Key words

Ewelina Frejlich

_{, Julia Rudno-Rudzińska}

_{, Kacper Janiszewski}

_,

Łukasz Salomon

_{, Krzysztof Kotulski}

_{, Oskar Pelzer}

_,

Zygmunt Grzebieniak

_{, Robert Tarnawa}

_{, Wojciech Kielan}

Caspases and Their Role in Gastric Cancer

Kaspazy i ich znaczenie w raku żołądka

1 _{2nd Department of General and Oncological Surgery, Wroclaw Medical University Hospital, Wrocław, Poland} 2 _{Department of Gastroenterology and Hepatology, Wroclaw Medical University Hospital, Wrocław, Poland} 3 _{Division of Surgical Oncology and General Surgery, Regional Hospital Center in Jelenia Góra,}

Jelenia Góra, Poland

A – research concept and design; B – collection and/or assembly of data; C – data analysis and interpretation;

D – writing the article; E – critical revision of the article; F – final approval of article; G – other

Abstract

Caspases (Cysteine Aspartate Specific Proteases) are a group of cysteine-containing proteolytic enzymes produced by the cells of living organisms. They participate in immunological functions, proliferation, cell migration and organiza-tion. Caspases also influence the secretion of various regulative factors. Moreover, they are responsible for cellular maturation and reconstruction, and for regulating the number and quality of cells initiating the apoptosis of old cells or those that cannot play their normal role due to abnormalities. Multiple pathological processes are associated with disorders in the activity of caspases. Changes in expression of individual caspases have been observed in gastric cancer. The expression of some caspases is also correlated with particular histological traits and the frequency of metastases, which suggests their possible use as a prognostic factor. It has also been discovered that some somatic mutations in caspase coding genes might lead to inhibition of apoptosis and the progression of the disease. Gene polymorphism may be a gastric cancer risk factor, but may also play a protective function. Considering the less than satisfactory effects of conventional therapeutic methods, the search for alternative ways to activate apoptosis – through gene therapy or selec-tive activation of individual elements of the apoptotic pathways – constitutes a promising direction for studies of new therapeutic strategies. Caspases, enzymes playing a central role in the process of programmed cellular death, may pos-sibly be a key to the development of a more effective anti-cancer therapy (Adv Clin Exp Med 2013, 22, 4, 593–602).

Key words: caspases, apoptosis, gastric cancer, prognostic factor.

Streszczenie

Kaspazy są grupą cysteinowych enzymów proteolitycznych wydzielanych przez komórki żywych organizmów. Pełnią funkcje immunologiczne, biorą udział w proliferacji, organizacji i migracji komórek, a także wpływają na sekrecję różnych czynników regulatorowych. Odpowiadają również za dojrzewanie i przebudowę komórkową oraz regulację liczby i jakości komórek przez uruchamianie apoptozy komórek starych, uszkodzonych lub wykazujących nieprawi-dłowości uniemożliwiające im spełnianie powierzonych zadań. W różnych procesach patologicznych dochodzi do zaburzeń ich działania. W nowotworach żołądka obserwowano zmiany ekspresji poszczególnych kaspaz w porów-naniu z tkankami zdrowymi, jak również korelację ekspresji niektórych z nich z cechami histologicznymi i częstością przerzutów, co sugeruje możliwość ich zastosowania jako czynnika prognostycznego. Wykryto również, że niektóre mutacje somatyczne w genach kodujących kaspazy mogą doprowadzić do zahamowania apoptozy i progresji cho-roby. Polimorfizm genowy poszczególnych kaspaz może natomiast być czynnikiem ryzyka raka żołądka, ale rów-nież pełnić funkcje ochronne. Z powodu niesatysfakcjonujących rezultatów konwencjonalnych metod leczniczych poszukiwanie alternatywnych form aktywacji apoptozy, czy za pomocą terapii genowej, czy selektywnego pobudza-nia poszczególnych elementów jej szlaków, jest obiecującym kierunkiem badań nad nowymi strategiami terapeu-tycznymi. Kaspazy jako enzymy pełniące centralną rolę w procesie zaprogramowanej śmierci komórki wydają się kluczem do rozwoju leczenia przeciwnowotworowego (Adv Clin Exp Med 2013, 22, 4, 593–602).

Słowa kluczowe: kaspazy, apoptoza, rak żołądka, czynnik prognostyczny.

