**3. Apoptosis and cancer**

Cancer is a multi-complex disease arising from mutation of tumor suppressor or oncogenes, which can be due to impairment of several signaling pathways and associated with apoptosis [80]. The evasion of apoptosis is an important sign of cancer, which mostly results in a very high level of resistance to many chemotherapeutic agents. Therefore, attention has been given to apoptotic proteins and became the most attractive anti-cancer target for effective treatment. The resistance against apoptosis is seen in many types of cancers, and required much higher doses of drugs to overcome the resistance, which increases the risk of "off-target" negative effects [81]. Apoptosis is also a very complex system with its regulation and/or function via several proteins, kinases, and links with several pathways might provide important therapeutic benefit for novel approaches to treat cancer. A better understanding of apoptotic pathways and their alteration in cancer cells had the attention of the development of pro-apoptotic agents. In cancer cells, cell death has been reported for a long time and associated with cancer therapy

#### *The Program Cell Death (Apoptosis) and the Therapy of Cancer DOI: http://dx.doi.org/10.5772/intechopen.97289*

due to the designation of killing malignant cells by chemo- and/or radiotherapy. Unfortunately, this also leads to the death of normal cells along with cancer cells [80]. Kerr et al. reported that radiation increases the rate of apoptosis in cancer cells. Therefore, apoptosis-associated directly with the development of cancer cells in humans. Most importantly, it can also promote resistance to the therapies which aim to kill cancer cell by targeting apoptosis [2]. In healthy cells, the recognition of damage leads to activation of program cell death, which ending up with their elimination by killing themselves. Such as the activation of tumor suppressor p53 can activate apoptosis inducers including Puma and Noxa, in normal cells [61, 62, 64]. Thus, the un-endowed cells which are not able to eliminate themselves may cause an accumulation of genetic damage. The oncogenes (e.g. Myc) induced apoptosis is a very crucial protective mechanism against cancer development, controlled by a p53-dependent pathway, activated in response to abnormal mitogenic signals. Such as the cells overexpressing the apoptosis inhibitor Bcl2 or have damage in the p53 gene are incapable of this removal process including cancer cells. The loss of function mutations in the p53 tumor suppressor gene causes defection in the apoptotic mechanism and also impacts its tumor-suppressive functions, which involving DNA repairment, cell-cycle arrest, and cell senescence [82, 83]. Also, the incapable cells with an additional mutation can lead to atypical cell proliferation. Such as chromosome translocation which ends up with over-expression of c-Myc, then will lead to quick malignant cell growth. Of note, this explains that the defects in apoptotic mechanism such as over-expression of apoptosis inhibitor Bcl-2 and unregulated cell proliferation due to c-Myc overexpression in the development of lymphoma [82, 84, 85].

Understanding of extrinsic apoptotic pathway on death receptors especially on TRIAL signaling, including TRAIL-1 and -2, shown to lower toxicity compared to TNF or FAS signaling were targeted. The lowering expression levels of TRAIL receptors via recombinant human TRAIL-R1 and -R2 opened up an important therapeutic window. The tumor cells pre-dominantly signal via TRAIL-R1 and -R2, and the expression of these receptors displayed that they do not behave as an indicator of favored receptor signaling [86]. The higher expression levels of these receptors have been found on brain tissue and hepatocytes [87, 88]. However, these findings opened up several windows about the use of TRIAL in the clinic which has been reported notable apoptotic cell death in primary hepatocytes. These findings underlining the importance of selective models to evaluate appropriate drug combinations and their potential on- and/or off-target adverse effects. A study by Von Karstedt et al. in KRAS mutant cancers, known to have an elevated level of TRAIL-R2 expression shown that triggering TRAIL signaling is worsen the development of cancer, and metastasis [89, 90]. Several other studies in different cancers such as pancreatic cancer also shown that TRIAL-R2 signaling interacts with the miRNA processing complex and leads to the inhibition of the maturation of mRNA, called let-7. The result of TRAIL R2-miRNA interaction displayed an up-regulated level of let-7 transcriptional target genes which also prompt cancer development [91]. The sublethal concentrations of TRAIL were shown to induce the activation of caspase-8 dependent apoptotic nucleases, which leads to increased mutations caused by DNA damage in surviving cells [92].

The IAP proteins which are downstream of DRs and Bcl2 family proteins are excessively expressed in cancer cells, therefore associated with cancer development and as well as resistance to therapies. The involvement of cIAPs has also been associated with the regulation of the NF-KB pathway. The lack of cIAPs leads to an increase of inflammatory cytokines including TNF family and IL6, IL8, IL10, and Mcl1, which are effectively involved in systemic inflammation [93]. Nevertheless,

these cytokines have also been reported to be increased in cancer progression due to their link with inflammation which causes to include immune cells to the cancer site and prompt the tumor development (**Figure 2**).

