Role of Dietary and Other Phytochemicals in Cancer Prevention, Growth and Metastasis

1. Introduction:

The world is full of fatal diseases and the most burdening one seems to be Cancer [1]. Cancer refers to abnormal cell growth resulting from multiple physiological changes in cell [2]. It is a global menace causing increase in mortality and morbidity worldwide. It is estimated that number of cancer cases will become 21 million by the year 2030 [3]. Cancer development takes place due to both external factors such as radiations, smoking, tobacco, pollutants in drinking water, food, air, chemicals, certain metals, infectious agents etc. and lifestyle and and internal factors such as genetic mutations, body immune system, hormonal disorders etc. depending on lifestyle and environment [3, 4].

The studies focused on ‘cancer prevention’ are directed with the aim to reduce the number of new cancer cases and deaths related to cancer. Primarily, the focus is to maintain a healthy lifestyle, avoid carcinogen exposure, and consume chemopreventive agents and drugs through diet so that cancer onset can be completely prevented or delayed. Secondarily, the aim is to help in the better management and treatment of tumors through early
detection of cancer in the precancerous or early stage tumors. The tertiary aim is to reduce the risk of metastases, development of secondary tumors and recurrence with the use of preventive agents. All these three aims can be fulfilled by natural
products (from botanicals, herbs, etc.) as well as minerals and vitamin. Phytochemicals are the class of compounds among natural products which have been extensively studied for their biological effects. [5]

The reversal, suppression or prevention of cancer development by use of natural or synthetic chemicals is known as cancer chemoprevention [6]. Phytochemicals are considered nature’s gifts for cancer control and management because of their inexpensiveness, readily applicability, acceptability and accessibility to inhibit the progression and development of cancer [3, 4]. In fact, 69% of anticancer drugs approved from 1980s to 2002 are natural products or developed based on their knowledge and three-quarters of plant-derived drugs clinically used today are originated from traditional phytomedicines. [7]

Approximately 250 000 plant species are present in our plant kingdom among which studies have been done on only 10% for treatment of different diseases. Different parts of a plant, e.g., flower, flower stigmas, pericarp, sprouts, fruits, seeds, roots, rhizomes, stem,
leaf, embryo, bark contain phytochemicals and their derived analogues which perform several pharmacological functions. Alkaloids, flavonoids, lignans, saponins, terpenes, taxanes, vitamins, minerals, glycosides, gums, oils, biomolecules, primary and secondary metabolites etc, plant products can inhibit cancer cell by targeting cellular molecules and molecular pathways such as reactive oxygen species, inflammation, cell cycle, apoptosis, invasion, angiogenesis, transcription factors, and protein kinases. [2, 3]

2. Biologically active dietary and other phytochemicals

1. Glycosides-

Cardiac– Bufalin; Xanthone– Gambogic acid; Quinone– Anthraquinone (Emodin), Napthaquinone (β-Lapachone, Plumbagin), Benzoquinone (Thymoquinone)

Sources: Apples, grapes, plums, blackberry, raspberry, red berries, cranberry, red onion and red cabbage. [3]

2. Phenols:

Phenolic acid– Hydroxybenzoic acid (Ellagic acid), Hydroxycinnamic acid (Ferulic acid), Flavonoids– Flavones (Apigenin, Luteolin, Nobiletin, Chrysin), Catechin flavones (Epigalletocatechin-3-gallate), Flavonol (Kaempferol, Morin, Quercetin), Flavonone (Hesperitin, Naringenin), Isoflavone (Genistein, Puerarin, Daidzen), Flaronolignan (Silibinin); Stillbenes– Resveratrol, Combrastatin-A

Sources: Green tea, red wine, cocoa, berries, cranberries, pomegranate, papaya, and others. [8, 9]

3. Resin– Curcumin

Source: Turmeric (Curcuma longa) [8]

4. Lignan-  Podophyllotoxin, Honokiol

Sources:Cereals and grain products, vegetables, fruit, berries, beverages. [10]

5. Alkaloids-

Quinoline– Camptothecin; Isoquinoline- Berbarine, Noscapine; Benzisoquinoline- Tetrandine; Piperidine– Piperine; Indole – Vincristine, Vinblastine

Sources: Chili peppers, black peppers, long peppers,  cruciferous vegetables, fenugreek. [9]