Adv Clin Exp Med 2013, 22, 4, 593–602 ISSN 1899–5276

REvIEWS

Caspases (Cysteine Aspartate Specific Proteas-es) are a group of cysteine proteolytic enzymes pro-duced by the cells of living organisms. Fifteen dif-ferent caspases have been discovered in mammals so far, and eleven human caspases are known [1].

Structure

There are several divisions of caspases, de-pending on the criteria applied: philogenetic ori-gin, substrates, homologies and prodomain length (Table 1). Common features of all caspases include presence in cells in the form of an inactive zymo-gene (procaspase), a structurally complex N-termi-nal prodomain, a large p29 subunit, a small C-ter-minal p10 subunit and a connection between the large and small subunits [2]. With regard to the length of the prodomain, caspases are divided into two subfamilies: Those with a long prodomain are pro-inflammatory and apoptosis initiators; and those with a short prodomain are “executioner caspases” (Table 1). The long N-terminal prodo-main contains a characteristic motif, the so called “death domain” or caspase activation and recruit-ment domain (CARD) or a pair of death-effec-tor domains (DEDs). That region is responsible for the oligomerization of caspases, as well as for interference with adaptation proteins and recep-tors of the cellular membrane [3]. Procaspase ac-tivation is associated with a change in structural configuration.

The Activation of Caspases

Four specific caspase-activating complexes have been identified. Those are:

– apoptosome, mediating caspase-9 activation by interacting with the apoptotic protease-activat-ing factor-1 (Apaf-1) adaptor in the presence of cy-tochrome c;

– death inducing signalling complex (DISC), mediating activation of caspase-8 through interac-tion with a Fas-Associated protein with Death Do-main (FADD) adaptor;

– inflammasomes, mediating in caspase-1 and caspase-5 activation by interacting with an apop-tosis-associated speck-like protein containing CARD (ASC) adaptor or the nucleotide-binding and oligomerization domain (NOD)-like receptor (NLR) family;

– and PIDDosome, mediating in the activation of caspase-2 through interaction with the adaptors RIP-associated ICH-1/CED-3 homologous pro-tein with a death domain (RAIDD) [4] and p53- -induced protein with a death domain (PIDD) [5].

Those complexes are highly specialized and become activated in response to various proapop-totic and proinflammatory signals. Apoptosome is created in response to the release of cytochrome c resulting from the disintegration of mitochon-dria. DISC is formed as a result of death receptor stimulation via their ligands, such as the Fas ligand (FasL) or tumor necrosis factor (TNF-a). More-over, inflammasomes participate in the activation

Table 1. Divisions of caspases

Tabela 1. Podział kaspaz

1. By amino acid sequence (Według sekwencji aminokwasów)

I initiators of apoptosis -2,-8, -9, -10, -15, II executioners of apoptosis -3,-6, -7

II proinflammatory -1, -4. -5, -11, -12, -13, -14 2. By prodomain length (Według długości prodomeny)

long -1,-2, -4, -5–8, 9, -10, -11, -12, -13, -14–15 short -3,-6, -7

3. Phylogenetically (Filogenetycznie)

CED-subfamily -2,-3,-6, -7, -8, -9, -10 ICE-subfamily -1, -4, -5, -12, -14

ICE-subfamily – Interleukin-1beta-Converting Enzyme-subfamily. CED-subfamily – Caenorhabditis elegans gene ced-3-subfamily.

of the innate immunological response to intercel-lular pathogens, and PIDDosome reacts to DNA damage [6] and cellular stress. This specialization of individual caspases is probably a result of the variability of harmful factors affecting cells and organisms.

Caspase activation takes place through a se-quence of interactions between subsequent critical points of apoptosis (Fig. 1). The two basic pathways leading to cellular apoptosis are the internal mito-chondria-related pathway, and the external path-way dependent on specific proteins belonging to the TNF family (TRAIL-R1, TRAIL-R2), membra-nous receptor Fas/CD95, and death receptor DR3, DR4, DR5, DR6 antigens. The extrogenous (exter-nal) pathway can be activated in various ways, e.g., in response to a lack of nutrients, as an effect of the impact of chemical substances, due to physi-cal factors or growth factor deficiency, as well as in response to hormones and cytokines (e.g. FasL, TNF, Apo3L, Apo2L), viruses and free radicals. The endogenous (internal) pathway is activated by

oncogenes as a result of hypoxia, general materi-al damage, the accumulation of abnormmateri-al proteins and cellular depletion of growth factors.

