**Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases**

Noemí Eiró, Belen Fernandez-Garcia, Antonio Altadill, Luis O. González and Francisco J. Vizoso

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/56877

### **1. Introduction**

Liver cancer is the fifth and seventh most frequently diagnosed cancer worldwide in men and women, respectively, but the second most frequent cause of cancer death in men [1]. In addition, hepatocellular carcinoma (HCC) represents the major histological subtype of primary liver cancers, accounting up to 90% of the total liver cancer burden worldwide [2]. The incidence of HCC has increased significantly over the past 10 years and is expected to increase because of the actual high prevalence of viral hepatitis C-seropositive individuals and also because of the known long latency period to HCC development from the initial hepatitis C virus (HCV) infection, which may take 2-3 decades [3]. Despite of the varied treatment options, the prognosis of HCC remains poor. Thus, estimated 5-year survival rates are in the range of 26% to 50%, and disease-free survival is 13% to 29% [4]. At present, systemic chemo‐ therapy is quite ineffective in HCC treatment, and is also known to express the multidrugresistance gene MDR-1 [5]. Therefore, is necessary to identify and characterize molecular abnormalities of clinical significance in HCC. Besides the heterogeneity of different HCC subtypes, these tumors may use different cellular pathways and oncogenic mechanisms at different development stages, and this is of essential importance to develop biologically-based clinical trials.

### **2. Steroid hormone receptors and hepatocellular carcinoma**

Although sex differences in liver cancer may be attributed to differences in lifestyle [6-8], there are several epidemiological and experimental studies suggesting that HCC might be, in part, hormone-related. Liver cancer is predominantly a male disease, with approximately three

© 2013 Eiró et al.; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

times higher incidence and mortality among men than women [9]. This male predominance is further supported by the fact that chronic liver disease progresses more rapidly to cirrhosis in males than in females, and therefore cirrhosis-derived HCC is largely seen in men and postmenopausal women disease [10].

**Reference Protein /mRNA Type of liver**

**tissue**

[127] ER protein Normal BA cytosolic and

[129] ER protein Non-cancerous BA cytosolic and

[135] ER protein Normal BA 9 9

[127] ER protein HCC BA cytosolic and

[139] ER protein HCC BA cytosolic and

[141] ER protein HCC BA cytosolic and

[135] ER protein HCC BA cytosolic and

**Method Localization n**

nuclear

nuclear

[125] ER protein Normal BA - 6 100 Germany 1982 [126] ER protein Normal BA - 6 100 United States 1983

[128] ER protein Surrounding BA cytosolic 30 43 Japan 1986

[130] ER protein Non-cirrhotic EIA cytosolic 12 12 Japan 1987 [131] ER protein Normal - 2 2 Japan 1988 [132] ER protein Surrounding BA cytosolic 17 65 Japan 1989 [133] ER protein Surrounding 22 64 Japan 1989 [134] ER protein Surrounding BA cytosolic 5 100 Japan 1990

[52] ER protein Non-cancerous BA cytosolic 26 42 Spain 1993 [136] ER mRNA Non-cancerous ISH 13 54 Italy 1993 [137] ER mRNA Peritumoral RT-PCR - 32 88 Korea 2006 [124] ER protein HCC BA cytosolic 5 100 United States 1982

[138] ER protein HCC IHC cytosolic 10 10 Singapore 1984 [128] ER protein HCC BA cytosolic 30 40 Japan 1986

[130] ER protein HCC EIA cytosolic 13 38 Japan 1987 [132] ER protein HCC BA cytosolic 19 37 Japan 1989 [24] ER protein HCC BA cytosolic 66 39 Japan 1990 [134] ER protein HCC BA cytosolic 6 17 Japan 1990 [140] ER protein HCC BA cytosolic 21 48 Japan 1991

nuclear

nuclear

nuclear

nuclear

9 89

9 89

**% positive cases**

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

**Country Year**

297

http://dx.doi.org/10.5772/56877

4 100 United Kingdom 1983

7 43 Japan 1986

United States, Italy

5 100 United Kingdom 1983

8 13 Japan 1986

United States, Italy

United States, Italy

1991

1991

1991

The role of estrogens in modulating morphological and physiological features of liver became evident in early 1970s when a possible correlation between occurrence of hepatic neoplasms and use of oral contraceptives was suggested [11-12]. On the basis of these data, several *in vitro* and *in vivo* studies have explored the importance of sex hormones in HCC. Animal modelbased studies indicate that sex hormones play a key role in tumor progression, showing that ovarian estrogens protect against tumor progression, whereas androgens promote tumori‐ genesis [13-15]. It has been reported that the protective effect of estrogens against chemically induced liver tumors is mediated by prolactin (PRL) through liver prolactin receptors (PRLR) [16-18]. Nevertheless, the precise role of male and female sex hormones and their receptors in HCC remains still poorly understood. There is not enough information regarding the mecha‐ nism of estrogen and androgen in HCC.

Estrogen and androgen mediate their biological functions through binding to their specific receptors, the estrogen receptor (ER) and the androgen receptor (AR). Both ER and AR belong to the nuclear receptors family that, as transcription factors, regulate the expression level of several genes such as those involved in triggering immune responses, cell proliferation and apoptosis [19-21]. Therefore, sex hormones play a key role in normal physiology of organs other than these of the reproductive system.

Variable expressions of ER and AR has been found in normal liver and HCC using different methods, such as immunohistochemistry, enzyme-immuno assays, or determining mRNA levels (Table 1 and Table 2 give the details of these studies for ER and AR, respectively), which indicate a relationship between sex hormones and pathogenesis of HCC. The percentage of positivity for these receptors varied between the different studies [22]. These differences may be due to several methodological aspects, such as differences in the origin of the studied patients populations, sample types or technologies determining the receptor expression. In a recent study, we determined the expression of sex hormones receptors in 31 HCC patients by immunohistochemistry using tissue micro-arrays technology [23]. Our results demonstrate a wide variability in the immunohistochemical values for steroid receptors among HCCs: 67.7% of tumors stained positively for AR, 51.6% for ER and 83.8% for progesterone receptor (PgR), but, among the positive cases, immunostaining score values for each protein were largely variable.



times higher incidence and mortality among men than women [9]. This male predominance is further supported by the fact that chronic liver disease progresses more rapidly to cirrhosis in males than in females, and therefore cirrhosis-derived HCC is largely seen in men and

