In Table 1, the various tumor types which have thus far been most strongly correlated with EMT are presented with a brief review of known EMT markers. Cancer types are ordered by the estimated percentage of diagnosed patients who have survived 5-year following cancer metastasis. While this table is by no means exhaustive, it helps highlight several interesting trends. For example, the Snail, Twist, Zeb, and E-cadherin axis as described above, has thus far been correlated with nearly every clinically significant tumor type. Furthermore, the striking commonalities between these distinct tumors reveal the profound clinical importance of EMT as a shared, ubiquitous mechanism that promotes metastasis. Fittingly, the field of oncology has seen a recent explosion of EMT-related research for both prognostication and treatment of metastatic cancers, and to date, numerous classical EMT markers have been significantly correlated with metastasis. Moreover, recent works suggest that assessing classical markers of EMT may help clinicians predict resistance to chemotherapy, and thus poor prognosis [56].

Table 1 Major tumor types organized by virulence, clinical significance, and epigenetic markers Full size table

Another exciting area of research is the use of EMT markers in the analysis of circulating tumor cells (CTC). Diagnostically, CTC has been a mainstay of clinical practice in assessment of metastasis and prognosis. The presence of CTC in a patient’s blood can be measured using the AdnaTest, a PCR assay for markers of EMT such as Twist, Akt, and Pi3k. The test employs a method for enriching the CTCs in a blood sample using antibodies conjugated to magnetic beads. Once the tumor cells have been pulled down, the mRNA can be isolated and expression of EMT markers determined. The test is reported to be sensitive enough to detect two CTCs in a 5 mL sample of blood [57]. Recent works have indicated that consideration of CTC EMT status is critical to achieve a more accurate prognosis. In studies of metastatic breast cancer, CTC were found to express known EMT regulators, including TGF-β pathway components and the FOXC1 transcription factor. These data support a role for EMT in the blood-borne dissemination of human breast cancer. Classical markers of EMT, Twist, and vimentin, have been identified in breast cancer patients and specifically show elevated expression in patients with metastatic cancer relative to patients with early-stage cancer, supporting the hypothesis that EMT controls the metastatic potential of CTCs [58]. Importantly, other work suggests that EMT-CTCs may be more likely to evade classical CTC detection by the AdnaTest as a result of down-regulation of EpCAM. As suggested by Gorges et al., this may explain why patients with late metastatic cancers may report low CTC numbers, suggesting the urgent need for a better understanding of EMT-CTC in prognosis [59,60].

Pancreatic cancer

Pancreatic cancer generally has a poor prognosis, in part because symptoms often do not appear until the cancer is too advanced for surgical treatment. Pancreatic exocrine tumors have an average 5 year survival of up to 14%. Neuroendocrine tumors have a 61% 5-year survival rate if detected at Stage 1, but these tumors are rarely detected at this phase [2,89]. Thus, early detection and inhibition of metastasis remain among the greatest challenges in the treatment of these tumors. Several genes related to EMT have been considered with respect to these clinical challenges. In one in vitro study, Hh inhibition with cyclopamine resulted in down-regulation of Snail and up-regulation of E-cadherin, as well as a striking reduction of invasive capacity. Combining gemcitabine and cyclopamine completely abrogated metastasis while also significantly reducing the size of “primary” tumors. These findings suggest that inhibition of the Hh pathway is a valid therapeutic strategy for pancreatic cancer that particularly targets metastasis [64,65]. Similarly, Resveratrol, which inhibits pluripotency-maintaining factors such as Kras (G12D), and EMT have been indicated in the management of pancreatic cancer [90,91].

Hepatocellular carcinoma

Hepatocellular carcinoma (HCC), which is among the most deadly forms of cancers worldwide, is the most common primary liver cancer and is the fastest growing cause of cancer death in men in the United States [92]. The dominant risk factors are chronic Hepatitis B or Hepatitis C infection. In addition, cirrhosis can have an effect on the tumor microenvironment as well as on tumorigenesis. Cirrhosis can lead to the activation of stellate cells, which increase production of extracellular matrix proteins, cytokines, and growth factors, many of which can alter hepatocyte proliferation and promote tumorigenesis [93,94]. HCC tends to have a poor prognosis due to late diagnoses and a lack of effective treatment options. While EGFR-targeted therapies have been successful in some types of cancers, erlotinib and cetuximab have not been very effective in clinical HCC trials, particularly in the treatment of mesenchymal HCC cells. In the case of hepatic carcinomas, Sorafenib, which inhibits STAT3 and phosphorylates TGF-β which are both up-regulated in EMT, is also being studied as a potential therapeutic agent [67].