Ligand binding to a specific receptor leads to the oligomerization of the death domain (DD) re-ceptors mentioned earlier. This change allows the attachment of an adaptor protein possessing do-mains homologous to the DD. In the case of the Fas receptor, the attachment of the FasL ligand leads to a conformation change allowing the attachment of the FADD protein. via DEDs, the protein com-bines with procaspase-8; in the case of receptors belonging to the TNF family, the RIP protein com-bines with procaspase-8 via the TRADD domain. Therefore, activation of caspase-8 as well as cas-pase-10 takes place as a result of the recruitment of caspase to the death receptor via adaptor pro-teins, leading to the formation of the aforemen-tioned DISC complex [3, 7].

In its N-terminal prodomain procaspase-8 has a pair of two DEDs to which FADD is bound, whereas the C-terminal prodomain has a homology

Fig. 1. The Caspase Activation Cascade (based on [12])

DISC – death inducing signalling complex; TNF – tumor necrosis factor; TNFR – tumor necrosis factor receptor; Fas – TNF receptor superfamily member 6; FasL – Fas-ligand; TRADD – TNF-associated death domain;

FADD – Fas-associated death domain; Apaf-1 – apoptotic protease-activating factor-1; IAPs – inhibitor of apoptosis proteins; Bad, Bid, Bax, Bcl-2 – BcL-family proteins; tBid – truncated BID; Smac – Second mitochondria-derived acti-vator of caspase protein; DIABLO – Direct Inhibitor of Apoptosis-Binding protein with LOw pI;

APOPTOSOME – large quaternary protein – explanation in the text

region. Immediately following recruitment, pro-caspase is transformed into the catalytically active form of heterotetramer, able to continue the cas-cade of reactions. Caspase-8 causes the stimulation and transformation of procaspases-3, -6 and -7 in-to corresponding caspases.

The internal pathway of caspase activation is carried out with the direct participation of mito-chondria. Chemical stimuli, Uv radiation, gam-ma-radiation or a lack of growth factors lead to an increase in the permeability of the external mito-chondrial membrane and the release of cytochrome c from the intra-membranous space, through spe-cial channels. The release of cytochrome c is the first stage in the formation of the apoptosome. ATP, Apaf-1 protein and procaspase-9 are also contained in the apoptosome. The formation of the complex is necessary for oligomerization and the subsequent auto-proteolysis of procaspase-9 into caspase-9, which in turn is a direct activator of procaspase-3 and -7 [8]. Procaspase-9 may also become activated as a result of interaction with cas-pase-8. If there is a low concentration of caspase-8, insufficient for activating caspase-3, the Bid pro-tein (a propro-tein belonging to the Bcl-2 family, be-lieved to serve as a regulator of cytochrome-c-asso-ciated apoptosis) becomes proteolytically modified. Modified tBid transmits a signal to the mitochon-dria, stimulating the internal pathway [9].

The mechanisms of the regulation of pro-in-flammatory caspases are not as clear; they are mul-tidirectional and their description exceeds the scope of this article. It should, however, be men-tioned that only caspase-1 and -5 become activated within the aforementioned inflammasome, lead-ing to the activation of proinflammatory cytokines IL-1β and IL18 [10].

The Natural Inhibitors

of Caspases

Caspases, as natural enzymes participating in cellular and systemic biological processes, are reg-ulated and inhibited by natural inhibitors. Apop-tosis via the internal path is prevented by, among other things, anti-apoptotic proteins belonging to the Bcl-2 family, including Bcl-2, Bcl-w, Bcl-xL, Boo/Diva, Mcl-1 and others [11, 12], and also by the TUCAN protein Akt kinase acting direct-ly or indirectdirect-ly on caspase-9. The external path-way is inhibited by such proteins as BAR, Bap 31 and Flip [13]. Proteins belonging to the IAP fam-ily – including 6 human proteins: cIAP1, cIAP2, XIAP, NAIP, survivin and livin – are antagonists of the common pathway. The group is equipped with a characteristic BIR domain with a structure

of zinc fingers capable of blocking the caspase cat-alytic center. Some of the proteins belonging to the IAP family play an apoptosis-regulating function, for example survivin.

Other natural caspase inhibitors include Crm A, a serpin of the vaccinia virus, which acts as a strong inhibitor of caspase-1 and -8, and as a less potent inhibitor of caspase-3 and -6, and the p35 protein of baculovirus, which is an inhibitor of cas-pases 1–8 [12].