The role of estrogens in modulating morphological and physiological features of liver became evident in early 1970s when a possible correlation between occurrence of hepatic neoplasms and use of oral contraceptives was suggested [11-12]. On the basis of these data, several *in vitro* and *in vivo* studies have explored the importance of sex hormones in HCC. Animal modelbased studies indicate that sex hormones play a key role in tumor progression, showing that ovarian estrogens protect against tumor progression, whereas androgens promote tumori‐ genesis [13-15]. It has been reported that the protective effect of estrogens against chemically induced liver tumors is mediated by prolactin (PRL) through liver prolactin receptors (PRLR) [16-18]. Nevertheless, the precise role of male and female sex hormones and their receptors in HCC remains still poorly understood. There is not enough information regarding the mecha‐

Estrogen and androgen mediate their biological functions through binding to their specific receptors, the estrogen receptor (ER) and the androgen receptor (AR). Both ER and AR belong to the nuclear receptors family that, as transcription factors, regulate the expression level of several genes such as those involved in triggering immune responses, cell proliferation and apoptosis [19-21]. Therefore, sex hormones play a key role in normal physiology of organs

Variable expressions of ER and AR has been found in normal liver and HCC using different methods, such as immunohistochemistry, enzyme-immuno assays, or determining mRNA levels (Table 1 and Table 2 give the details of these studies for ER and AR, respectively), which indicate a relationship between sex hormones and pathogenesis of HCC. The percentage of positivity for these receptors varied between the different studies [22]. These differences may be due to several methodological aspects, such as differences in the origin of the studied patients populations, sample types or technologies determining the receptor expression. In a recent study, we determined the expression of sex hormones receptors in 31 HCC patients by immunohistochemistry using tissue micro-arrays technology [23]. Our results demonstrate a wide variability in the immunohistochemical values for steroid receptors among HCCs: 67.7% of tumors stained positively for AR, 51.6% for ER and 83.8% for progesterone receptor (PgR), but, among the positive cases, immunostaining score values for each protein were largely

**Method Localization n**

[122] ER protein Normal BA cytosolic 4 100 United Kingdom 1978 [123] ER protein Normal BA - 3 100 Germany 1978 [124] ER protein Normal BA - 2 100 United States 1982

**% positive cases**

**Country Year**

postmenopausal women disease [10].

296 Hepatocellular Carcinoma - Future Outlook

nism of estrogen and androgen in HCC.

other than these of the reproductive system.

**Reference Protein /mRNA Type of liver**

**tissue**

variable.


**Reference Protein /mRNA Type of liver**

**tissue**

[127] AR protein HCC BA cytosolic and

[150] AR protein HCC BA cytosolic and

[139] AR protein HCC BA cytosolic and

[135] AR protein HCC BA cytosolic and

transcriptase-polymerase Chain reaction; IHC: Immunohistochemistry.

overall survival in patients with resectable HCC [30,26].

[151] AR protein HCC BA cytosolic 13 100

**Method Localization <sup>n</sup> % positive**

5 100

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

5 100

9 100

nuclear

nuclear

nuclear

nuclear

[138] AR protein HCC IHC cytosolic 10 50 Singapore 1984 [149] AR protein HCC BA cytosolic 19 74 Japan 1985

[132] AR protein HCC BA cytosolic 19 37 Japan 1989 [27] AR protein HCC BA cytosolic 45 69 Japan 1989 [134] AR protein HCC BA cytosolic 11 64 Japan 1990 [140] AR protein HCC BA cytosolic 21 86 Japan 1991

[152] AR protein HCC BA cytosolic 5 100 Japan 1992 [52] AR protein HCC BA cytosolic 26 54 Spain 1993 [28] AR protein HCC BA cytosolic 43 65 Spain 1995 [23] AR protein HCC IHC nuclear 31 68 Spain 2007 [129] AR mRNA HCC ISH - 22 73 Italy 1994 [153] AR mRNA HCC RT-PCR - 38 89 Italy 2002 [137] AR mRNA HCC RT-PCR - 32 100 Korea 2006

AR: Androgen receptor; HCC: Hepatocellular carcinoma; BA: Binding assay; ISH: *In situ* hybridization; RT-PCR: Reverse

Several studies have analyzed the relationship between ER or AR and clinicopathological parameters from HCCs patients and their clinical outcome (Table 3). In general, ER expression was associated with higher tumor aggressiveness and/or worse prognosis in HCC patients [24-26], whereas AR expression was negatively associated with recurrence [27-29]. It is remarkable the finding that the presence of the ERα variant has been associated with shortened

**Table 2.** Androgen receptor expression in normal liver and hepatocellular carcinoma tissue samples.

**cases Country Year**

http://dx.doi.org/10.5772/56877

United Kingdom

United Kingdom

United Kingdom

United States, Italy

8 50 Japan 1986

1983

299

1985

1988

1991

ER: Estrogen receptor; HCC: Hepatocellular carcinoma; BA: Binding assay; EIA: Enzyme immunoassay; ISH: *In situ* hybridization; RT-PCR: Reverse transcriptase-polymerase Chain reaction for ERα wild type; IHC: Immunohistochemistry.

**Table 1.** Estrogen expression in normal liver and hepatocellular carcinoma tissue samples.



**Reference Protein /mRNA Type of liver**

298 Hepatocellular Carcinoma - Future Outlook

**Reference Protein /mRNA Type of liver**

**tissue**

[146] ER protein HCC IHC cytosolic and

**Table 1.** Estrogen expression in normal liver and hepatocellular carcinoma tissue samples.

[135] AR protein Normal - - 9 100

**tissue**

[127] AR protein Normal BA cytosolic and

[139] AR protein Non-cancerous BA cytosolic and

**Method Localization n**

nuclear

**Method Localization <sup>n</sup> % positive**

4 0

nuclear

nuclear

[132] AR protein Non-cancerous BA cytosolic 17 65 Japan 1989 [133] AR protein Surrounding 21 33 Japan 1989 [134] AR protein Surrounding BA cytosolic 10 80 Japan 1990

[137] AR mRNA Peritumoral RT-PCR 32 100 Korea 2006

[147] ER protein HCC IHC - 28 39 China 2004 [147] ER mRNA HCC RT-PCR - 28 89 China 2004 [137] ER mRNA HCC RT-PCR - 32 100 Korea 2006 [148] ER protein HCC IHC - 66 5 Mexico 2007 [23] ER protein HCC IHC Nuclear 31 52 Spain 2007

ER: Estrogen receptor; HCC: Hepatocellular carcinoma; BA: Binding assay; EIA: Enzyme immunoassay; ISH: *In situ* hybridization; RT-PCR: Reverse transcriptase-polymerase Chain reaction for ERα wild type; IHC: Immunohistochemistry.

