Targets and Mechanism of Action of Chemical constituents from Plants with Potential Anti-leukemic Activity

The plant kingdom has been the most signi icant source of anticancer drugs. These include alkaloids, diterpenes, tannins, phenolics, lignans, glycosides which have exhibited lesser toxicity than conventional drugs. In leukaemia, the human body is susceptible to infections due to the replacement of normal leucocytes replaced by a large number of immature cells. Chemotherapy for leukaemia is associated with side effects and drug resistance by the leukemic cells. Cytotoxic agents with higher ef icacy and lesser side effects are good candidates in cancer therapy, and plant metabolites serve as potential bioactive agents in anticancer drug formulations. This review article discusses the anti-leukemic properties of compounds obtained from plants and the mechanism of anti-leukemic activity induced by each of these plants. Effect of plants and their metabolites on different leukemic cell lines such as HL60, Kasumi-1, CCRF-CEM, K-562, U-937, THP-1 and MOLT-3 was compared. The indings showed anti-leukemic activity through cell cycle arrest, DNA damage, destruction of mitochondrial function, suppression of tumour genes, apoptosis-inducing enzymes and cytotoxic activities of plants and their derivatives. Based on extensive research indings from this review, phytochemicals and their derived analogues possess the most promising option for the better and less toxic anti-leukemic treatment. Identi ication of themechanism of action by the plant bioactive compounds helps in developing standard herbal medicines for effective leukaemia treatment.


INTRODUCTION
In leukaemia, normal white blood cells (WBCs) are replaced by a large number of immature cells, thus leaving the body highly susceptible to infections (Bennett et al., 1976). Based on cell lin-eage and evolution of the disease, leukaemia is classi ied into lymphoid or myeloid and acute or chronic, respectively (Javed et al., 2012). The most effective method of leukaemia treatment involves chemotherapy, but the side effects associated with chemotherapy and drug resistance of leukemic cells makes it unhealthy treatment method.
Further, most leukemic children and adults die despite the signi icances in chemotherapy. Signi icant attention is paid to cytotoxic agents due to the signi icant health and economic impact of leukaemia. Scienti ic investigations in the direction of modifying treatment options or treatment modalities are being carried out to avoid the side effects of cytotoxic agents which are currently used in the treatment of leukaemia (Shaked et al., 2005). Cancer preventive agents with higher ef icacy as well as reduced side effects are need of the hour, which could be used as cytotoxic agents in cancer therapy.
Identifying new agents for treatment leukaemia lead to intensi ied efforts from the scienti ic community globally. Among the novel agents, natural products from plant origin have been used as alternative drugs for malignant tumours due to their wide range of biological activities and lack of toxicity in animal models. Anticancer agents from natural origin with mild to moderate side effects makes them as good candidates during drug identi ication. Phytochemicals, due to their complex nature and interaction with cancer cell receptors, in luences various biochemical and molecular cascades thus serve as potential bioactive agents in anticancer drug formulations. Hence treatment of cancer involving using bioactive compounds from plants has attracted considerable attention worldwide, and plant-derived drugs against leukemic cell types were reported earlier by Aboul-Soud et al. (2016).
Plants due to their bioactive compounds play an essential role in drug development, thus hold key promises for cancer prevention and treatment as explored by many scientists throughout the world. Both in vitro and in vivo anticancer activities of several plants were reported by many researchers using various cell lines and animal models. Based on the anticancer properties of plants, this review focuses on various plants that exhibited potential anti-leukemic activity.

SOURCES AND METHODOLOGY
Detailed search using the keywords such as antileukemic potential, cytotoxicity and medicinal plants were done in PubMed, Scopus, ScienceDirect, Web of Science and Google Scholar to retrieve the published information between 1976 and 2019. The inclusion was based on anti-leukemic activities of plants and their parts along with the mechanism of action. The obtained data were extracted in the form of Table 1, and further explained under subheadings.