The Function of Caspases

Caspases play various biological functions. The basic nomenclature and division into proin-flammatory, apoptosis-inducing and apoptosis-ex-ecuting caspases does not reflect the whole scope of functions of these substances. The division is al-so not completely precise.

Caspases play immunological functions, par-ticipate in the proliferation, organization and mi-gration of cells, and also influence the secretion of various regulating factors. They are also responsi-ble for cellular maturation and reconstruction, as in the elimination of the cellular nucleus in eryth-roblasts, the reduction of cytoplasm in gameto-genesis or dendrite connections in the brain. Their proapoptotic role controls the number and qual-ity of cells by activation of programmed death of old or abnormal cells that cannot play their normal roles [14]. The biological functions of caspases may be therefore defined as (broadly understood) ho-meostasis at the cell-tissue-organ-organism levels. Disturbances of caspase-involving process-es are the basis of numerous pathologiprocess-es. Diseasprocess-es of the immunological system, degenerative condi-tions, proliferative and atrophic diseases may be in-cluded in that group. Reduced activity is observed in those diseases where cells characterized by low susceptibility to apoptosis accumulate, for example in cancer. The ability to selectively activate apopto-sis in cancer cells would be of immense value.

The Role of Caspases

in Gastric Cancer

basic directions of research is the search for critical features determining neoplastic cells’ resistance to apoptosis and favoring their proliferation and ex-tra-tissue migration. Caspases appear to be an in-teresting research subject in the effort to identify the causes of the proliferation of abnormal clones of cells.

Gastric cancers are in the top ten of both de novo cases and cancer-induced deaths in Poland: Among females, gastric cancer is the 7th _most common cancer and the 6th _{most common cause} of death; among males it is in 5th _{place in terms} of frequency and the 4th _{most common cause of} death [16]. These cancers constitute a serious ther-apeutic problem, considering their aggressiveness and the low survival rate. Gastric cancer cases are also a special object of interest for Asian research-ers, because that type of neoplasm is of the highest prevalence in Asia [17].

Caspases have been the subject of research for many years, but due to their extensive functions they appear to be a virtually inexhaustible topic. An example of this may be the findings of Jee et al. [18], who demonstrated a correlation between pro-inflammatory caspase-1 and human gastric cancer. In a study of 301 cases of gastric cancer, a loss of expression of caspase-1 was found in al-most in 20% of the cases. At the same time a signif-icant association was found between loss of expres-sion of caspase-1, pTNM, lymph node metastases and poor patient survival. These researchers also found that in three cell lines the loss of caspase-1 expression at the protein and mRNA level could be reversed using the gene expression modifying drugs trichostatin A (TSA) and/or 5-aza-2’-deoxy-cytidine (5-aza-C)[18].

Anticancer drug related apoptosis appears to be associated with caspase-1 activity. A study on gastric cancer cell lines showed a strong increase in caspase-1 expression after the administration of cisplatin (CDDP), suggesting that changes in cas-pase-1 activity after exposure to CDDP may be useful in predicting apoptosis following CDDP treatment in gastric cancer [19]. This may be par-ticularly important in the context of high-stability genes encoding caspase-1.

Soung et al. found only two somatic muta-tions in 54 cases of gastric cancer, one missense mutation in exon 7 and the other mutation in in-tron substitution. There were no significant cor-relations with the degree of differentiation, TNM stage, histologic subtype of gastric cancer and the mutation [20].

There are also reports indicating possible uses for caspase-2, an apoptosis initiator, in cancer treat-ment. The expression of caspase-2 in gastric cancer tissue seems to be decreased. In 120 cases studied

by Yoo et al., caspase-2 expression was observed in all normal tissue samples and only in 35% of the cancer tissue. The researchers observed a correla-tion between the loss of caspase-2 expression and a relatively early stage of gastric carcinogenesis, in-dicating that caspase-2 could be a new immuno-histochemical marker for gastric cancer [21]. The caspase-2 gene has also been under examination. Somatic mutation of this gene is rare, but may lead to a loss of function. The mutant showed signifi-cant decreases in apoptosis induction compared to the wild-type caspase-2, confirming the theory that disorders at this level can interfere with apoptosis and as a consequence promote neoplasia [22].