[52] ER protein HCC EIA cytosolic 26 15 Spain 1993 [136] ER protein HCC IHC - 15 0 Italy 1993 [136] ER mRNA HCC ISH - 15 73 Italy 1993 [142] ER mRNA HCC RT-PCR - 14 57 Italy 1995 [25] ER protein HCC EIA cytosolic 33 39 Germany 1997 [143] ER protein HCC IHC cytosolic 71 24 China 1997 [144] ER mRNA HCC RT-PCR - 40 70 Italy 1998 [145] ER mRNA HCC RT-PCR - 42 60 Italy 2003

**% positive cases**

45 71

**Country Year**

United States/ Koreans

**cases Country Year**

6 17 Japan 1986

United Kingdom

United States, Italy

1983

1991

2004

AR: Androgen receptor; HCC: Hepatocellular carcinoma; BA: Binding assay; ISH: *In situ* hybridization; RT-PCR: Reverse transcriptase-polymerase Chain reaction; IHC: Immunohistochemistry.

**Table 2.** Androgen receptor expression in normal liver and hepatocellular carcinoma tissue samples.

Several studies have analyzed the relationship between ER or AR and clinicopathological parameters from HCCs patients and their clinical outcome (Table 3). In general, ER expression was associated with higher tumor aggressiveness and/or worse prognosis in HCC patients [24-26], whereas AR expression was negatively associated with recurrence [27-29]. It is remarkable the finding that the presence of the ERα variant has been associated with shortened overall survival in patients with resectable HCC [30,26].

### **3. Steroid receptors and hepatitis virus**

It is of special interest the interaction between sex estrogen receptors and viral proteins in hepatitis B virus (HBV) and hepatitis C virus (HCV)-induced HCC. It has been reported that a HBV protein (HBx) interacts with ERα. HBx is a multifunctional protein involved in neoplastic transformation in cultured cells and in HCC induction in transgenic mice. Both HBx and vERα (Delta 5 deletion variant of ERα) have additive effects on suppressing ERα trans‐ activation [31]. In addition, it was reported that tamoxifen inhibits ERα actions and suppresses HCV genome replication [32], which may be of potential interest to develop new anti-HCV strategies based on anti-ER drugs. On the other hand, higher serum androgen concentrations or a specific AR gene, which leads to higher AR activities, have also been associated to higher risk in HBV-mediated HCC [33-34]. Likewise, it has been reported that the combination of male gender and HBV infection had a significant synergistic effect on HCC progression [35]. Most recently, Ming-Heng *et al*. found that hepatic AR increases the HBV viral titer by enhancing HBV RNA transcription through direct binding to the androgen response element near the viral core promoter. This activity forms a positive feedback mechanism with the cooperation of its downstream target, the HBx protein, to promote hepatocarcinogenesis. In addition in these same study administration of a chemical compound that selectively degrades AR, ASC-J9, was able to suppress HCC tumor size in a transgenic HBV mouse model that developed HCC upon exposure to a low dose of N'-N'-diethylnitrosamine (DEN). These results demonstrate that targeting the AR, rather than the androgen, could be developed as a new therapy to battle HBV-induced HCC [36].

**Reference**

**Protein / mRNA**

[28] AR protein

outcome of patients with HCC.

[132] AR protein Recurrence and survival rate

[29] AR protein Tumor size and survival time

**4. Endocrine therapy in hepatocellular carcinoma**

ER: Estrogen receptor; AR: Androgen receptor

**clinicopathological parameters**

Tumor size and tumor recurrence

**Table 3.** Relationship between estrogen or androgen receptors with clinicopathological parameters and clinical

Considering both the epidemiologic and the experimental data supporting the sex steroids influence in growth and progression from HCCs, different clinical studies analyzed if endo‐ crine therapy could be interesting in advanced disease. Table 4 gives details of some of these studies. Trials using several agents, such as the anti-estrogen tamoxifen, megestrol acetate, progestin, the gonadotropin releasing hormone (GnRH) agonist leuprorelin and the antiandrogen flutamide have been uniformly disappointing [37-40]. At present, experts have concluded that hormonal manipulation should not be a part of the current management of patients with HCC [41-42]. However, it is possible that the rational design of an endocrine therapy requires a more complete understanding of the role of sex hormones in the tumorigenic process and how hormones and organ systems interact during this process [18], in addition to careful selection of the new studies patient populations, since patient stratification based on gender may uncover signals of activity of hormonal therapy in these settings. Notably, it has been shown that the anti-estrogens tamoxifen and raloxifene, behave differently in different tissues, producing estrogen-agonist activity in one tissue while behaving as an antagonist in another [43-44]. In addition, *in vitro* studies in the liver cell line Hep3B, indicate that raloxifene induces the insulin-like growth factor (IGF-I) gene transcription, whereas estradiol or tamox‐ ifen inhibits it [45]. On the other hand, the lack of hormone efficacy in these clinical studies, in terms of tumor growth and survival, could be due to either a low expression of ER or AR in HCC or to mutated receptors expression. Therefore, further studies will be necessary to

[26] ER mRNA Survival Wild type ERs tumors showed

**Correlation n year country**

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

96 2000 Italy

http://dx.doi.org/10.5772/56877

301

45 1989 Japan

43 1995 Spain

32 1998 China

long survival in patients

AR+ patients showed higher recurrence rates. AR- patients showed better survival rates.

AR expression correlated with smaller tumor size. Higher tumor recurrence in AR+ surrounding tissues.

AR+ tumor correlated with higher tumor size and lower survival rates.



**Table 3.** Relationship between estrogen or androgen receptors with clinicopathological parameters and clinical outcome of patients with HCC.

### **4. Endocrine therapy in hepatocellular carcinoma**

**3. Steroid receptors and hepatitis virus**

300 Hepatocellular Carcinoma - Future Outlook

therapy to battle HBV-induced HCC [36].