6. Terpenoid-

Diterpenoid– Triptolide (Paclitaxel, Docitaxel); Triterpenoid– Cucurbitacin B, Betulinic acid, Oleanolic acid, Ursolic acid; Sesquiterpenoid– Artemisinin, Zerumbone, Famesiferol C

Sources: Sweet fennel, Turmeric, Ginseng, Citrus fruits, Cherries, grapes, mango, strawberry, Rosemary, basil [11]

7. Indoles and glucosinolates-

Sulforaphane [12]

Source: Vegetables such as broccoli, cabbage, cauliflower, mustard greens, Brussels sprouts, and kale [8]

8. Carotenoids-  Carotene, Lycopene, Fucoxanthin

Sources– Carrots (alpha- and beta-carotenes) and tomatoes (lycopene)[8, 12]

3. Bioavailability of phytochemicals

 Phytochemicals face some common barriers prior to reaching their target organ or tissue. Non-specific distribution, Intestinal mucosa barrier,quick metabolism, indigestibility, phytochemicals and transport molecule interaction are included in them. This results in limited potential of many phytochemicals in pre-clinical or clinical settings despite their therapeutic potential both in in-vitro and animal models. [13] The problem of bioavailability is the most fundamental challenge in drug designing. During initial phase of drug discovery, the bioavailability can be determined by kinetic simulation using animal models. [13]

The process of bioavailability has different phases such as liberation, absorption, distribution, metabolism and elimination. [13] In case of oral administration of some phytochemicals, the process of absorption and distribution in body is hampered by Phase I and Phase II metabolism.  Phase I metabolic enzyme cytochrome P450 increases drug polarity and the ability of phytochemicals to impact CYP enzymes affects its metabolism [14]. Many Phytochemicals such as Curcumin, Quercetin etc. have low oral bioavailability due to rapid metabolism and require higher doses. [15]. Again low solubility of Ginsenoside lowers its oral bioavailability [16].

Effective ways of improving bioavailability include:

(i) Increasing the efficacy and bioavailability of phytochemicals by chemical syntheses of novel analogues,

(ii) Delivery of phytochemicals to their intended target organs by selective and effective novel formulation designing and

(iii) Pharmacokinetic modulation with novel delivery system formulations [13]

Enhancement of bioavailability can be done with Nanotechnology, liposomes, micelles, various coating materials, and phospholipid complexes.

The following are some of the ways bioavailability of phytochemicals can be enhanced:

1. Increasing water solubility of Curcumin can enhance its bioavailability [14]. 

2. A synthetic analogue (CDF-diflourinated-curcumin) is effective in increasing bioavailability of Curcumin. [17].

3. Bioavailibility of Flavonoids can be increased by converting glycoside to glucosides. [18].

4. 5% bioavailability of Berberine can be increased by changing its structure or using an absorption enhancer. [19].

5. Use of hesperetin-7-glucoside instead of free hesperetin results in increased bioavailability. [13]

6. Complexation of genistein with high-amylose corn starch increases bioavailability of Genistein. [14]

4. Preventive and therapeutic Mechanisms of Phytochemicals

Anti-oxidant activity:

Oxidative stress causes an imbalance in normal cell metabolism by-products Reactive oxygen species (ROS) and Reactive Nitrogen species (RNS). Hydrogen peroxide (H2O2), superoxide radical (%O²⁻), hydroxyl radical (%OH^–), peroxyl radical (ROO.) are the main culprits of oxidative stress termed as ROS. This can induce Cancer through direct DNA damage, alteration of protein conformation and function. Phytochemicals can counteract the damaging effects of oxidation by a direct quenching of ROS through upregulation of gene expression which cause metabolism of toxic compounds, cellular homeostasis and detoxification of reactive species. Ex- Phytochemicals such as caffeic acid, chlorogenic acid, curcumin, resveratrol and ursolic acid inhibit/delay the progression of known tumor promoter TPA-induced carcinogenesis is indicative of their antioxidant effects as TPA is a known tumor promoter which generates potent superoxide anions.

