IL8-CXCRI signaling pathway antagonist. In certain embodiments, the subject is administered a loading dose of about 35-45 mg of the 118 CXCRi signaling pathway antagonist (e.g. subcutaneously), and then daily doses of about 10 mg (e.g. subcutaneously) for about 4-6 weeks. 71 Additional dosing considerations relate to calculating proper target levels for the agent being administered, the agent's accumulation and potential toxicity, stimulation of resistance, lack of efficacy, and describing the range of the agent's therapeutic index. In certain embodiments, the present invention contemplates using routine methods 5 of titrating the agent's administration. One common strategy for the administration is to set a reasonable target level for the agent in the subject. In sore preferred embodiments, agent levels are measured in the subject's plasma. Proper dose levels and frequencies are then designed to achieve the desired steady-state target level for the agent. Actual, or average, levels of the agent in the subject are monitored (e.g., hourly, daily, weekly, etc. 10 such that the dosing levels or frequencies can be adjusted to maintain target levels. Of course, the pharmacokinetics and pharmacodynamics (e.g., bioavailability, clearance or bioaccumulation, biodistribution, drug interactions, etc.) of the particular agent or agents being administered can potentially impact what are considered reasonable target levels and thus impact dosing levels or frequencies. 15 Target-level dosing methods typically rely upon establishing a reasonable therapeutic objective defined in terms of a desirable range (or therapeutic range) for the agent in the subject. In general, the lower limit of the therapeutic range is roughly equal to the concentration of the agent that provides about 50% of the maximum possible therapeutic effect. The upper limit of the therapeutic range is usually established by the 20 agent's toxicity and not by its efficacy. The present invention contemplates that the upper limit of the therapeutic range for a particular agent will be the concentration at which less than 5 or 10% of subjects exhibit toxic side effects. In some embodiments, the upper limit of the therapeutic range is about two times, or less, than the lower limit. Those skilled in the art will understand that these dosing consideration are highly variable 25 and to some extent individualistic (e.g., based on genetic predispositions, immunological considerations, tolerances, resistances, and the like). Thus, in some embodiments, effective target dosing levels for an agent in a particular subject may be 1, .. , 5, . . . I0, . 15, . . . 20, . . . 50, . . . 75, . . . 100, . . . 200, . . . X%, greater than optimal in another subject. Conversely, some subjects may suffer significant side effects and toxicity related 30 health issues at dosing levels or frequencies far less (1, .... , . . . 1 5, . . . 20. .. 50, . . . 75, . . . 100, . . . 200, . . . X%) than those typically producing optimal therapeutic 72 levels in some or a majority of subjects. In the absence of more specific information, target administration levels are often set in the middle of the therapeutic range. In preferred embodiments, the clinician rationally designs an individualized dosing regimen based on known pharmacological principles and equations. In general, 5 the clinician designs an individualized dosing regimen based on knowledge of various pharmacological and pharnacokinetic properties of the agent, including, but not limited to, F (fractional bioavailability of the dose), Cp (concentration in the plasma), CI (clearance/clearance rate), Vss (volume of drug distribution at steady state) Css (concentration at steady state), and ti/2 (drug half-life), as well as information about the 10 agent's rate of absorption and distribution. Those skilled in the art are referred to any number of well known pharmacological texts (e.g. , Goodman and Gilman's, Pharmaceutical Basis of Therapeutics, 10th ed., Hardman el a., eds., 2001) for further explanation of these variables and for complete equations illustrating the calculation of individualized dosing regimes. Those skilled in the art also will be able to anticipate 1s potential fluctuations in these variables in individual subjects. For example, the standard deviation in the values observed for F, CL, and Vss is typically about 20%. 50%, and 30%, respectively. The practical effect of potentially widely varying parameters in individual subjects is that 95% of the time the Css achieved in a subject is between 35 and 270% that of the target level. For drugs with low therapeutic indices, this is an 20 undesirably wide range. Those skilled in the art will appreciate, however, that once the agent's Cp (concentration in the plasma) is measured, it is possible to estimate the values of F, CL, and Vss directly. This allows the clinician to effectively fine tune a particular subject's dosing regimen. In still other embodiments, the present invention contemplates that continuing 25 therapeutic drug monitoring techniques be used to further adjust an individual's dosing methods and regimens. For example, in one embodiment, Css data is used is to further refine the estimates of CL/F and to subsequently adjust the individual's maintenance dosing to achieve desired agent target levels using known pharmacological principles and equations. Therapeutic drug monitoring can be conducted at practically any time during 30 the dosing schedule. In preferred embodiments, monitoring is carried out at multiple time points during dosing and especially when administering intermittent doses. For 73 example, drug monitoring can be conducted concomitantly, within fractions of a second, seconds, minutes, hours, days, weeks, months, etc., of administration of the agent regardless of the dosing methodology employed (e.g., intermittent dosing, loading doses, maintenance dosing, random dosing, or any other dosing method). However, those 5 skilled in the art will appreciate that when sampling rapidly follows agent administration the changes in agent effects and dynamics rnay not be readily observable because changes in plasma concentration of the agent may be delayed (e.g., due to a slow rate of distribution or other pharmacodynamic factors). Accordingly, subject samples obtained shortly after agent administration may have limited or decreased value. 10 The primary goal of collecting biological samples from the subject during the predicted steady-state target level of administration is to modify the individual's dosing regimen based upon subsequently calculating revised estimates of the agent's CL/F ratio. However, those skilled in the art will appreciate that early postabsorptive drug concentrations do not typically reflect agent clearance. Early postabsorptive drug 15 concentrations are dictated principally by the agent's rate of absorption, the central, rather than the steady state, volume of agent distribution, and the rate of distribution. Each of these pharmacokinetic characteristics have limited value when calculating therapeutic long-term maintenance dosing regimens. Accordingly, in some embodiments, when the objective is therapeutic long-term 20 maintenance dosing, biological samples are obtained from the subject, cells, or tissues of interest well after the previous dose has been administered, and even more preferably shortly before the next planned dose is administered. In still other embodiments, where the therapeutic agent is nearly completely cleared by the subject in the interval between doses, then the present invention 25 contemplates collecting biological samples from the subject at various time points following the previous administration, and most preferably shortly after the dose was administered. VI. Repertaxin and Other Small Molecule CXCRI Inhibitors 30 In certain embodiments, the methods, kits, and compositions of the present invention employ small molecule inhibitors of CXCR 1. One exemplary agent is 74 repartaxin. In certain embodiments, the in vivo dose of repartaxin is between 3 and 60 ng per kilogram (e. 3 ... 30 ... 50 ... 60 mg/kg). In particular embodiments, the dose of repartaxin is about 30 rng per kilogram. The chemical formula for repartaxin is shown below: In other embodiments derivatives of the repertaxin are employed. Other small molecule CXCRI antagonists include SB26561 0 (Glaxo SmithKline Beecham; Benson et al., 2000, 151:196-197), as well as SCH 527123 (2-hydroxy-N,N-dimethvl-3-{2-[[(R)-1-(5 10 methylfuran-2-yl)propyl amino]-3,4-dioxocyclobut-1 -enylamino} beuzamide (SCHIlI 527123), an orally bioavailable CXCR2/CXCR1 receptor antagonist (Schering Plough)). Other small molecule inhibitors can be identified by the screening methods described above. 15 EXAMPLES The following example is provided in order to demonstrate and further illustrate certain preferred embodiments and aspects of the present invention and is not to be construed as limiting the scope thereof. 20 EXAMPLE 1 CXCRI Identifies Cancer Stem Cells This example describes the identification of CXCRI, as well as other proteins (e.g., FBXOZ 1), as cancer stem cell markers. 25 Cell culture. Breast cell lines (BCL) were obtained from the ATCC ("http://www." followed by "lgcpromochem-at.cccom/ common/catalog/cellBiology/ 7S5 cellBiologyIndex.cfm") or from collections developed in the laboratories of Drs, S. Ethier (now available on "http://www." followed by "asterand.coi/asterand/BI OREPOSITORY /hbreastcancercel lines.aspx, SUM44, SUM52, SUM149, SUM159, SUM 185, SUM190, SUM225, SUM229"), VJ. M6bus (BrCa-MZ-01), and V. Catros (S68). All BCLs tested 5 were derived from carcinomas except MCF0I GA, which is derived from fibrocystic disease, and the HMEC-derived 184A1, which was derived from normal mammary tissue. The cell lines were grown using the recommended culture conditions. All experiments were done with subconfluent cells in the exponential phase of growth. 10 A LDEF LUOR assay and' separation of the A LD H-positive population by FAI C& ALDH activity was assessed in 33 BCLs representing the main molecular subtypes of human breast cancer. The ALDEFLUOR kit (StemCell technologies, Durham, NC, USA) was used to isolate the population with high ALDH enzymatic activity (17). Cells obtained from subconfluent cell lines after trypsinization or from freshly dissociated 15 xenografts were suspended in ALDEFLUOR assay buffer containing ALDH substrate (AAI pImol/l per 1x106 cells) and incubated for 40 minutes at 37 9 C. In each experiment a sample of cells was stained under identical conditions with 50mmol/L of diethylaminobenzaldehyde (DEAB), a specific ALDH inhibitor, as negative control. Flow cytometr; sorting was conducted using a FACStarPLUS (Becton Dickinson). 20 ALDEFLUOR fluorescence was excited at 488 nm and detected using standard fluorescein isothiocyanate (FITC) 530/30 band pass filter, For xenotransplanted tumors, incubation with an anti-H2Kd antibody (BD biosciences, 1/200, 20 min on ice) followed by a secondary antibody labeled with phycoerythrin (Jackson labs, 1/250, 20 min on ice) was used to eliminate cells of mouse origin. The sorting gates were established using PI 25 stained cells for viability, ALDEFLUOR-stained cells treated with DEAB, and those stained with secondary antibody alone, Prior to RNA profiling or NOD/SCID mice injection, the purity of sorted populations was checked using double sorting of 10,000 ALDEFLUOR-positive and negative cells in BrCa-MZ-01 and SUM159 cell lines. For both cell lines, sorted ALDEFLUOR-positive populations contained more than 98% of 30 ALDEFLUOR-positive cells and no ALDEFLUOR-positive cells were detected in the ALDEFLUOR-negative population. 76 TuPorigenicity in NOD/SCID rnice. Tumorigenicity of ALDELFUOR-positive, negative and unseparated SUM 159, MDA-MB-453 and BrCa-MZ-0I cells was assessed in NOD/SCID mice. Fat pads were cleared of epithelium at 3 weeks of age prior to 5 puberty and huniized by injecting human fibroblasts (1:1 irradiated:non-irradiated, 50,000 cells/i 0001 Matrigel/fat pad) as described (17). The animals were euthanized when the turnors were 1.2 em in the largest diameter, in compliance with regulations for use of vertebrate animal in research. A portion of each fat pad was fixed in formalin and embedded in paraffin for histological analysis. Another portion was assessed by the 10 ALDEFLUOR assay, followed by sorting and serial transplantation. Anchorageindependent culture. ALDEFLUOR-positive, -negative and unseparated cells from 184A1, SUM 149 and SUM159 were plated as single cells in ultra low attachment plates (Corning, Acton, MA) at low density (5000 viable cells/ml). Cells 15 were grown in serum-free mammary epithelial basal medium (Cambrex Bio Science, Walkerville, MD) for 3-7 days, as described (18). The capacity of cells to form spheres was quantified after treatment with different doses of IL8 (GenWay Biotech, San Diego, CA) added to the medium. 