### **3.1 p53 phosphorylation: is it a driver to cell death?**

DNA damage comprises different causes that involve cell stress, mutations, toxic agents, or genomic lesions. Upon damage, the confrontation of cells to damage lead to their potential responses to repair the damage or eliminate the harmed cells by apoptosis. Usually, the response of the damaged cell fate differs from strength to the

#### **Figure 2.**

*Molecular mechanisms of intrinsic and extrinsic apoptotic signaling pathways activation.* **a.** *the activation of intrinsic apoptotic signaling pathway through cellular stress (ER), and deprivation of growth factor (DNA damage). This alteration promoting the activation of pro-apoptotic members of the BCL-2 family, including BIM, PUMA, BAD, or NOXA. The BH3-only proteins bind and therefore, neutralize BCL-2 pro-survival proteins, involving BCL-2, BCL- XL, MCL-1, and BCL-2- related protein A1. Thus, releasing the important effectors of apoptosis signaling, such as BAK and BAX, which then unite into big complexes and lead to incompliance in the mitochondrial outer membrane. Afterward, this results in the free off cytochrome c and SMAC, which are apoptogenic factors. Some of BH3- only proteins (BIM and PUMA) recently have been reported to bind and promote the activation of MOMP. Also, cytochrome c binding to APAF1 leads to the formation of the apoptosome complex. Caspase-9 is activated in this apoptosome complex, and later proteolytically causes the activation of both caspase- 3 and caspase-7. The caspase inhibitor XIAP activity is blocked by SMAC.* **b.** *Extrinsic apoptotic signaling is stimulated through the activation of DRs, for example, the activation of FAS via FAS ligand (FASL). This cause the pro-caspase 8 recruitment to the death receptor by the adaptor protein FADD, and ending up with the formation of the DISC, which is an important catalyzer for the caspase-8 activation. The activation of caspase 8 promotes cell killing, in two ways: i) via direct activation of caspases 3 and 7, or ii) by indirect activation of the BH3 only protein BID into tBID, thus linking with the intrinsic apoptotic signaling pathway. Effector caspases can be activated in two ways, either by cleavage of hundreds of intracellular proteins to promote the conventional apoptotic morphology and to avoid the efflux of intracellular DAMPs which can result in inflammatory responses. In addition, effector caspases either directly or indirectly can lead to activation of ROCK-1 kinase, which is important for plasma membrane blebbing by actin contraction and caspase-activated DNAse (CAD), which cause inter-nucleosomal DNA cleavage and thus chromatin condensation. These effector caspases also can proteolytically lead to the inactivation of lipid flippases including ATP11, and also can activate the lipid scramblase such as XKR8. Altogether, this results in the exposure of PtdSer on the cell membrane. Therefore, this is called an 'eat me signal' for phagocytic cells and ending up with the engulfment of cells that are undergoing apoptosis [94].*

#### *The Program Cell Death (Apoptosis) and the Therapy of Cancer DOI: http://dx.doi.org/10.5772/intechopen.97289*

content of DNA damage, and different signaling pathways are activated. The different cellular decision-making process is guided with specific signaling molecules at the molecular level [95]. The tumor suppressor p53 plays a key role in response to DNA damage and involve in the cellular decision-making process, in turn, found to prompt DNA repair by shutoff the cell cycle to allow time for the repairment and restore genome stability [96]. However, it remains unknown still that how p53 does different cell-fate decisions, activate different target genes, and at a non-transcriptional level have an effect in the cytoplasm of MOMP [97]. The phosphorylation of p53 at Ser15 and Ser20 occurs over immense DNA damage and these are critical sites for stabilization of p53 [98]. Importantly, there is a specific phosphorylation site which is Ser46 and selectively associated with the activity of cell killing caused by genotoxic stress. These crucial different phosphor-sites of p53 might improve the understanding of the function of specific phosphorylation sites. Many studies focused on the phosphorylation of p53 at Ser46 which modulates the target gene transactivation and it has been implicated to potentiate cell death transcriptionally and also, non-transcriptionally [95]. The transactivation of significant pro-apoptotic p53 target genes including BAX, NOXA, p53AIP1, PTEN, and AKT kinase, which are all up-regulated over p53 Ser46 phosphorylation [63, 99–101]. How this different transcriptional effect occurs and which mechanisms are evolved is an open question? The specific role of the p53 Ser46 phosphorylation site in making a cellfate decision upon cell death leads to the thought of its disturbed regulation might cause cancer cell resistance to therapy. The deregulation of p53 Ser46 kinases in cancer cells could be caused by many factors involving mutations, mis-localization, lower expression level, and disturbed kinase activity. The mutations of p53 at Ser46 have been reported with 1.4–1.6% rates in different human cancer types including breast, colorectal, lung, and stomach. Besides, the down-regulation of several p53 Ser46 kinases are reported such as ATM, DYRK2, HIPK2, and p38 in different tumor types [63, 102–104]. Although, since the regulation of the kinase relies on p53 Ser46 activity, their specific potentials and activities is a challenging task.