Squamous cell carcinoma

Vimentin positive tumor cells have been detected among squamous cell carcinomas; although, high epithelial vimentin has not been correlated with tumor grade. Squamous cell carcinomas tend to have periostin rich stroma. Periostin is usually localized to the periphery of stromal cells surrounding carcinoma cells. Expression of versican and periostin were frequently accentuated toward the pseudo-basement membrane of the extracellular matrix around these carcinomas, and high stromal vimentin is associated with higher grade [71]. Since EMT plays a large role in the development and spread of lung cancer, numerous drugs that specifically target EMT are being developed or are in use in the treatment of lung cancer. For example, Sorafenib has been show to increase HAT expression in adenocarcinoma, therefore positively influencing the epigenetic profile of the cancer cells [95]. Furthermore, an immunotherapeutic approach to target a major driver of EMT, the T-box transcription factor T, also known as brachyury, is currently in Phase I clinical trial as a potential new therapy for patients with advanced lung cancer carcinomas [96,97].

Pulmonary adenocarcinoma

Adenocarcinoma is a type of cancerous tumor that forms from glandular structures [98]. Stromal periostin protein is associated with versican collagen, and tumor cell epithelial periostin is associated with both versican and vimentin. Each of these associations suggests that cancer cells have undergone EMT and become more metastatic, but surprisingly, this study did not find a correlation between vimentin up-regulation and morphological trans-differentiation. However, the authors observed that the up-regulation of stromal vimentin, periostin, and versican is associated with higher cancer grades. As vimentin is the constituent of the cytoskeleton network, it is possible that stromal populations go through certain changes during the induction of EMT. Similar results were found in breast carcinoma [68,69].

Urothelial carcinoma

Urothelial carcinoma makes up the majority of bladder cancers and has a high likelihood of returning after treatment. The most common treatment is surgery if the carcinoma is detected in an early stage. Urothelial cancers are further classified as either superficial or muscle invasive.

Renal cancer

Via blood filtration, the kidneys are exposed to a disproportionately high concentration of toxins. Thus, perhaps it is not surprising that renal cancer is one of the 10 most common cancers. EMT has also been observed in mature epithelial tubular cells and has been linked to the pathogenesis of renal interstitial fibrosis. Furthermore, in mouse models it has been demonstrated that the selective blockade of EMT-associated TGF-β, hepatocyte growth factor, and BMP-7 expression reduces fibrotic lesions after obstructive injury [74].

Colorectal cancer

Colorectal cancers tend to start as a small growth in the inner lining of the colon known as a polyp, ultimately giving rise to adenocarcinomas. Colorectal cancer is one of the most common cancers, and yet it is not among the most lethal cancers as early clinical detection via routine screenings has dramatically improved overall mortality [99]. Still, careful study of EMT markers has revealed additional clinically relevant information. A clear link has been established between CD44, enhancement of EMT, and colon cancer invasion [100]. Furthermore, FGFR4 has also been shown to play a crucial role in tumorigenesis, invasion, and survival in colorectal cancer, and its specific targeting marks a new avenue of colorectal cancer therapy [101]. Vimentin is highly expressed in the stroma of colorectal cancer cells compared to healthy cells, but interestingly, not in the cancer cells themselves. Higher levels of stromal vimentin have been correlated with poor prognosis of colorectal cancer. Specifically, since vimentin is expressed in mesenchymal cells and not epithelial cells, it indicates that EMT has taken place [102].

Cervical cancer

Perhaps the most significant recent breakthroughs with respect to cervical cancer have come from the understanding that human papilloma virus (HPV) silences tumor suppressor genes through production of proteins E6 and E7. However, as worldwide immunization campaigns evolve, cervical cancer persists as a clinical challenge, and stage IV cervical cancer is still generally considered untreatable, though chemotherapy is recommended which uses platinum drugs [103]. Several EMT genes have recently been explored as potential biomarkers or targets of drug treatment in cervical cancer. For example, FTS silencing was found to reduce EMT and cell migration by EGF treatment [104]. Importantly, Twist2 has been identified as the key Twist isoform coupling aberrant signals from EMT to senescence, with significant implications on its potential utility as a biomarker of cervical cancer prognosis [75,76].