20 RANA extraction. Total RNA was extracted from frozen ALDEFLUOR-positive and -negative cells using DNA/RNA All Prep Maxi Kit, according to the manufacturer's instructions (Qiagen, Sample and Assay technologies, The Netherlands). Eight BCLs were used for transcriptional analysis: 184A1, BrCa-MZ-01, HCC1954, MDA-MB-231, MDA-MB-453, SK-BR-7, SUM149, and SUM159. RNA integrity was controlled by 25 denaturing formaldehyde agarose gel electrophoresis and micro-analysis (Agilent Bioanalyzer, Palo Alto, CA). Gene expression profiling with DNA microarrays. Gene expression analyses used Affymetrix U1 33 Plus 2.0 human oligonucleotide microarrays containing over 47,000 30 transcripts and variants inc I uding 38,500 well -characterized human genes. Preparation of cRNA, hybridizations, washes and detection were done as recommended by the supplier 77 ("http://www." followed by "affynetuix.coin/index.affx"). Expression data were analyzed by the RMA (Robust Multichip Average) method in R using Bioconductor and associated packages (19), as described (20, 21). RMA did background adjustment, quantile normalization and summarization of 11 oligonucleotides per gene. 5 Before analysis, a filtering process removed from the dataset genes with low and poorly measured expression as defined by expression value inferior to 100 units in all the 16 samples, retaining 25,285 genes/ESTs. A second filter, based on the intensity of standard deviation (SID), was applied for unsupervised analyses to exclude genes showing low expression variation across the analyses. SD was calculated on log2-transformed 10 data, in which lowest values were first floored to a minimal value of 100 units, i.e. the background intensity, retaining 13,550 genes/ESTs with SD superior to 0.5. An unsupervised analysis was done on 16 ALDEFLUOR-positive, -negative cells on 13,550 Scenes. Before hierarchical clustering, filtered data were log2-transformed and submitted to the Cluster program (22) using data median-centered on genes, Pearson correlation as 15 similarity metric and centroid linkage clustering. Results were displayed using TreeView program (22). To identify and rank genes discriminating ALDEFLUOR-positive and negative populations, a Mann and Whitney U test was applied to the 25.Z285 genes/ESTs and false discovery rate (FDR, (23) was used to correct the multiple testing hypothesis. The classification power of the discriminator signature was illustrated by classifying 20 samples by hierarchical clustering. A LOOCV was applied to estimate the accuracy of prediction of the identified molecular signatures and the validity of supervised analysis; each sample was excluded one by one and classified with the linear discriminate analysis (LDA, (24) by using model defined on the non-excluded samples. 25 Real-timne RT-PCR. After ALDEFLUOR-positive and ALDEFLUOR-negative populations from different cell lines were sorted, total RNA was isolated using RNeasy Mini Kit (QIAGEN) and utilized for real-time quantitative RT-PCR (qRT-PCR) assays in a ABI PRISM® 79001]T sequence detection system with 384-well block module and automation accessory (Applied Biosystemns). Primers and probes for the Taqman system 30 were selected from the Applied Biosystems website. The sequences of the PCR primer pairs and fluorogenic probes used for CXCR 1, FBXO21, NFYA, NOTCH2, RAD51 Ll 78 and TBP are available on the Applied Biosystems website (CXCR 1 assay ID: [Is_00174146_mi; FBXO21 assay ID: Us_00372141 ni, NFYA assay ID: Hs_00953589 mi, NOTCH2 assay ID: Hs_01050719 mi, RAD51L1 assay ID: Hs00172522_ni, TBP assay ID: Hs_00427620 mi). The relative expression nRNA 5 level of CXCRI, FBXO21, NFYA, NOTCH2, RAD5 ILl was computed with respect to the internal standard TBP gene to normalize for variations in the quality of RNA and the ariount of input cDNA, as described previously (25). Invasion assay. Assays were done in triplicate in transwell chambers with 8um 10 pore polycarbonate filter inserts for I 2-well plates (Corning, NY). Filters were coated with 30 ul of ice-cold 1:6 basement membrane extract (Matrigel, BD-Bioscience) in DMEM/F12 incubated 1 hour at 37 0 C. Cells were added to the upper chamber in 200 ul of serum-free medium. For the invasion assay, 5000 cells were seeded on the Matrigel coated filters and the lower chamber was filled with 600 ul of medium supplemented with 15 10% human serum (Cambrex) or with 600 ul of serum-free medium supplemented with IL8 (i00ng/mL). After 48 hours incubation, the cells on the underside of the filter were counted using light microscopy. Relative invasion was normalized to the unseparated corresponding cell lines under serum condition. 20 Lentivirus infection. For luciferase gene transduction, 70% confluent cells from 11CC1954, MDA-MB-453, and SUM159 were incubated overnight with a 1:3 precipitated mixture of lentiviral supernatants Lenti-LUC-VSVG (Vector Core, Ann Arbor, MI) in culture medium. The following day the cells were harvested by trypsin/EDTA and subeultured at a ratio of 1:6. After I week incubation, cells were 25 sorted according to the ALDEFLUOR phenotype and luciferase expression was verified in each sorted population (ALDEFLUOR-positive and ALDEFLUOR-negative) by adding 2 ml D-luciferin 0.0003% (Promnega, Madison, WI) in the culture medium and counting photon flux by device camera system (Xenogen, Alameda, CA). 30 Intracardiac inoculation. Six weeks-old NOD/SCID mice were anesthetized with 2% isofluorane/air mixture and injected in the heart left ventricle with 100,000 cells in 79 100 pL of sterile Dulbecco's PBS lacking Ca2+ and Mg2+. For each of the three cell lines (HCC 1954, MDA-MIB-453, SUM159) and for each population (ALDEFLUOR positive, ALDEFLUOR-negative and unsorted), three animals were injected. 5 Bioluiniesccnce detection. Baseline bioluminescence was assessed before inoculation and each week thereafter inoculations. Mice were anesthetized with a 2% isofluorane/air mixture and given a single i.p. dose of 150 mg/kg D-hLciferin (Prornega, Madison, WI) in PBS. Animals were then re-anesthetized 6 minutes after administration of D-luciferin. For photon flux counting, a charge-coupled device camera system 10 (Xenogen, Alameda, CA) was used with a nose-cone isofluorane delivery system and heated stage for maintaining body temperature. Results were analyzed after 2 to 12 minutes of exposure using Living Image software provided with the Xenogen imaging system. Signal intensity was quantified as the sum of all detected photon flux counts within a uniform region of interest manually placed during data postprocessing. 15 Normalized photon flux represents the ratio of the photon flux detected each week after inoculations and the photon flux detected before inoculation. Statistical analysis. Results are presented as the mean ±SD for at least three repeated individual experiments for each group. Statistical analyses used the SPSS 20 software (version 10.0.5). Correlations between sample groups and molecular parameters were calculated with the Fisher's exact test or the one-way ANOVA for independent samples. A p-value *0.05 was considered significant. The majority of breast cell lines contain an ALDEF LUOR-positi'e population. 25 The ALDEFLUOR assay (17) was used to isolate CSC from 33 BCLs representing the diverse molecular subtypes and features of breast cancer (20). It was found that 23 out of the 33 cell lines contained an ALDEFLUTOR-positive cell population that ranged from 0.2 to nearly 100%. All 16 basal/mesenchymal BCLs contained an ALDEFLUOR-positive population whereas 7 out of the 12 luminal BCLs did not contain any detectable 30 ALDEFLUOR-positive cells (p::::0.0006, Fischer's exact test). 80 ALDEFL UOR-positive cells have tunorsphere-briing capacity. It has previously been reported that mammary epithelial stem and progenitor cells are able to survive and proliferate in anchorage-independent conditions and form floating spherical colonies which are termed mammospheres (18). Data fror breast tumors, as well as cell 5 lines, have demonstrated that cancer stern-like cells or cancer-initiating cells can also be isolated and propogated as "tumorspheres" in similar assays (26). All marnmosphere initiating cells in the normal human mammary gland are contained within the ALDEFLUOR-positive population (17). To characterize the ALDEFLUOR-positive population from BCLs, the ability of ALDEFLUOR-positive and -negative populations 10 from 184A1, SUM149 and SUM159 to form tumorspheres were compared. In each cell line, the ALDEFLUOR-positive population showed increased tumorsphere-forming capacity compared to ALDEFL UOR-negative cells. ALDEFL UOR-positive BCL cells have cancer stem cell properties in vivo. To 15 determine the hierarchical organization of BCL, the stem cell properties of the ALDEFLUOR-positive and -negative populations of MDA-MB-453, SUM159, and BrCa-MZ-01 cell lines were analyzed. The ALDEFLUOR-positive populations of these three BCLs constituted between 3.54±1.73% and 5,49±3,36% of the total cell populations (F'ig. IA-B. G-H; Fig. 2A-B). As shown in Fig. IF, L the size and latency of tumor 20 formation correlated with the number of ALDEFLUOR-positive cells injected. Remarkable, 500 ALDEFLLUOR-positive cells from MDA-MB-453 and 1,000 ALDEFLUOR-positive cells from SUM159 were able to form tumors. The tumor generating capacity was maintained through serial passages demonstrating the self renewal capacity of these cells. In contrast, ALDEFLUOR-negative cells failed to 25 generate tumors, although limited growth was produced when 50,000 ALDEFLUOR negative MDA-MB-453 cells were injected. H&E staining of the fat pad sections confirmed that tumors formed by ALDEFLUOR-positive cells contained malignant cells whereas only residual Matrigel, apoptotic cells and nouse tissue were seen at the sites of ALDELFUOR-negative cell injections (Fig. 1 E, K). Consistent with the ALDEFLUOR 30 positive population having cancer stein cell characteristics, tumors generated by this population recapitulated the phenotypic heterogeneity of the initial tumor, with a similar 81 ratio of ALDEFLUOR-positive and -negative cells (Fig. IC, I). This indicates that ALDEFLUOR-positive cells were able to self-renew, generating ALDEFLUOR-positive cells and were able to differentiate, generating ALDEFLUOR-negative cells. When BrCa-MZ-0I cells were separated into A LDEFLUOR-positive and 5 negative components, both were capable of tumor generation. Turnors generated by the ALDEFLUOR-positive population consisted of both ALDEFLUOR-positive and negative cells recapitulating the phenotypic heterogeneity of the initial tumor. In contrast, tumors generated by ALDEFLJOR-negative cells gave rise to slowly growing tumors containing only ALDEFLUOR-negative cells. In contrast to the ability of 10 ALDEFLUOR-positive cells to be serially transplanted, serial passages of ALDEFL UOR-negative tumors produced decreasing tumor growth with no growth following three passages. This suggests that the ALDEFLUOR-positive component of the BrCa-MZ-01 cells contain cells with stem cell properties, whereas the ALDEFLUOR negative cells contain progenitor cells able to undergo limited growth but not self 15 renewal. Gene expression profiling of ALDELFUOR-positive and -negative cell populations. To determine whether ALDEFLUOR-positive cells isolated from different BCLs expressed a common set of "cancer stem cell" genes, the ALDEFLUOR-positive 20 and -negative cell populations isolated from eight BCLs (I 84A1, BrCa-MZ-01, TCC1954, MDA-MB-231, MDA-MB-453, SK-BR-7, SUM49, and SUM159) were analyzed using Affymetrix whole-genome oiigonucleotide microarrays. Unsupervised hierarchical clustering, applied to the 16 samples and the 13,550 filtered genes/ESTs, did not separate ALDEFLUOR-positive and -negative populations. Instead, ALDEFLUOR 25 positive and -negative populations clustered with the parental cell line. This suggests that the differences in mRNA transcripts between clonal cell lines supersede differences between ALDEFLUOR-positive and ALDEFLUOR-negative cells. This further suggests that only a limited number of genes are differentially expressed between putative cancer stern cells and their progeny. 30 To determine which genes discriminated ALDEFLUOR-positive and -negative populations, the Mann and Whitney U test was applied to all genes but those with low 82 and poorly measured expression, i.e. 25,285 probe sets. This test identified and ranked after FDR correction, 413 genes/ESTs that discriminated the ALDEFLUOR-positive and -negative cell populations. The 28 overexpressed genes corresponding to unique genes are shown in Table 1, and the most frequently underexpressed genes are shown in Table 2 . TABLE I - Up Regulated Genes Category Symbol Descripton Cytband Probe set 1: Funchon GexLs precisusly seiN-ed c av rara~ecl R k Laani-ep.t~e ar-aham bcxs-ake cht3Z25i 2321_acc tt~ rlmr:clnamn icipi~ n ec TP RXL -tsspn STCH2 a homrdnag 2 ra sgila c'dp-rl11 2 t3_xat Sese-- nexn: erm R~ 1 NAh--.-cc ci ST3bt-palackside aa- hi4 1M sca at m:-~-- '.tmc hn ei sialf:ha a race: arc-sn~e "' ::-' a N. A circasiptzrrnc'Ys id a~p13 2d1, 7_ ae-nmip PCNX p xcmixe .Dxmhilar -a1M4. 221-3_ ac. .. d rr $Sgaa:irgcEC F-b xcc1reai21 283242 12221 t I c-aran WWG ( WW5 dcelaktc mcccta-'n--' c>cxre -- sase 61%1 -3. c15_s a: '>miF :cCA inK2l epm"'rn iae hit 4 6a ao'i :Ia -NL2 aa-like phospha edarwma ''innga 'h5 'SSt254rc-c:criehy -: ICictle+ icacsic-iar Tha-x'ce 5 :hc&<'--1. 2 Ta 3cc5e c-7_risd F5 -L5 15-icxic arS :aine--ch :eae p-sair' 'r 4h 22225655 atr:#cr:aScc AD156:B ~ cecc gi. esh. eci!cimace.1 1h-r33 47_a Phosh.,.!.d cc e~ - x-ried :scmprc-s 3i.C3EA2 Se:-ie cccrar tas 6 y 35 moderia 1 sht255519243 cat+uaita a<icm ca tramp-ayrac M rabsaci prntain Scercakinc S 'aearcar.aigsia ch'--'C 20794_at Isctlaan y ceaenc TA-52Rf4 Tastereccsieccya 2.mcembeac 4 hsp32416 s7_t Cite:perc-ct-a CDS200LS C 32-caaed-4c ainiyaxish hr Ei 55417.?_a Imiccrxce spersa DNlAcepair 1A5i511) 15ADI-53-ik 1 3.caressae::. t6514Z23<-42 1575 ; cc_at rcejzsccciianati 5)hcccma.ic {3 7 cc~ ini:aracia sana1E 3SWI-tske) chscii) 225184O cets:asStg1.515N Citsieleten EPPK511 arspialc1: -chd-S2 4 25?156_x -a:tx Sae-narc iW aciic.a. ;ys anr ce dic m gen -hS 43 215-22_x_at RNA snierece ElC aEda:'ytc :i'cskai cin Nt .tim (N2 2 edg2r4 I213TNA-mGace UnknorwicZ:P4 zinc armai- m<-ec em lgr I -e -h,2. S 2 2 22_ sa :Jlnccn FA11sl.