Keywords

4.1 Introduction

Cancer is the second major noncommunicable disease and it caused about 8.2 million deaths in 2012 (Torre et al. 2015). Cancer is defined as a group of diseases in which normal tissue or organ is invaded by abnormal dividing cells. If left untreated, it spreads throughout the human body and even results in loss of human life (Eid et al. 2015). Carcinogenesis is the metabolic process, in which normal cells are transformed into cancer cells through three major stages such as initiation, promotion, and progression. The initiation (first) stage occurs in the normal cells due to exposure to carcinogenic (procarcinogens, epigenetic carcinogens, genotoxic carcinogens, etc.) or mutagenic (physical and chemical) agents. However, this first stage alone is not enough for tumor formation. In promotion (second) stage, tumor promoter agents help to convert the initiated cells into tumor cells. This second stage occurs very slowly, and it even takes several months to years depending upon changes in diet and lifestyle of the individual person. Progression (third/final) stage converts the tumor cells into high-degree malignant cells. At this progression stage, human diet has less impact on tumor progression (Reddy et al. 2003; Rajesh et al. 2015). The cancer incidence cases have been reported more from the developed countries compared to that of developing countries. Similarly, in the developing countries, breast, lung, and colorectal cancers are reported more among females (Jemal et al. 2011). Various risk factors have been reported in the cancer development which includes age, geographic area, and race (Millimouno et al. 2014). Chemotherapy is the major treatment approach; however, in recent years 30–80% of cancer patients are developing resistance to chemotherapeutic drugs. Therefore, plant-based substances (phytoconstituents) have attained much attention among the researchers. The present chapter deals with the anticancer effect of several compounds of plant origin and their mechanisms of action.

4.2 Mechanisms Involved in Cancer Chemoprevention and Treatment

Plant-based substances have been reported to induce cell cycle arrest and apoptosis by targeting multiple cellular signaling pathways such as (1) p53 pathway, (2) nuclear factor-kappaB transcription factor pathway, (3) nuclear factor-related factor 2 signaling pathway, (4) growth factors pathway, (5) signal transducers and activators of transcription (STAT) pathway, (6) Wnt/β-catenin pathway, (7) hedgehog (SHH) signaling pathway, (8) phosphatidylinositol 3 kinases (PI3K) pathway, (9) cyclooxygenase 2/prostaglandin E2 (COX 2/PGE2) pathway, (10) mitogen-activated protein kinase (MAPK) signaling pathway, (11) Cripto 1 protein signaling pathway, and (12) hypoxia signaling pathway. Thus, by targeting/modulating the abovementioned cellular signaling pathways, anticancer activity has been successful achieved by plant-based substances (Millimouno et al. 2014).

4.2.1 Cell Cycle Arrest

The cell cycle is the metabolic process by which cell progress and division consist of several biochemical and molecular signaling pathways. It exhibits four important stages or phases.

  1. 1.

    G1 (Gap 1) phase in which the cell grows and prepares to synthesize deoxyribonucleic acid (DNA)

  2. 2.

    S (synthesis) phase in which the cell synthesizes DNA (genetic material)

  3. 3.

    G2 (Gap 2) phase in which the cell prepares to divide

  4. 4.

    M (mitosis) phase in which cell division occurs and finally phase is termed as G0 (resting phase) in which cell leaves the cell cycle and quit dividing them

Cyclin-dependent kinases (CDKs) are group of enzymes which regulate the cell cycle transitions. These enzymes contain two subunits, namely, catalytic subunit (CDK) and regulatory subunit (cyclin). Each phase of the cell cycle has individual CDK cyclin enzymatic activity as shown below:

  1. 1.

    CDK2 cyclin E and A regulates G1 to S phase transition.

  2. 2.

    CDK1 cyclin A regulates late S to G2 phase transition.

  3. 3.

    CDK1 cyclin B regulates G2 to M phase transition.