Cell Cycle Arrest
Wattakaka Volubilis leaves were investigated for the in vitro anti-leukemic activities using U-937, HL-60 and K-562 cell-lines (Nandi et al., 2012). The experiments reported the IC 50 values of 13.5, 10.8, and 13.2(µg/ml) in U-937, K-562, and HL-60 cell lines by kaempferol-3-O-[α-l-rhamnopyranosyl-(1→4)-O-α-l-rhamnopyranosyl-(1→6)-O]-βd-glucopyranoside present in the leaf extract. Inhibition of cell proliferation was associated with cell cycle arrest at G1 phase in U-937 and K-562, whereas it was G2/M phase in HL60 cell lines. Caxito et al. (2015) reported the inhibitory effect of Xanthosoma sagittifolium on leukaemia cell proliferation by decreasing the cell number at the G2/M phase. Further, DNA fragmentation led to the higher leukaemia cell numbers in the sub-G1 phase in the presence of leaf extract.
Induction of apoptosis by Bidens pilosa in HTLV-1-infected T-cell lines was associated with a cell cycle arrest in G1 phase (Nakama et al., 2011). Water extracts of B. pilosa inhibited the phosphorylation of inhibitor of nuclear factor κB (IκB) kinase β and α, DNA binding of NF-κB and protein expression reduction during G1/S cell cycle transition. Reactive oxygen species-mediated suppression of NF-κB activity and AP-1-DNA binding suppression was through inhibition of expression of JunB and JunD. Vitek et al. (2017) investigated the biological activities of phytocompounds from leaves and stem bark of Eugenia dysenterica. The presence of quercetin-3-O-(6 ′′ -O-galloyl)β-d-glucopyranoside in E. dysenterica promoted cell cycle arrest and exerted cytotoxic effects on Kasumi-1 and CCRF-CEM cells.
Eurycomanone from Eurycoma longifolia exhibited anti-proliferative and apoptotic potentials in K-562 leukemic cell lines both in vitro and in vivo (Al-Salahi et al., 2014). Does dependent cytotoxic effects were con irmed by observing chromatin fragmentation, uniform condensation and forming clusters against the nuclear periphery. Cell cycle arrest in both G1 and S phases was reported in a dose and time-dependent manner. In a study by Zhamanbayeva et al. (2016), different combinations of Hippophae rhamnoides, Rosa canina, Salvia of icinalis and Origanum vulgare extracts had reduced viable cell counts on HL60 cells. A marked decrease in G1/S ratios indicated that HL-60 cells were more prone to cell cycle changes after treatment with combined plant extracts. Reduction in G0/G1-phase populations and relatively enlarged S-phase populations resulted in anti-proliferative effects in HL-60 cells as induced by the plant extracts.
Rosa cymosa fruit extract prepared from ethanol displayed reactive oxygen species and endoplasmic reticulum stress-mediated apoptosis (Wang et al., 2019). This led to overexpression of PTEN and the dysregulation of PI3K/Akt and Jak/Stat 3 signalling pathways which ultimately resulted in Molt-4 cell death. Besides, suppression of protein expressions involved in signalling pathways by the R. cymosa extra was observed. The anti-leukemic activity of Pseuduvaria rugosa alkaloids in human promyelocytic (HL-60) cells resulted in cell cycle  (Jannet et al., 2017) Glycine max Seeds Genistein Loss of clonogenicity (Raynal et al., 2008).
Aerial parts of Corchorus acutangulus were investigated for their anti-leukemic activity by Mallick et al. (2010). Solvent fractions of the aerial parts were prepared in methanol and butanol with the identi ication of corchorusin-D. A triterpene saponin, corchorusin-D in the solvent fraction was found to release apoptosis-inducing factors from mitochondria, thereby producing mitochondrial dysfunction and cell death. Activation of caspase-3 and PARP cleavage due to translocation of Bax from the cytosol to mitochondria by corchorusin-D resulted in DNA fragmentation in U-937 and HL-60 cell lines.
Jamaican boll moss was examined against various leukemic cell lines, and the highest inhibition with an IC 50 value of 1.83 µM was observed against K562 cell lines (Lowe et al., 2014). In another study, root extracts of Vernonia amygdalina had completely destructed the lymphoblastic leukaemia cells, whereas 70% cytotoxicity was observed in myeloid leukaemia cells (Khalafalla et al., 2009). Chiang et al. (2003) demonstrated the cytotoxic potential of linalool, oleanolic acid, ursolic acid and luteolin against human leukaemia and lymphoma cell lines. Linalool exhibited the strongest activity against U-937, P3HR1 cell lines with IC 50 values of 3.51 and 4.21 µg/ml, respectively. Cell proliferation of P3HR1 and K-562 was inhibited by ursolic acid with IC 50 of 2.5 and 17.8 µg/ml. Afolabi et al. (2017) isolated a tetranorditerpene from Polyalthia longifolia to treat human leukemic-60 cells after methyl esteri ication of the compound. After treatment, changes in the cellular morphology as indicated by cell shrinkage, blebbing and formation of apoptotic bodies were observed. Morphological changes were due to the synergistic relationship between native tetranorditerpene and methyl esteri ication reaction in the process of discovering novel cytotoxic and chemopreventive agents.