- Sany 'atch accccx---8ny 4, membrae "xa$42 251182_at teknownc SO S ca:1+ra-sis n-ecp:ty cc ccidi 2 nreli 220035_a: Ucc:ma: 10 S-183 TABLE 2 - Down Regulated Genes Probe set Category Sym blC4 Descripbon Cytcxand F uction Pro!§in synfhe'Sis 148?"142"rkne s ie is:s~ L42 dc2 2 i s S a Fd spa Mfsi ~n t izcn IvntAi-eais MRE24W raih oet scLr: 4 : 3.3 22-"7 a Frot-? - s&. nsin k 'iN e:-ritx2c tz MR~~tm r .inh-an :i b-.sclm-z :nr L ? d g&3 240s at Pretis: n si wif:temixc re M S3 r.'inh---c--., x. -ncs'm-. :p- i - 323 'v 2 322,q23 23-'' at - -'ie r: y nsi- vnst& vth-me~hr~i : a a 3a22 Inat a ? - Si- ynrTh e -"s unz' c *. kDa p74 at : n p x) SMTTal:ian ALG5', as rge:eam ne~a k nK 3.2 183 at Pre:Ais 2 syac E:--33 ::-6330 M megsuhzr,:liy 1%, -'=a-352 at12 kiln.. 2 -n'-'s -s': -22ster tzxsenesis +53Sc --'3-s -- 1ra iS -li-l pT P ha 7 c sftesy c in mide syn', pha . e 0.ism: -na sn e3 2 '-Yd '.- 2 -,X, -t irn. b hi synhes I-re I.- ::l--ar pas!--' :: iiis2.: 1a15-223S 033+ar NUPF nc5le 3:- 21Sa22 -t oaibaC' sS.r3. 25 : D 3 -NET- 3- l'2 er- 3 in2 , -Nch:.:02-324 43 02 2rp 3 '2tau lipesel EC2' C23 p-c t ii o-r4S 22c3'2at poten gre fi ApctFWi uMlisPiPepi se5 ssag 22'.15 s81i 2re.; Pra and :2Wara * -M-- D'3 GI mi-- ;.om esria3sng5 22512at InM- R -Ke' -:-aa Sa4oiSanin z4 p23 prden, - cycle Apcpts3 s M 3 '' rnj 1, is M- s m-l- B L -n.iR2 L2-,42%_s. arresi p21MAFiiCiP asdp27z:' Repende:j:'. -ceneia . ao -i -s-.id- 3ssm..tisempla 3332223- . Eyth 23dffe ('stion iinIa, iRr ,aii at3-3 ir-y ciin' i3eq e-einila 4 . 2 a.e z Cei cydze :M2 B at220 159ie Feycn2-t:3 flmiYCmU ,e dyriat*: RNA splichrg: CONL 1P iyop Lk3 i2.32.S 1525M11 a at Pre--RN\A premsS- FRF3 ar-mN pf!messNg iadr 3 PRFF2' hmsnk"'., 3F3. rviaa di3V142321. 22 25231a r-eN oes: LS3M3, LSM-3 'nrraz--c' US. sr3-: n.-d2ear RN*A -hG2 2'22.nsat Pre-itt A ps-s:c Oc~~sassociaae AT. r,, d A s'LN 2. -- Si33 -t-flt PPF4h' hem-t-- -'yas' 'en25.2 2*22127 at Pte-rrRNIA p 3 - 3.,s 3 s- TxPdative AT F'n2,s c-s 2 2sca ing siplnpharyiat A TPSS' r~~.Ai-.h--d . ~ errs...ser zI ub (faelov '-ir4"22 - 252_2 s <a' subm - oiihcoPai A TP ssPhass 1NDUFowat s-a." m p 1tS2-niass s at 3nm- ' ATP$2" rFi.che-dct s -- .or3 e su-o- F2 e A-D,2. 242-1 s at s rmi ccnd-a: A TP spiase 1sixe nzshodP 4 r r2 :WD1Letissa-i 'di22.333 *2 22223 2325< 4 at F'2nsis nE PAas i3-3Ics -1*id22.2 224211 at Upirncvcr. 4<34 2 MGCMW~~ hStsnlpia2C17 3.R 22325 at Usknem WDJRt245' WDcrep-a - 32 22-31-4 R Unnm wFPriM 2 tt P 224-!M s at Uikncw. F0t342745A' Sc-s--I :'ztaal s-ti 3 -FL -- n 2327 .:n 3 23544. a U n 2*35 P12 22,'S-snirtd'- 10scain t.Jct1 2335 2 Uknciis X3.T 21X 2i2TP 22S5--uss ese gmtet A ' di_____R2 2 _____at Uttnn-sr 84 The classification power of this discriminating signature was illustrated by classifying the 16 ALDEFLUOR-positive and -negative samples with the 413 differentially expressed genes/ESTs. Hierarchical clustering ranked 15 out of the 16 samples (Fig. 2A). 5 A number of genes known to play a role in stein cell biology were upregulated in the ALDEFLUOR-positive populations (Table 1), including NFYA, NOTCH2, PCNITX, RBM15, ST3GAL3, and TPRXL. Other genes encode proteins that have putative or uncharacterized role in stern cell function, such as ARID 1B, RAD51 LI, and the chemokine receptor CXCR1/IL8RA (27). Genes underexpressed in the ALDEFLUOR 10 positive population are involved in cell differentiation, apoptosis, RNA splicing, and mitochondrial metabolism. To increase the stringency of analysis, the threshold of the Mann and Whitney analysis was raised to the 0.5 risk and obtained a list of 49 genes/ESTs that discriminated ALDELFLUOR-positive and -negative populations (genes with asterisk in Tables 1-2). 15 With this list, all of the ALDEFLUOR-positive cells, except from SK-BR-7, clustered together. Among these 49 genes/ESTS, 45 corresponded to identified unique genes; only 3 of these 45 were overexpressed in the ALDEFLUOR-positive group while 42 were underexpressed. Characterized overexpressed genes code for an F-box protein FBXO21 and CXCRI/IL8RA. Underexpressed genes include those coding for mitochondrial 20 proteins (MRPL4 1, MRPL42, MRPL47. MRPL54, MRPS23. IMMP I L and differentiation (INACA) and pre-mRNA splicing factors (LSM3, pre-mRNA processing factor PRPF39 and PRPF4B). Leave-one-out cross-validation (LOOCV) at 0.5% risk estimated the accuracy of prediction of the identifier molecular signature and 88% of the samples were predicted in the right class with this "cancer stem cell signature" 25 confirming the supervised analysis. Quantitative RT-PCR assessment confirmed a significant increase of CXCR1 and FBXO2 I in ALDEFLUOR-positive cells. Quantitative RT-PCR analysis of five discrininator genes overexpressed in ALDEFLUOR-positive populations (CXCRI1IL8RA, FBXO21, NFYA, NOTCH2 and RAD5 IL1) was performed. Three cell 30 lines used in the profiling analysis (BrCa-MZ-01, MDA-MB-453, SU1 59) and two additional luminal cell lines (MCF7, S68) were sorted by ALDEFLUOR-assay and 85 ALDEFLUOR-positive and -negative populations were processed separately for quantitative RT-PCR analysis. The quantitative RT-PCR expression level of CXCRl and FBX021 are presented in Fig. 2 B and C. Gene expression levels measured by quantitative RT-PCR confirmed the results obtained using DNA microarrays with an 5 increase of CXCR1 and FBXO2 I mRNA level in the A LDEFLUOR-positive population compared to the ALDEFLUOR-negative population (p<0.05). IL8 prornotes cancer stem cell self renewal. The profiling studies suggested that the IL8 receptor CXCRi/IL8RA was consistently expressed in the ALDEFLUOR 10 positive cell population. To confirm this association, the protein expression of CXCR1/IL8RA was measured by flow cytometry in ALDEFLU OR-positive and negative populations. The ALDEFLUOR-positive and -negative populations from four different cell lines were isolated by FACS, fixed, and stained with a CXCR1 monoclonal antibody labeled with phycoerythrin. As shown in Fig. 3A, ALDEFLUOR-positive cells 15 were highly enriched in CXCRi -positive cells compared to the ALDEFLUOR-negative populations. To determine whether IL8 signaling is important in stein cell function, four BCLs were treated with human recombinant IL8 to determine its effect on the cancer stem cell population as measured by the formation of tumorspheres and by ALDH enzymatic 20 activity. As shown in Fig. 3B, addition of IL8 increased the formation of primary and secondary tumorspheres in a dose-dependent manner. Furthermore, 11.8 increased the ALDEFLUOR-positive population in a dose-dependent manner in each of the four BCLs analyzed (Fig. 3C). This illustrates the power of the "CSC signature" to identify pathways that may play a role in stem cell function. 25 The IL8/CXCR1 axis is involvedl in cancer stein cell invasion. The 1L8/CXCRi axis has been reported to play a role in cancer stem cell invasion (28, 29). A Matrigel invasion assay was utilized, using serum as attiactant, to examine the ability of ALDEFLUOR-positive and -negative cell populations from three different cell lines 30 (H1CC1954, MDA-MB-453, SUMI 59) to invade. As shown in Fig. 4A, ALDEFLUOR positive cells demonstrated 6- to 20-fold higher invasion through Matrigel than the 86 ALDEFLUOR-negative population (p<0.01). When used as a chemo-attractant IL8 (100 ng/rnl) increased invasion of the ALDEFLUOR-positive cells (p<0.05) (Fig. 4A). In contrast to its effects on ALDEFLUOR-positive cells, IL8 did not have any effect on the invasive capacity of ALDELFLUOR-negative cells. These results indicate that cancer 5 stem cells exhibited invasive behavior and furthermore that IL8 facilitates this process. ALDE L UI? O-positive cells have increased metastatic potential. It has been proposed that CSCs play a crucial role in cancer metastasis (30, 31). The above experiments demonstrated that ALDEFLUOR-positive cells have increased invasive 10 capacity compared to ALDEFLUOR-negative cells. To determine the relationship between ALDEFLUOR-positivity and metastatic capacity, HiCC 1954, MDA-MB-453, and SUM 159 were infected with a hiciferase lentivirus reporter system. Luciferase infected cells were sorted using the ALDEFLUOR assay and introduced into NOD/SCID mice by intracardiac injection. A suspension of 100,000 cells from each population was 15 injected and metastasis was assessed by bioluminescent imaging. Mice inoculated with ALDEFLUOR-positive cells developed metastases at different sites and displayed a higher photon flux emission than mice inoculated with unseparated cells, which developed no more than one metastasis per mouse, or mice inoculated with ALDEFLUOR-negative cells, which developed only occasional metastases limited to 20 lymph nodes (Fig. 4B-J). Histologic sections confirmed the presence of metastases at these sites (Fig. 4K-M), Thus, the metastatic capacity of BCLs is predominantly mediated by CSCs contained in the ALDEFLUOR-positive population. The hypothesis that tumors are organized in a cellular hierarchy driven by CSCs 25 has fundamental implications for cancer biology as well as clinical implications for the early detection, prevention and treatment of cancer. Evidence for CSCs has largely relied on primary and early passage xenograft models (32-34). However, the success of establishing breast tumor xenograft has been low particularly for certain molecular subtypes. In contrast to primary tumors, cell lines are available in unlimited quantities 30 and provide only carcinomatous populations for molecular analysis without normal tissue and stroma. In breast cancer, a large number of immortalized cell lines have been 87 produced which represent the different molecular subtypes found in primary human breast cancers (2, 20). However, a fundamental question remains as to how closely these cell lines are able to recapitulate the biology of human breast cancer. In vivo evidencefir stein cells in cell lines. Recent studies have suggested that 5 although cell lines may be clonally derived, they contain a cellular hierarchy representing different stages of cellular differentiation. Several studies have utilized markers such as CD44+/CD24- to identify CSC within breast cancer cell lines. However, their utility is limited by the observation that frequently a large percentage of cells within a cell line express these putative stem cell markers. For example, greater than 90% of cells in basal 10 breast cancer cell lines display the CD44+/CD24- phenotype. Indeed, the CD44+/CD24 phenotype did not isolate the tumorigenic population of these cell lines (Ginestier et al. Cell Stem Cell 1:555-567., herein incorporated by reference in its entirety). An alternative approach has been to use the SPJ from cell lines. However, functional studies utilizing Hoechst staining are limited by the toxicity of this agent (35). There is also 15 evidence that the functional stem cell activity is not contained within the SP(36). ALDH activity assessed by the ALDEFLUOR assay isolates cells with stem cell properties from various cancers (14, 37). In this Example it was demonstrated that 23 out of 33 BCLs (predominantly basal cell lines) contain an ALDEFLUOR-positive population. Lack of an ALDEFLUOR-positive population in some luminal BCLs may 20 indicate that these luminal BCLs are derived from ALDEFLUOR-negative progenitor cells. This Example utilized in vivo assays in NOD/SCID mince to demonstrate the stem cell properties of the ALDEFLUOR-positive populations. Self-renewal was demonstrated by serial passage in NOD/SCID m'ice and differentiation was demonstrated 25 by the ability of ALDEFLUOR-positive but not ALDEFLUOR-negative cells to regenerate the cellular heterogeneity of the initial tumor. A breast cancer sten cell signature. Utilizing eight breast cell lines, this Example identified 413 genes whose expression discriminates ALDEFLUOR-positive and 30 negative cells. This signature contained a number of genes known to play a role in stem cell biology. Genes overexpressed in the ALDEFLUOR-positive population include 88 Notch homolog2 (NOTCH2). which regulates self-renewal and differentiation of narary stem cells (18, 38), NFYA, known to regulate self-renewal and differentiation of stern cells. (39, 40), pecanex hoinolog PCNX, RBM15/OTT, which plays a pleiotropic role in hematopoietic stem cells (41) and affects myeloid differentiation via NOTCH 5 signaling (42), homeobox-like factor TPRXL involved in embryonic development, ST3GAL3, which codes for a stage-specific embryonic antigen-4 synthase, associated with fetal development and renal and gastric carcinogenesis (43). Notably, stage-specific embryonic aitigen-4 protein (SSEA-4) is expressed in stem cell populations such as CXCR4+/CD133+/CD34-+/lin- stem cells in human cord blood and quiescent manmary 10 stem cells (44). Genes underexpressed in the ALDEFLUOR-positive population are involved in cell differentiation, apoptosis, and mitochondrial oxidation. They include genes coding for nascent polypeptide-associated complex alpha subunit NACA, programmed death proteins PDCD5 and PDCD 10, mitochondrial ribosomal protein L41 (MRPL4 1), which 15 induces apoptosis through P153-dependent and independent manner via BCL2 and caspases, and proteins involved in mitochondrial processes such as oxidative phosphorylation (NDUFA2, ATP5J2, IMM IL) and protein synthesis in the mitochondrion (MRPL42, MRPL47, MRPL54, MRPS23). Downregulation of apoptotic genes in CSCs may play a role in the resistance of these cells to radiation and 20 chemotherapy (45. 