Cannibalism, cell survival, and endocrine resistance in breast cancer

Breast cancer cells often respond to an endocrine therapy by altering expression of specific estrogen-responsive genes and inducing autophagy, a cannibalistic lysosomal pathway. Autophagy eliminates damaged or other organelles, allowing the recovery of the energy stored in their macromolecules to attempt restoration of metabolic homeostasis. Induction of autophagy can result from activation of the unfolded protein response following metabolic stress, the final cell fate often being determined by the extent and duration of autophagy. A study by Gonzalez-Malerva and colleagues builds upon this extensive knowledge, adding HSPB8 to the list of altered genes associated with endocrine resistance in breast cancer and describing the ability of HSPB8 to regulate autophagy and confer tamoxifen resistance.


Background
Approximately 70% of newly diagnosed breast cancers express detectable levels of estrogen receptor alpha (ERα). Patients with these tumors generally receive an endocrine-based intervention, usually in the form of an aromatase inhibitor or antiestrogen. While these interven tions are associated with a signifi cant increase in overall survival [1], many ERα-positive breast cancers recur.
Breast cancer cells often respond to endocrine therapies by altering the expression of a subset of estrogenresponsive and nonresponsive genes and inducing autophagy [2-7]. Autophagy is a lysosomal self-digestion pathway, where selected subcellular components are fi rst segregated into double membrane-bound structures called autophagosomes [8]. Subsequent fusion with lysosomes forms autolysosomes, which enables the materials sequestered therein to be digested by lysosomal hydrolases. Th ree primary forms of autophagy have been described: macroautophagy, microautophagy, and chaperonemediated autophagy. Th e comments in this viewpoint relate primarily to macroautophagy.
Autophagy can be prodeath or prosurvival and refl ects an attempt by stressed cells to eliminate damaged or other organelles and recover the energy stored in their macromolecules to restore metabolic homeostasis. Autophagy can result from activation of the unfolded protein response (UPR) [9] in response to the metabolic stress of endocrine therapies [4]. In endocrine manipulated cells, prosurvival autophagy signaling is driven at least partly by UPR activation of the uncon ven tional splicing of the estrogen-responsive XBP1 mRNA [3,10] and, downstream, increased expression of BCL2 family members including BCL2 and BCLW (BCL2L2) [11]. Prodeath signaling can involve induction of a caspasedependent apoptotic cell death (programmed cell death 1) [12] when there is adequate energy, and an autophagyassociated necrotic cell death that is probably energy independent [11]. An autophagic cell death, often referred to as programmed cell death 2, also occurs. Th e extent of autophagy induced in response to stress may be controlled by the respective abilities of BCL2 and BCLW, and perhaps other BCL2 family members, to bind and sequester BECN1 [11], a key inducer of autophagy [8]. Regulation of the mammalian target of rapamycin (mTOR) by AKT also aff ects the induction of autophagy, and AKT is associated with UPR signaling [9].
An individual cell's fate can be determined by the extent and duration of autophagy. Prolonged exposure to a high level of autophagy can leave the cell with insuffi cient mitochondria and/or other organelles to survive (prodeath autophagy). Adequate recovery of energy by autophagy, with a suffi cient remaining subcellular infrastructure, can allow the cell to survive (prosurvival autophagy) [13]. In the face of endocrine therapies, this survival is manifest as drug resistance.

The article
A recent study by Gonzalez-Malerva and colleagues used gene expression microarray analysis, and a functional

Abstract
Breast cancer cells often respond to an endocrine therapy by altering expression of specifi c estrogenresponsive genes and inducing autophagy, a cannibalistic lysosomal pathway. Autophagy eliminates damaged or other organelles, allowing the recovery of the energy stored in their macromolecules to attempt restoration of metabolic homeostasis. Induction of autophagy can result from activation of the unfolded protein response following metabolic stress, the fi nal cell fate often being determined by the extent and duration of autophagy. A study by Gonzalez-Malerva and colleagues builds upon this extensive knowledge, adding HSPB8 to the list of altered genes associated with endocrine resistance in breast cancer and describing the ability of HSPB8 to regulate autophagy and confer tamoxifen resistance. genomic approach of overexpressing >500 full-length human kinase cDNAs, to study resistance in a series of MCF-7 cells with diff ering tamoxifen responses [14]. Th irty-one kinases were identifi ed that both enhanced cell proliferation in the presence of 4-hydroxytamoxifen and were validated in at least four of six independent experiments. ERBB2 and IGF1R were among those genes that scored at least twice in these experiments, were diff erentially regulated in the microarray dataset, and are known to aff ect endocrine responsiveness [2]. Among the genes identifi ed, the authors selected HSPB8 (heat shock 22 kDa protein 8; E2-induced gene protein 1; α-crystallin C chain) for further evaluation because this gene gave one of the strongest responses in their primary assay, validated in fi ve out of six screens, and was present in their microarray signature. A role for HSBP8 in tamoxifen resistance has not previously been reported.
Subsequent analyses in vitro showed that HSBP8 reduced the antiproliferative eff ects of 4-hydroxy tamoxifen and blocked its ability to induce autophagy. Th e eff ects on autophagy were not mediated by chemical inhibi tion of the autophagy regulator mTOR, although the mTOR pathway was aff ected as assessed by changes in p70S6k P and 4-EBP1 P . HSBP8 similarly aff ected these two proteins but independently of mTOR. An association of HSPB8 (α-crystallin C chain) expression with a poor clinical outcome was described in one clinical gene expression dataset [14], an association previously reported for the α-crystallin B chain protein [15]. Overall, the authors concluded that autophagy may be an important player in aff ecting tamoxifen sensitivity and noted that the mechanism of HSBP8 action requires further study.

Viewpoint
Activation of autophagy is clearly a key determinant of the nature of the response to antiestrogens in some breast cancer cells. In cells that are forced (and able) to activate an energy-dependent programmed cell death, it seems probable that autophagy provides both the energy and part of the cell destruction machinery. Th ere are advantages to an orderly cell death that results in the release of at least partly digested cellular components, which may be less likely to induce the destructive infl ammatory response associated with an apparently chaotic necrotic cell death. Adjacent cells in the micro environment may be able to scavenge and more effi ciently use released macromolecules to support their own attempt to evade the shared metabolic stress. For endocrine therapies, the prolonged duration of autophagy -even when prodeath is not immediately lethal -may explain why cell death is generally slow in vitro and why complete pathological responses are infrequent even among highly responsive ERα-positive breast tumors. While the study by Gonzalez-Malerva and colleagues primarily addressed prodeath autophagy, a prosurvival autophagic adaptation to the nutrient deprivation present in poorly vascularized and/or endocrine-resistant tumors may explain the some times decades-long dormancy associated with breast cancer development and recurrence.
Gonzalez-Malerva and colleagues clearly add HSBP8 to the list of genes associated with endocrine resistance and autophagy [14]. How HSBP8 infl uences autophagy is unclear but the mechanism does not seem to involve mTOR. Th e researchers did not look in detail at other mechanisms such as those that aff ect BECN1 action, a pathway known to aff ect autophagy in endocrine responsive ness. Th e authors also implicate pathways indepen dent of HSBP8 in antiestrogen resistance. Whether HSBP8 will be validated as an independent prognostic or predictive marker is not clear, since this was not seen in the independent dataset [14]. Nonetheless, HSBP8 is an interesting and important new addition to our understanding of cannibalistic autophagy signaling in endocrine resistance.