Based on the biological mode of action, Anti-oxidants are of following types:

a)      Anti-oxidants which boost the biosynthesis of endogenous antioxidants by hindering the release or formation of ROS

b)      Anti-oxidants which attack and inactivate previously generated ROS

c)      Anti-oxidants which remove and check the accumulation of bio-molecules damaged by oxidation

d)      Anti-oxidants which cause immunological modulation of ROS generating signaling pathways [5, 13]

Induction of Apoptosis:

The process of programmed cell death such as blebbing, cell shrinkage, and nuclear fragmentation through multiple signal transduction pathways is known as apoptosis. Apoptosis is induced by phytochemicals when apoptotic effectors such as mitochondrial derived activator of caspases (SMACs) are leaked from the mitochondria due to mitochondrial swelling and membrane permeability increase by apoptotic proteins. These effectors prevent apoptosis arrest by binding to inhibitor of apoptosis proteins (IAPs) and preventing caspase suppression. [14]

Mitochondrial apoptosis-induced channel (MAC) is formed in the outer mitochondrial membrane. It facilitates release of cytochrome C and it’s binding with apoptotic protease activating factor-1 (Apaf-1) and ATP, which then binds to pro-caspase-9. [14] This leads to release of active form of caspase-9 creating a protein complex apoptosome and cleaving pro-caspase. Active caspase-9 causes activation of effector caspase-3. MAC and Mitochondrial Outer Membrane Permeabilization Pore (MOMPP) is regulated by Bcl-2 family proteins. Pro-apoptotic Bax and/or Bak cause and anti-apoptotic Bcl-2, Bcl-xL or Mcl-1 inhibit the pore formation. [14]

Another mediator of phytochemical induced apoptosis is a cytokine Tumor Necrosis Factor (TNF) which is produced by activated macrophages. Binding to its receptor, TNF initiates survival and inflammatory responses.  Fas ligand (FasL), a transmembrane protein of the TNF family and Fas receptor (Apo-1 or CD95) is responsible for death-inducing signaling complex (DISC) formation. Fas-associated death domain protein (FADD), caspase-8, and caspase-10 are contained in DISC. [14]

Modulation of DNA damage/repair:

Some phytochemicals can act as chemopreventive agents by triggering DNA repair and damage mechanisms in cancer cells.  Cell death by apoptosis can be caused by phytochemical induced sustained DNA damage response if repair is inadequate. Curcumin, Green tea catechin etc. can induce DNA damage and inhibit its repair in cancer cells. [13]

Anti-inflammatory activity:

Inflammation is closely related to onset and progression of cancer which occurs through a number of signaling pathways. The most studied one is nuclear factor kappa-light-chain-enhancer of activated B cells (NF-_B) pathway which is affected by many phytochemicals like curcumin, resveratrol, EGCG, plumbagin and honokiol. Other mechanisms of anti-inflammatory activity of phytochemicals are growth factor beta1 (TGF-b1) transformation, up-regulation and down-regulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX2), modulation of toll-like receptors (TLR)/interleukin-1 receptor (IL-1R) and Keap1/Nrf2 pathway. Some inflammatory mediators are  interleukins and cytokines (i.e., IL-1, IL-6 and TNF-α) [5, 13]

Inhibition of cell proliferation and cell cycle arrest:

Many phytochemicals can cause arrest of cell cycle at different cell cycle checkpoints which withdraw cell cycle progression such as at G1, G2M and probably during S phase. p53 is a vital component of G1 checkpoint is functioning of which is linked to cellular fates due to intrinsic and extrinsic stress. It is also responsible for initiation of repairing pathways, senescence or cell cycle arrest and normal cell propagation and homeostasis. [13] The mutated form of p53 under influence of oncogenic drivers e.g. PI3K/Akt/ mammalian target of rapamycin (mTOR) and (Ras/Raf/MEK/ERK) pathways, c-Myc (oncogene) and CDKs, cause tumorigenesis. By modulating p53, the phytochemicals Andrographolide, Curcumin, Ginsenoside etc. can inhibit abnormal proliferation of cancer cells and cause cell arrest. [13]

Tumor angiogenesis inhibition:

Many phytochemicals act as anti-cancer agents by inhibiting angiogenesis. The fundamental step in the transition of tumors from a dormant to a malignant state i.e. tumor spreading is angiogenesis, the process of the growth of new blood vessels from pre-existing vessels. It is induced by secretion of various growth factors, such as vascular endothelial growth factor (VEGF). [14] VEGF stimulate vessel permeability by producing NO, proliferation/survival, migration and differentiation into mature blood vessels via binding to VEGF receptor-2 (VEGFR-2) and initiating tyrosine kinase signaling cascade. Phytochemicals such as Andrographolide, EGCG, Di-indollylmethane, Genistein etc. are used because of their ability to inhibit or reduce the creation of new blood vessels and help to combat tumor. [14]

Prooxidant Activity:

The ability of some phytochemicals to induce apoptosis in cancer cells by ROS production in the presence of transition metal ions, especially copper is known as their prooxidant activity. These phytochemicals generate ROS in the presence of redox active transition ions through redox recycling of the ions due to the sensitivity of cancer cells with endogenously elevated copper, iron or zinc ion levels to electron transfer. Curcumin, EGCG, plumbagin, resveratrol, vitamin A and vitamin C possess prooxidant activity [5].

Tumor Metabolism Modulation:

The regulation of metabolites glucose and glutamine required for the growth and survival of cancer cells by certain phytochemicals can lead to altered tumor cell metabolism and eventual death. By direct binding and inhibition of glucose transporter 1 (GLUT1), the phytochemicals cause hamper basal transport of glucose in cancer cells and alter glutathione and lipid metabolism. Curcumin, Resveratrol and plumbagin are known to modulate tumor metabolism. [5]

 

5. Phytochemicals on modulating cancer promoting cell signalling pathways:

1.                MAPK signalling pathways:

The MAPK/ERK pathway (Mitogen-activated protein kinase/ Extracellular signal regulated kinase) is a downstream effector for ER (estrogen receptor), EGFR (epidermal growth factor receptor), and TNFα (tumor necrosis factor alpha) etc. membrane receptors. Being an essential route in cell survival and growth regulation, this pathway is a popular target for pro-apoptosis, anti-invasion, and anti-proliferation by photochemicals. [9]

It is also known as Ras-Raf-MEK (mitogen-activated protein kinase kinase)-ERK transduction cascade. Activator Ras and Raf-MEKERK comprises of a MAPK kinase kinase (MAPKKK, for instance Raf), a MAPK kinase (MAPKK, for instance MEK), and a MAPK (ERKs, JNKs i.e. c-Jun amino-terminal kinases). It causes cell proliferation by ERK1/2 phophorylation by RAS stimulation and Raf-1 pro kinase activation functioning as both as tumor suppressor or pro-oncogenic signal separately [9]

2.                PI3K/AKT signalling pathway

PI3K/Akt (PI3K for phosphoinositide 3-kinase, Akt for protein kinase B) pathway regulates cell growth, proliferation, differentiation, survival and intracellular trafficking by phosphorylating the 3 position hydroxyl group of the inositol ring of phosphatidylinositol (Ptdlns). It activates receptor tyrosine kinases (RTKs) and causes Ptdlns (3, 4, 5)P3 and Ptdlns (3,4)P2 production by PI3Ks (a lipid kinase family) at the inner side of the plasma membrane. [9] Akt induces these phospholipids to phosphorylate and translocate to the inner membrane after activation by PDK (phosphoinositide-dependent kinase) 1 and 2. Complete activation of Akt takes place upon serine phosphorylation by the TORC2 complex of the mTOR protein kinase. In Cancer, mutation of PI3k causes its activation in absence of its antagonist PTEN. [9, 14].

3.                 NF-κB/IkB signalling pathway

Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) is a primary transcription factor family which initiates inflammatory and innate immune responses. It is induced by Reactive oxygen species (ROS), TNF alpha, IL-1 beta, lipopolysaccharide (LPS) etc. NF-κB1 (p50), NF-κB2(p52), RELA(p65), RELB, c-REL are the 5 proteins in this family. In presence of harmful stimuli, IκB kinases (IKK) modify and degrade IκBs by phosphorylation. This releases NF-kB into the nucleus and induce the expression of specific genes that have DNA-Binding sites for NF-kB (such as cyclin D1 and apoptosis suppressor proteins Bcl-2 and Bcl-XL). IkB alpha acts as NF-kB repressor to form an auto feedback loop. NF-kB is activated in cancer cells so phytochemicals inhibiting NF-kB are used as anti-proliferating agents. [14, 20]