46). ALDH1Ai was not identified as a differentially-expressed gene in the ALDEFLUOR-positive signature. However, examination of gene expression profile of individual BCLs revealed that although some showed differential expression of ALDIHIA] in the ALDEFLUOR-positive population, others showed differential expression of ALDUI A3, a different ALDH isoforn in this population. This suggests 25 that the expression of different ALDIH isoforms could contribute to the ALDEFLUOR positive phenotype. From chemokines to "stemokines. " The expression of CXCRI, a receptor for IL8, is increased in a variety of cancers (47-50). Although IL8 expression is associated with ER-negative breast cancer (51), this chemokinie has riot previously been reported to 30 play a role in stem cell function. Its implication in the regulation of growth and metastasis is well -established in anidroger-independent prostate cancer (52). 89 Furthermore, the expression level of IL8 is associated with tumorigenicity and metastasis through VEGF production and angiogenesis (53, 54). The gene expression data was validated in three ways. First, quantitative RT-PCR analysis confirmed a significant increase of CXCRI mrRNA in ALDEFLUOR-positive population from cell lines both 5 included and not included in profiling analysis. Second, it was demonstrated using flow cytometry that CXCRi-containing cells were found exclusively within the ALDEFLUOR-positive population. Third, recombinant IL8 increased manmosphere formation and the percent of ALDEFLUOR-positive cells in BCLs. The IL8/CXCRI axis thus appears to regulate mammary stem cell proliferation or self-renewal. Since 10 endothelial and stromal cells secrete IL8 this chemokine appears to play a role in mediating interactions between tumor stem cells and the tumor microenvironment. Recent studies have suggested a role for interleukines/chemokines in the regulation of CSCs (55, 56). This includes a role for IL6 in breast CSCs and IL4 in mediating chemoresistance of colon CSCs (56-59). These factors may be involved in the 15 association between inflammation and cancer. This also includes a role for CCL5 (RANTES), a chemokine secreted by mesenchymal stem cells, which acts as a paracrine factor and enhance breast cancer cells motility, invasion and metastasis(55). The roots ofmetaistasis. CSCs may be responsible for mediating tumor 20 metastasis. A link between CSC and metastasis was first suggested with the identification of stein cell genes in an 11-gene signature generated using comparative profile of metastastatic and primary tumors in transgenic mouse model of prostate cancer and cancer patients (60). This signature was also a powerful predictor of disease recurrence, death after therapy and distant metastasis in a variety of cancer types. This 25 Example has demonstrated that ALDEFLUOR-positive cells are more metastatic than ALDEFLUOR-negative cells and that IL8, previously reported to play a role in tunor metastasis, promotes the invasion and chemotaxis of cancer stein cells which preferentially express the IL8 receptor CXCRI. The ability to isolate metastatic cancer stern cell from cell lines should facilitate studies of the molecular mechaniisins by which 30 cancer stem cells mediate tunor metastasis. 90 EXAMPLE 2 CXCRI Inhibition and Combination Therapy This example describes various methods employed to test the effect of CXCR 1 5 inhibition on tumor cells, as well as the combination of CXCRI inhibition in combination with an anti-mitotic agent (docetaxel). Efct of CXCRI inhibition on the cell growth and on the ALDE FL UOR-positive vovulation of SUA/1,59 cell line. 10 The SUM 159 cell line was cultured in adherent condition and treated the cells using the CXCR1/CXC(R2 inhibitor Repertaxin or two specific blocking antibodies for CXCRI or CXCR2. After 4 days of treatment, the effect on cell growth was analyzed using the MTT assay (Figure 5A) and on the cancer stem cell population using the ALDEFLUOR assay (Figure 5B). More than 95% of cell growth inhibition was observed 15 in the cells treated with Repertaxin or the CXCR1 blocking antibody, whereas no effect was observed for the cells treated with the CXCRz blocking antibody (Figure 5A). Interestingly similar effect was observed on the ALDEFLUOR-positive population with a decrease of 80% and 50% of the ALDEFLUOR-positive population in the cells treated with Repertaxin and CXCRi blocking antibody respectively (Figure 5B). 20 Repertaxin treatment induces a bystander effect mediated by the FAS/FAS ligand signaling SUTM1 59 cell line cells were cultured in adherent conditions and then treated with Repertaxin alone or in combination with a FAS antagonist. Interestingly, the cell growth 25 inhibition induced by the Repertaxin treatment was partially rescued by the addition of a FAS antagonist (anti/Fas-ligand from BD pharmingen (cat# 556371)). Moreover, the cells treated with a FAS agonist displayed a similar cell growth inhibition than the cells treated with Repertaxin. These results suggest that Repertaxin treatment induces a bystander effect mediated by the FAS/FAS ligand signaling. 30 91 Eject of Repertaxin treatment on F AK, AKT and FOXOA3 activation. In order to evaluate the effect of Repertaxin treatment on the CXCRI downstream signaling, SUM 159 cells were cultured, during 2 days, in adherent condition in the 5 absence or in presence of 1 00nM of Repertaxin and stained by inuunofluorescence with antibodies against p-FAK, p-AKT, and FOXOA3. In the non-treated cells (Fig. 7A), it was detected that 30% of cells expressing p-FAK and 10% of cells expressing p-AKT displayed inactivation, while cells treated with Repertaxin displayed a complete inactivation of p-FAK and p-AKT (Fig. 7B). The non-treated SUM159 cells presented 10 80% of cells positive in the cytoplasm for FOXOA3. Interestingly, SUM159 cells treated with Repertaxin presented 80% of cells positive in the nucleus for FOXOA3. The change in FOXOA3 cellular localation from the cytoplasm to the nucleus indicates an activation of FOXOA3 protein. 15 Tumors growth curves following the treatment with Repertoxin, docetaxel or the combination The effect of Repertaxin, docetaxel, or the combination thereof was evaluated using one breast cancer cell lines (8A, SUM159) and three human breast cancer xenografts generated from different patients (8B, MCI; 8C, UM2; and 8D, UM3). For 20 each sample, 50,000 cells were injected into the mammary fat pad of NOD-SCID mice which were monitored for tumor size. Injections were started when the tumor size was about 4 mm. Repertaxin was injected (15mg/Kg) twice a day for 28 days or once a week, docetaxal was I.P. injected (10mg/Kg), or the combination (Repertaxin/Docetaxel) was employed. Figure 8 shows the tumor sizes before and during the course of each indicated 25 treatment (arrow, beginning of the treatment). Similar results are observed for each sample (SUM159, MCI, UM2, UM3) with a statistically significant reduction of the tumor size when treated with Docetaxel alone or the combination Repertaxin/Docetaxel compared to the control (p<0.0i) whereas -no significant difference are observed between the growth of the control tumors and the tumors treated with Reperataxin. 30 92 Effect of Repertaxin, docetaxel, or the combination treatment on the cancer stem cell population as assessed by the A LDEFL UOR assay ALDH activity was assessed by the ALDEFLUOR assay for analyzing the cancer stern cell populations size in each turnor (9A. SUM159, 9B. MCI, 9C. UM2, 9D. UM3) 5 treated with Repertaxin, docetaxel or the combination. Similar results are observed for each sample. Docetaxel treated tumor xenografts showed similar or increase percentage of ALDEFLUOR-positive cells compare to the control, whereas Repertaxin treatment alone or in combination with docetaxel produced a statistically significant decrease in ALDEFL UOR-positive cells with 65% to 85% less cancer stein cells compare to the 10 control (p<G.01 Effect of Repertaxin, docetaxel, or the combination treatment on the cancer stem cell population as assessed by implantation in secondarY mice. Serial dilutions of cells obtained from primary tumors (1A. SUM159, 10B. 15 MCI, 1 GC. UM2, 10D. UM3) non treated (control) and treated with Repertaxin, docetaxel or the combination were implanted in the mammary fat pad of secondary NOD-SCID mice, Control and docetaxel treated primary tumors formed secondary tumors at all dilutions whereas, only higher concentration of primary tumors treated with Reperatxin or in combination with docetaxel were able form delayed secondary tumors 20 which were significantly smaller in size than the control or docetaxel treated tumors (p<0.01). Moreover, 1000 and 100 primary treated cells with the combination failed to form secondary tumors for 3 out of 4 samples (SUMI 59, UM2, UM3). Repertaxin treatment reduces the metastatic potential of SUA59 cell line 25 A SUM 159 cell line was infected with a lentivirus expressing luciferase and inoculated 250,000 luciferase infected cells in the heart of NOD/SCID mice. The mice were organized into two groups. The two groups of mice were treated 12 hours after the intracardiac injection either with s.c. injection of saline solution or s.c. injection of Repertaxin (1 5mg/kg), twice a day during 28 days. Metastasis formation was monitored 30 using bioluminescence imaging (I IB: Mice treated with saline solution; 11 C: Mice treated with Repertaxin). Quantification of the normalized photon flux measured at 93 weekly intervals following inoculation revealed a statistically significant increase of metastasis formation in the group of mice treated with saline solution compare to the group of mice treated with Repertaxin ( IIA). EXAMPLE 3 Treatment of cancer stem cells by CXCR1 blockade This example demonstates the effect of CXCR1 inhibition on tumor cells, through both in vitro assays and mouse models. 10 Dissociation ofmamnarv tissue. 100-200 g of normal breast tissue from reduction mammoplasties was minced with scalpels, dissociated enzymatically, and single cells were cultured in suspension to generate mammospheres or on a collagen substratum in adherent condition to induce cellular differentiation (Dontu et al. Genes Dev. 17:1253 1270., herein incorporated by reference in its entirety). 15 Cell culture. Breast cancer cell lines were grown using recommended culture conditions (Charafe-Jauffret et al. Cancer Res. 69:1302-1313. , herein incorporated by reference in its entirety). Breast cancer cell lines were treated in adherent condition with repertaxin (Sigma-Aldrich), anti-human CXCR1 mouse monoclonal antibody (Clone 42705, R&D 20 systems), anti-human CXCR2 mouse monoclonal antibody (clone 48311, R&D systems), anti-human CD95 mouse monoclonal antibody (Clone DX2, BD Pharmingen) utilized as a FAS signaling agonist, anti-human FAS-Ligand mouse monoclonal antibody (Clone NOK-1, BD pharmingen) utilized as a FAS signaling antagonist, or with docetaxel (Taxotere, Sanofi-Aventis). 25 Cell viability. For MTT assays, cells were plated in adherent condition in 96-well plates at 5,000 cells per well. After one day, treatment with repertaxin was started. The effect of repertaxin treatment on cell viability was estimated at different time points by addition of 20 hl of MTT solution (5 mg/mL in PBS) in each well. Cells were then incubated for 1 30 hour at 37 'C followed by addition of 50 L of DMSO to each well. Absorbance was measured at 560 nm in a fluorescence plate reader (Spectrafluor, Tecan). For TUNEL 94 assays, cells were plated in adherent conditions in 6-well plates at 50,000 cells per well. After one day, treatment with repertaxin was started. The number of apoptotic cells was estimated after four days treatment. Cells were fixed in 3.7% formaldehyde and stained utilizing the TACS TdT kit (R&D systems). Nuclei were counterstained with 5 DAPI/antifade (Invitrogen). Sections were examined with a fluorescent microscope (Leica, Bannockborn, IL, USA) with apoptotic cells detected in green. ALDEFL UOR assaY. The ALDEFLUOR kit (StemCell technologies) was used to isolate the population with high ALDH enzymatic activity using a FACStarPLUS (Becton 10 Dickinson) as previously described (Ginestier et al. Cell Stem Cell 1:555-567., herein incorporated by reference in its entirety). In order to eliminate cells of mouse origin from the xenotransplanted tumors, cell population was stained with an anti-H2Kd antibody (BD biosciences, 1/200, 20 min on ice) followed by staining with a secondary antibody labeled with phycoerythrin (PE) (Jackson labs, 1/250, 20 min on ice). 15 EL15A assav. To measure the level of soluble FAS-ligand secreted in the culture medium of cells treated or not with repertaxin, Human sFAS Ligand Elisa (Bender Medsystems) was utilized. Absorbance was read on a spectro-photometer using 450 nm as the primary wave length. 20 Western blotting. Cells were lysed in a laemmli buffer and loaded onto SDS polyacrylamide gels. Blots were incubated with the respective primary antibodies diluted in TBST (containing 0.1 %Tween20 and 2% BSA) either overnight at 4o, or 2 hours at room temperature. Blots were washed and incubated with appropriate secondary 25 antibodies (GE Healthcare, UK) and detected using SuperSignal West Pico Chemiluminescent Substrate (Pierce). Inmunostaining. For immunofluorescent staining, sorted CXCRI-positive cells were fixed with 95% methanol at -20 0 C for 10 minutes. Cells were rehydrated in PBS and 30 incubated with respective antibodies at room temperature for 1 hour. Primary antibodies used were P-FAK (1:50, Cell Signaling Technology), P-AKT (1:300, Cell Signaling 95 Technology), and FOXO3a (1:250, Cell Signaling Technology). Slides were then washed and incubated 30 minutes with PE conjugated secondary antibodies (Jackson labs). The nuclei were counterstained with DAPI/antifade (Invitrogen) and coverslipped. Sections were examined with a fluorescent microscope (Leica, Bannockborn, IL, USA). 