Application prospects of plant and fungal compounds in antitumor therapy

INTRODUCTION

Although a wide range of effective antitumor drugs have been developed to date, this field continues to remain relevant due to the search for optimal drug combinations with the least number of side effects. In this connection, plant- and fungus-derived compounds are attracting particular attention [1]. The available data suggest that natural compounds affecting autophagic and apoptotic pathways are effective mediators of cancer therapy with specificity for target cancer cells. The multidirectional antitumor effect of natural compounds in combination with their low toxicity are significant prerequisites for the development of drugs for cancer prevention and cancer treatment [1].

Cancer inhibitors of plant and fungal origin (IPFOs) have slightly different mechanisms of action in antitumor therapy. The plant-derived cancer inhibitors mainly affect the cellular signaling pathways of carcinogenesis, while exhibiting an anti-inflammatory effect. The fungus-derived cancer inhibitors are capable of stimulating the immune response with subsequent tumor recognition or preventing cancer cell division. An important advantage of IPFOs as candidate substances consists in the presence of properties essential for all drug medications, such as gastrointestinal absorption and metabolism action. In addition, IPFOs demonstrate a high chemical diversity necessary to study correlations between activity and structure [2]. Some first plant-derived drugs (paclitaxel, vinblastine, vincristine, topotecan, irinotecan, and teniposide), have been extensively studied, partially modified, and approved by the U.S. Food and Drug Administration (FDA, USA). These medicines are not the subject of this review [3].

In this study, we analyze promising directions in the development of anticancer drugs, review the current data on the mechanisms of IPFO action in the context of a combined approach to cancer treatment.

MATERIALS AND METHODS

In addition to the Google Scholar and PubMed search system, the Naturally Occurring Plant-based Anti-cancerous Compound-Activity-Target Database (NPACT, http://crdd.osdd.net/raghava/npact/) was used. NPACT features about 1980 experimentally confirmed interactions of compounds and targets. The search queries included the following keywords: anticancer therapy; plant and fungal cancer inhibitors; apoptosis; programmed cell death; angiogenesis; autophagy; ferroptosis; cell cycle regulation (in Russian and English). The search depth was 10 years. The inclusion criteria were the relevance and practical significance of the publications, as well as the availability of preclinical and clinical trial data.

RESULTS AND DISCUSSION

The search for compounds with antitumor potential is currently underway. Anticancer IPFOs represent a promising direction in cancer treatment, offering a variety of mechanisms of action. There is a large number of conventional chemotherapy drugs with high effectiveness, which are also of plant origin. This circumstance justifies the relevance of searching for new compounds. Solid tumors exhibit a pronounced ability to both proliferate and induce angiogenesis, driving the need to develop new plant-derived compounds with antiangiogenic properties, along with the investigation of other IPFOs. As a rule, anticancer IPFOs affect several metabolic pathways simultaneously, exerting a combined effect on different targets in the cancer cell and reducing the risk of drug resistance development.

Among the main methods used to identify compounds with a certain activity are gene cloning, DNA and RNA sequencing, studying the compounds effect on the enzymes activity involved in the relevant metabolic pathways, evaluating differential gene expression using microarrays, flow cytometry, the use of various cell cultures, including those of tumor origin, the use of animal models to assess the systemic effect of the compound, as well as its pharmacokinetics and pharmacodynamics, multidimensional statistical analysis to assess the reliability of the results obtained. The largest amount of information on natural anticancer compounds in NPACT concerns those derived from plants [4]. Many anticancer IPFO have been isolated from herbs used in traditional Chinese medicine [5]. However, it should be noted that not only plant-derived, but also fungus-derived, medications have been found to possess anticancer potential. The key classes of inhibitory compounds for plants and fungi are phenolic compounds and terpenoids. At the same time, in plants, it is alkaloids, flavonoids, and coumarins that demonstrate similar anticancer properties, while in fungi, these are polysaccharides, glucans, steroids, cerebrosides, and proteins. The diverse mechanisms of action of such compounds include apoptosis induction, inhibition of angiogenesis and the cell cycle, immunomodulation, reprogramming of cellular signaling pathways involved in carcinogenesis, as well as various antioxidant and anti-inflammatory effects (Table 1).

Induction of programmed cell death

Apoptosis

Apoptosis, i.e., programmed cell death, occupies a special place among various processes associated with cell cycle regulation, correct functioning of the immune system, hormone-dependent atrophy, and embryo development [9]. The ability of chemical compounds to induce apoptosis determines their significant therapeutic potential. The following IPFOs can serve as such examples.