In mammalian cells, about 10 CDKs and 20 cyclins have been reported; interestingly not all participate in cell cycle regulation. In normal cells cell cycle machinery controls the cell proliferation, which also includes repair mechanism with them. In the case of cancer cells, cell cycle machinery loses control and results in uncontrolled cell proliferation (Collins et al. 1997; Hwang and Clurman 2005).

4.2.2 Apoptosis

Apoptosis is the process of active cell death resulting in the breakdown of cellular structures, without causing any immune or inflammatory response to the host. It is also referred as “programmed cell death,” which occurs in the several physiological and pathological conditions (Cummings et al. 1997; Iannolo et al. 2008). It is characterized by biochemical and morphological hallmarks such as cell shrinkage, chromatin condensation, cytoplasmic membrane blebbing, and nuclear DNA fragmentation (Fulda and Debatin 2006). It is essential for embryonic development, tissue homeostasis, immune function, and tumor suppression especially in multicellular organisms (Iannolo et al. 2008). It usually maintains balances between pro-apoptotic (BAD or BAK, BAK and BID) and anti-apoptotic (Bcl-2 and Bcl-Xl) signals. The accumulation of pro-apoptotic signals leads to apoptosis induction. Defects in apoptotic pathways have been observed in several diseases, including tumor and neurodegenerative disorders (Lowe and Lin 2000). Three important biochemical events occur during apoptosis; they are (1) activation of caspases activity, (2) breakdown of DNA and protein, and (3) membrane modifications due to phagocytes (Wong 2011). The process of apoptosis is mainly divided into two pathways: (1) intrinsic pathway mediated by molecules released from mitochondrial membrane and (2) extrinsic pathway triggered by death receptor.

4.2.2.1 Intrinsic Pathway of Apoptosis

The intrinsic pathway is activated by physical or chemical stimuli such as cell detachments, cytokines, DNA damage, growth factor deprivation, hypoxia, and/or other stress signals. These stimuli modulate mitochondrial functions such as increases the mitochondrial membrane potential (MMP) and releases the cytochrome C into the cytoplasm. This cytosolic cytochrome C in turn interacts/binds with apoptotic protease-activating factor 1 (Apaf 1) and pro-caspase 9 (zymogen). Pro-caspase 9 activates caspase cascade such as caspases 3, 6, and 7, leading to DNA fragmentation and cell death. B-cell leukemia/lymphoma 2 (Bcl2) family proteins play an important role in regulating the intrinsic pathway by inducing or preventing the release of cytochrome C (Lowe and Lin 2000; Iannolo et al. 2008; Millimouno et al. 2014).

4.2.2.2 Extrinsic Pathway of Apoptosis

The extrinsic pathway is activated, when a death ligand binds to the extracellular domains of the death receptor and leads directly to caspase activation. For instance, Fas ligand (FasL) binds with its respective receptor Fas receptor (also called as Apo 1 or CD95), which forms death-inducing signaling complex (DISC) which contains the specific adaptor protein, namely, Fas-associated death domain protein (FADD) and caspase 8. Caspase 8 in turn activates caspase 3 and apoptosis in type 1 cells. The extrinsic pathway is quite similar to the intrinsic apoptotic pathway, which is also caspase-dependent; the one and only difference is that apoptotic signaling is initiated through membrane-bound death receptors (Wajant 2002; Iannolo et al. 2008).

4.2.2.3 Caspase-Independent or ROS-Mediated Apoptosis Pathway

Reactive oxygen species (ROS) are generated due to physiologic stress which is associated with the production of oxidative species through intracellular damage to DNA, lipids, proteins, and RNA. During cellular redox the excessive generation of ROS in turn induces oxidative stress, loss of cell function, and apoptosis. Granzyme A (enzyme belonging to serine proteases family) directly induces the ROS generation which in turn results in caspase-independent mitochondrial damage. Then ROS drives the endoplasmic reticulum (ER)-associated SET complex into the nucleus, where it activates apoptosis. ROS also mediates poly(ADP-ribose) polymerase 1 (PARP-1) activation, which is needed for apoptosis-inducing factor (AIF) release from mitochondria. Thus, AIF is the main pro-apoptosis factor involved in caspase-independent apoptosis pathway (Martinvalet et al. 2005; Lieberman 2010).