**clinicopathological parameters**

Serum alpha-fetoprotein, HBV profile, tumor histology, carcinoembryonic antigen

Serum alpha-fetoprotein,

Sex, age, alcohol consumption, hepatic function, other liver disease, tumor size, hepatic resection, tumor recurrence, long-term survival rate

> Survival after curative resection

**Correlation n year country**

Histopathology, HBV markers - <sup>19</sup> <sup>1989</sup> Japan

Local recurrence AR expression was associated

ER- patients showed higher rate of resection and larger tumor size

with intrahepatic recurrence

ER+ tumor have a negative effect on patient survival after curative resection


66 1990 Japan

78 1995 Japan

28 1997 Germany

**Protein / mRNA**

ER and AR protein

ER and AR protein

[128] ER protein

[24] ER protein

[25] ER protein

**Reference**

[132]

[49]

It is of special interest the interaction between sex estrogen receptors and viral proteins in hepatitis B virus (HBV) and hepatitis C virus (HCV)-induced HCC. It has been reported that a HBV protein (HBx) interacts with ERα. HBx is a multifunctional protein involved in neoplastic transformation in cultured cells and in HCC induction in transgenic mice. Both HBx and vERα (Delta 5 deletion variant of ERα) have additive effects on suppressing ERα trans‐ activation [31]. In addition, it was reported that tamoxifen inhibits ERα actions and suppresses HCV genome replication [32], which may be of potential interest to develop new anti-HCV strategies based on anti-ER drugs. On the other hand, higher serum androgen concentrations or a specific AR gene, which leads to higher AR activities, have also been associated to higher risk in HBV-mediated HCC [33-34]. Likewise, it has been reported that the combination of male gender and HBV infection had a significant synergistic effect on HCC progression [35]. Most recently, Ming-Heng *et al*. found that hepatic AR increases the HBV viral titer by enhancing HBV RNA transcription through direct binding to the androgen response element near the viral core promoter. This activity forms a positive feedback mechanism with the cooperation of its downstream target, the HBx protein, to promote hepatocarcinogenesis. In addition in these same study administration of a chemical compound that selectively degrades AR, ASC-J9, was able to suppress HCC tumor size in a transgenic HBV mouse model that developed HCC upon exposure to a low dose of N'-N'-diethylnitrosamine (DEN). These results demonstrate that targeting the AR, rather than the androgen, could be developed as a new

> Considering both the epidemiologic and the experimental data supporting the sex steroids influence in growth and progression from HCCs, different clinical studies analyzed if endo‐ crine therapy could be interesting in advanced disease. Table 4 gives details of some of these studies. Trials using several agents, such as the anti-estrogen tamoxifen, megestrol acetate, progestin, the gonadotropin releasing hormone (GnRH) agonist leuprorelin and the antiandrogen flutamide have been uniformly disappointing [37-40]. At present, experts have concluded that hormonal manipulation should not be a part of the current management of patients with HCC [41-42]. However, it is possible that the rational design of an endocrine therapy requires a more complete understanding of the role of sex hormones in the tumorigenic process and how hormones and organ systems interact during this process [18], in addition to careful selection of the new studies patient populations, since patient stratification based on gender may uncover signals of activity of hormonal therapy in these settings. Notably, it has been shown that the anti-estrogens tamoxifen and raloxifene, behave differently in different tissues, producing estrogen-agonist activity in one tissue while behaving as an antagonist in another [43-44]. In addition, *in vitro* studies in the liver cell line Hep3B, indicate that raloxifene induces the insulin-like growth factor (IGF-I) gene transcription, whereas estradiol or tamox‐ ifen inhibits it [45]. On the other hand, the lack of hormone efficacy in these clinical studies, in terms of tumor growth and survival, could be due to either a low expression of ER or AR in HCC or to mutated receptors expression. Therefore, further studies will be necessary to

improve patient selection on the basis of gender, ER tumor expression and their functional status. With regard to this latter aspect, considering that PgR is an estrogen inducible protein, it led us to consider that this steroid receptor could be a possible marker of ER functional status in HCC, to select candidate patients to an anti-estrogenic therapy. In addition, we demon‐ strated that PgR expression have a prognostic implication in patients with resected HCCs, being a factor associated with better prognostic [23], which is a clinical finding previously demonstrated in breast cancer [46-47]. This finding is very important considering also our reported positive association between PgR expression and HCV infection [23], which repre‐ sents an increasing aetiology of HCC in our patient's population [3].

Several data suggest that blocking AR may be interesting in HCC therapy. Studies in animal models suggest that increased hepatic AR expression correlated with accelerated tumor development [48], or even AR expression was associated with intrahepatic recurrence in HCC [49]. However, AR expression has also been reported to be associated to small tumor size, but not with a higher rate of recurrence [28]. More recent data indicate that suppression of androgen or AR signals also led to an increase in the number of infiltrating cells, such as macrophages as well as B or T-cells [50], which shows diverse and important roles in the promotion of HCC tumorigenicity [51]. Although there is few data on the effect of antiandrogens in HCC patients, it has been suggested that anti-androgen therapy may have some benefit in patients with androgen-positive tumors [52]. Likewise, recently it has been reported that a multicenter trial with anti-androgens in HCC male patients, has been interrupted because of digestive side effects [39]. Nevertheless, further studies will be necessary to assess whether AR status may be a useful marker to select more accurately candidate patients to further anti-androgenic therapies. Notably, it has been reported that expression of androgeninduced proteins, such as apolipoprotein D or Zinc-alpha2-glycoprotein, is associated with poor prognosis in HCC.

Apolipoprotein D (ApoD) is an androgen-induced protein increased in both prostate and breast cancer cells [53-55]. A report of Utsunomiya *et al*. showed that low ApoD expression correlated significantly with less-differentiated HCCs and therefore with a worse prognosis of patients [56]. However, we did not found any relationship between ApoD expression and the hormonal receptor status in HCC [23]. Nevertheless, further studies will be also necessary in order to assess the possible value of ApoD as a biological marker of androgen response and/ or other hormonal pathways in HCC. Thus, with regard to this latter aspect, although ApoD is an androgen-regulated protein, it can also been induced by other hormonal steroids or substances such as glucocorticoids, retinoic acid or 1,25-dihydroxyvitamin D3, that might be involved in the regulation of this ApoD in human tumors [54,57-58]. This is relevant since it has been described that both retinoid and glucocorticoid receptors have been also detected in HCC [59-60], and that preliminary data show that retinoids are considered of clinical interest as cancer chemotherapeutic agents in advanced HCC [61].

Zinc-alpha2-glycoprotein (ZA2G) is also an androgen-induced protein increased in breast cancer [58,62]. Recently, ZA2G expression was found to be decreased in HCC tissues at both mRNA and protein levels. Moreover, low expression of ZA2G was notably more prevalent in patients with poor tumor differentiation, advanced liver cirrhosis, high serum alpha fetopro‐ tein (AFP) level and shorter survival time. These results suggest that ZA2G may be a promising novel prognostic biomarker for HCC [63], and advocate for the need for validation studies.