 

4.                Nrf2/Keap1 signalling Pathway

Nuclear factor (erythroid-derived 2)-like 2 (Nrf2, or NFE2L2) is a basic leucine zipper (bZIP) transcription factor regulating antioxidant responses. Normally, Nrf2 remains attached with Kelch like-ECH-associated protein 1 (Keap1) in the cytoplasm. During stress, removal of cysteine residue causes Keap1 to release Nrf2 into the nucleus where it heterodimerizing with small Maf proteins, binds to the anti-oxidant response element (ARE). [14] This results in the transcription of many antioxidative genes into cytoprotective roteins like NAD(P)H-quinone oxidoreductase 1 (NQO1); heme oxygenase-1 (HO-1),  glutathione S-transferase (GST), UDP-glucuronosyltransferase (UGT), or phase III transporters, such as multidrug resistance-associated proteins (MRPs). [14]

 

5.                Wnt/β-Catenin signalling pathway

Wnt proteins are group of secreted lipid modified (palmitoylation) signaling proteins containing 350-400 amino acids in length which carry a conserved pattern of 23-24 cysteine residues on which palmitoylation occurs on a cysteine residue. It is discovered in liver, lung, gastric, ovarian, breast, colon, leukemia, endometrial, brain etc. cancers. [14] The Wnt proteins cause the activation of Dishevelled (DSH) family proteins by binding to cell-surface receptors of the Frizzled family and change amount of β-catenin reaching the nucleus. The DSH complex also inhibit the proteolytic degradation of the β-catenin and stabilize cytoplasmic β-catenin by protein complex such as axin, GSK-3 and APC inhibition. This results in promotion of specific gene expression by interaction with TCF/LEF family transcription factors. Various pathways such as Wnt, β-catenin, cadherin, etc are activated by Wnt proteins for embryonic development, cell differentiation, and cell polarity generation [9, 14].

6.                 JAK/STAT signalling pathway

JAK/STAT (Janus kinase/signal transducer and activator of transcription) pathway regulates expression of genes required for cellular proliferation, growth, and migration through a cascade of signals. Activation of JAK by ligand binding to membrane receptor subunits causes phosphorylation and translocation of STATs to the nucleus. This results in regulation of transcription of apoptosis/survival-related Bcl-2, p53, and survivin, growth-related PCNA (proliferating cell nuclear antigen), cyclin D and myc, and premetastatic MMP, IL-6 (interleukin-6), and CXCL3 (chemokine ligand 3 or C-X-C motif ligand 3) etc. genes. [9, 14, 21]

 

7.                VEGF signalling pathway

Vascular endothelial growh factor (VEGF) receptor regulates the formation of blood and lymphatic vessels which plays a significant role in invasion and migration of solid malignancies.

Tyrosine kinase members VEGFR‑1, VEGFR‑2, and VEGFR‑3 which are also called Flt‑1, KDR/Flk‑1, and Flt‑4 of the VEGFR family regulate angiogenesis in endothelial cells normally and during tumor growth, migration, and chemoresistance in malignant situations. [14] In cancer cells, VEGF-A causes formation of new vessels and promotion of autocrine signal‑mediated tumor cell proliferation by binding to VEGFR‑2 present on endothelial cells surface. Phytochemicals blocking the VEGF/VEGFRs pathway can be used as anti-angiogenic and anti-proliferating agent in Cancer. [14, 22]

 

8.                TGF-β/SMAD signalling pathway

Tumor Growth Factor (TGF-β)/SMADs signaling pathway is one of the regulators of cell cycle arrest. Tumor suppressor TGF-β causes inhibition of cell growth, apoptosis, and differentiation through expression of SMAD proteins. [23]

9.                  p53 signalling pathway

p53 is a tumor suppressor protein which can activate the apoptotic cascade and other anticancer advances by binding to DNA. Cancer causes mutation or disturbed upstream signaling of p53 rendering it dysfunctional. p53 signalling is cross-linked with pathways such as  Akt, JNK and MAPK which is seen in ECG (Epicatechin gallate) . Phytochemicals such as Quercetin, resveratrol, EGCG, and piceatannol can induce expression and phosphorylation of p53 gene leading to cell cycle arrest and apoptotic cancer cell death. [9]