Apoptosis
Curcumin was also known to induce apoptosis by an increase of phosphatase and tensin homolog (PTEN) in a dose-dependent manner (Taverna et al., 2015). Further, it also decreased protein kinase B and vascular endothelial growth factor expression and release. Derivatives of curcumin are more active than natural curcumin. For example, 4-(4-Pyridinyl methylene) curcumin, inhibited the heat shock protein 90 (Fan et al., 2018) in K-562 cells by induced apoptosis through a stimulated mitochondrial pathway.
Induced apoptosis and inhibition of cell proliferation in human leukaemia HL-60 cells by a member of Meliaaceae family, Toona Sinensis is reported by Kakumu et al. (2014). Nuclei fragmentation, chromatin condensation and membrane blebbing were due to the activity of loropetalin D isolated from T. Sinensis. Similarly, T. Sinensis extract, along with gallic acid had induced apoptosis through the release of cytochrome c, caspase 3 activation and speci ic proteolytic cleavage of poly (ADP-ribose) polymerase (PARP). Besides, the apoptosis was associated with reduced levels of cell death inhibitor (Bcl-2) and increase of Bax protein (Yang et al., 2006). Kabeel et al. (2018) investigated the effect of Arctium lappa on leukaemia in vivo. The plant contains antagonists for sphingosine-1-phosphate receptor-1 that regulates its expression in dimethyl Benz(a)anthracene treated rats. It was also reported that caspase-mediated apoptotic cell death in MH 60 cells by A. lappa (Matsumoto et al., 2006). Pardede et al. (2016) isolated 4-methoxylanceoletin from methanolic extract of Coreopsis lanceolate and claimed its apoptotic potential in human leukaemia HL-60 cells.

Mitochondrial function destruction
The potent anti-leukemic activity of Zingiber of icinale Roscoe and Nerium oleander L characterized by apoptosis resulted through destruction of mitochondrial function of both K562 and MOLT-4 cells is reported by Bhargava et al. (2015). Cell viability assays revealed the IC50 values up to 11.2 and 27.8 µg/ml for N. oleander and Z. of icinale, respectively.

Kinase activity inhibition
Hyperforin from Hypericum perforatum activates pro-apoptotic proteins, targets matrix metalloproteinase-2 and vascular endothelial growth factor in leukemic cells (Zaher et al., 2012;Billard et al., 2012). This is, in turn, inhibits the kinase activity and resulted in cell death of acute myeloid and chronic lymphoid leukaemia cells. Adfa et al. (2016) studied the utility of Cinnamomum parthenoxylon to treat leukaemia. Hinokinin and cubebin from C. parthenoxylon woods displayed strong cell proliferation inhibition and induced apoptotic morphology in leukemic cell lines.

Tumour suppressor genes
Soybean iso lavone, genistein was effective against leukemic cell lines as reported by Raynal et al. (2008). Anti-proliferative assays revealed MOLT-3 lymphoid cells as the most sensitive cell lines, followed by HL-60 and Raji cell lines. The antileukemic potential of genistein is also associated with reactivation of tumour suppressor genes and its reduced effect on promoter demethylation.

CONCLUSION
This review summarizes the anti-leukemic potential of various plants, their bioactive compounds and mode of action against leukemic cell lines. The collective information presented in this review helps to identify new, effective therapeutics from plant origin to treat leukaemia by understanding the mode of action of each bioactive compound.