**Treatment n Clinical outcomes Year Reference**

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

No effect on the tumor, in 8 remained stable between 5 and 13 months. Prolonged survival in 4 patients (over 18 months).

No effect on the tumor. No effect on survival.

Tumor regression to half size in patients with wild type ERs following tamoxifen treatment. Megestrol slowed down tumor growth in tumors with variant ERs.

No effects on survival, regardless of the type of receptor expressed.

Significantly slowed down tumor growth and improved survival in treated patients than placebo group.

No effect on survival or on quality of life. Deleterious effects with the higher dose.

1990 [154]

http://dx.doi.org/10.5772/56877

303

1995 [155]

1996 [30]

2000 [156]

2001 [157]

2002 [158]

Tumor growth Progestin 5 Tumor regression in 2 1982 [124]

33

120 (placebo = 62)

8 patients, 4 with wild ERs and 4 with variant ERs.

> 119 (placebo = 58)

45 (placebo = 24).

329

**Hormone Receptor/ mRNA**

ER (previous patient group)

NA

NA

Wild type and variant ER RNA m

> Variant ER mRNA.

> > NA

**Valuation parameters**

Anti-tumoral effect. Survival.

Anti-tumoral effect. Survival.

Tumor growth.

ER y PR Survival. Tamoxifen.

Tumor growth. Survival.

Survival. Quality of life. Tamoxifen 20 mg twice daily.

Tamoxifen 20 mg daily.

Tamoxifen 80 mg daily in patients with wild-ERs

or megestrol 160 mg daily in patients with variant

ERs.

Megestrol 160 mg daily.

Tamoxifen 120 mg or 60 mg

ER= Estrogen receptor. NA=information not available. PR= Progesterone Receptor.

daily.

**Table 4.** Results of hormonal treatment in hepatocellular carcinoma


ER= Estrogen receptor. NA=information not available. PR= Progesterone Receptor.

**Table 4.** Results of hormonal treatment in hepatocellular carcinoma

improve patient selection on the basis of gender, ER tumor expression and their functional status. With regard to this latter aspect, considering that PgR is an estrogen inducible protein, it led us to consider that this steroid receptor could be a possible marker of ER functional status in HCC, to select candidate patients to an anti-estrogenic therapy. In addition, we demon‐ strated that PgR expression have a prognostic implication in patients with resected HCCs, being a factor associated with better prognostic [23], which is a clinical finding previously demonstrated in breast cancer [46-47]. This finding is very important considering also our reported positive association between PgR expression and HCV infection [23], which repre‐

Several data suggest that blocking AR may be interesting in HCC therapy. Studies in animal models suggest that increased hepatic AR expression correlated with accelerated tumor development [48], or even AR expression was associated with intrahepatic recurrence in HCC [49]. However, AR expression has also been reported to be associated to small tumor size, but not with a higher rate of recurrence [28]. More recent data indicate that suppression of androgen or AR signals also led to an increase in the number of infiltrating cells, such as macrophages as well as B or T-cells [50], which shows diverse and important roles in the promotion of HCC tumorigenicity [51]. Although there is few data on the effect of antiandrogens in HCC patients, it has been suggested that anti-androgen therapy may have some benefit in patients with androgen-positive tumors [52]. Likewise, recently it has been reported that a multicenter trial with anti-androgens in HCC male patients, has been interrupted because of digestive side effects [39]. Nevertheless, further studies will be necessary to assess whether AR status may be a useful marker to select more accurately candidate patients to further anti-androgenic therapies. Notably, it has been reported that expression of androgeninduced proteins, such as apolipoprotein D or Zinc-alpha2-glycoprotein, is associated with

Apolipoprotein D (ApoD) is an androgen-induced protein increased in both prostate and breast cancer cells [53-55]. A report of Utsunomiya *et al*. showed that low ApoD expression correlated significantly with less-differentiated HCCs and therefore with a worse prognosis of patients [56]. However, we did not found any relationship between ApoD expression and the hormonal receptor status in HCC [23]. Nevertheless, further studies will be also necessary in order to assess the possible value of ApoD as a biological marker of androgen response and/ or other hormonal pathways in HCC. Thus, with regard to this latter aspect, although ApoD is an androgen-regulated protein, it can also been induced by other hormonal steroids or substances such as glucocorticoids, retinoic acid or 1,25-dihydroxyvitamin D3, that might be involved in the regulation of this ApoD in human tumors [54,57-58]. This is relevant since it has been described that both retinoid and glucocorticoid receptors have been also detected in HCC [59-60], and that preliminary data show that retinoids are considered of clinical interest

Zinc-alpha2-glycoprotein (ZA2G) is also an androgen-induced protein increased in breast cancer [58,62]. Recently, ZA2G expression was found to be decreased in HCC tissues at both mRNA and protein levels. Moreover, low expression of ZA2G was notably more prevalent in

sents an increasing aetiology of HCC in our patient's population [3].

as cancer chemotherapeutic agents in advanced HCC [61].

poor prognosis in HCC.

302 Hepatocellular Carcinoma - Future Outlook

patients with poor tumor differentiation, advanced liver cirrhosis, high serum alpha fetopro‐ tein (AFP) level and shorter survival time. These results suggest that ZA2G may be a promising novel prognostic biomarker for HCC [63], and advocate for the need for validation studies.