Icaritin induces programmed cell death of ovarian cancer cells by activating the apoptosis pathway through p53 and inhibiting the Akt/mTOR signaling pathway [10]. The curcumin anticancer activity may directly depend on the effect on the p53 pathway in human osteosarcoma (HOS) cells [11]. The triggering of internal and external apoptosis pathways is also responsible for the curcumin anticancer effects in monocytic leukemia (SHI-1) cells [12]. It was shown that matrine is capable of stimulating the main apoptotic cascades by increasing the accumulation of Fas and FasL, Bax proteins, while reducing the amount of Bcl-2 apoptosis regulator, which leads to activation of caspases-3, -8, and -9 in human osteosarcoma MG-63 cells, as well as U-2OS, Saos-2, and MNNG/HOS [6, 13]. The molecular mechanism of action of tetrandrine in cancer cells is aimed at increasing the number of Bax, Bak, Bad, and Apaf-1 apoptotic proteins, while the number of Bcl-2 and Bcl-xl antiapoptotic proteins in the cell decreases with the release of cytochrome c (cyt c) and activation of caspase-3 and caspase-9 in the mitochondrial pathway of apoptosis [14][15].

A number of studies on animal models showed that epigallocatechin gallate (EGCG) can inhibit the growth of malignant cells and induce apoptosis even in cancer cell lines resistant to CD95-mediated apoptosis [16]. Saikosaponin A has proapoptotic activity, namely, it positively regulates the pathway mediated by Bax/Bcl-2/caspase-9/caspase-7/PARP [17], causing apoptosis of human colon cancer cells SW480 and SW620 (colon cancer cell lines derived from primary tumors and lymph node metastases, respectively) in a dose-dependent manner, which is obviously related to the inhibition of the PI3K/Akt/mTOR signaling pathway [18]. Bavachinin can influence the expression of Bcl-2, Bax, caspase-3/9, and the peroxisome proliferator γ (PPARγ) receptor. The bavachinin-induced generation of reactive oxygen species (ROS) depends on the PPARγ activation, which is capable of inducing A549 cell death. The effect caused by an increase in the level of reactive oxygen species (ROS) highlights the potential role of bavachinin as a chemotherapeutic agent against non-small cell lung cancer [19]. Gossypol can interact with the BH3 domain binding groove of the Bcl-xL and Bcl-2 antiapoptotic proteins. Simultaneous incubation of non-Hodgkin’s lymphoma Ramos cells with gossypol and etoposide enhances apoptosis due to the intensive release of cytosolic cyt c and activation of caspase-3 signaling depending on time intervals. These results are the basis for future preclinical and clinical studies of gossypol in the treatment of non-Hodgkin’s lymphoma [20]. It was shown that the action mechanism of resveratrol involves the blocking of certain transcription factors, such as B cell nuclear factor (NF-kB), AP-1 and Egr-1, as well as a decrease in the expression of antiapoptotic genes and activation of caspases. Its ability to influence the immune response mediated by B cells and increase the serum level of antibodies, exerting an antitumor effect, was previously revealed [21].

Autophagy and ferroptosis

Autophagy is a cellular degradation process leading to the removal of improperly folded or aggregated proteins, as well as the degradation of damaged organelles such as mitochondria, endoplasmic reticulum (EPR), and peroxisomes [22]. Autophagy can inhibit the growth and progression of malignant tumors, since the removal of damaged or non-functioning organelles prevents oncogenesis. At the same time, stimulation of autophagy remains an effective approach in antitumor therapy.

The mammalian target of rapamycin, mTOR, which has the properties of a modulator of cell growth and proliferation, and AMP-activated protein kinase (AMPK), responsible for signal conversion in response to various metabolic stresses, are regulators of autophagy initiation [23]. This process prevents tumor development only provided the possibility of selective autophagy, targeting certain cell organelles [24]. In cases where a tumor has already formed, autophagy suppression often leads to the development of less aggressive cancer forms [25]. It was shown that synthetic quinine analogues — chloroquine (CQ), isolated from the cinchona bark (Cinchona officinalis), and hydroxychloroquine (HCQ) — are the most common drugs used to treat acute and chronic inflammatory diseases. These drugs are also used in cancer treatment based on autophagy inhibition mechanisms, which consist in interrupting the fusion of autophagosomes and lysosomes [26].