4.2.3 Necrosis

Necrosis is the process of passive cell death resulting in the breakdown of cellular structures, caused by specific physiological and pathological stimuli such as tumor necrosis factor (TNF), TNF-related apoptosis-inducing ligand (TRAIL), lipopolysaccharides (LPS), oxidative stress, and DNA damage (via PARP). Necrosis is characterized by biochemical and morphological hallmarks such as loss of membrane integrity, cell swelling, permanent loss of mitochondrial membrane potential, and DNA fragmentation (post-lytic/late stage). Specific physiological and pathological stimuli activate receptor-interacting protein (RIP1) kinase. This RIP1 kinase directly/indirectly transduces signal to mitochondria which leads to mitochondrial permeability and transition. This mitochondrial damage enhances enzymes such as protease (calpains, cathepsin) and phospholipase activities and finally results in plasma membrane destruction (sign of necrotic cell death). In contrast to apoptosis process, the mode of cell death is required for tumor regression, and thus necrosis plays important in anticancer therapy (Proskuryakov and Gabai 2010).

4.2.4 p53 Pathway

p53 (tumor suppressor gene) is stimulated by cellular stress like carcinogens, hypoxia, ionizing radiation, oxidative stress, and UV radiation. It regulates the apoptosis, genomic integrity, cell cycle, and DNA repair caused due to genotoxic stresses. p53 binds to regulatory DNA sequences as a tetramer and transactivates genes involved in apoptosis in response to DNA damage (ASPP1/2, BAX, Fas, NOXA, p53AIP1, PERP, PIDD, PUMA), cell cycle arrest (p21, cyclin G1, GADD45, 14-3-3, reprimo), angiogenesis (BA11, GD-AIF, maspin, TSP1), and genetic stability (DDB2, MSH2, p21, XPC). About 40 different isoforms of the p53 family members have been reported so far. Among these, p73 plays a significant role during neurogenesis, whereas p63 is essential in skin, limb, and craniofacial development. Interestingly, some p53 isoforms have oncogenic potential, while others have tumor suppressor activity (Millimouno et al. 2014; Pflaum et al. 2014).

4.2.5 NF-κB Pathway

Nuclear factor-kappaB (NF-κB transcription factor) is stimulated by cellular stress like carcinogens, cytokines, endotoxins, ionizing radiation, oxidative stress (free radicals), and UV radiation. It is involved in tumor initiation and progression. NF-κB is usually present in the cytoplasm via association with its IκB (endogenous inhibitor of NF-κB), which further phosphorylated by IκB kinase (IKK). IKKa activates metastasis in prostate cancer by inhibiting maspin (mammary serine protease inhibitor) and stimulates angiogenesis via activating interleukin 8 (IL8) and vascular endothelial growth factor (VEGF). The accumulation of the IκBα leads to activation of anti-apoptotic NF-κB, resulting in apoptosis. NF-κB pathway plays significant role in carcinogenesis by transactivating genes involved in angiogenesis, apoptosis, cell proliferation, metastasis, and tumor cell invasion (Millimouno et al. 2014).