Recent data indicates that estrogens attenuate tumor progression in HCC *in vivo* by reducing tumor cell invasion, arresting cell cycle progression, and promoting apoptosis, which is characterized by an increased expression of cleaved caspase-3, and by a decrease in the expression of Proliferating Cell Nuclear Antigen (PCNA), cyclin A, cyclin D1, Bcl-2, and matrix metalloproteases (MMP) 2 and 9 [64]. It is relevant the data indicating that steroid hormones, such as estrogens, may inhibit MMPs in several experimental models [65-67]. This may be of great importance due to MMPs are proteolytic enzymes which participate in the degradation of the stromal connective tissue and basement membrane component, therefore facilitating tumor invasion and metastasis. Here, we show a negative relationship between ER expression and MMP2, MMP-9 or MMP-11 expression in HCC (Figure 1).

sized as inactive zymogens, which are then activated predominantly pericellularly by other MMPs or by serine-proteases. MMPs' activity is specifically inhibited by the so-called tissue inhibitors of metalloproteases (TIMPs). Currently, four different TIMPs are known to exist: TIMP-1, -2, -3 and -4. In addition, there are other two additional aspects conferring relevance to this enzymatic system in cancer biology. First, MMPs are able to impact on tumor cell behavior *in vivo* as a consequence of their ability to cleave growth factors, cell surface receptors, cell adhesion molecules, or chemokines/cytoquines [72-77]. Furthermore, by cleaving proa‐ poptotic factors, MMPs may induce a more aggressive phenotype via generation of apoptotic resistant cells [78]. MMPs may also regulate angiogenesis in cancer, both positively through their ability to mobilize or activate proangiogenic factors [79-80], and negatively via generation of angiogenesis inhibitors, such as angiostatin and endostatin, cleaved from large protein precursors [81-83]. Second, it is now assumed that TIMPs are multifactorial proteins also

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

http://dx.doi.org/10.5772/56877

305

involved in the induction of proliferation and the inhibition of apoptosis [84-85].

stromal cell) was not specifically considered.

by human mesenchymal cells but also by hepatoma cells [87].

Previous studies have shown the expression of several MMPs in HCC [86-100]. These findings have a great interest because HCC is characterized by a disposition for vascular invasion and high metastatic potential, thus leading to high incidence of early postoperative recurrence and poor survival [101-102]. Although there is no basal membrane in the liver, HCC cancer cells grow surrounded by extracellular matrix proteins secreted as a consequence of cirrhosis, and therefore proteolytic activity is required to allow HCC cells to penetrate and cross over such tissue boundaries [103]. In fact, several studies have suggested the significance of some MMPs in the malignant behavior of HCC, such as MMP-2 [86-93], MMP-7 [88,94], MMP-9 [95-97,91,93,104], MMP-12 [105] or MMP-14 (MT1-MMP) [98,91,99-100]. In these studies, MMPs were associated with several parameters indicative of tumor aggressiveness and poor prognosis, although the cellular type expressing each factor (tumor cell and/or peritumor

There are few available data referring to the integrated expression of these factors in relation with HCC. In this context, we have described new findings about MMPs and TIMPs expressions in HCC, together with an important expression by tumor stromal cells, as well as their clinical relevance. It is known that MMPs expression in neoplastic tissues is high due to regulation, in a paracrine manner, by growth factors and cytokines secreted by either tumor or stromal cells [106]. Nevertheless, high MMPs and TIMPs expression by HCCs tissues may be also due to the interplay between transformed cells and their microenviron‐ ment, particularly the surrounding extracellular matrix. In human livers, fibrogenesis underlies the development of HCCs in at least 90% of cases [107], and HCC is typically surrounded by a fibrous capsule at an early stage [108]. In addition, several studies have shown that some MMPs (MMP-1, -2, -9 and -13) and TIMPs (TIMP-1 and -2) are in‐ volved in the liver fibrosis processes [90,109-112]. It has been demonstrated that extracellu‐ lar matrix deposition induces overexpression of MMPs and TIMPs, such as MMP-2, secreted

Recently we reported that immunostaining for MMPs and TIMPs was localized predominantly in tumor cells, but also in peritumor stromal cells in a significant percentage of cases. However,

**Figure 1.** Relationship between ER expression and median score values of MMP2, MMP-9 or MMP-11 expression in HCC.

#### **5. Metalloproteases and their inhibitors**

Degradation of the stromal connective tissue and basement membrane components are key elements in tumor invasion and metastasis. Some components of the extracelluar matrix, particularly interstitial collagens, are very resistant to proteolytic attacks, being degraded only by matrix metalloproteases (MMPs) [68].

The human MMP family currently consists of 28 members of homologous zinc-dependent endopeptidases, which can be divided into eight structural classes or, based on their substrate specificity and primary structure, into the more familiar subgroups of collagenases (MMP-1, -8 and -13), gelatinases (MMP-2 and -9), stromelysins (MMP-3, -10, -11), membrane-associated MMPs (MMP-14, -15, -16, -17, -23, -24, -25) and other novel MMPs [69-71]. MMPs are synthe‐ sized as inactive zymogens, which are then activated predominantly pericellularly by other MMPs or by serine-proteases. MMPs' activity is specifically inhibited by the so-called tissue inhibitors of metalloproteases (TIMPs). Currently, four different TIMPs are known to exist: TIMP-1, -2, -3 and -4. In addition, there are other two additional aspects conferring relevance to this enzymatic system in cancer biology. First, MMPs are able to impact on tumor cell behavior *in vivo* as a consequence of their ability to cleave growth factors, cell surface receptors, cell adhesion molecules, or chemokines/cytoquines [72-77]. Furthermore, by cleaving proa‐ poptotic factors, MMPs may induce a more aggressive phenotype via generation of apoptotic resistant cells [78]. MMPs may also regulate angiogenesis in cancer, both positively through their ability to mobilize or activate proangiogenic factors [79-80], and negatively via generation of angiogenesis inhibitors, such as angiostatin and endostatin, cleaved from large protein precursors [81-83]. Second, it is now assumed that TIMPs are multifactorial proteins also involved in the induction of proliferation and the inhibition of apoptosis [84-85].

Recent data indicates that estrogens attenuate tumor progression in HCC *in vivo* by reducing tumor cell invasion, arresting cell cycle progression, and promoting apoptosis, which is characterized by an increased expression of cleaved caspase-3, and by a decrease in the expression of Proliferating Cell Nuclear Antigen (PCNA), cyclin A, cyclin D1, Bcl-2, and matrix metalloproteases (MMP) 2 and 9 [64]. It is relevant the data indicating that steroid hormones, such as estrogens, may inhibit MMPs in several experimental models [65-67]. This may be of great importance due to MMPs are proteolytic enzymes which participate in the degradation of the stromal connective tissue and basement membrane component, therefore facilitating tumor invasion and metastasis. Here, we show a negative relationship between ER expression

**Figure 1.** Relationship between ER expression and median score values of MMP2, MMP-9 or MMP-11 expression in

Degradation of the stromal connective tissue and basement membrane components are key elements in tumor invasion and metastasis. Some components of the extracelluar matrix, particularly interstitial collagens, are very resistant to proteolytic attacks, being degraded only

The human MMP family currently consists of 28 members of homologous zinc-dependent endopeptidases, which can be divided into eight structural classes or, based on their substrate specificity and primary structure, into the more familiar subgroups of collagenases (MMP-1, -8 and -13), gelatinases (MMP-2 and -9), stromelysins (MMP-3, -10, -11), membrane-associated MMPs (MMP-14, -15, -16, -17, -23, -24, -25) and other novel MMPs [69-71]. MMPs are synthe‐

and MMP2, MMP-9 or MMP-11 expression in HCC (Figure 1).