#### **3.2 The pro-oncogenic roles of apoptosis**

The deregulation of apoptosis is a feature of cancer cells. Apoptosis is a tumor suppressor mechanism in cancer cells. Hence, the dysregulation of apoptosis causing uncontrollable cancer cell-survival [105]. Besides, the tumorsuppressive effects of apoptosis, also, suggested having pro-oncogenic functions. The maintenance of aggressive tumors by apoptosis has been described in the literature. The term tumor aggressiveness defined with increased proliferation, and metastatic features of cancer cells. Such as apoptotic stresses are maybe the reason for aggressive tumor development. Therefore, have pro-oncogenic roles in cancer development and aggressiveness [106, 107]. Importantly, this opens several questions about the beneficial effects of apoptosis in cancer therapy, since strategies aim to trigger apoptosis [108]. Apoptosis-induced proliferation (AIP) is an important compensatory proliferation form, which helps to eliminate damaged cells and maintain tissue homeostasis. Caspase-dependent inflammation and repressed AIP activity might lead to tumor development. One important mediator of AIP is Prostaglandin-E2 (PGE-2), and functioning as a trigger of tumor proliferation in apoptotic cells [109]. The regulation of tumor cells by PGE-2 through caspase-3 dependent manner is an important strategy to treat melanoma. Here, the dying cells trigger the growth of living-cells after cytotoxic therapy [110, 111]. Therefore, targeting PGE-2 to enhance the response to therapy might help to overcome the resistance in cancer cells, when designing cancer therapies [112].

#### **3.3 The oncogenic roles of apoptosis**

Apoptotic cells release the "eat-me" signal which is a signal for phagocytosis. These dynamic signals are including fractalkine (FTK), phosphatidylserine (PS), nucleotides ATP, and UTP [113, 114]. These signals are responsible for immune cell activation. Such as the effects of FTK associated with cell migration, and angiogenesis. Also, these signals by apoptotic cells are thought to induce inflammation. Somehow, in the occurrence of normal apoptotic cell death, inflammation is prevented by efferocytosis. The mechanism of efferocytosis is to eliminate the apoptotic cells by immune cells, macrophages. Importantly, it also plays a key role in cancer cells by creating an immunosuppressive effect to organize several signalization events between tumor cells [115]. For example, a study by Ford and colloquies, in 2015, showed that apoptotic cancer cells induce tumor growth and angiogenesis in B-cell lymphoma and also, malignant melanoma [116]. In conclusion, the effects of apoptotic cells in their microenvironment is changed upon their features. The studies that suggest apoptotic cell involvement might play important role in signaling events due to their environment and features. Of note, it remains unknown how they do these roles, therefore, more studies are needed to clarify this point.

### **4. The therapeutic approaches to cancer**

The question what happens when apoptotic machinery is impaired? The answer is impairment of apoptosis results in uncontrolled cell proliferation, tumor development and progression, and resistance to anticancer therapeutics [117, 118]. The deregulation of apoptosis is not only causing the development and progression of the tumor but also lead to resistance to therapeutic approaches [80]. Cancer cells can promote drug resistance to several treatments, which target several different molecular signaling. So many cancers present an internal resistance to chemotherapy, without previous exposure to anti-cancer agents. Therefore, the beginning response to cancer treatment is weak. Furthermore, several tumors might also gain drug resistance because of chemotherapy [119–122]. Among many intrinsic and gained drug resistance are increased drug leakage, defective inflow transporters, changes in the function of drugs and their targets, drug inactivation metabolically, drug compartmentalization, increased activity of DNA repairment [120, 123–126]. To improve therapeutically new approaches, the understanding of these signaling pathways is required to progress cancer therapy and more importantly get over the resistance. Besides, targeting the apoptotic signaling events that are involved in resistance may improve a valid strategy to increase the cell sensitivity of cancer cells to apoptosis and overcome the failure of therapeutic difficulties.

#### **4.1 The role of Bcl2 family proteins in cancer and drug resistance**

The Bcl-2 family proteins contains at least one Bcl-2 homology (BH) domain [127–129]. The different BH-domains exist, but especially BH-3 is crucial to form several BCL-2 protein complexes that govern the result between survival and death [130–133]. These interactions between BH3-BH3 are also important to antagonize pro-survival complexes. The increased level of anti-apoptotic Bcl-2 family proteins is associated with carcinogenesis. Such as the transgenic mice model that overexpress Bcl-2 shown to develop tumors spontaneously [134]. However, the high expression of Bcl-2 was also reported to cause a shift in the balance of cancer cells through survival and also lead to resistance to chemotherapy [135]. Overexpression of Bcl-xL, another member of the Bcl2-family is also associated with colorectal

cancer [136]. This unbalanced expression of Bcl-2 family proteins confers failure of apoptosis and resistance to cancer therapies [137]. Especially, overexpression of anti-apoptotic proteins such as Bcl-2 and Bcl-xL is correlated with several cancers including ovarian and breast cancer [138, 139]. On the other side, an increased ratio of Bcl-2/Bax is associated with chronic lymphocytic leukemia (CLL) [140].