4.2.6 Nuclear Factor-Related Factor 2 (Nrf2) Signaling Pathway

Nrf2 (belongs to the Cap ‘N’ Collar family) plays an important role in transcriptional activation of phase II detoxification enzymes (glutathione S-transferases) and plays as an important regulator of cell survival both in normal and cancer cells. It protects against chemical carcinogen by decreasing the ROS content and DNA damage in cells. Its defense role has been reported against various diseases such as acute pulmonary injury, aging, cancer, cardiovascular disease, diabetes, inflammation, photooxidative stress, and pulmonary fibrosis. Similarly, relationships between p21, p62, and Nrf2 have been reported. Nrf2 activators (from phytochemicals) have been shown to induce the Nrf2-mediated defense mechanism by enhancing phase II detoxification enzymes, antioxidants, and (ABC) transporters; this in turn protects the carcinogenic stimuli (Jaramillo and Zhang 2013; Millimouno et al. 2014).

4.2.7 Growth Factor Pathway

Growth factors (GFs) and growth factor receptors (GFR) play a vital role in physiological conditions such as growth and differentiation, wound healing, etc. Growth factors like colony-stimulating factor (CSF), fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), insulin-like growth factor (IGF), and transforming growth factor (TGF) are few growth factors involved in carcinogenesis. Growth factor receptor activation leads to downstream signaling of PI3K-Akt and Ras-MAPK pathways and thus acts as target for numerous anticancer/antitumor agents. For instance, suramin (polysulfonated drug) inhibits the binding of growth factors like epidermal growth factor (EGF), fibroblast growth factor (FGF), insulin-like growth factor (IGF1 and IGF2), nerve growth factor (NGF), platelet-derived growth factor (PDGF), and transforming growth factor (TGF-α) to their receptors and induces disassociation of bound growth factors from their respective receptors and thereby has exhibited anticancer activity against adrenal, prostate, and renal cancer (Rajkumar 2001; Millimouno et al. 2014).

4.2.8 Signal Transducers and Activators of Transcription (STAT) Pathway

STAT belongs to family members of interferon (IFN) signaling complex and plays a dual role as signal transduction and transcription activators. STATs are cytoplasmic proteins that are activated by tyrosine kinases resulting in phosphorylation and dimer formation and translocate into the nucleus to regulate the transcription of genes. Seven STAT family members have been reported so far; they are STATs 1, 2, 3, 4, 5a, 5b, and 6. STAT3 activation has been reported by both Janus kinase (JAK) and also by non-receptor tyrosine kinases (c-Src kinases). Thus, inhibition of JAK/STAT pathway has been recognized as novel chemopreventive and chemotherapeutic target (Millimouno et al. 2014; Xiong et al. 2014).

4.2.9 Wnt/β-Catenin Pathway

Wnt binds with frizzled family receptor (FZD) and leads to the cytoplasmic accumulation of β-catenin. Then β-catenin translocates into the nucleus, where it interacts with T-cell factor/lymphoid enhancer factor 1 (TCF/Lef 1) transcription complex and regulates the transcription of genes. Wnt/β-catenin pathway has been reported in several cancers, and Wnt inhibition has been recognized as new target for colorectal cancer treatment (Pai et al. 2017).

4.2.10 Hedgehog (SHH) Signaling Pathway

SHH (glycoprotein) binds and inactivates the patched 1 (PTCH1) receptor, which inhibits the protein smoothened (SMO) activity. This in turn leads to activation of glioma-associated (GLI) transcription factors. Then GLI translocates into the nucleus and regulates the transcription of genes. Abnormal hedgehog (SHH) signaling that has been reported in several cancers includes colorectal, glioma, pancreatic, and prostate carcinoma. GLI 1 small interfering RNA (siRNA) has been used to induce apoptosis in prostate cancer, and thus hedgehog (SHH) signaling has been recognized as new target for cancer treatment (Wang et al. 2012; Rimkus et al. 2016).

4.2.11 Phosphatidylinositol 3 Kinases (PI3K) Pathway

Activation of phosphatidylinositol 3 kinases (PI3K) leads to the production of phosphatidylinositols [P2 and P3 (Ptdlns 3, 4) and P3 (Ptdlns 5)], which are bound by Akt. Phosphatidylinositols are further activated by phosphoinositide-dependent protein kinase 1 (PDK1) and Akt. These activated phosphatidylinositols are translocated into the plasma membrane and regulate the cellular process. Phosphatidylinositol 3 kinases (PI3K) play a significant role in cellular transformation and cancer development, and thus inhibition of PI3K serves a new target for cancer therapy (Liu et al. 2009; Wang et al. 2012).

