Research Grants Awarded
Critical Role Of Integrin-Associated Cd151 In Progression Of Er-Negative Mammary Tumors
Career Catalyst Research
Laminin-binding integrins (alpha6beta4, alpha6beta1, alpha3beta1) play critical roles during the initiation, progression and malignancy of mammary tumors. CD151, a 28 kDa member of the tetraspanin protein family, strongly and directly associates with laminin-binding integrins (alpha6beta4, alpha6beta1, alpha3beta1, alpha7beta1) via a critical extracellular protein interaction site. CD151 modifies integrin-dependent morphology, migration, invasion, and adhesion strengthening for laminin-binding integrins, but not other integrins. Surprisingly, CD151-null mice develop normally, although they show deficiencies in pathological angiogenesis, wound healing, and growth of tumor xenografts. CD151 expression has consistently been linked to invasion, metastasis, and poor prognosis in breast human cancer and other tumor types.
Our preliminary data show significant elevation in CD151 expression in total human ER-negative tumors, in basal-like tumors (ER-/HER- compared to ER+/HER-), and in high grade mammary tumors, regardless of subtype. Furthermore, in CD151-null mice crossed with mice expressing the MMTV-ErbB2 transgene, formation of spontaneous mammary tumors was markedly delayed. Also, ablation of CD151, via shRNA, from human basal-like MDA-MB-231 mammary carcinoma cells caused a considerable delay in ectopic tumor growth in nude mice. These findings suggest that CD151, in concert with laminin-binding integrins, supports the progression of both ErbB2-driven and basal-type breast cancer.
Additional preliminary experiments provide insights into the CD151 mechanism of action. When CD151 was ablated, migration and invasion of mammary tumor cell lines was severely impaired in vitro. Notably, defects in EGF-dependent migration and invasion assays could not be overcome by adding more EGF, suggestive of impaired integrin-EGFR synergy. In addition, laminin-binding integrin-dependent signaling through focal adhesion kinase (FAK), Lck, and Rac1 was markedly diminished. Furthermore, the absence of CD151 essentially disrupted the molecular neighborhood of laminin-binding integrins - without CD151, molecular links were severed between laminin-binding integrins and several other known and unknown membrane proteins. From these results we suggest that CD151-dependent effects on mammary cell signaling, motility, and tumor progression are largely due to CD151 playing a central role in the organization of laminin-binding integrins into multicomponent molecular complexes.
To test our hypotheses we will carry out the following specific aims:
Aim I. We will examine the putative tumor-promoting role of CD151 in vivo. A) From human patient data we will look for a correlation between CD151+ breast cancer and diminished survival. B) Effects of CD151 on ErbB2-driven mouse mammary tumor formation will be examined in vivo. C) Xenografts of human basal-like tumors, with or without CD151, will be examined in mammary fat pads, and upon tail vein injection. These experiments will establish definitively in vivo roles for CD151 in ER-negative mammary cancers in vivo.
Aim II. We will determine the extent to which manipulation of CD151 (either by knockdown or by inhibitory agents) affects A) integrin alpha6beta4, ErbB2 and EGFR-dependent mammary cell proliferation, migration and signaling, and B) mammary cell sensitivity to ErbB inhibitors such as Iressa, Herceptin, and Cetuximab. These results will determine the extent to which CD151 enhances synergy between laminin-binding integrins and ErbB receptors.
Aim III. To investigate molecular mechanisms for CD151 function we will A) analyze mammary tumor signaling pathways (involving FAK, Lck, Akt, Rac 1) and B) identify and prioritize proteins linked, via CD151, to laminin-binding integrins in ErbB2 and basal type mammary tumor cells. These results should provide insights into the roles of particular molecules, recruited by CD151, that may account for its functions.
It may be therapeutically beneficial to target laminin-binding integrins on breast cancer cells. However, laminin-binding integrins also make major contributions to normal physiology. By contrast, the contribution of CD151 appears to be restricted to pathological conditions. Hence, CD151 may be a fundamentally better target than the laminin-binding integrins. Our proposed experiments will determine the extent to which CD151 contributes to mammary tumor progression, and will provide mechanistic insights into how this is achieved. The role of CD151 during breast cancer progression has never before been definitively addressed. Hence, our approach is novel.
Human breast cancer is an enormous world-wide problem. We have obtained preliminary data showing that some of the worst types of human breast cancers contain a small protein called CD151. Furthermore, we have discovered that CD151 protein is not simply present, but also is contributing to breast cancer having a bad outcome. To make this discovery, we used a mouse, genetically engineered to undergo spontaneous breast cancer formation. When the CD151 gene was removed from that mouse, spontaneous formation of breast cancer was considerably delayed. In another experiment, removal of CD151 from human breast cancer cells caused them to grow more slowly when implanted into a mouse. These preliminary findings strongly suggest that CD151 protein contributes to the progression of multiple subtypes of breast cancer.
Additional preliminary experiments were aimed at discovering what CD151 is doing in breast cancer cells. When we removed CD151 from these cells, several things happened. First, the cells were no longer able to respond to growth factors ? small soluble proteins that stimulate tumor growth and movement. Second, the breast tumor cells no longer were able to invade into artificial membranes. Third, the breast cancer cells lost their ability to transmit key molecular signals to the inside of the cell. Fourth, several important proteins that are normally clustered on the surface of breast tumor cells became disorganized and disconnected. These preliminary results give good initial insights into how CD151 functions on mammary tumor cells.
Though suggestive, our preliminary results are not definitive. Now we seek to establish whether the presence of CD151, in a statistically significant manner, correlates with loss of survival in humans with breast cancer. Also, we will determine whether the absence of CD151 protein causes multiple types of breast cancer to be significantly delayed in mice. In addition, we will investigate whether interference with CD151 can make breast cancer cells more sensitive to drugs that inhibit growth factors. Finally, we will identify specific molecules and pathways that are organized by CD151 protein, while contributing to breast cancer growth and invasion.
Our proposed experiments may lead to improved therapy for breast cancer. If we can show that CD151 is indeed contributing to breast cancer progression, then the next step would be to design drugs that inhibit CD151, thereby inhibiting breast cancer progression. Identification of key molecules that interact with CD151 should enable the intelligent design of anti-CD151 drugs that are specific and effective. If interfering with CD151 does indeed make breast cancer cells more sensitive to drugs that inhibit growth factors, that would represent a major step forward in improving the effectiveness of those drugs, which are increasingly being utilized clinically. Finally, because CD151 is not needed for normal life, it may be possible to design drugs that inhibit CD151 without causing too many unwanted side effects.