According to Solomko et al., matrine-induced signals in tumor cells can lead to ferroptosis (has a protective effect against cervical cancer) [6][13]. Matrine exhibits considerable antitumor activity both in vitro and in vivo, along with other beneficial properties, e.g., antianxiety and antidepressive effects, relieving neuroinflammation in the brain caused by severe diseases. The mechanism of action of this compound is based on the suppression of cell proliferation and apoptosis induction, e.g., the highly metastatic breast cancer cell line MDA-MB-231 uses the VEGF-Akt-NF-kB signaling pathway. Unfortunately, numerous anticancer drugs (such as etoposide, tyrosine kinase inhibitors, arsenic trioxide, 5-fluorouracil) that cause ferroptosis are cardiotoxic [27]. To address this issue, non-toxic cardioprotective antitumor plant-derived medications with anticancer activity, such as berberine, epigallocatechin gallate, and resveratrol, have been developed and are used in combination with conventional chemotherapeutic agents [16][21][28].

Angiogenesis inhibition

Tumors induce the growth of new blood vessels due to releasing various growth factors, including vascular endothelial growth factor (VEGF). This results in the formation of blood capillaries inside the tumor. Protein Kinase G (PKG) regulates beta-catenin levels in healthy cells, promoting angiogenesis. Angiogenesis, in turn, is an important factor in the spread of tumor metastases. Fennel extracts — Trianthema portulacastrum and Spatholobus suberectus — inhibit tumor growth and angiogenesis, as well as alter the expression of HSP90 heat shock protein and its co-chaperone interactions in mouse models of breast cancer. These findings on the role of HSP90 in breast cancer biology and therapy are consistent with the effects described in the current literature. In fact, tumor growth and angiogenesis decrease when HSP90 is suppressed by the interaction of the KU-32 inhibitor with the C-terminal domain of this chaperone in trastuzumab-resistant HER2-positive breast cancer cells [29]. Morelloflavone blocks injury-induced neointimal hyperplasia by inhibiting vascular smooth muscle cell migration without causing apoptosis or cell cycle arrest [30]. Thus, the use of certain compounds of natural origin suppresses the formation of new blood vessels that require a significant amount of oxygen and nutrients for their growth, which can increase the antitumor effect [7].

Modulation of cellular signaling pathways

MAPK paths. Phytochemicals can affect both the cascade pathway kinase regulated by extracellular signals (ERK) and mitogen-activated protein kinases (MAPK) regulating cell growth and cell survival. Phytochemicals such as ursolic acid, kaempferol, resveratrol, gingerol, sulforaphane, genistein, and isothiocyanates have been reported to induce cancer cell apoptosis through the MAPK and ERK pathways [31]. It was shown that curcumin exerts the anticancer effect on retinoblastoma cells (RB, Y79) by activating exclusively the MAPK pathway.

Akt signaling pathways. The Act/PI3 signaling pathway plays an important role in cancer development. Epidermal growth factor (EGF) regulates a number of molecular mechanisms, including NF-kB activation and Akt phosphorylation. This promotes resistance to apoptosis and uncontrolled cell proliferation, which in turn leads to effects on caspases, Bcl-2 and glycogen synthase 3-β (GSK3b) kinases, as well as mTOR. Alkaloids and phenolic compounds make a significant contribution to the control over the expression of these factors. Resveratrol, curcumin, luteolin, flavone, and sulforaphane exhibit anticancer properties by ceasing the cell cycle and apoptosis, interfering with Akt/PI3K signaling [32]. In addition, Saikosaponin A inhibits the invasion and migration of SK-N-AS cells (human neuroblastoma cells) by regulating the angiogenesis-associated VEGFR2/Src/Akt pathway and protein expression associated with epithelial-mesenchymal transition (EMT) [17].

JAK/STAT signal transmission paths. By inhibiting the activity of JAK/STAT signaling and activating apoptotic cascades, the curcumin, resveratrol, and EGCG phytochemical compounds inhibit the translocation and collection of β-catenin in the nucleus by stimulating glycogen synthase kinase 3 (GSK3), which can lead to cell death in some forms of cancer [33].