4.2.12 Cyclooxygenase 2/Prostaglandin E2 (COX 2/PGE2) Pathway

Inflammatory stimuli activate COX 2 activity, which in turn activates PGE2. This extracellular PGE2 binds with prostaglandin E2 (EP) receptors and initiates multiple intracellular signaling pathways. COX 2 is one of the pro-inflammatory mediators, found to be elevated at the early stage of tumorigenesis (Reader et al. 2011; Wang et al. 2012).

4.2.13 Mitogen-Activated Protein Kinase (MAPK) Signaling Pathway

The RAS (small G protein)/RAF (rapidly accelerated fibrosarcoma)/MEK (mitogen-activated protein kinase)/ERK (extracellular signal-regulated kinase) signaling pathway involves complex network that regulates the proliferation, differentiation, cell survival, and apoptosis. The ligand (e.g., cytokine, growth factor, or hormone) binds with receptor tyrosine kinase (RTK). In this pathway, three protein kinases play a major role, namely, mitogen-activated protein kinase kinase kinase (MAPKKK or BRAF) that phosphorylates and activates mitogen-activated protein kinase kinase (MAPKK or MEK), which in turn activates mitogen-activated protein kinase (MAPK or ERK). This activated ERK translocates into the nucleus and regulates the gene expression. Numerous inhibitors of components of this pathway have been reported and recognized as good therapeutic approach for melanoma treatment (Kolch 2000; McCain 2013). Similarly, genistein (soy-derived isoflavone) has been reported to inhibit cervical cancer (HeLa and CaSki) cells via by inhibiting ERK1/2 activity and activating p38 MAPK pathway (Kim et al. 2009).

4.2.14 Cripto 1 and Its Allied Protein Signaling Pathway

Cripto 1 (CR1) mediates several cellular processes such as angiogenesis, cell growth, fetal development, inflammation, invasion, migration, tumor formation, and wound repair. CR 1 is a mitogenic protein, also known as teratocarcinoma-derived growth factor 1 (TDGF 1). CR 1 binds with growth factor receptors such as 78 kD glucose-regulated protein (GRP 78) or heat shock 70 kD protein 5 (HSPA5) and mediates several signal transduction pathway that includes nodal-dependent (Smad2/3) and nodal-independent (Src/p44/42/Akt) pathways. CR 1 functions as a chaperone protein by inducing cellular signaling via Wnt/β catenin and Notch/Cbf 1 signal transduction pathways. In cancer cells, activin is inhibited by the complex formed between CR1, activin, and activin receptor type II (ActRII). Alantolactone (Inula helenium) has been reported to exhibit antitumor activity via by inhibiting the activin signaling pathway (Wang et al. 2012; Klauzinska et al. 2015).

4.2.15 Hypoxia Signaling Pathway

Hypoxia plays a significant role in the number of pathological conditions including aging, cancer, diabetes, and ischemia/stroke. Hypoxia-inducible factor (HIF) is the main transcriptional factor in nutrient stress signaling and otherwise known as oxygen-sensitive transcription factor. HIF also plays a major role in autophagy, cell invasion, intracellular pH regulation, and metabolism. HIF also regulates the expression of two key angiogenic factors, namely, VEGF-A and angiopoietin 2 (Ang 2). HIF activity is predominately regulated through posttranslational modifications and stabilization of HIF 1α and 2α proteins. HIF-α protein activity is regulated by two key oxygen sensing/depending enzymes, namely, hypoxia-inducible factor prolyl hydroxylase 2 (HIF-PH2 or PHD) and factor inhibiting HIF (FIH or asparaginyl hydroxylase). Stabilization of HIF 1α activates transcription of genes that involved in angiogenesis, dedifferentiation, and invasion which are all important factors/pathways in carcinogenesis. Thus HIF pathway components serve as potential therapeutic targets against cancers (Pouysségur et al. 2006; Brocato et al. 2014; Elks et al. 2015). Apart from the important mechanism listed above, there are few other mechanisms not discussed. For example, plants/plant-based substances are known to have multitarget activity against multidrug resistance (MDR)-related proteins, thus providing solution to overcome drug resistance problem (Eid et al. 2015).