5 Inununohistochemistry for the detection of ALDHI 1 (1:100, lBD biosciences), P-FAK, P AKT, FOXO3a expression was done on paraffin section (Ginestier et al. Arn. J Pathol. 161:1223-1233., herein incorporated by reference in its entirety). Staining was done utilizing the Histostainplus kit (Zyrned laboratories). Diaminobenzidine (DAB) or 3 amino-9-ethylearbazole (AEC) was used as chromogen and sections were counterstained 10 with hematoxylin. Animal model. Tumorigenicity of ALDELFUOR-positive/CXCR1-positive and ALDEFLUOR-positive/CXCRl -negative SUMI 59 cells was assessed in NOD/SCID mice (Ginestier et al. Cell Stern Cell 1:555-567., herein incorporated by reference in its 15 entirety)., The SUM159 cell line and three primary human breast cancer xenografts generated from three different patients (MCI, UM2, UM3) were utilized to determine the efficiency of repertaxin treatment on tumor growth (Ginestier et al. Cell Stem Cell 1:555 567., herein incorporated by reference in its entirety). Cells from these tumors were transplanted orthotopically in the humanized cleared fat-pad of NOD/SCID mice. without 20 cultivation in vitro. Fat pads were prepared as described previously (Ginestier et al. Cell Stem Cell 1:555-567., herein incorporated by reference in its entirety). 50,000 cells from each xenotransplants were injected in the humanized fat pad of NOD/SCID mice and monitored the tumor growth. When the tumor size was approximately 4mm, treatment with repertaxin alone (s.c., 15mg/Kg, twice a day, during 28 days), docetaxel alone (i.p., 25 10mg/Kg, once a week, during 4 weeks), in combination (repertaxin/docetaxel), or a control group injected with saline (i.p., once a week and sc. twice a day, during 28 days) was initiated. The animals were euthanized when the tumors were approximately 1.5 cm in the largest diarneter, to avoid tumor necrosis and in compliance with regulations for use of vertebrate animal in research. A portion of each fat pad injected was fixed in 30 formalin and embedded in paraffin for histological analysis. The rest of the tumor cells were re-implantated into secondary NOD/SCID mice. Serial dilutions of cells were 96 utilized for the re-implantation with injection of 10,000, 1,000, and 100 cells for each treated turnor. Anchorage- independent cultre. BCLs treated, in adherent conditions, with repertaxin 5 (1OOnM), anti-CXCR1 antibody (I Ogg/ml), or anti-CXCR2 (l Og/nil) were dissociated and plated as single cells in ultra-low attachment plates (Corning, Acton, MA) at low density (5,000 viable cells/mI). Cells were grown as previously described (Charafe Jauffret et al. Cancer Res. 69:1302-1313., herein incorporated by reference in its entirety). Subsequent cultures after dissociation of primary tumorospheres were plated on 10 ultra-low attachment plates at a density of 5,000 viable cells/ml. The capacity of cells to form tumorspheres was quantified after the first (primary tumorospheres) and second (secondary tumorospheres) passage. RNA extraction and qRT-PCR. After SUMI59 cells were treated, total RNA was isolated 15 using RNeasy Mini Kit (QIAGEN) and utilized for real-time quantitative RT-PCR (qRT PCR) assays in a ABI PRISM® 7900HT sequence detection system. Primers and probes for the Taqman system were selected from the Applied Biosystems website (wxwww dot appliedbiosystems dot com) (FAS-Ligand assay ID: Hs 00899442 mi; IL8 assay ID: Hs 00174103 mi. TBP assay ID: Hs_ 00427620. mi). The relative expression mRNA 20 level of FAS-Ligand and IL8 was computed with respect to the internal standard TBP gene to normalize for variations in the quality of RNA and the amount of input cDNA, as described previously (Ginestier et al. Clin. Cancer Res. 12:4533-4544., herein incorporated by reference in its entirety). 25 Flow cvtoietry analysis. CD44/CD24/Lin staining was performed (Ginestier et al. Cell Stein Cell 1:555-567., herein incorporated by reference in its entirety). CD95/FAS staining were performed utilizing an anti-CD95 labeled APC (1:20, BD biosciences). For CXCR1 and CXCR2 staining, primary antibodies anti-CXCRI (1:100, Clone 42705, R&D systerns) and anti-CXCR2 (1:100, clone 48311, R&D systems) were followed by a 30 staining with a secondary antibody anti-mouse labeled with PE (dilution 1:250, Jackson Labs). Fresh cells were stained with I pg/ml PI (Sigma) for 5 min for viability. 97 Virus infection. Two different lentiviral constructs were produced for the expression of Luciferase gene (Lenti-LUC-VSVG) (Charafe-Jauffiet et al. Cancer Res. 69:1302-1313., herein incorporated by reference in its entirety) and for the inhibition PTEN expression 5 (Lenti-PTEN-SiRNA-DsRed) (Korkaya et al. PLoS Biolog. 7:e1000121., herein incorporated by reference in its entirety), respectively. All lentiviral constructs were prepared by the University of Michigan Vector. An adenoviral construct for the overexpression of FAK (Ad-FAK-GFP) was also utilized (Luo et al. Cancer Res. 69:466 474, herein incorporated by reference in its entirety). Cells infection with different 10 vectors was performed as previously described (Charafe-Jauffret et al. Cancer Res. 69:13021-1313., herein incorporated by reference in its entirety). Efficiency of infection was verified by measuring the percentage of DsRed or GF P expressing cells. intracardiac inoculation. Six weeks-old NOD/SCID mice were anesthetized with 2% 15 isofluorane/air mixture and injected in the heart left ventricle with 250,000 cells in 100 pL of sterile Dulbecco's PBS lacking Ca2+ and Mg2+. For each of the three cell lines (HCC1954, MDA-MB-453, and SUM159) and for each treatment (saline or repertaxin) six animals were inj ected. Twelve hours after intracardiac injections, mice were begun on twice per day repertaxin injections or saline for the controls. 20 Bioluminescence detection. Baseline bioluninescence was assessed before inoculation and each week thereafter inoculations. Bioluminescence detection procedures was performed as previously described (Charafe-Jauffret et al. Cancer Res. 69:1302-1313., herein incorporated by reference in its entirety). Normalized photon flux represents the 25 ratio of the photon flux detected each week after inoculations and the photon flux detected before inoculation, CXCR] expression subdivides cancer stem cell populations. Identi fying cell signaling pathways that regulate cancer stem cells (CSC) provides potential therapeutic targets in a 30 cell population. A breast CSC signature based on gene expression profiling that contained several genes potentially involved in breast CSC regulatory pathways has been identified 98 (Charafe-Jauffret et al. Cancer Res. 69:1302-1313., herein incorporated by reference in its entirety). Among the genes overexpressed in the breast CSC population, CXCRI a receptor that binds the proinflannatory chemokine IL-8/CXCL8 appeared to be a promising candidate since recombinant IL-8 stimulated the self-renewal of breast CSC 5 (Charafe-Jauffret et al. Cancer Res. 69: 1302- 1 313 ., herein incorporated by reference in its entirety). Utilizing flow cytometry, CXCRI protein expression was measured in the breast CSC population as assessed by the A LDELFTOR assay in the human breast cancer cell lines HCC1 954, MDA-MB-453, and SUM 159. Cells with functional stem cell properties in NOD/SCD mouse xenographs were contained within the ALDEFLUOR 10 positive cell population (Charafe-Jauffret et al. Cancer Res. 69:1302-1313., herein incorporated by reference in its entirety). The CXCRi -positive population, which represents less than 2% of the total population, was almost exclusively contained within the ALDEFL UOR-positive population (SEE FIG.12A and Table 4). 15 TABLE 4. ALDEFLUOR CXCRi Csver:ap CXCR1IALDEFUOR Breast cancer cell lines HCCI964 3.42 172 0.94 MDA-MB-453 4.22 0 03 SUM159 5.24 0.2 0.48 Human breast cancer xenngmfts MCl 12-3 1.8 132 UM2 '.4 1.23 Mb UM3 9.7 0.84 0.76 CXCR2 expression was also assessed. CXCR2 is a receptor that can also bind IL 8/CXL8 although with reduced affinity compared to CXCRL In contrast to CXCR1 positive cells, CXCR2-positive cells were equally distributed between the 20 ALDEFLUOR-positive and ALDEFLUOR-negative populations (SEE FIG 1 2A). To 99 determine the hierarchical organization of the cancer stem cell population according to CXCRI expression, ALDEFLUOR-positive/CXCR i-positive and ALDEFLUOR positive/CXCRi -negative cell populations were sorted and injected in NOD/SCID mice (SEE FIG. 13). Both cell populations generated tumors. Tumor growth kinetics 5 correlated with the latency and size of tumor formation and the number of cells injected. Turnors generated by the ALDEFLUOR-positive/CXCRi -positive population reconstituted the phenotypic heterogeneity of the initial tumor upon serial passages whereas the ALDEFLUOR-positive/CXC RI -negative population gave rise to tumors containing only ALDEFLUOR-positive/CXCRI -negative cells. These results suggest that 10 CSC cellular hierarchy is organized according to CXCR1 expression, however both cell populations displayed similar tumorigenic capacity. CXCR1 blockade decreases the breast cancer stein cell population in vitro. Three different cell lines were treated with with repertaxin (IOOnM), a CXCR1/2 inhibitor, to 15 evaluate the effect of CXCRi blockade on the breast CSC population (Bertini et al. Proc. Nati. Acad. Sci. U. S A 101:11791-11796., herein incorporated by reference in its entirety). For SUM! 59, after three days of treatment a five-fold reduction in the proportion of ALDEFLUOR-positive cells was observed (SEE FIG, 12B). A similar effect was observed after treatment of SUM159 cells with an anti-CXCRi blocking 20 antibody. In contrast, no effect was observed after treatment with an anti-CXCR2 blocking antibody, suggesting that the effects of repertaxin on the ALDEFLUOR-positive population were mediated by CXCR . Data from breast tumors, as well as cell lines, demonstrate that cancer stem-like cells or cancer-initiating cells can also be isolated and propagated as "tumorspheres" in 25 suspension culture (Ponti et al. Cancer Res. 65:5506-5511., herein incorporated by reference in its entirety). After three days of treatment with repertaxin or with the anti CXCR 1 blocking antibody, when cells were detached and cultured in suspension, an 8 fold decrease in primary and secondary tumorsphere formation was observed compared to controls. In contrast, anti-CXCR2 blocking antibody had no effect on tumorsphere 30 formation (SEE FIG. 14). 100 Surprisingly, after five days of treatment with repertaxin we observed a massive decrease in viability of the entire cell population as assessed by MTT assay, with only 3% of cells remaining viable (SEE FIG. 12C). Similar results were observed with the anti CXCRI blocking antibody but not the anti-CXCR2 blocking antibody, thus indicating 5 that this effect was dependent on CXCRI blockade. This effect of repertaxin was delayed with loss of cell viability beginning three days after treatment (SEE FIG. 15A). Repertaxin treatment induced a similar effect on the 11(CC1954 breast cancer cell line whereas no effect was observed on M DA-MB-453 cells which harbor a PTEN mutation (Hoilestelle et al. Cancer Res. 5:195-201., herein incorporated by reference in its entirety) 10 (SEE FIG. 14, 15B-C, and 16). Utilizing a TUNEL assay, SUM159 cells were stained after 4 days of treatment with repertaxin and a massive decrease in cell viability, due to induction of apoptosis with 36% apoptotic cells detected after repertaxin treatment, was observed (SEE FIG. 12D). Results suggest that CXCRi blockade results in a decrease of the breast CSC 15 population followed by induction of massive apoptosis in the remaining bulk tumor population. CXCRI blockade induces cell death in CXCRi-negative cells via a bystander effect. The observation that repertaxin or anti-CXCRl blocking antibody induced massive cell death 20 despite the fact that the CXCRl-positive population represented less than 2% of the total cell population suggested that CXCR1 blockade in CXCRI -positive cells induced CXCR i-negative cell death via a bystander effect. The sorted CXCRI -positive and CXCRI -negative populations were treated with repertaxin (SEE FIG. 12E). Repertaxin decreased cell viability in the CXCR I-positive population within three days whereas no 25 effect was observed in the CXCR I -negative population. Repertaxin induced massive cell death in unseparated cells. The effect of repertaxin on cell viability of the unseparated and CXCRI -positive populations was dose-dependent (SEE FIG. 12E). The results are consistent with repertaxin treatment targeting the CXCR1 -positive population that in turn induces CXCR I-negative cell death via a bystander effect. 30 To determine whether this effect was mediated by a soluble factor induced by repertaxin, conditioned medium was collected from the CXCR I-positive population after 101 three days of repertaxin treatment and dialyzed this medium utilizing a membrane with 3.5 KDa exclusion in order to remove repertaxin frorn the medium while retaining molecules larger than 3.5 KDa. The dialyzed conditioned medium induced a massive decrease in cell viability in both CXCRi -negative and unseparated populations but not in 5 the CXCRi -positive population (SEE FIG. 12F). These results demonstrate that CXCRI blockade in the CXCRI -positive population induces cell death in the CXCRI -negative population via a soluble non dialyzable factor. Although the CXCR I-positive population is sensitive to repertaxin it is resistant to the dialyzable death factor. 