carcinoma (defined by the caspase-3 expression index – C3EI) with clinical and pathological fea-tures, and obtained statistically significant results. It was demonstrated that a low level of caspase-3 expression in neoplastic cells is correlated with a better prognosis and may constitute an indepen-dent prognostic factor; and that neoplastic cells are able to inhibit apoptosis through the inhibition of caspase-3 activation. A strong expression of sur-vivin as an inhibitor of executioner caspase-3 may be important in the process. Immunohistochem-ical analysis demonstrated a strong expression of survivin in neoplastic cells, with almost completely inhibited survivin expression in healthy tissue. In the same specimens a reverse correlation with cas-pase-3 was observed: Patients with a higher expres-sion of survivin had a higher level of histological and pathological advancement (grading and stag-ing). Caspase-3 expression correlated with histo-logical but not pathohisto-logical properties. It was also observed that survivin (–), caspase-3 (+) patients had higher apoptotic indexes [27].

Selective activation of the caspase cascade lead-ing to the apoptosis of neoplastic cells may be one of the future trends in gastric cancer therapy. At-tempts are being made to introduce recombinant caspase-3 into cells. Fu et al. constructed an eukary-otic vector pcDNA/Rev-Caspase-3 and evaluated its effect on apoptosis of cellular lines of gastric can-cer SGC7901 in laboratory conditions [28]. The re-sults of those experiments are promising: Reduced growth of neoplastic cell clones and increased apop-tosis were observed. Apopapop-tosis was evaluated by identifying its products compared to control cells. The results suggest a possible application of gene therapy in the case of that cancer, but that possibili-ty still has to be carefully evaluated and observed.

Somatic mutation in a caspase-3 encoding gene appears to be rare. In a study by Soung et al., mutation was found in two out of 165 cases of gas-tric cancer – one in intron 4 and the second in ex-on. No significant abnormalities were detected in either case, and they were silent mutations [29].

Caspase-3 activation does not necessarily have to take place at the molecular level, and many sci-entists around the world are looking for activators and regulators of its function among various bio-logical and chemical substances. One of the can-didates is trioxide As2O3, which causes inhibition of cancer cell growth and activation of apoptosis through its effect on caspase-3 (via both the inter-nal and exterinter-nal pathways). The study also demon-strated a high level of gastric cancer cell suscepti-bility to that substance [30]. However, the harmful effect of arsenic on the organism as a whole con-stitutes a rather important limitation to its appli-cation. The compound is used, though, in some

particularly severe cases of cancer. Similarly, the plant-derived alkaloid matrine causes dose- and exposure time-dependent inhibition of gastric cancer cell proliferation and a significant increase in the expression level of the surface antigen Fas and caspase-3 [31].

Gastric cancer belongs to the group of chemo-susceptible cancers. However, numerous clones of cells demonstrate resistance to conventional treatment based mainly on the cisplatin deriva-tives FAP (5-fluorouracyl, adriamycin and cisplat-in) and EAP (etoposide, adriamycin and cisplatcisplat-in), and 5-year survival rates in Poland are at the lev-el of 14.9% among men and 18.2% among wom-en [32]. Thymoquinone (KT), which makes can-cer cells susceptible to 5-fluorouracyl (5-FU), may set a new trend in chemotherapy. Studies dem-onstrated that pre-treatment with KT significant-ly increased the apoptotic effect of 5-FU in gastric cancer cell lines. Moreover, it was demonstrat-ed that KT-reinforcdemonstrat-ed 5-FU causes gastric cancer cell death by reducing the activation of the anti-apoptotic protein bcl-2, the activation of the pro-apoptotic protein Bax, and the activation of both caspase-3 and caspase-9. It was also demonstrated that this polytherapy is more effective compared to monotherapy with each of the agents [33].

The available literature holds no evidence that caspase-4 and -5 have a significant effect on the process of apoptosis and carcinogenesis in gas-tric cancer. Somatic mutations in the gene encod-ing caspase-5 in gastric cancer tissue – up to 16.7% – have been reported, but there was no relation to clinicopathologic characteristics, including gen-der, age, stage or histological type [20].