10.                  Hypoxia/ HIF‑1α  signalling pathway

Hypoxia is the state of oxygen and nutrients depravation which solid malignancies face due to less blood supply. Hypoxia‑inducible Factor‑1α (HIF‑1α) regulates angiogenesis, invasion and metastasis, cell survival, oxygen transport, iron metabolism, glycolysis, and glucose transport of tumor cells. Genes important in angiogenesis such as VEGFA, PIGF, VEGFR1, Tie‑2, angiopoietin 1 and 2, MMPs, prolyl‑4‑hydroxylase, uPAR, FGF2, MCP‑1, PDGF, SDF‑1, and CXCR4 etc. are targets of HIF‑1α. [22]

11.                Hedgehog signalling pathway

The hedgehog signaling pathway regulates cell development with hedgehog signaling proteins in a concentration-depended manner. Abnormal activation of this pathway causes cancer stem cell formation. Binding to the Patched-1 (PTCH1) receptor, sonic hedgehog (SHH) activates the downstream protein Smoothened (SMO) leading to the activation of the GLI transcription factors. The GLI accumulated in the nucleus causes the transcription of hedgehog target genes resulting in increase of angiogenic factors, cyclins, anti-apoptotic genes and decrease of apoptotic genes.

[14]

12.                  Autophagy signalling pathway

The process in which cells shut down and degrade unnecessary cellular processes or dysfunctional cell components through lysosome action to adapt in adverse situations. Tumor cells undergo autophagy in poor nutrient and hypoxic conditions by blocking glucose uptake in mitochondria and through mTOR signaling. [9] Cancer cell death can be caused by intervening with ATP (adenosine triphosphate) synthesis or the mitochondrial oxidative phosphorylation chain. Chemoprevention can be done by both promoting and inhibiting autophagy as autophagy promotion leads to limitation of genome damage and cancer progression whereas autophagy inhibition promotes apoptosis in cancer cells. [9]

 

13.             Other cell signalling pathways

Plk1 expression: The proto-oncogene enzymePolo-like kinase 1 (Plk1) overexpression causes progression of cell cycle in tumor cells such as lung cancer. It is also related to tumor suppressor p53 related pathways. [14]

Poly-ADP-ribosylation: Poly(ADP-ribosylation) is a post-translational process in which β-NAD(+) is converted into ADP-ribose via polymer synthesis by pro-survival factor poly(ADP-ribose) polymerase (PARP) enzyme.  It is significant in DNA replication, repair, and transcription processes. Inhibition of PARP can be chemopreventive and poly(ADP-ribosylation) inactivation can limit injury to cell. [14]

Human epidermal growth factor receptor (HER2) pathway- This pathway isactivated in breast cancer [14].

 

6.                Phytochemicals on epigenetic modulation:

Epigenetic modulation in cancer cells cause erratic expression of genes which affects oncogene and tumor suppressor gene [24]. Such epigenetic modification effects include inhibition of tumor suppressor, apoptosis-inducing and DNA repair genes, blocking of signal transducers and transcription factors by methylation of promoter, inhibition of translation and targeted degradation of mRNA, histone and non-histone protein (p53, NF-kB, and the chaperone HSP90 ) modifications by acetylation or methylation etc. [25]

Epigenetic modifications are reversible and the affected genes remain intact. Phytochemicals acting as potential reactivators of the affected genes are able to prevent or reverse gene expression inactivation and can be used against cancer. [25]

Gene expression gets modified by following mechanisms:

1. DNA methylation 2. Histone modification 3. MicroRNa changes [26]

1.                DNA methylation

Expression of tissue specific genes is regulated by specific DNA methylation mediated by DNA methyltransferases (DNMT) which transfer methyl groups from S-adenosyl-L-methionine (SAM) to the 5-position of cytosines.  During carcinogenesis, DNA hypermethylation causes silencing of Tumor suppressors and other biologically important genes functions [25].