10 The bystander efect induced by CX'CRI blockade is mediated by FAS-ligand/F AS signaling. FAS-ligand/FAS interaction is activated in different physiologic states such as mammary gland involution or in conditions of tissue injury including that induced by chemotherapy (Chhipa et al. J Cell Biochem. 101:68-79., Song et al. J Clin. Invest 106:1209-1220., herein incorporated by reference in their entireties). The level of soluble 15 FAS-ligand in the medium of SUM 159 cells treated with repertaxin using an ELISA Assay to evaluate the role of FAS-ligand/FAS interaction in mediating the apoptotic bystander effect induced by CXCR1 blockade. More than a five-fold increase of soluble FAS-ligand in the medium of cells treated for four days with repertaxin compared to non treated cells was observed (SEE FIG. 17A). The transcriptional regulation of FAS-ligand 20 by repertaxin treatment by measuring FAS-ligand mRNA level was confirmed by RT PCR (SEE FIG. 17B). A 4-fold increase of the FAS-ligand mRNA level in the repertaxin treated cells was observed compared to non-treated cells. Similar results were observed after treatment with a FAS agonist that activates FAS signaling, indicating that FAS ligand is a target of FAS signaling generating a positive feed-back loop. 100% of the 25 SUM159 cells expressed FAS protein as determined by flow cytometry. Treatment of the SUM159 cells with the FAS agonist reproduced the killing effect observed with the repertaxin treatment with massive reduction in cell viability (SEE FIG. 17C). The effect of repertaxin treatment on cell viability was partially reversed by an anti-FAS-Ligand blocking antibody, with 44% of cells remaining viable after treatment with repertaxin and 30 anti-FAS-ligand antibody compared to only 3% with repertaxin alone (SEE FIG. I 7C). 102 Results suggest that the massive cell death induced by repertaxin is due to a bystander effect mediated by the FAS-Ligand/FAS pathway. Treatment of SUM159 cells with the FAS agonist resulted in a ten-fold and three fold increase in the percent of CXCR 1-positive and ALDEFLUOR-positive cells, 5 respectively (SEE FIG. 17D/E and 18). The effects of repertaxin on both populations were not rescued by anti-FAS-ligand (SEE FIG. 17D/E), suggesting that the ALDEFLUOR-positive population that contains the CXCRi -positive population, while directly sensitive to CXCR I blockade which in turn induces FAS-ligand production by these cells is resistant to FAS-ligand/FAS pro-apoptotic signaling. In contrast, the 10 ALDEFLUOR-negative bulk cell population does not express CXCR1 but is sensitive to FAS-ligand mediated cell death. FAS-ligand/FAS signaling plays an important role during mammary gland involution (Song et al. J Clin. Invest 106:1209-1220., herein incorporated by reference in tis entirety). The effect of CXCRi blockade on human normal mammary epithelial cells 15 obtained from reduction mammoplasties was examined. As observed in breast cancer cell lines, CXCR1 -positive normal manmnary cells were almost exclusively contained within the ALDEFLUOR-positive population (SEE FIG. 19A). To determine whether IL-8 signaling is important in normal breast stem/progenitor function, normal mammary epithelial cells cultured in suspension were treated with human recombinant IL-8 and 20 determined its effect on the CSC population as measured by the formation of mammospheres (Dontu et al, Genes Dev. 17:1253-1270., herein incorporated by reference in tis entirety), Addition of IL.-8 increased the formation of primary and secondary mamiospheres in a dose-dependent manner (SEE FIB. 19B), suggesting that the IL-8/CXCR1 axis may be involved in the regulation of normal mammary 25 stem/progenitor cells proliferation or self-renewal. Treatment with repertaxin or the FAS agonist had no effect on the viability of normal mammary epithelial cells cultured in adherent conditions, even when high concentrations of repertaxin (500nM) were utilized (SEE FIG. 16A). However, as observed for breast cancer cell lines, an increase of soluble FAS-ligand was detected in the medium of normal mammary epithelial cells treated with 30 repertaxin (SEE FIG. 2013). This observation may be explained by the absence of FAS expression in the normal epithelial cells cultured under these conditions (SEE FIG. 20C). 103 This is consistent with studies that demonstrate that expression of FAS in the mammary gland occurs only during the involution process following lactation (Song et al. J Clin. Invest 106:1209-1220., herein incorporated by reference in its entirety). In contrast to its lack of effect on the bulk population of normal mammary epithelial cells, repertaxin 5 significantly decreased mamnosphere formation by these cells (SEE FIG. 20C). These results suggest that the IL-8/CXCRI axis plays an important role in the regulation and the survival of normal and malignant marnmary epithelial stem/progenitor cell populations. The ability to affect bulk cell populations via a FAS-ligand mediated bystander effect may relate to the level of FAS expression in these cells. 10 CXCR1 blockade effects on cancer stem cells are mediated by the FA KIAKYY1FOXO3A pathway. CXCRi acts through a signal transduction pathway involving the phosphorylation of the focal adhesion kinase (FAK) resulting in activation of AKT (Waugh et al. Clin. Cancer Res. 14:6735-6741., herein incorporated by reference in its 15 entirety). To evaluate the impact of CXCRI blockade on the FAK and AKT activation the level of FAK and AKT phosphorylated proteins was measured by western blot for the three different cell lines. For SUM159 and HCC1954. we detected a decrease in FAK Tyr' and AKT Ser 4 ' phosphorylation in cells treated with repertaxin compared to untreated cells suggesting that repertaxin effects may be mediated by the FAK/AKT 20 pathway (SEE FIGS. 21A and 22). The observation that MDA-MB453 is resistant to repertaxin treatment may be explained by the presence of a PTEN mutation (919G>A) that activates the P13K/AKT pathway (Hollestelle et al. Mol. Cancer Res. 5:195-201., herein incorporated by reference in its entirety). No modification in FAK Tyr- and AKT Ser 4

'

3 phosphorylation was detected after repertaxin treatment in MDAMB453 cell line 25 (SEE FIG. 22). To confirm a functional role of the FAKIAKT pathway in mediating the effects of the CXCRI blockade, two viral constricts were used, one knocking down PTEN expression via a PTEN shRNA and the other leading to FAK overexpression. PTEN, through its lipid phosphatase antagonizes PI3-K/AKT signaling (Vivanco et al. Nat. Rev. Cancer 2:489-501., herein incorporated by reference in its entirety). PTEN 30 knockdown resulted in AKT activation as demonstrated by an increase of AKT Ser4 phosphorylation (SEE FIGS. 21A and 22). PTEN knockdown blocked the effect of 104 repertaxin treatment on FAK and AKT activity. FAK overexpression also blocked the effects of repertaxin and induced an activation of FAK and AKT, measured by increased expression of FAK Tyr 397 and AKT Ser 73 phosphorylation. These results indicate that CXCRI blockade effects are mediated by FAKIAKT signaling. 5 Utilizing inununofluorescence staining on CXCRI -posi tve cells confirmed that repertaxin treatment results in a dramatic decrease of phospho-FAK and phospho-AKT expression compared to untreated cells (SEE FIG. 21B). AKT regulates the activity of the forkhead transcription factor FOXO3A via a phosphorylation event resulting in cytoplasnic FOXO3A sequestration (Brunet et al. Mol. Cell Biol. 21:952-965., herein 10 incorporated by reference in its entirety). In contrast, the non-phosphorylated form of FOXO3A transits to the nucleus where it acts as a transcription factor that regulates the synthesis of FAS-ligand (Jonsson et al. Nat. Med. 11:666-671.), herein incorporated by reference in its entirety. Repertaxin induces cell death via a FAS-ligand mediated bystander effect; the effects of repertaxin on this signal transduction pathway were 15 examined by inununofluorescence staining. FOXO3A was present in a cytoplasmic localization in untreated cells but shuttled to the nucleus upon repertaxin treatment (SEE FIG. 21B). This indicates that CXCRi blockade induces FOXO3A activity through inhibition of the FAK/AKT pathway. Cells with PTEN deletion or FAK overexpression display a high level of phospho-FAK and phospho-Akt expression, detected by 20 immunofluorescence, in both repertaxin-treated and untreated cells. Repertaxin treatment did not induce FOXO3A activation in cells with PTEN deletion or FAK overexpression, as shown by the cytoplasinic location of FOXO3A (SEE FIG. 21B). As a consequence of the constitutive activation of the FAK/AKT pathway, cells with PTEN deletion or FAK overexpression displayed resistance to repertaxin treatment. 25 Cells with PTEN deletion or FAK overexpression did not display any decrease in cell viability with repertaxin treatment. It has been proposed that AKT signaling plays a critical role in the biology of CSC (SEE FIGS. 21B and 22) (Dubrovska et al. Proc. Nati. Acad. Sci. U. S A 106:268-273., Korkaya et al. PLoS Biolog. 7:e100012 1., Yilmaz et al. Nature 441:475-482., herein incorporated by reference in their entireties). Activation of 30 the FAK/AKT pathway blocked the repertaxin effects on the CSC populations, as shown by the maintenance of the ALDELFUOR-positive populations after treatment with the 105 inhibitor (SEE FIG. 21 B). All the results indicate CXCR1 blockade directly affects the FAK/AKT/FOXO3A pathway. Repertaxin treatment inhibits AKT signaling which is crucial for CSC activity and subsequently induces a bystander effect on the bulk turnor cells mediated by CS(C7-generated FAS-ligand. 5 Repertaxin treatment retc/ees the breast cancer stein cell population in vivo. Recent evidence suggests that breast CSC are relatively resistant to chemotherapy and radiation and may contribute to tumor regrowth following therapy (Phillips et al. J Natl. Cancer Inst. 98:1777-1785., Yu et al. Cell 131:1109-1123., Li et al. J Nati. Cancer Inst. 100:672 10 679., herein incorporated by reference in their entireties). The CSC concept suggests that significant improvements in clinical outcome will require effective targeting of the CSC population (Reva et al. Nature 414:105-111., herein incorporated by reference in its entirety). Several factors are synthesized and secreted during the apoptotic process when the bulk tumor cells are targeted by chemotherapy. Among these factors, FAS-ligand 15 amplifies chemotherapy effects by mediating a bystander killing effect (Chhipa et al. J Cell Biochem. 101:68-79. herein incorporated by reference in its entirety).Chemotherapy may also induce IL-8 production in injured cells. The commonly utilized chemotherapeutic agent, docetaxel, induced both IL-8 and FAS-ligand mRNA in SUM159 cells (SEE FIG. 10a/B). We also detected a 4-fold increase of IL-8 mRNA level 20 after FAS agonist treatment (SEE FIG. 10B). We have shown that IL-8 is able to regulate the CSC population. This indicates that the addition of repertaxin to cytotoxic chemotherapy may block this effect and target the cancer stem cell population. The Slit 59 cell line and three primary human breast cancer xenografts generated from three different patients (MCI, UM2, UM3.) were used to explore the 25 efficiency of repertaxin treatment on tumor growth. Cells from these tumors were transplanted orthotopically into the humanized cleared fat-pad of NOD/SCID mice, without cultivation in vitro. For each of these xenotransplants the CSC population was exclusively contained within the ALDEFLUOR-positive population (Ginestier et al. Cell Stern Cell 1:555-567., Charafe-Jauffret et al. Cancer Res. 69:1302-1313., herein 30 incorporated by reference in their entireties). In each of the tumors, the CXCRi -positive 106 population was almost exclusively contained within this ALDEFLUOR-posi ive population (SEE Table 5) and the PTEN/FAK/AKT pathway is activated (SEE FIG. 25). Table 5 ALDEFLUOR CXCRI Oveiaop CXCR IALDEFLUOR Breast cancer ceI lines HCC1954 3.42 1.72 0.4 MDA-MB-453 4.22 0.8 0 6 SUMiS9 5_24 I.52 0148 Human breast cancer xenografts MC1 12.3 i1 1.32 UM2 8.4 1.23 R88 UM3 97 C. 84 0.76 5 50,000 cells from each xenotransplant were injected into the humanized fat pad of NOD/SCID mice and monitored tumor growth. When the tumor size was approximately 4mm, treatment was initiated with repertaxin alone (15ng/Kg, twice a day, during 28 10 days), docetaxel alone (1 0rng/Kg, once a week, during 4 weeks), or a combination of both drugs. Tumor growth was compared to saline injected controls. For each xenotransplant, a significant inhibition of tumor growth induced by docetaxel treatment or the combination repertaxin/docetaxel was observed (SEE FIGS 26A and 27). Repertaxin treatment alone had a moderate impact on tumor growth. After four weeks of 15 treatment, animals were sacrificed and the residual tumors were analyzed utilizing the ALDEFLUOR assay. Residual tumors treated with docetaxel alone contained either an unchanged or increased percent of ALDELFUOR-positive cells compared to untreated controls (SEE FIGS 26B and 27). In contrast, repertaxin treatment alone or in combination with docetaxel reduced the ALDEFLUOR-positive population by over 75% 107 (SEE FIGS 26B and 27) The results were confirmed by immunohistochemistry of ALDHIl expression in the different xenotransplants. A decrease in ALDHi1 -positive cells was detected in repertaxin-treated tumors compared to untreated tumnors, whereas the percent of ALDIHI -positive cells was unchanged or increased in tumors treated with 5 docetaxel alone (SEE FIGS 26D). The presence of CD44'/CD24- cells in these tumors was evaluated. Markers have previously been shown to be expressed in breast cancer stem cells (Al Iajj et al. Proc. Natl. Acad. Sci. U. S A 100:3983-3988., herein incorporated by reference in its entirety). The overlap between the CD44 t /CD24- phenotype and CXCR1 expression was measured. 