Chemokine Receptor Signaling in Breast Cancer

Institution: University of California, San Diego
Investigator(s): Morgan O'Hayre, B.S. -
Award Cycle: 2008 (Cycle 14) Grant #: 14GB-0147 Award: $74,660
Award Type: Dissertation Award
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease



Initial Award Abstract (2008)

Chemokines are critical mediators of leukocyte cell migration during routine immune surveillance, lymphocyte development and homing, and inflammation. In parallel with their role in controlling cell migration during immune responses, chemokines and their receptors have been implicated in the cancer metastatic process by directing the migration of tumor cells. The chemokine receptors CXCR4 and CXCR7 are not expressed on normal breast epithelia but they are frequently expressed on breast cancer cells, and there is preferential metastasis of breast cancer cells to sites where their ligand, called CXCL12, is constitutively expressed. However, the exact mechanisms of how CXCR4 and CXCR7 enhance metastasis and/or tumor growth and their full implications on disease progression are unknown. Yet it is likely that chemokine receptor signaling may provide more than just a migrational advantage by also helping the metastasized cells establish and survive in secondary environments.

The goal of the proposed studies is to characterize the mechanisms and functional roles of CXCL12, and its receptors, CXCR4 (Aim 1) and CXCR7 (Aim 2), in breast cancer. In particular, we plan to characterize how these receptors promote the survival, proliferation, and metastasis of these cancer cells. Two breast cancer cell lines (MDA-MB 231 and MCF-7) will be selected for high expression levels of CXCR4 or CXCR7. The selected cells and the starting population compared in terms of their ability to directly or indirectly promote cell proliferation and survival. They will also be studied using a mouse model of tumor metastasis to gain a better understanding of how receptor expression influences tumor growth and rate of metastasis. Cells from the primary breast tumors and subsequent metastatic sites will be collected to more extensively explore how the chemokine receptor levels have changed and how the cells respond to CXCL12 ligand. Specific inhibitors of CXCR4 and CXCR7 will be used to help dissect the individual contributions of these receptors to breast cancer and standard survival and proliferation pathways (e.g., Akt, ERK1/2, NF-?B) will be analyzed for their contribution to chemokine-dependent metastasis.

The research is distinct from the traditional approaches of understanding the genetic mechanisms of breast cancer malignancy as it is aimed at examining the roles of signals from the local environment in metastasis, which has received much less attention. The potential impact of the proposed research is in making current chemotherapies more effective in destroying metastatic lesions, by combining them with molecules that reduce the inherent viability and growth of cancer cells. Such molecule inhibitors of CXCR4, are already in clinical trials for AIDS, and thus there is potential for such drug combinations to be translated rapidly into cancer therapeutics.




Final Report (2010)

Chemokines and their receptors play an important role in the immune system by guiding the migration of cells involved in routine immune surveillance and inflammatory responses. However, they can also be exploited by cancer cells to facilitate metastasis and enhance tumor growth. Our work has focused on elucidating the roles of the chemokine, CXCL12, and its receptors, CXCR4 and CXCR7, in breast cancer progression. Although CXCR4 and CXCR7 are not normally expressed in breast tissue, they are often upregulated (increased) in breast cancer and contribute to metastasis and primary tumor growth, respectively. While there is strong evidence for the role of CXCL12/CXCR4 in metastasis, little is known about how CXCR4 contributes to the growth and survival of primary tumors or metastasized cells. Additionally, little is known regarding the involvement of CXCR7 in breast cancer progression and whether it functions through redundant or distinct mechanisms and/or in conjunction with CXCR4. Thus, our goal has been to explore how these two chemokine receptors function independently and in conjunction in breast cancer.

Over the past two years, we have made considerable progress towards understanding the individual and collective contributions of CXCR4 and CXCR7 to breast cancer tumor growth/survival and metastasis, although the complexity of the system has also generated a number of new questions. Results thus far suggest that CXCR4 and CXCR7 do not serve redundant functions. This is supported by findings from other groups demonstrating that CXCR7 does not signal like CXCR4 through traditional chemokine receptor mechanisms, and from our findings of their distinct localizations in the cell where CXCR7 exhibits intracellular accumulation while CXCR4 is mostly surface expressed. We demonstrated that while both CXCR4 and CXCR7 independently could accelerate primary tumor growth of breast cancer cells, CXCR4 appeared to have a stronger effect. Additionally, expression of CXCR4 but not CXCR7 in these cells enhanced rates of metastasis to the lungs and lymph nodes. Intriguingly, high expression of CXCR7 with CXCR4 delayed and reduced tumor progression as well as metastasis. High levels of CXCR7 when co-expressed with CXCR4 could also reduce or delay some of the signaling functions of CXCR4, including calcium mobilization. Furthermore, we identified the tumor suppressor protein, programmed cell death factor 4 (PDCD4), as a novel target of CXCL12 signaling. PDCD4 is an important modulator of cell growth and so provides a potential mechanism by which CXCL12/CXCR4 may contribute to breast cancer cell growth. High expression of CXCR7 with CXCR4 also delayed and reduced the phosphorylation of PDCD4 by CXCL12 in breast cancer cells. However, the inhibitory effects of CXCR7 on CXCR4 function appears to strongly depend on the levels of CXCR7; thus, low or moderate levels of CXCR7 may not be inhibitory and could potentially even enhance breast cancer tumor growth.