304 Hepatocellular Carcinoma - Future Outlook

**5. Metalloproteases and their inhibitors**

by matrix metalloproteases (MMPs) [68].

HCC.

Previous studies have shown the expression of several MMPs in HCC [86-100]. These findings have a great interest because HCC is characterized by a disposition for vascular invasion and high metastatic potential, thus leading to high incidence of early postoperative recurrence and poor survival [101-102]. Although there is no basal membrane in the liver, HCC cancer cells grow surrounded by extracellular matrix proteins secreted as a consequence of cirrhosis, and therefore proteolytic activity is required to allow HCC cells to penetrate and cross over such tissue boundaries [103]. In fact, several studies have suggested the significance of some MMPs in the malignant behavior of HCC, such as MMP-2 [86-93], MMP-7 [88,94], MMP-9 [95-97,91,93,104], MMP-12 [105] or MMP-14 (MT1-MMP) [98,91,99-100]. In these studies, MMPs were associated with several parameters indicative of tumor aggressiveness and poor prognosis, although the cellular type expressing each factor (tumor cell and/or peritumor stromal cell) was not specifically considered.

There are few available data referring to the integrated expression of these factors in relation with HCC. In this context, we have described new findings about MMPs and TIMPs expressions in HCC, together with an important expression by tumor stromal cells, as well as their clinical relevance. It is known that MMPs expression in neoplastic tissues is high due to regulation, in a paracrine manner, by growth factors and cytokines secreted by either tumor or stromal cells [106]. Nevertheless, high MMPs and TIMPs expression by HCCs tissues may be also due to the interplay between transformed cells and their microenviron‐ ment, particularly the surrounding extracellular matrix. In human livers, fibrogenesis underlies the development of HCCs in at least 90% of cases [107], and HCC is typically surrounded by a fibrous capsule at an early stage [108]. In addition, several studies have shown that some MMPs (MMP-1, -2, -9 and -13) and TIMPs (TIMP-1 and -2) are in‐ volved in the liver fibrosis processes [90,109-112]. It has been demonstrated that extracellu‐ lar matrix deposition induces overexpression of MMPs and TIMPs, such as MMP-2, secreted by human mesenchymal cells but also by hepatoma cells [87].

Recently we reported that immunostaining for MMPs and TIMPs was localized predominantly in tumor cells, but also in peritumor stromal cells in a significant percentage of cases. However, we found no stromal expression of MMPs or TIMPs in normal liver samples [113]. We also found a positive and noteworthy association between MMP-1 expression by fibroblasts or by mononuclear inflammatory cells (MICs) and a larger tumor size. In addition, our data showed that MMP-1 expression by stromal fibroblasts, as well as the expression of MMP-13, TIMP-1 and 2, by MICs, were significantly associated with a shorter overall survival (Figure 2) [113].

On the other hand, a decrease on TIMP-2 and -3 expression has been reported to be associated with invasion and metastases in HCC [115-116]. Certainly, if TIMPs inhibit MMPs *in vivo*, it should be expected that high levels of these inhibitors would prevent tumor progression and thus to be related with good outcome in cancer patients. However, TIMPs are multifunctional proteins that in addition to its MMP-inhibitory effect also shown distinct tumor-stimulatory functions, such as the induction of proliferation and the inhibition of apoptosis [84]. Thus, it is of note that TIMP-1 and -2 expressions by stromal cells were associated with shortened overall survival[113]. Accordingly, it has been reported that TIMP-1 overexpression leads to an increase of hepatoma cells migration [117], and also that it is associated with invasion and

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

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307

**Figure 3.** Graphical representation of two-dimensional unsupervised hierarchical clustering results on immunohisto‐ chemistry expression (score values) profile in 10 proteins (MMPs and TIMPs) in 30 samples of HCC tissue. Rows: tumor samples; columns: MMPs and TIMPs. Protein expressions are depicted according to a color scale: red: positive staining; green: negative staining; gray: missing data. At right bank of the figure is represented the overall survival status for

each patient (0, alive; 1, death due tumor progression).

metastases in HCC [115,117].

**Figure 2.** Kaplan-Meier survival curves as function of the expression by fibroblasts cells of MMP-1 (A); expression by MIC of MMP-13 (B), TIMP-1 (C) and TIMP-2 (D).

There are discrepancies between different studies regarding the prognostic significance of several of these factors in HCC, such as MMP-9 and MMP-2 expression in stromal compartments [113,93]. Nevertheless, it is worth considering the existence of possible differences in MMPs or TIMPs expression between different patient's populations. Thus, for example, in Asian countries HCC originates more frequently in healthy liver tissues, with less incidence of cirrhosis, than in Europe, whereas in the Mediterranean area as well as in the North American countries liver HCC only develops in chronically injured livers, where altered turn-over and increased deposition of ECM proteins has been described. In a such environment, upregulation of MMPs and TIMPs has been reported and it is possible for them to play an important role in the rearrangement of the liver tissue architecture an aspect that could influence the different expression pattern of MMPs and TIMPs in stromal HCC cells [110,114].

On the other hand, a decrease on TIMP-2 and -3 expression has been reported to be associated with invasion and metastases in HCC [115-116]. Certainly, if TIMPs inhibit MMPs *in vivo*, it should be expected that high levels of these inhibitors would prevent tumor progression and thus to be related with good outcome in cancer patients. However, TIMPs are multifunctional proteins that in addition to its MMP-inhibitory effect also shown distinct tumor-stimulatory functions, such as the induction of proliferation and the inhibition of apoptosis [84]. Thus, it is of note that TIMP-1 and -2 expressions by stromal cells were associated with shortened overall survival[113]. Accordingly, it has been reported that TIMP-1 overexpression leads to an increase of hepatoma cells migration [117], and also that it is associated with invasion and metastases in HCC [115,117].

we found no stromal expression of MMPs or TIMPs in normal liver samples [113]. We also found a positive and noteworthy association between MMP-1 expression by fibroblasts or by mononuclear inflammatory cells (MICs) and a larger tumor size. In addition, our data showed that MMP-1 expression by stromal fibroblasts, as well as the expression of MMP-13, TIMP-1 and 2, by MICs, were significantly associated with a shorter overall survival (Figure 2) [113].