Caspase-6 and -7 are activated by both the apoptosis extrinsic and intrinsic pathway and will be analyzed together. In the previously mentioned study by Yoo et al. [21] it was found that there is a significant difference between the expression of caspase-6 and -7 in normal gastric mucosal cells and in tumor cells. In gastric cancer the levels of caspase-6 and -7 expression were 53% and 33% re-spectively, whereas in healthy tissue the expression was considerably higher. It was also found that there are higher levels of caspase-6 and -7 expres-sion in the early stage of gastric cancer. The conclu-sion seems to be the same as in the case of caspas-2: It may be possible to use caspase-6 and caspase-7 as immunohistochemical markers of gastric cer. Mutations in the caspase-6 gene in gastric can-cer are rare – in a study by Lee et al. the rate was 2% (1/50). However there is still the possibility that genetic alternation coexisting with other mutated apoptosis-related genes may disrupt apoptosis and lead to carcinogenesis [34].

caspases, and disturbance at this stage may result in the withholding of the apoptosis cascade. Nor-mal gastric mucosal cells present weak or no ex-pression of those caspases. In gastric cancer cells the expression of caspase-8 and -9 was 93% and 90% respectively, with high co-expression. There was no correlation between caspase-8 and -9 ex-pression the histological type of the cancer (diffuse or intestinal types).

High expression of caspase-8 and -9 should promote apoptosis and cancer cell elimination, but the damaged cell clones proliferate and avoid apoptosis. There must be a mechanism by which tumor cells are resistant to apoptosis even when the activity of caspase-8 and -9 is maintained [35].

The potential role of caspase-8 in gastric can-cer treatment has been investigated. Kanehara et al. investigated the status of caspase-3 and -8 af-ter treatment with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). The results revealed that 24 hours after treatment with TRAIL, caspase-8 activity was correlated with the antican-cer effect of TRAIL. The effects of anticanantican-cer ther-apy may be predicted by caspase-8 activation after TRAIL treatment [36]. Cancer cell apoptosis may be disrupted in several ways, including somatic mutation, the loss of expression of proapoptic pro-teins and the overexpression of inhibitors of apop-tosis. Gene polymorphism and somatic mutations of caspase-8and -9are interesting. Somatic muta-tions of the caspase-8 coding gene have been iden-tified in cell lines of advanced gastric cancer. In one study, as many as eight out of nine mutations reduced the cell’s susceptibility to apoptosis. Those were typical mutations of function loss related to gene frame shift and the production of abnormal protein.Theauthors suggest that mutations of the caspase-8 gene may influence the pathogenesis of gastric cancer, especially in the late stage [37].

The situation is different in case of polymor-phisms. Liamarkopoulos et al. demonstrated that some of them, for example CASP8-652 6N ins/ del and CASP (-1263 A>G, may have a protective character, reducing the risk of developing gastric cancer. Other polymorphisms play no important role [38].

The phosphorylation of caspase-9 (p-cas-pase-9) through the mitogen-activated protein ki-nase (MAPK) pathway is associated with a loss of caspase-9 function. Normal mucosal cells present no p-caspase-9 expression, but in gastric cancer cells the expression was frequent. This may suggest that p-caspase-9 expression could be one way cas-pase-9-mediated apoptosis is resisted. P-caspase-9 expression has no correlation with the presence of metastases in gastric cancer, suggesting a role in early cancerogenesis [39].

As in the case of caspase-8 and-9, the expres-sion of caspase-10 was higher in the gastric tumor cells, with weak or no expression in normal mu-cosal cells. Caspase-10 is a member of the same CED-subfamily and also plays a role in activating apoptosis [35].

Caspase-10 may cause disturbances in the apoptosis of gastric cancer cells as a result of loss- of-function mutations in the genetic background. A missense mutation in a region coding the sec-ond death effector domain leads to a change in the amino acids, a structural change in the caspase-10 prodomain, and prevents DED-DED interaction between FADD and caspase-10. Consequently, there is no catalytic activation of caspase-10 and thus this pathway of apoptosis is inhibited. Alter-natively, a nonsense mutation in the caspase-10 coding gene leads to a loss of the catalytic center because of the shorter truncated protein and may be similar to a short form of FLICE-inhibitory pro-tein (FLIPs). This may be a typical loss-of-function mutation. In the same study, the researchers ob-served that the somatic missense mutation, which seems to be identically to common Danish poly-morphism in the caspase-10 gene, leads to a sig-nificant defect in caspase-10-mediated apoptosis. The researchers concluded that somatic mutation in the caspase-10 coding gene might contribute to pathogenesis in a subset of gastric cancers through the loss of their apoptotic function [40].