Further experiments to investigate how varying levels of CXCR7 in conjunction with CXCR4 may affect breast cancer tumor growth and metastasis are currently being pursued. Future experiments to determine if the inhibitory effects of high CXCR7 observed were due to sequestration of CXCL12 and/or due to heterodimerization of CXCR4 and CXCR7 are also planned. We will use selective antagonists to the receptors to better dissect the functions of these two chemokine receptors in breast cancer progression and continue to evaluate their individual and collective effects on breast cancer tumor growth and metastasis.




Symposium Abstract (2010)

Morgan O’Hayre (PI), Catherina Salanga, Jing Yang, and Tracy Handel (mentor)

Chemokines and their receptors play an important role in the immune system by guiding the migration of cells involved in routine immune surveillance and inflammatory responses. However, they can also be exploited by cancer cells to facilitate metastasis and enhance tumor growth. Our work focuses on elucidating the roles of the chemokine, CXCL12, and its receptors, CXCR4 and CXCR7, in breast cancer. Although CXCR4 and CXCR7 are not normally expressed in breast tissue, they are often expressed on breast cancer cells and are known to contribute to metastasis and primary tumor growth, respectively. However, the extent by which CXCR4 and CXCR7 serve redundant verses distinct functions has not been established, nor the underlying mechanisms of how they contribute to disease. Thus, our goal is to explore how these two chemokine receptors function independently and in conjunction in breast cancer progression.

In order to address these questions, we derived sublines from the MDA-MB231 human breast cancer cell line that express low levels of both receptors, high levels of CXCR4, high levels of CXCR7 or high levels of both receptors. Consistent with published observations, our results indicate that cells with high expression of either CXCR4 or CXCR7 resulted in accelerated tumor growth compared to those with low expression. Furthermore, we found that high CXCR4 but not high CXCR7 resulted in increased metastasis, consistent with previous findings from studies of the individual receptors. However, we observed striking differences in the effects of dual receptor expression on tumor growth depending on “dose” of the receptors. When both CXCR4 and CXCR7 are expressed at moderate levels, an increase in tumor growth and metastasis is observed compared to cells with either receptor expressed alone. However, upon high expression of CXCR7 with CXCR4, an inhibitory effect is observed, whereupon tumor growth rate is significantly reduced, even compared to the control cells with very low levels of receptor expression. In order to better understand the molecular mechanisms underlying these contrasting results, we are currently comparing how differential levels of receptor expression influences intracellular signaling events involved in cell survival and proliferation. Additionally, we have demonstrated that a key distinction between these two receptors is that CXCR4 primarily localizes at the surface of MDA-MB231 breast cancer cells, while CXCR7 largely localizes inside the cell. Therefore, receptor localization may be important for understanding the dose-dependent differences of CXCR4 and CXCR7 expression on tumor growth/metastasis.

Results thus far suggest that CXCR4 and CXCR7 do not serve redundant functions although they influence each others functions both at the cellular level and in breast cancer growth and metastasis. Understanding the mechanisms by which CXCR7 cooperates with CXCR4 when expressed at moderate levels, and how it is inhibitory when expressed at high levels will be an important goal of future studies. These findings could have significant implications in terms of disease aggressiveness and the effectiveness of targeting the receptors and downstream signaling pathways for the treatment of breast cancer.



Modulation of chemokine receptor activity through dimerization and crosstalk.
Periodical:Cell and Molecular Life Sciences
Index Medicus: Cell Mol Life Sci
Authors: Salanga CL, O'Hayre M, Handel T
Yr: 2009 Vol: 66 Nbr: 8 Abs: Pg:1370-86

Elucidating the CXCL12/CXCR4 signaling network in chronic lymphocytic leukemia through phosphoproteomics analysis.
Periodical:PLOS One
Index Medicus: PLOS One
Authors: O'Hayre M, Salanga CL, Kipps TJ, Messmer D, Dorrestein PC, Handel TM
Yr: 2010 Vol: 5 Nbr: 7 Abs: Pg:e11716

Phosphoproteomic analysis of chemokine signaling networks.
Periodical:Methods in Enzymology
Index Medicus: Methods Enzymol
Authors: O'Hayre M, Salanga CL, Dorrestein PC, Handel TM
Yr: 2009 Vol: 460 Nbr: Abs: Pg:331-46