10 CXCRI-positive cells were present in the CD44+/CD24- cell population and the cell population expressing CD24 or CD44-negative (SEE Table 6). Table 6 CD241CD44+ CXC.R Ovedap £D241CD44+JCXCRi %%) Human breast cancer xenografts MCI 1)B 03 UM2 31.7 1 2 03 UM3 43 0.3 0.2 15 In residual tumors treated with docetaxel alone, either an unchanged or increased percent of CD44 t /CD24- cells was observed, whereas repertaxin treatment alone or in combination with docetaxel resulted in a reduction of the CD44*/CD24- cell population (SEE FIG 28). 20 A functional in vho assay consisting of re-implantation of cells from treated tumors into secondary NOD/SCID mice provided a direct test assessing the tumor initiating and self-renewal capacity of CSC remaining after treatment. Tumor cells derived from control or docetaxel-treated animals showed similar tumor regrowth at all dilutions in secondary NOD/SCID mice. In contrast, repertaxin treatment with or without 25 docetaxel, reduced tumor growth in secondary recipients (SEE FIG. Z26C). When equal 108 numbers of cells were injected, those from repertaxin-treated animals showed a 2-5-fold reduction in tumor growth compared to cells from control or docetaxel-treated animals (SEE FIG. 26C). For each xenotransplant model, 1000 or 100 tumor cells obtained from animals treated with a combination of repertaxin and docetaxel failed to form any 5 secondary tumors in NOD/SCID mice (SEE FIG. 26C, 27, and Table 7). These studies demonstrate that repertaxin treatment specifically targets and reduces the CSC population. Table 7 TumorsAnjections number of celis injected 10.00 50 2 500 1 000 500 250 100 Control 2 -- 8/8-- -- 68 Repertaxin 4/4 2/2 2/2 4/6 1/3 0/2 0/ Docetaxel: 2/2 44 22 6 6 3/4 2/3 8/9 Repertaxin/Docetaxel 2/2 3V4 2/2 1/ 1/4 0:4 0/9 10 Repertaxin treatment inhibits FAK/AKT signaling and activates FOXO3A in vivo. The expression of phospho-FAK and phospho-AKT was examined by immunohistochemistry in each of the xenotransplants after treatment. Membranous phospho-FAK expression 15 was detected in 50% of cells from the control and docetaxel-treated tumors whereas the phospho-FAK expression was abolished in the tumors treated with repertaxin alone or in combination with docetaxel (SEE FIG. 26D). Similar results were observed for the phospho-AKT expression. with 70% of cells expressing phospho-AKT in the untreated tumors, 20% phospho-AKT-positive cells in docetaxel-treated tumors and a complete 20 inhibition of phospho-AKT expression in the tumors treated with repertaxin alone or in combination with docetaxel (SEE FIG. 26D). Nuclear FOXO3A was detected in the cells from the tumors treated with docetaxel alone, repertaxin alone, and the combination 109 repertaxin/docetaxel. These in vivo data are consistent with the in vitro data and confirm that repertaxin treatment inhibits FAK/AKT signaling and activates FOXO3A. Repertaxin treatment reduces the development of systernic metastasis. To determine 5 whether repertaxin reduces systemic metastasis we infected HCCl 954, MDA-MB-453, and SU/M159 breast cancer cell lines with a luciferase lentivirus reporter systern and introduced the cells into NOD/SCD rice by intracardiac injection. A suspension of 250,000 cells for each cell line was injected and metastasis formation was monitored once per week by bioluminescent imaging. Twelve hours after intracardiac injection, 10 mice were treated twice per day by repertaxin injection or saline for the controls. Repertaxin treatment in mice injected with HCC 1954 and SUM 159 cells significantly reduced metastasis formation with a lower photon flux emission in the treated compared to the untreated mice (SEE FIG. 29A/B). Histologic sections confirmed the presence of metastases at several sites in untreated animals (SEE FIG. 29D). Repertaxin treatment 15 did not have any effect on metastasis formation in mice injected with MDA-MB-453 cells (SEE FIG. 29C). The photon flux emission and the number of animals that developed metastasis were similar in both repertaxin-treated and untreated group. This result is consistent with data that described MDA-MB-453 as a cell line resistant to repertaxin due to the presence of a PTEN mutation. These results indicate that CXCR1 20 blockade with agents such as repertaxin may be able to reduce metastasis which is mediated by the CSC population (Charafe-Jauffret et al. Cancer Res. 69:1302-13 13., herein incorporated by reference in its entirety). Experiments conducted during development of embodiments of the present 25 invention indicate that cellular subcomponents with stem cell properties drive tumor growth and metastasis Visvader et al, Nat. Rev. Cancer 8:755-768., herein incorporated by reference in its entirety). By virtue of their relative resistance to current therapeutic modalities, these cells may contribute to treatment resistance and relapse (Reya et al. Nature 414:105-111., herein incorporated by reference in its entirety). The present 30 invention provides an approach based on blocking the CXCR I cytokine receptor, which is expressed on breast cancer stern cells, to effectively target the cancer stem cell 110 population and to improve therapeutic outcome. Experiments conducted during development of embodiments of the present invention in a number of systems have demonstrated that cytokine networks play an important role in tumorigenesis. There is evidence that several of these cytokines may regulate stem cell behavior. IL-4 is capable 5 of regulating self-renewal of pancreatic cancer stein cells and IL-6 of regulating cancer stern cells in colon and breast cancer (Todaro et al. Cell Stem Cell 1:389-402., Sansone et al. J Clin. Invest 117:3988-4002., herein incorporated by reference in their entireties). The role of IL-8 in mediating tumor invasion and metastasis has previously been demonstrated (Waugh & Wilson. Cancer Res. 14:6735-6741., Inoue et al. Clin. Cancer 10 Res. 6:2104-2119., herein incorporated by reference in their entireties). In addition, IL-8 increases neural stem cell self-renewal during wound healing in the brain (Beech et al. J Neuroinununol. 184:198-208., herein incorporated by reference in its entirety). Lung cancer stem cells were described as expressing the chemokine receptor CXCRi (Levina et al. PLoS. ONE. 3:e3077., herein incorporated by reference in its entirety). Experiments 1s conducted during development of embodiments of the present invention demonstrated that the CXCRi-positive population is almost exclusively contained within the ALDEFLUOR-positive population in breast cancer cell lines and primary xenografts as well as in normal mammary cells. The chemokine receptor is overexpressed in ALDEFLUOR-positive breast cancer cell populations (Charafe-Jauffret et al. Cancer Res. 20 69:1302-1313., herein incorporated by reference in its entirety). In breast cancers, IL-8 is produced in the tumor microenvironinent by a number of cell types including inflammatory cells, vascular endothelial cells, tumor-associated fibroblasts and mesenchymal stem cells (Waugh et al. Clin. Cancer Res. 14:6735-6741., herein incorporated by reference in its entirety). Cytokine networks mediate interaction between 25 these cell types, therefore cancer stem cells can be targeted through the blockade of the IL-8 receptor CXCR1. Utilizing in vitro assays, it was demonstrated that CXCR1 but not CXCR2 (an alternative IL-8 receptor) blockade reduced the breast cancer stern cell population. This was followed by induction of apoptosis in the entire remaining cell population, which 30 lacks CXCR1 expression. In addition to CXCR1 blocking antibodies, experiments performed during development of embodiments demonstrate that repertaxin, a CXCR 1/2 111 inhibitor, induced similar effects by targeting the CXCR I -positive population. In contrast to its direct effects on the CXCRi-expressing cancer stem cell population, repertaxin had no direct effect on the bulk tumor cell population that lack CXCRI expression. This indicates that CXCRI blockade in CXCR1 -positive cells induced cell death in CXCRi 5 negative cells via a bystander effect. Experiments described herein demonstrate that the FAS-ligand/FAS pathway is the mediator of this bystander killing effect. This phenomenon explains the efficacy of repertaxin treatment in inducing massive apoptosis in the entire cell population despite the fact that the CXCRi -positive population represents less than 1% of the cell population. The role of FAS-ligand was demonstrated 10 by the effective blocking of bystander killing by anti-FAS-ligand antibody. Experiments conducted during development of embodiments of the present invention indicate that similar cytokine interactions may occur in tumors exposed to cytotoxic chemotherapy. Chemotherapy may directly induce cellular apoptosis in differentiated tumor cells as well as inducing the production of FAS-ligand by these 15 dying cells that in turn induces apoptosis in surrounding tumor cells via a FAS mediated bystander effect. Concomitant with the production of FAS-ligand, these injured cells also secrete increased levels of IL-8 in a process resembling mammary involution or wound healing. As is the case in the involuting mammary gland, this IL-8 may stimulate breast cancer stem cells as well as protecting them from apoptosis. This may contribute to the 20 relative increase in cancer stem cells observed after chemotherapy in preclinical models (4) and neo-adjuvant clinical trials (5). The effects of chemotherapy on apoptosis and self-renewal pathways in tumors are shown in Figure 30. To determine whether CXCR1 blockade could target breast cancer stem cells in vivo, the effects of the cytotoxic agent docetaxel were compared with repertaxin on the 25 cancer stein cell compartment and on tumor growth in NOD/SCID mice. Docetaxel is one of the most effective chemotherapeutic agents currently used to treat women with breast cancer. The cancer stem cell populations were assessed by the ALDEFLLUOR assay and by serial transplantation in NOD/SCID mice. Utilizing these assays it was determined that chemotherapy treatment alone resulted in either no change or a relative increase in 30 the cancer stem cell populations. In contrast, repertaxin treatment alone or with chemotherapy significantly reduced the cancer stem cell population. Despite the 112 significant reduction in t tumor-initiating populations, use of repertaxin alone did not result in significant umnor shrinkage. The combination of repertaxin plus chemotherapy resulted in significant reduction in tumor size as well as in the cancer stem cell population. Combining these agents to target both cancer stem cells and bulk tumor cell 5 populations maximizes the efficacy of these treatments. To elucidate the mechanism of action of repertaxin, the pathways downstream from CXCR I were analyzed. The interaction between CXCR 1, FAK and AKT was confirmed. CXCRl blockade acts specifically through FAK and AKT activation. Experiments conducted during development of embodiments of the present invention 10 indicate that AKT activation regulates normal and malignant breast stem cell self-renewal through phosphorylation of GSK3p resulting in the activation of the WNT pathway (Korkaya et al. PLoS Biolog. 7:e 1000121, herein incorporated by reference in its entirety). These results indicate why cells with PTEN knockdown are resistant to repertaxin. An additional function of AKT is the regulation of cell survival through 15 phosphorylation of the forkhead transcription factor FOXO3A. ART phosphorylation of FOXO3A results in its cytoplasmic sequestration. In contrast, it was demonstrated that CXCRi blockade leads to decreased AKT activation resulting in the translocation of FOXO3A in the nucleus whence it induces a number of genes including FAS-ligand (Jonsson et al. Nat. Med. 11:666-671., herein incorporated by reference in its entirety). 20 FAS-ligand induced via CXCRi blockade in turn is responsible for the observed bystander killing effects (SEE FIG. 30). In addition to its role in CXCR1 signaling, FAK mediates the interactions of cells with extracellular matrix components through integrin receptors (Waugh et al. Clin. Cancer Res. 14:6735-6741., herein incorporated by reference in its entirety). FAK 25 signaling plays a role in regulating the self-renewal of normal and malignant mouse mammary stem cells in transgenic models (Luo et al. Cancer Res. 69:466-474., herein incorporated by reference in its entirety). FAK activation also promotes cell survival by blocking FADD and RIP-inediated apoptosis (Kurenova et al. Mol. Cell Biol. 24:4361 4371., Xu et al. J Biol. Chem. 275:30597-30604., herein incorporated by reference in 30 their entireties). This provides an explanation for the resistance of the cancer stern cell population to the FAS/FAS-ligand induced apoptosis. 113 It has been demonstrated that breast cancer stem cells play an important role in turnor invasion and metastasis (Croker et al. J Cell Mol. Med. 2008, Charafe-Jauffret et al. Cancer Res. 69:1302-1313., herein incorporated by reference in their entireties). It is shown herein that IL-8 and CXCRI also play important roles in these processes. The 5 effects of CXCRI blockade was analyzed utilizing repertaxin on the formation of experimental metastasis. It was demonstrated that CXCRi blockade reduces the development of metastasis when administered subsequent to intracardiac injection of breast cancer cells. Clinical studies utilizing repertaxin have demonstrated a lack of toxicity. 