2.                Histone modification

[27, 28]A globular core containing two molecules of H2A, H2B, H3, and H4 each (histone octomer) with approximately 146 DNA base pairs form a nucleosome [29]. Acetylation and deacetylation of chromatin histone tail controls expression of genes [30]. Genes get acetylated for the transcription factors to access the DNA and deacetylated for condensation of chromatin and transcriptional repression [29]. Epigenetic modifications in chromatin and histone results in pro-apoptotic, anti-apoptic and tumor suppressor gene/protein upregulation, control of Wnt/β-catenin, soni hedgehog, DKK-1, Notch, CDK6 pathways, self-renewal activity induction and arrest of cell cycle [24]. The activity and expression of enzymes Histone Acetyl Transferases (HAT) and Histone deacetylases (HDAC) regulate these reactions and inhibition of these enzymes can cause cell cycle arrest and eventual apoptosis in cancer cells by reactivation of epigenetically silenced genes [24, 30]. Phytochemicals which can inhibit HDACs could overcome cancer invasion, relapse and drug resistance. [24]

3.                 Oncogenic and tumour suppressor miRNAs

MicroRNAs (miRNAs) are non-coding post-transcriptional gene expression inhibiting RNAs consisting of 20–22 nucleotides[25]. They control many important physiological processes such as cell differentiation, apoptosis, proliferation, and metabolism etc. By binding to the 3’-untranslated regions (3′-UTR) of the target mRNAs, miRNAs induce repression of protein synthesis during translation and degradation of mRNA. Epigenetic modulations during carcinogenesis can cause downregulation of miRNA leading to initiation and progression of cancer [25, 29]

Fig: Common Cancer-Associated miRNAs

Phytochemical	Effect on miRNA
Apigenin	miR-138 ↑[29]
Boswellic acid	let-7b and -7i expression ↑ miR-220b and miR-220c expression ↑ two tumor suppressor genes,  [31]
Diallyl disulphide	miR-17↓[29] p21 mRNA ↑[28]
Diindolylmethane	miR-200b, miR-200c, let-7b, let-7e ↑ [32]
Folate	miR-222 ↑[32]
Retinoic acid (Vitamin A)	miR-15a, miR-15b, miR-16–1, let-7a-3, let-7c, let-7d, miR-223,miR-342, miR-107, miR-10a, miR-10b ↑, miR-181b ↓, tumor suppressor miRNAs ↑  [17, 32]
Vitamin D	miR-182 expression alteration, p53 and PCNA levels modulation [17]
Vitamin E	miR-122 and miR-125b Folate expression alteration [17]
Vitamin B (Folate)	potential chemoprevention by affecting miRNAs [17]
Sodium Selenite	p53 activation and targeting of  miR-34 family ↑  [17]
n-3-Polyunsaturated fatty acids (n-3 PUFAs)	5 tumor suppressor miRNA ↓   [17]
Curcumin	11 miRNAs ↑, 18 miRNAs ↓, MiR-22 ↑, oncogenic miR-196 ↓, tumor suppressor PTEN induction by miR-200 and miR-21 expression attenuation [17], miR-9, let-7a↑[27], miR-21, miR-34a↑[29], loss of let-7 family and MiRNA-143 expression, along with increased expression of MiRNA-21, MiRNA-143 and decreased expression of MiRNA-21
Oleanic acid	miR-122 expresssion ↑  [31]
Oleic acid	miR-21 ↑ [32]
Ursolic acid	miR-21 expression ↓  [31]
Genistein	miR-1260, miR-27a and miR-155↓[29] miR-146a ↑, expression of miR-1296 ↑ up to fivefold [17], miR-221 ↓,  miR-222 ↓[32]
Ginsenoside	44- miRNAs ↑ including let-7 and miR196, 24 miRNAs ↓ including miR-193, miR-15b expression ↓[31]
Epigallocatechingallate	13 miRNAs ↑ including  tumor suppressor miR-16 [17], miR-210↑, miR-25, miR-92, miR-141 and miR-200↓[29]
Ellagitannin	17 miRNAs ↑ and 8 miRNAs ↓ including let-7 family members, miR-370, miR-373, and miR-526b [17]
Indole-3-carbinol	miR-21(I3C)↓, miR-21(DIM), let-7↑[29]
isoflavone	miR-200, let-7 family miRNAs ↑ [17, 32]
PEITC	miR-192 ↑ (Ras activation), let-7a, let-7c ↑ (cell proliferation, angiogenesis, Ras activation), miR-146 ↑ (NFkB activation), miR-123, miR-222 ↑ (angiogenesis, cell proliferation) miR-99b ↑ (apoptosis) [17]
Quercetin	miR-142-3p↑[29]
Resveratrol	miR-328↑, miR-200c,miR-129, miR-204, and miR-489↑ miR-16-1 and miR-15a↑[29] miR-150 ↑ , miR-296–5p ↑, miR-7 ↓, miR-17 ↓, miR-20a ↓, miR-18b ↓, miR-20b ↓, miR-92b ↓, miR-106a ↓, miR106b ↓ [32] miR-155  ↓, miR-663 ↑[27]
SCFA Butyrate	miR-17 ↓, miR-20a ↓, miR-20b ↓, miR-93 ↓, miR-106a ↓, miR-106b ↓ [32]
Sulforaphane	miR-200c↑[29]
Terpene	let-7, miR-200, miR-34a↑, miR-27a↓[29]
Tanshinone I	miR135a-3p↑[29]
Tanshinone IIA	miR-155↓[29]

