Extracellular signalregulated kinases 1 and 2 mitogenactivated protein

Our previous studies, using antibodies raised to the dually phosphorylated pTEpY region within the catalytic core of the active form of ERK1 and ERK2 MAP kinase, are supportive of a pivotal role for exaggerated ERK1/2 MAP kinase activation (pMAP kinase) in ER-positive and -negative endocrine-resistant tumor growth.103 Immunocytochemistry using such antibodies generated heterogeneous nuclear immunostaining for pMAP kinase within formalin-fixed, paraffin-embedded human breast tumor specimens (Figure 10.3), and two distinct subgroups of patients were readily identifiable based on the resultant staining.

The first group of patients demonstrated very low pMAP kinase. Many such patients exhibited objective responses (i.e. complete (CR) or partial (PR) responses) to tamoxifen as measured at 6 months after initiation of antihormonal therapy. Responses were of extended duration with a longer patient survival, as measured from the initiation of endocrine therapy. These low levels of pMAP kinase may be critical to the growth of such tumors since, using sequential tamoxifen-treated samples obtained from primary elderly breast cancer patients, we have been able to detect further decreases in pMAP kinase in parallel with the clinical tamoxifen response profile. Moreover, as stated above, we have demonstrated some cell growth inhibition of the endocrine responsive breast cancer line MCF-7 with the MEK1 inhibitor PD098059, despite only minimal activation of ERK1/2 MAP kinase. Such data again suggest that the diminished pMAP kinase levels detectable in endocrine responsive disease may be reflective of highly efficient regulation of enzyme phosphorylation. Some link has been made between pMAP kinase regulation and the IGF-1R pathway in ER-positive well-differentiated tumors, although it should be noted that there was some conflict with our own observations since higher pMAP kinase levels were reported for this patient group.104 The second patient subgroup in our study (72% of patients) exhibited quite substantial pMAP kinase. Such patients invariably exhibited de novo endocrine resistant (i.e. progressive) disease, or at best disease stabilization. There was a poorer survival from initiation of endocrine therapy (by univariate analysis) and a shortened time to disease relapse on such treatment (by univariate and multivariate analysis103). Biochemical studies using tissue homogenates confirm that hyperexpression and anomalous MAP kinase activation is a feature of a proportion of breast neoplasms.105-108 Furthermore, a biochemical study by Mueller et al109 reported a relationship between elevated MAP kinase activity and shortened disease-free survival in primary breast cancer, which is complementary to our own data. However, such observations remain controversial since links between increased pMAP kinase and good outcome have also been reported.110 The reasons underlying such variations in results remain unknown; however, localization appears critical to the observations since, while nuclear ERK correlates with shortened patient survival, cytoplasmic staining is favorable.111

What mechanisms might underlie any exaggeration of pMAP kinase in poorer prognosis de novo endocrine resistant clinical breast cancer? Mutation, overexpression or constitutive activity of ERK1/2 MAP kinase,105'106'108'112 or indeed of any of the key regulators identified in cell models of endocrine resisitance such as growth factor receptors (EGFR, HER2) and phosphatases (MKP3), might feasibly explain this phenomenon. Indeed, an increasing number of anomalies in erbB/ERK1/2 MAP kinase signaling have been identified within such tumors.113 Associations have been made between endocrine independence and exaggeration of EGFR and HER2,113 and very recent studies have shown that activation of HER2 correlates with pMAP kinase (and also with the alternative kinase p38114), while inhibition of EGFR and/or HER2 with agents such as lapatinib and gefitinib can deplete pMAP kinase, and thereby proliferation in breast cancer, suggesting interlinked path-ways.115 It is also interesting that elevated levels and/or activity of many additional intracellular molecules impinging on the ERK 1/2 MAP kinase signaling pathway (including pp60c-src, Grb2, RHAMM, Ras, Raf, protein kinase C) have been observed in malignant breast cells, commonly associating with a poorer patient prognosis.106,116-119

Interestingly, we noted that increased pMAP kinase was particularly common in the poorer prognosis, endocrine-unresponsive ER-negative patient subgroup.103 Such tumors are reported to employ elevated EGFR signaling for their expansion.44'45'113'120'121 Interestingly, these findings parallel the evidence derived from cell models implicating MAP kinase signaling in driving ER negativity. In these tumors, we observed associations between pMAP kinase, EGFR positivity, and the activated AP-1 component c-Jun,122 data implicating pMAP kinase as a key intermediary of elevated EGFR signaling which impinges on AP-1-mediated events, and thereby growth of ER-negative disease. Our in vivo observations are complemented by in vitro studies, which have similarly demonstrated enhanced tyrosine phosphorylation,74 and marked ERK1/2 MAP kinase activation in ER-negative MDA-MB-231 breast cancer cells.123

Importantly, however, we noted that associations between increased pMAP kinase and hormone refractory disease in the clinic were also retained within ER-positive patients.103 Multivariate analysis confirmed significant associations with earlier relapse on endocrine therapy and poorer survival time in these patients. Further studies have also linked increased MAPK activity and impaired tamoxi-fen response in ER-positive patients,124 although, as stated above, observations linking MAPK activation to adverse outcome remain controversial. Our clinical data are complimented by many in vitro observations equating enhanced MAP kinase activity with the acquisition of steroid hormone independence or antihormone resistance by ERpositive breast cancer cells, including our own panel of endocrine resistant MCF-7 sub-lines.7879 Indeed, we have recently observed increased pMAP kinase at the time of acquisition of tamoxifen resistance and disease relapse in ER-positive, initially responsive, clinical disease. In total, these data offer considerable support of a central role for exaggerated pMAP kinase in sustaining anti-hormonal-resistant ER-positive tumor growth. As observed in the model systems, there is also emerging clinical evidence that pMAP kinase may be able to promote ER activity in

ER-positive breast cancer. Prominent activation of ER serine 118 has been reported to correlate with more differentiated disease and better clinical outcome on tamoxifen.113,125,126 However, such ER activity is also readily detectable in ER-positive de novo and acquired tamoxifen-resistant breast cancer (moreover at elevated levels in relapse samples). Interestingly, both increased EGFR and pMAP kinase correlate with phosphorylation of serine 118 ER in clinical disease,113,126 while Poly-chronis et al127 observed that neoadjuvant gefitinib treatment of ER-positive/EGFR-positive disease depletes both pMAP kinase and serine 118 ER phosphorylation. In total, these data suggest EGFR/MAP kinase regulation of ER activation may be important to tamoxifen-resistant phenotypes in vivo as in breast cancer cell models.113,125 Surprisingly, our study showed that there was a lack of direct correlation between pMAP kinase and expression of a panel of "classically" estrogen regulated genes (i.e. PgR, pS2 and bcl-2), although some association was noted between TGFa, a known estrogen responsive gene, and pMAP kinase.103 These data indicate that the mechanisms involving pMAP kinase priming of ER activity are far from simple, and may selectively influence specific subsets of estrogen-regulated genes more integral in tumor growth processes. Moreover, while initially entering into positive crosstalk with ER, at its most extreme, hyperactivation of MAP kinase may ultimately act to inhibit ER expression, thereby producing completely endocrine refractory growth.

10 Ways To Fight Off Cancer

10 Ways To Fight Off Cancer

Learning About 10 Ways Fight Off Cancer Can Have Amazing Benefits For Your Life The Best Tips On How To Keep This Killer At Bay Discovering that you or a loved one has cancer can be utterly terrifying. All the same, once you comprehend the causes of cancer and learn how to reverse those causes, you or your loved one may have more than a fighting chance of beating out cancer.

Get My Free Ebook

Post a comment