10 Strategies aimed at interfering with cytokine regulatory loops such as IL-8 and CXCRi represent methods to target breast cancer stem cells. REFERENCES The following references are herein incorporated by reference in their entireties, 1s as if fully set forth herein. 1. Hanahan D and Weinberg R A. The hallmarks of cancer. Cell 2000; 100: 57-70. 2. Neve R M. Chin K, Fridlyand J et al. A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell Z2006; 10: 515-527. 3. Bonnet D and Dick J E. Human acute myeloid leukemia is organized as a 20 hierarchy that originates from a primitive hematopoietic cell. Nat.Med. 1997; 3: 730-737. 4. Glinsky G V. Stein cell origin of death-from-cancer phenotypes of human prostate and breast cancers. Stem Cell Rev. 2007; 3: 79-93. 5. Jaiswal S, Traver D, Miyamoto T, Akashi K, Lagasse E, and Weissman I L. Expression of BCRIABL and BCL-2 in myeloid progenitors leads to myeloid leukemias. 25 Proc.Natl.Acad.Sci.U.S.A 2003; 100: 10002-10007. 6. Krivtsov A V, Twomey D, Feng Z et al. Transformation from committed progenitor to leukemia stein cell initiated by MLL-AF9. Nature 2006; 4 4 2: 818-822. 7. Christgen M, Ballmaier M, Bruchhardt I, von Wasielewski R, Kreipe 11, and Lehmann U. Identification of a distinct side population of cancer cells in the Cal-51 30 human breast carcinoma cell line. Mol.Cell Biochem. 2007; 306: 201-212. 114 8. Fillmore C M and Kupeirwasser C. Human breast cancer cell lines contain stem like cells that self-renew, give rise to phenotypically diverse progeny and survive chemotherapy. Breast Cancer Res. 2008; 10: R25. 9. Kondo T, Setoguchi T, and Taga T. Persistence of a small subpopulation of 5 cancer stem-like cells in the C6 glioma cell line. Proc.Natl.Acad.Sci.U.S.A 2004; 101: 781-786. 10. Setoguchi T, Taga T, and Kondo T. Cancer stem cells persist in many cancer cell lines. Cell Cycle 2004; 3: 414-415. 11. Patrawala L, Calhoun T, Schneider-Broussard R, Zhou J, Claypool K, and Tang D 10 G. Side population is enriched in tumorigenic, stem-like cancer cells, whereas ABCG2+ and A. Cancer Res. 2005; 65: 6207-6219. 12. Chute J P, Muramoto G G, Whitesides J et al. Inhibition of aldehyde dehydrogenase and retinoid signaling induces the expansion of human hematopoietic stem cells. Proc.Natl.Acad.Sci.U.S.A 2006; 103: 11707-11712. 15 13. Duester G. Families of retinoid dehydrogenases regulating vitamin A function: production of visual pigment and retinoic acid. Eur.J Biochem. 2000; 267: 4315-4324. 14. Cheung A M, Wan T S. Leung J C et al. Aldehyde dehydrogenase activity in leukemic blasts defines a subgroup of acute mycloid leukemia with adverse prognosis and superior NOD/SCID engrafting potential. Leukemia 2007: 2 1: 1423-1430. 20 15. Corti S, Locatelli F. Papadimitriou D et al. Identification of a primitive brain derived neural stein cell population based on aldehyde dehydrogenase activity. Stem Cells 2006; 24: 975-985. 16. Pearce D J, Taussig D, Simpson C et al. Characterization of cells with a high aldehyde dehydrogenase activity from cord blood and acute myeloid leukemia samples. 25 Stem Cells 2005; 23: 752-760. 17. Ginestier C, Hur M HI, Charafe-Jauffret E et al. ALDI Is a Marker of Normal and Malignant Human Mammary Stem Cells and a Predictor of Poor Clinical Outcome. Cell Stern Cell 2007; 1: 555-567. 18. Dontu G, Abdallah W M, Foley J M et al. In vitro propagation and transcriptional 30 profiling of human mammary stemn/progenitor cells. Genes Dev. 2003; 17: 1253-1270. 115 19. Irizarry R A, I-obbs B, Collin F et a]. Exploration, normalization, and summaries of high density oligonucleotide array probe level data. Biostatistics. 2003; 4: 249-264. 20. Charafe-Jauffret E, Ginestier C, Monville F et al. Gene expression profiling of breast cell lines identifies potential new basal markers. Oncogene 2006; 25: 2273-2284. 5 21. Finetti P, Cervera N, Charafe-Jauffret E et al. Sixteen-kinase gene expression identifies lurninal breast cancers with poor prognosis. Cancer Res. 2008; 68: 767-776. 22. Eisen M B, Spellman P T, Brown P 0, and Botstein D. Cluster analysis and display of genome-wide expression patterns. Proc.Natl.Acad.Sci.1.S.A 1998; 95: 14863 14868. 10 23. Reiner A, Ytkutieli D, and Benjamini Y. Identifying differentially expressed genes using false discovery rate controlling procedures. Bioinformatics. 2003; 19: 368 24. Hua J, Balagurunathan Y, Chen Y et al. Normalization benefits microarray-based classification. EURASIP.J Bioinform.Syst.Biol. 2006; 43056. 15 25. Ginestier C, Cervera N, Finetti P et al. Prognosis and gene expression profiling of 20ql3-amplified breast cancers. Clin.Cancer Res. 2006; 12: 45334544. 26. Ponti D, Costa A, Zaffaroni N et al. Isolation and in vitro propagation of tumorigenic breast cancer cells with stem/progenitor cell properties. Cancer Res. 2005; 65: 5506-5511. 20 27. Ringe J, Strassburg S. Neumann K et al. Towards in situ tissue repair: human mesenchymal stem cells express chemokine receptors CXCR 1, CXCR2 and CCR2, and migrate upon stimulation with CXCL8 but not CCL2, J Cell Biochem. 2007; 101: 135 146. 28. Hughes L, Malone C, Chumsri S, Burger A M, and McDonnell S. 25 Characterisation of breast cancer cell lines and establishment of a novel isogenic subelone to study migration, invasion and tumourigenicity. Clin.Exp.Metastasis 2008. 29. Itoh Y, Joh T, Tanida S et al. IL-8 promotes cell proliferation and migration through metalloproteinase-cleavage proH-IB-EGF in hurnan colon carcinoma cells. Cytokine 2005; 29: 275-282. 30 30. Gupta G P, Perk J, Acharyya S et a]. I D genes imediate tumor reinitiation during breast cancer lung metastasis. Proc.Natl.Acad.Sci.J.S.A 2007; 104: 19506-19511. 116 31. Li F, Tiede B, Massague J, and Kaing Y. Beyond tumorigenesis: cancer stem cells in metastasis. Cell Res. 2007; 17: 3-14. 32. Al JHajj M, Wicha M S, Benito-[Hernandez A, Morrison S J, and Clarke M F. Prospective identification of tumorigenic breast cancer cells. Proc.Natl.Acad.Sci.U.S.A 5 2003; 100: 3983-3988. 33. Li C, Heidt D G, Dalerba P et al. Identification of pancreatic cancer stem cells. Cancer Res. 2007; 67: 1030-1037. 34. Ricci-Vitiani L, Lombardi D G, Pilozzi E et al. Identification and expansion of human colon-cancer-initiating cells. Nature 2007; 445: 111-115. 10 35. Montanaro F, Liadaki K, Schienda J, Flint A, Gussoni E, and Kunkel L M. Demystifying SP cell purification: viability, yield, and phenotype are defined by isolation parameters. Exp.Cell Res. 2004; 298: 144-154. 36. Stingl J, Eirew P, Ricketson I et al. Purification and unique properties of mammary epithelial stem cells. Nature 2006; 439: 993-997. 15 37. Matsui W, Huff C A, Wang Q et al. Characterization of clonogenic multiple myeloma cells. Blood 2004; 103: 2332-2336. 38. Farnie G and Clarke R B. Mammary stem cells and breast cancer--role of Notch signalling. Stem Cell Rev. 2007; 3: 169-175. 39. Krstic A, Mojisin M, and Stevanovic M. Regulation of SOX3 gene expression is 20 driven by multiple NF-Y binding elements. Arch.Biochem.Biophys. 2007; 467: 163-173. 40. Zhu J, Zhang Y, Joe G J, Pompetti R, and Emerson S G. NF-Ya activates multiple hematopoietic stein cell (HSC) regulatory genes and promotes HSC self-renewal. Proc.Natl.Acad.Sci.U.S.A 2005; 102: 11728-1173 41. Raffel G D, Mercher T, Shigematsu H et al. Ottl (Rbmi 5) has pleiotropic roles in 25 hematopoietic development. Proc.Nati.Acad.Sci.L.S.A 2007; 104: 6001-6006. 42. Ma X, Renda M J, Wang L et al, Rbm15 modulates Notch-induced transcriptional activation and affects myeloid differentiation. Mol.Cell Biol. 2007; 27: 3056-3064. 43. Peiffer 1, Eid P, Barbet R et al. A sub-population of high proliferative potential quiescent human mesenchymal stem cells is under the reversible control of interferon 30 alpha/beta. Leukemia 2007; 21: 714-724. 117 44. Villadsen R, Fridriksdottir A J, Ronnov-Jessen L et al. Evidence for a stem cell hierarchy in the adult human breast. J Cell Biol. 2007; 177: 87-101. 45. Harnbardzumyan D, Becher O J, and Holland E C. Cancer stern cells and survival pathways. Cell Cycle 2008; 7. 5 46. Jagani Z and Khosravi-Far R. Cancer stern cells and impaired apoptosis. Adv.Exp.Med.Biol. 2008; 615: 331-344. 47. Maxwell P J, Gallagher R, Seaton A et al. HIF-i and NF-kappaB-inediated upregulation of CXCR 1 and CXCR2 expression promotes cell survival in hypoxic prostate cancer cells. Oncogene 2007; 26: 7333-7345. 10 48. Murphy C, McGurk M, Pettigrew J et al. Nonapical and cytoplasmic expression of interleukin-8, CXCR1, and CXCR2 correlates with cell proliferation and microvessel density in prostate cancer. Clin.Cancer Res. 2005; 11: 4117-4127. 49. Trentin L, Miorin M, Facco M et al. Multiple myclioma plasma cells show different chemokine receptor profiles at sites of disease activity. Br.J Haematol. 2007; 15 138: 594-602. 50. Varney M L, Johansson S L, and Singh R K. Distinct expression of CXCL8 and its receptors CXCRI and CXCR2 and their association with vessel density and aggressiveness in malignant melanoma. Am.J Clin.Pathol. 2006; 125: 209-216. 51. Freund A, Chauveau C, Brouiliet J P et al. IL-8 expression and its possible 20 relationship with estrogen-receptor-negative status of breast cancer cells. Oncogene 2003; 22: 256-265. 52. Inoue K, Slaton J W, Eve B Y ei al. Interleukin 8 expression regulates tumorigenicity and metastases in androgen-independent prostate cancer. Clin.Cancer Res. 2000; 6: 2104-2119. 25 53. Balbay M D, Pettaway C A, Kuniyasu H et al. Highly metastatic human prostate cancer growing within the prostate of athymic mice overexpresses vascular endothelial growth factor. Clin.Cancer Res. 1999; 5: 783-789. 54. Kirn S J, Uehara H, Karashirna T, Mccarty M, Shih N, and Fidler I J. Expression of interleukin-8 correlates with angiogenesis, tumorigenicity, and metastasis of human 30 prostate cancer cells implanted orthotopically in nude mice. Neoplasia. 2001; 3: 33-42. 118 55. Karnoub A E, Dash A B, Vo A P et al. Mesenchymal stem cells within tumour stroma promote breast cancer metastasis. Nature 2007; 449: 557-563. 56. Schafer Z T and Brugge J S. IL-6 involvement in epithelial cancers. J Clin.lnvest 2007; 117: 3660-3663. 5 57. Todaro M, Alea M P, Di Stefano A B et al. Colon cancer stem cells dictate tumor growth and resist cell death by production of interleukin-4. Cell Stem Cell 2007; 1: 389 402. 58. Landi S, Bottari F, Gemignani F et al. Interleukin-4 and interleukin-4 receptor polymorphisms and colorectal cancer risk. Eur.J Cancer 2007; 43: 762-768. 10 59. Sansone P, Storci G, Tavolari S et al. IL-6 triggers malignant features in mamimospheres from human ductal breast carcinoma and normal mammary gland. J Clin.Invest 2007; 117: 3988-4002. 60. Glinsky G V, Berezovska 0, and Glinskii A B. Microarray analysis identifies a death-from-cancer signature predicting therapy failure in patients with multiple types of 15 cancer. J Clin.Invest 2005; 115: 1503-1521. 61. Golub T R. Slonim D K, Tamayo P et al. Molecular classification of cancer: class discovery and class prediction by gene expression monitoring. Science 1999: 286: 531 537,. 20 All publications and patents mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described method and system of the invention will be apparent to those skilled in the art without departing fTom the scope and spirit of the invention. Although the invention has been described in 25 connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention that are obvious to those skilled in the relevant fields are intended to be within the scope of the present invention. 30 119

Claims (9)

1. A method of detecting solid tumor stem cells in a subject comprising detecting CXCR1+ cells in a tissue sample taken from a tumor of said subject.

2. The method of claim 1, wherein said detecting comprises contacting said tissue sample with an antibody or antibody fragment. 3 The method of claim 2, wherein said antibody or antibody fragment comprises a signal molecule.

4. The method of claim 3, wherein said signal molecule comprises a flourescent molecule or an enzyme that can catalyze a color producing reaction in the presence of a colorimetric substrate.

7. The method of claim 1, wherein said tumor is selected from the group consisting of: a prostate cancer tumor, an ovarian cancer tumor, a breast cancer tumor, a melanoma, a non-small cell lung cancer tumor, a small-cell lung cancer tumor, and an esophageal adenocarcinoma tumor.

8. A method for treating a subject having a tumor comprising administering a compound to said subject, wherein said compound is a CXCR1 antagonist or an IL8-CXCR1 signaling pathway antagonist.

9. A method of claims 8 wherein both cancer stem cells and non-tumorigenic cancer cells in a subject having a tumor are reduced by the administration of the CXCR1 antagonist or an IL8 CXCR1 signaling pathway antagonist.

10. The method of claim 8, wherein said compound is Repertaxin or a Repertaxin derivative. 121

11. The methods of claim 8, wherein said CXCR1 antagonist, or IL8-CXCR1 signaling pathway antagonist comprises an antibody or antibody fragment.

12. The method of claim 8, wherein said tumor comprises cancer stem cells selected from the group consisting of: prostate cancer stem cells, ovarian cancer stem cells, breast cancer stem cells, skin cancer stem cells, non-small cell lung cancer stem cells, small-cell lung cancer stem cells, and esophageal adenocarcinoma stem cells. The Regents of the University of Michigan Patent Attorneys for the Applicant/Nominated Person SPRUSON & FERGUSON