7.                Phytochemical-loaded nanomedicines for cancer treatment

A great number of Phytochemicals were found to be effective in treatment and prevention of different types of cancer. But their uses in anti-cancer therapy were disturbed as they lacked target specificity, had low bioavailability and were poorly soluble in aqueous medium. Many were highly unstable with short circulation time and low circulation concentrations due to degradation and metabolism by enzymes in the gastrointestinal tract, the liver and other tissues [33]. Biocompatible and biodegradable nanoparticles such as Liposomes, Nanostructured lipid carrier, Lipid micelles, Polymeric micelle, Dendrimer, Polymeric Nanoparticles, Metal nanoparticles and Nanoemulsions are used as nano drug delivery systems 1. [34, 35]

8.                Phytochemicals as potential carcinogens:

The use of phytochemicals as chemoprotective agents is widely discussed but their potential toxic, genotoxic and carcinogenic effects are less known. There are quite a few phytochemicals which apart from having anti-cancer activity, can act as carcinogens [13]. Some of them are:

Capsaicin-   Promotes neuronal desensitization and  skin carcinogenesis mediated by TRPV1, EGFR and COX2 especially in presence of tumor promoter. [13, 36]

Cycasin and metabolite methylazoxymethanol (MAM) – Induces neurodegeneration and cancer development by DNA damage and changes in gene expression.

Dietary phytoestrogens-  increases colorectal cancer risk and promote tumor growth by negating the effectiveness of aromatase inhibitors

Phorbol esters: Phorbol 12-myristate 13-acetate (also known as TPA) promotes tumor in skin

Pyrrolizidine alkaloids cause liver cancer by binding to cellular protein and DNA and skin cancer by lipid peroxidation through ROS generation when metabolized to dehyro-PA. [36]

Ptaquiloside (Bracken fern) : causes tumor development when activated to a dienone.

Safrole– Act as liver carcinogen and increases oral cancer, esophageal cancer and hepatocellular carcinoma in rats [13, 36].

Methyleugenol, Aristolochic acid I and II, coumarin, lasiocarpine, lucidin, reserpine and symphytine: reported to be a genotoxic and carcinogenic compound

Thus, it is evident that phytochemicals may be toxic in certain doses, and so, t is essential to evaluate the safety of phytochemicals in terms of exposure time, dosing amount and drug delivery system etc. prior to use as chemotherapeutic agents [13].

 

9.                Conclusion and future direction

As Cancer has spread out throughout the world, it is now a necessity to develop alternative, co-friendly, biocompatible and cost-effective anticancer drugs to overcome the high cost and many limitations of conventional therapy. In that purpose, phytochemicals in anti-cancer therapy is a promising and effective research area with scope for future development [3]. Dietary and other phytochemicals are able to regulate and modify metabolic programming and cell homeostasis for control of disease etiology along with human health improvement. Several studies have established the chemopreventive effects of various phytochemicals which are included in this article. After absorption at cellular level, phytochemicals activate different metabolic processes interacting with specific signaling pathways and influencegene and protein expression as endogenous cellular mediators [37]. In order to further develop phytochemicals as anticancer agents, clinical trials on different models are required for their safety, efficacy and toxicological evaluation in order to determine the optimum dosage regimen [3]. Also, further investigation is required to understand the casuality between phytochemicals and cell signaling transduction targets [9]

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