4.3 Plants Having Anticancer Activity

Whole/crude extracts from plants (more than 120,000 plant extracts belongs to 6000 genera) have been widely studied in the past few decades, and several plants found to have anticancer/antitumor activity. Some of these plants have been clinically proven as anticancer agents, while few others have been used as tools to elucidate the biochemical/molecular mechanisms involved in the growth and regulation of cancers/tumors. Plants are known to exhibit a wide range/spectrum of mechanisms of action such as blockers of cell cycle progression, antagonists of mitogenic signaling, anti-metastasis, upregulators of the immune system, and inhibitors of blood vessel formation (Mans et al. 2000). Among different plant families reported, the 11 predominant plant families such as Fabaceae, Asteraceae, Zingiberaceae, Euphorbiaceae, Apocynaceae, Meliaceae, Solanaceae, Rutaceae, Moraceae, Liliaceae, and Myrtaceae have been shown to exhibit anticancer activity (Table 4.1).

Table 4.1 List of medicinal plants with anticancer potential
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4.4 Phytoconstituents Having Anticancer Activity

Phytoconstituents are potent in the treating several cancers/tumors. Alkaloids, flavonoids, polyphenols, and terpenes are the few chemical classes that have been reported for anticancer/antitumor activity. According to Batra and Sharma (2013), flavonoids have modulated several cellular events in cancer cells such as apoptosis, cell differentiation, cell proliferation and vascularization, etc. Vinblastine, honokiol, magnolol, wedelolactone, oridonin, alantolactone, and costunolide are the few classical examples for phytoconstituents having anticancer activity (Millimouno et al. 2014). Apart from phytoconstituents, some marine-based substances also known to have anticancer activity include aragusterol A, ascididemin, bryostatin1, discodermolide, fascaplysin, indanone, jaspamide, lyngbyabellin A, melophlins A and B, salinosporamide A, and spisulosine (Mayer and Gustafson 2003). In the present chapter, Table 4.2 summarizes those 165 phytoconstituents shown to have anticancer activity by modulating several cellular signaling pathways.

Table 4.2 List of phytochemicals with anticancer potential
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4.5 Conclusions and Future Prospects

Despite much therapeutic advancements in the understanding of carcinogenesis processes, cancer still remains as a major health issue around the global. Plants have been recognized as rich source of anticancer drugs. At present more than 60% of commercially available anticancer agents are directly or indirectly obtained from natural sources including plants. New technologies in isolating bioactive compounds and screening for anticancer activities (high throughput screening) have been developed and studied for natural products. Further, scientific evidences have revealed that anticancer phyto-drugs prevent and destroy cancerous cells through the involvement of various kinds of molecular mechanisms of action. Simultaneously, new challenges are emerging due to safety concern, increased cases of drug resistance, and cost of tumor diagnosis. Apart from these, rapid growing obesity rate and increasing addict to smoking has been recognized as two more challenges/risk factors leading to high cancer incidence. The gold standards for assessing the safety and efficacy of drugs must be followed strictly and uniformly across the globe. In this regard, more number of in vivo studies should be encouraged to access the potential of lead plant molecules in future. Similarly, the placebo-controlled clinical trials must be carried out universally to have statistical significance value. In conclusion, natural product research is fascinating approach for discovering novel bioactive compounds with unique chemical structure and unique mode of action against different cancer types.