Melanoma

TGF-β and EMT regulation markers such as MITF have been shown to play a critical role in melanoma progression. Furthermore, up-regulation of N-cadherin has been correlated with an increase in cell migration and invasion. Recent works have demonstrated the causal role of TGF-β-induced EMT-like changes on downstream activation of PI3K in human melanoma cells, which may ultimately yield new therapeutic options for these highly aggressive cancers [77]. Another significant recent insight has been that the EMT-like switch in phenotype is associated with a concomitant change in the expression of multiple tumor antigens, ultimately allowing cells to evade T-cell killing. This may have important implications for future immune therapies such as cancer vaccination, and careful selection of target antigens may help circumvent the problem of T-cell evasion by metastatic melanoma cells [78].

Ovarian cancer

Mutations in the BRCA1 and BRCA2 genes may contribute to development of ovarian cancer. PTEN and STK11 (a tumor suppressor protein related to EMT) may also be risk factors. Furthermore, CCR7, which can be induced in response to hypoxia and is often constitutively expressed in epithelial ovarian cancer cells, has been shown to participate in EMT development, leading to cell migration and invasion. This suggests that CCR7 may be an effective target for limiting cell invasion in certain ovarian cancers [80]. Other recent work has linked hTERT to Slug expression in norepinephrine-induced ovarian cancer EMT and metastasis. This suggests that these genes may serve as novel biomarkers and potential therapeutic targets for ovarian cancer [81,82].

Breast cancer

CTCs that have undergone EMT have been found in patients with HER2 (+) metastatic breast cancer. CD326 (−) and CD45 (−) cells show an enrichment of circulating stem cells (CSCs), and have been shown to be correlated with classical markers of EMT such as Snail1 and Zeb1 [84]. Therefore, assessing EMT-CTCs and CSCs in HER2 (+) breast cancer patients could be of great prognostic value [84]. Additionally, high levels of CD44 and low levels of CD24 have been linked to chemotherapy resistance and cancer relapse in metastatic breast cancer. Clinically, Lapatinib in combination with conventional therapy, was demonstrated as a possible therapeutic strategy for eliminating these cells to decrease recurrence and improve long-term survival [105].

Prostate cancer

During prostate cancer progression, as the cells undergo EMT, the stroma undergoes structural rearrangement in order to accommodate the tumor cell. Tumor cells can evade apoptosis by changing their relationship to the ECM. One marker of a reactive stroma is the presence of myofibroblasts, which is a cellular intermediate between fibroblasts and smooth muscle cells [106]. These cells secrete fibronectin, collagen, and proteoglycans such as versican and decorin [107-111]. The reactive stroma is not only responsible for assisting in EMT but also contributes to tumor vascularization [112]. Aberrant glycosylation also impacts such EMT and cell adhesion [113]. Several patterns of gene silencing have been documented in advancing prostate cancer. Genes such as APC, RASSF1A, CCND2, and RAR-β2 are silenced even in less virulent (low Gleason score) tumors, and loss of E-cadherin, GSTP1, and ER tend to be silenced in more aggressive tumors [85-88]. Approximately half of prostate cancers carry TMPRSS2-ERG translocations; however, the clinical impact of this genomic alteration remains unclear. Recent studies have suggested that ILK is a therapeutically targetable mediator of ERG-induced EMT and transformation in prostate cancer [114].

Glioblastoma

Arising from astrocytes, glioblastoma is the most common primary and most aggressive CNS tumor subtype. A particular challenge to treat, the tumors are generally very heterogeneous, and thus, some cells may respond to treatment while others may not. Glioblastomas are highly malignant in part because they reproduce quickly and have access to many blood vessels, but rarely spread to distant locations in the body. In glioblastoma multiform, EMT has been shown to cooperate with MMP activity, allowing cells to gain access to lymph vessels. Preliminary data suggest this new EMT-associated drug target in combination with stereotactic radiosurgery may provide potential targets for future treatment [115].

Smoking

Though beyond the scope of this paper, the role of smoking in the pathogenesis of EMT is also of high clinical significance. Recent works have established direct connections between cigarette smoke and acute inflammatory mechanisms such as NF-kB and EMT [116]. Although the bulk of the evidence for this relationship has been considered with respect to lung cancers, it is likely that these mechanisms will be more explicitly implicated for other tumor types as well. Thus, epidemiologically, smoking cessation may ultimately prove among the most important clinical interventions relevant to EMT.