The role of caspase-14 in gastric cancer was in-vestigated in a large international study of 180 cas-es of gastric cancer [41]. The rcas-esearchers observed that caspase-14 expression was associated with his-tological type and cellular differentiation in gastric cancer tissue. Caspase-14 expression was signifi-cantly higher in well-differentiated intestinal-type gastric cancers and lower in poorly differentiated diffuse-type cancers. Also, signet ring cells con-taining cancers show lower levels of caspase-14 ex-pression compared with non-signet ring cell tu-mors. Caspase-14 phylogenic analysis showed that its catalytic domain is most similar to the pro-inflammatory caspases -1, -4 and -5. The role of caspase-14 in gastric cancer might be related to chronic inflammation and intestinal metaplasia as result of cytokine activation during the gener-ation of protective immunity against the bacterial pathogens [41].

Conclusions

role in maintaining homeostasis in an organism. Disorders occurring during a cascade of intercel-lular reactions lead to loss of regulation of tissues, organs or entire organisms. In carcinogenetic cesses, cells become resistant to apoptosis and pro-liferation and migration become disturbed, which consequently leads to proliferation of abnormal clones of cells.

Deregulation of expression, somatic mutations and genetic polymorphisms are the main mecha-nisms of gastric cancer cells’ resistance to apopto-sis. This review of caspases’ structure, function and importance in gastric cancer indicated a strong correlation between each caspase and the process of carcinogenesis.

Apoptosis initiating caspases -8, -9 and -10 show increased expression in gastric cancer cells, while their expression in normal mucosal cell is low or even absent. Interestingly, there was no signif-icant relationship between the level of expression and clinico-pathological features. All of these cas-pases are susceptible to genetic alterations, somatic mutations and other inactivation processes leading to a loss or reduction of their biological function. The phosphorylation of caspase-9 and convention-al mutation of caspase-10 involve significant ab-normal apoptosis encouraging abab-normal cell clone proliferation. In the case of caspase-8 the relation-ship between carcinogenesis and loss of function is less pronounced.

Caspases-3, -6 and -7 deal present common problems. In all of these cases, normal expression is observed in healthy cells and is reduced in gas-tric cancer cells. Genetic disorders may lead to that dysfunction, but there is not yet sufficient ev-idence. Caspase-6 and -7 expression also appears to correlate with clinical and pathological features, especially at an early stage of gastric cancer devel-opment. In the case of caspase-3, investigations of the relationship seem to be inconclusive.

Caspase-1 and -14 belong to the pro-in-flammatory caspases, but also appear to play an

important role in gastric cancer. This may be as-sociated with chronic inflammation as one of the reasons for the development of gastric cancer [42]. Interestingly, there is a significant relationship be-tween both of these caspases and clinico-patholog-ical features, and with the survival prognosis. The role of caspases -4 and -5 is not clear, but the pres-ence of somatic mutations suggest that proinflam-matory caspases play a part in apoptosis and more detailed studies are required.

Caspases may therefore be a key to the devel-opment of more effective anti-cancer therapies. However, defining a particular abnormality typ-ical for a particular disease is difficult, especially considering the complex nature of the activation pathways and their unclear relations. There are still remaining many unanswered questions, and verification of specific abnormalities as character-istic of the disease requires continuing research in-volving as large a number of patients as possible.

The example of gastric cancer presented in this paper provides an insight into the multi-direction-al character of the research being done in pursuit of an effective future anticancer treatment. It should be remembered that carcinogenesis is and always will be a problem requiring the combined efforts of biologists, pathologists, oncologists, geneticists, surgeons and other specialists in various areas. The expected benefits associated with continued re-search include supplementation of surgical therapy, which is still the basic therapy for the majority of neoplasms; early and effective selection of patients for appropriate therapeutic protocols (e.g. neoadju-vant therapy) depending on the potential adneoadju-vantag- advantag-es. Continued research can also extend prognostic possibilities, improving predictions of the effective-ness of treatment and the risks of local recurrence or extra-tissue cancer cell migration. It may also lead to the introduction of new treatment strategies (so-called targeted therapy) to oncological practice and to the exclusion of systemic therapies that are burdened with numerous complications.

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Address for correspondence:

Ewelina Frejlich

2nd Department of General and Oncological Surgery Wroclaw Medical University Hospital

Borowska 213 50-556 Wrocław Poland

Tel.: +48 695 559 563 E-mail: [email protected]

Conflict of interest: None declared