Antioxidants with anti-inflammatory effects

Harmala extract (Peganum harmala) can reduce the viability of cervical carcinoma and colon cancer cells due to the action of alkaloids contained in high concentrations therein. In a study aimed at investigating cytotoxicity towards normal and tumor cells, the antioxidant activity of these alkaloids against human breast cancer cells was noted [34]. Three main epigenetic changes occur in tumor cells treated with plant polyphenols, i.e., a change in the structure of chromatin, DNA methylation, and, more importantly, alterations in the microRNAs level. For the same microRNAs, expression is increased in some tumors, while in others, on the contrary, it is reduced. Thus, the expression of the mi-Let-7 cluster increases in breast tumors and, conversely, decreases in lung tumors. Notably, EGCG, curcumin, and resveratrol modulate several classes of microRNAs that are involved in all stages of cancer development and regulate oncogenes or tumor suppressors of various cancers [16][21]. In particular, tetrandrine was shown to exhibit antiproliferative effects and cytotoxic activity against breast cancer (MDA-MB-231, HCC1937, MCF7) [14][15].

Cell cycle regulation

The cell cycle is a sequence of intracellular events, leading to cell division. The stages of the cell cycle are mediated by cyclin-dependent kinases (CDK) and their regulatory cyclin subunits [35]. Vindoline and catharanthine have antitumor effects due to their exposure in the M-phase cell cycle. They contribute to the cancer cell death by shortening microtubules and disrupting their function, which leads to the disappearance of mitotic spindle, thereby suppressing cell proliferation [34].

Quercetin can affect the cell cycle at the G1/S and G2/M control points by inducing the CDK p21 inhibitor and reducing the phosphorylation of the key regulatory pRb protein and indirectly blocking E2F, which are important factors of transcription and DNA synthesis [36]. The roscovitin synthetic compound, produced from the natural substance olomucine isolated from the Raphanus sativus (Brassicaceae) daikon, has passed clinical trials, showing high activity against various types of cancer. This medication is currently at the stage of clinical evaluation of efficacy in the treatment of Cushing’s disease and rheumatoid arthritis [37]. This drug is an inhibitor of cyclin-dependent kinases, preventing their activation and DNA repair due to non-homologous end joining (NHEJ). One of the most noticeable effects of the drug is the inhibition of formation of CDK2/cyclin E complexes, which causes a decrease in the pRb phosphorylation level and subsequent inactivation of members of the E2F family, leading to suppression of cyclin transcription and, ultimately, to cell cycle arrest. In this case, cell cycle arrest leads to the initiation of apoptotic death [6]. The modes of flavopiridol action are associated with the phosphorylation of cyclin-dependent kinases that block cell proliferation in G1 and G2 phases, and the apoptosis induction by increasing the level of E2F synthesis and Mcl-1 protein inactivation. A study in which the effect of EGCG on oncogenesis was tested on oral cancer (NOE) cell lines together with curcumin showed the EGCG ability to block cell division in G1, whereas curcumin blocked cell division in S/G2/M phases. The antagonistic interaction between curcumin and etoposide is caused by cell cycle arrest, which gives time for DNA damage to repair and prevents cell death. Another polyphenol, quercetin, may limit the effect of etoposide. Quercetin has a protective effect on HL-60 cells from etoposide, reducing the level of ROS generated in drug-treated cells (Table 2) [38][39].

Fungus-derived compounds

Cancer fungotherapy and the search for new anticancer agents are not limited to such fungi species as Fomitopsis pinicola, Hericium erinaceus, Trametes versicolor, and Inonotus obliquus from the Basidiomycota class. However, these four species can serve as typical representatives of medicinal fungi, which are widely used both in conventional medicine and in modern biomedical research. They belong to three different groups and are a rich source of bioactive compounds such as polyphenols, polysaccharides, glucans, terpenoids, steroids, cerebrosides, and proteins that show potential for treating various types of cancer [8]. Ergosterol is an active Fomitopsis pinicola fungus-derived compound, which is the main component affecting SW-480 cells and causing their apoptosis. Interestingly, the use of this extract in combination with cisplatin, a common chemotherapeutic agent, in mice showed their synergistic effect of impeding tumor growth. Taken together, these results provide solid evidence that, in addition to nonspecific cytotoxic compounds, F. pinicola contains substances with a specific antioncogenic potential, probably acting through the induction of apoptosis [48].

Krestin, isolated from the mycelium of the Trametes versicolor wood fungus, belongs to the class of polysaccharides. This compound shows significant antitumor activity against allogeneic and syngeneic animal tumors [45].