**Figure 2.** Kaplan-Meier survival curves as function of the expression by fibroblasts cells of MMP-1 (A); expression by

There are discrepancies between different studies regarding the prognostic significance of several of these factors in HCC, such as MMP-9 and MMP-2 expression in stromal compartments [113,93]. Nevertheless, it is worth considering the existence of possible differences in MMPs or TIMPs expression between different patient's populations. Thus, for example, in Asian countries HCC originates more frequently in healthy liver tissues, with less incidence of cirrhosis, than in Europe, whereas in the Mediterranean area as well as in the North American countries liver HCC only develops in chronically injured livers, where altered turn-over and increased deposition of ECM proteins has been described. In a such environment, upregulation of MMPs and TIMPs has been reported and it is possible for them to play an important role in the rearrangement of the liver tissue architecture an aspect that could influence the different expression pattern of MMPs and TIMPs in stromal

MIC of MMP-13 (B), TIMP-1 (C) and TIMP-2 (D).

306 Hepatocellular Carcinoma - Future Outlook

HCC cells [110,114].

**Figure 3.** Graphical representation of two-dimensional unsupervised hierarchical clustering results on immunohisto‐ chemistry expression (score values) profile in 10 proteins (MMPs and TIMPs) in 30 samples of HCC tissue. Rows: tumor samples; columns: MMPs and TIMPs. Protein expressions are depicted according to a color scale: red: positive staining; green: negative staining; gray: missing data. At right bank of the figure is represented the overall survival status for each patient (0, alive; 1, death due tumor progression).

The unsupervised hierarchical cluster analysis of MMPs/TIMPs expressions in HCCs led us to identify 2 well-defined clusters of cases (Figure 3): one with low expression of MMPs and TIMPs (Group 1), and another with high expression of these factors (Group 2). This classifi‐ cation could have biological interest in order to identify patient candidates to new therapies based on enzymatic MMP inhibition. With regard to this, although there are no published data about use of MMP inhibitors in HCC, recently it has been reported that decreased MMP activity mediated by statins reduced progression and limits metastatic diffusion of established HCC [118]. Likewise, it was demonstrated that blocking the tumor-related glycoprotein HAb18G/ CD147 by gene silence in HCC cells or with HA18 monoclonal antibody, resulted in a sup‐ pressive effect on MMP secretion and cell invasion [119]. More recent studies have shown an antimetastatic effect of Norcantharidin on HCC by transcriptional inhibition of MMP-9 [120], as well as a metastasis inhibition of HCC cells due to down-regulation of Osteopontin via a mechanism involving MMP-2 and urinary plasminogen activator (uPA) [121].

[4] Poon RT, Ng IO, Fan ST, Lai EC, Lo CM, Liu CL, Wong J (2001) Clinicopathologic features of long-term survivors and disease-free survivors after resection of hepato‐ cellular carcinoma: a study of a prospective cohort. J Clin Oncol 19 (12):3037-3044.

Hepatocellular Carcinoma, Steroid Hormones and Metalloproteases

http://dx.doi.org/10.5772/56877

309

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### **6. Conclusion**

In summary, we consider that expression analysis of steroid hormone receptors, MMPs and TIMPs, contributes to a better knowledge in the biological characterization of HCC and highlights the need for further studies exploring new therapeutic targets for this common tumor.

### **Author details**

Noemí Eiró, Belen Fernandez-Garcia, Antonio Altadill, Luis O. González and Francisco J. Vizoso\*

\*Address all correspondence to: fjvizoso@terra.es

Unidad de Investigación. Fundación Hospital de Jove. Gijón, Asturias, Spain

### **References**


[4] Poon RT, Ng IO, Fan ST, Lai EC, Lo CM, Liu CL, Wong J (2001) Clinicopathologic features of long-term survivors and disease-free survivors after resection of hepato‐ cellular carcinoma: a study of a prospective cohort. J Clin Oncol 19 (12):3037-3044. [PubMed: 11408499]

The unsupervised hierarchical cluster analysis of MMPs/TIMPs expressions in HCCs led us to identify 2 well-defined clusters of cases (Figure 3): one with low expression of MMPs and TIMPs (Group 1), and another with high expression of these factors (Group 2). This classifi‐ cation could have biological interest in order to identify patient candidates to new therapies based on enzymatic MMP inhibition. With regard to this, although there are no published data about use of MMP inhibitors in HCC, recently it has been reported that decreased MMP activity mediated by statins reduced progression and limits metastatic diffusion of established HCC [118]. Likewise, it was demonstrated that blocking the tumor-related glycoprotein HAb18G/ CD147 by gene silence in HCC cells or with HA18 monoclonal antibody, resulted in a sup‐ pressive effect on MMP secretion and cell invasion [119]. More recent studies have shown an antimetastatic effect of Norcantharidin on HCC by transcriptional inhibition of MMP-9 [120], as well as a metastasis inhibition of HCC cells due to down-regulation of Osteopontin via a

In summary, we consider that expression analysis of steroid hormone receptors, MMPs and TIMPs, contributes to a better knowledge in the biological characterization of HCC and highlights the need for further studies exploring new therapeutic targets for this common

mechanism involving MMP-2 and urinary plasminogen activator (uPA) [121].

Noemí Eiró, Belen Fernandez-Garcia, Antonio Altadill, Luis O. González and

Unidad de Investigación. Fundación Hospital de Jove. Gijón, Asturias, Spain

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**6. Conclusion**

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**Author details**

Francisco J. Vizoso\*

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## *Edited by Ahmed O. Kaseb*

Hepatocellular carcinoma (HCC) develops as a consequence of underlying chronic liver disease, most commonly cirrhosis. Therefore, HCC management draws on the expertise of a range of medical specialists. Many components of current novel therapeutic modalities for HCC are discussed in the current version of the book within the framework of a multidisciplinary approach with special emphasis on emerging treatment approaches and research strategies. This book is the essential clinical guide for oncologists, hepatologists, surgeons, and all physicians and researchers involved in the care of patients with HCC. I would like to thank the authors for their significant efforts in bringing this edition to life. This book is a tribute to their continued dedication to improving HCC outcome.

Hepatocellular Carcinoma - Future Outlook

Hepatocellular Carcinoma

Future Outlook

*Edited by Ahmed O. Kaseb*

Photo by Ralwel / iStock