Polysaccharide-K (PSK) demonstrates a similar activity in various types of cancer, especially in gastrointestinal cancer. This medicine has been approved in Japan and China for use in cancer treatment [49]. Lentinan (isolated from Lentinus edodes) also belongs to the class of polysaccharides. This compound prevents neoplastic transformations caused by chemical carcinogens and viruses, while also suppressing the development of allogeneic and some syngeneic tumors. This polysaccharide is most often used in the treatment of tumors of the stomach, bones, and breast. The action mechanism of lentinan involves stimulation of CD8⁺ T lymphocytes, induction of interleukins 1 and 3, as well as the production of nitric oxide by immune cells. This leads to an increase in the synthesis of colony-stimulating factor (CSF) and the accumulation of proteins of the inflammation acute phase in combination with direct and indirect (through T lymphocytes) effects on macrophages. Lentinan showed clinical efficacy in various types of cancer, including stomach and lung cancer [47]. Another study by the same scientific group was devoted to the use of conk extracts from the Hericium erinaceus fungus with various solvents and testing for cytotoxicity against human monocytic leukemia U937 cells. The results showed that both aqueous and ethyl extracts can induce apoptosis. The suggested mechanism of action is through the suppression of anti-apoptotic factors (Bcl-2, Bcl-xL(S), XIAP and cIAPs) in combination with the absence of an increase in the level of pro-apoptotic proteins.

Erinacin A, a mycelial derivative of H. erinaceus, demonstrates activity that suppresses the growth of various gastrointestinal tumor lines [50]. Extracts of H. erinaceus or their fractions/components were shown to exhibit immunostimulating activity; antimetastatic activity by inhibiting matrix metalloproteinases; activity promoting the growth of probiotic intestinal flora; antioxidant potential; proapoptotic activity; and angiogenesis inhibition. This range of anticancer properties is provided by various compounds, including polysaccharides, lipids, terpenoids (unique erinacins), and even proteins. Thus, there are two possible strategies for the investigation of H. erinaceus anticancer effects, such as studying the complex effects of extracts with their further use as preventive dietary supplements and a detailed study of the mechanisms of individual fungus-derived compounds for use in targeted personalized anticancer therapy [51]. The anticancer potential of the Inonotus obliquus fungus is represented by several groups of components. Unique triterpenoids such as lanostane, inotodiol, and inonotsuoxides act in vivo on preparations of mouse skin and tumors of mouse xenografts derived from human chronic lymphocytic leukemia. Low molecular weight polyphenolic compounds of this fungus can inhibit topoisomerase II, which leads to a decrease in the growth of cultured HCT116 human colon carcinoma cells. Similar to H. erinaceus, cinder conk (Inonotus obliquus) is extremely rich in polysaccharides, which can perform immunomodulatory functions and inhibit oncogenesis [52].

The Cordyceps militaris fungus showed promising results in preclinical studies of antiproliferative and antimetastatic effects against various types of cancer. Ganoderma lucidum, known as reishi, contains ganoderic acids, which have antitumor and immunomodulatory properties. Grifola frondosa, also known as maitake, contains such compounds as maitake polysaccharides, which demonstrated anticancer activity in preclinical studies [53].

CONCLUSION

Plants contain a diverse range of biologically active compounds, including alkaloids, flavonoids, terpenoids, and polyphenols, which have been traditionally used in medical practice. Despite promising laboratory results in suppressing tumor growth and metastasis, many plant compounds require further study and clinical trials to confirm their effectiveness and safety. Among the anticancer IPFOs analyzed in this review, which are not a substitute for conventional drugs but are promising for concomitant therapy, matrine, etoposide, resveratrol, and ergosterol have attracted the greatest research interest [39][30][48]. Establishing optimal dosages and compositions of potential drugs is essential for assessing the prospects for their practical use, taking into account possible interactions with other medicinal products. Sustainable cultivation and harvesting of plant sources should be ensured.

It is important to note that treatment methods based on promising plant-derived medicines do not replace conventional approaches in cancer treatment. In general, a comprehensive approach to assessing the therapeutic potential of plant-derived cancer inhibitors is required. Fungi are a vast and diverse group of organisms known for their complex chemistry and unique biological activity. Their potential to produce powerful anticancer compounds has been known for decades, leading to significant research efforts aimed at identifying and characterizing cancer inhibitors of fungal origin. On the other hand, some fungus-derived compounds can be toxic to healthy cells. Similar to plant-derived compounds, fungus-derived medicines require optimization to ensure safe and effective dosage, as well as targeted delivery. Currently, clinical trials face a number of issues associated with underfunding and regulatory obstacles. The development of safe and effective drugs based on compounds of fungal origin requires complex purification methods and the creation of appropriate formulations, as well as the introduction of innovative achievements to solve these problems. Further research is needed to elucidate the action mechanisms of these compounds and identify targeted therapeutic strategies, similar to plant-derived preparations.