Folate Mediated One Carbon Metabolism

Cancer metabolism is typically characterized by metabolic demands, nutrient supply, and the regulation of metabolic enzymes. While many of these features are linked to specific genetic factors, increasing evidence suggests that metabolic regulation, in line with genetic factors, significantly impacts cancer incidence and biological progression. As a result, specifically targeting tumor metabolism has emerged as a primary therapeutic strategy in oncology.

Folic acid, a water-soluble B vitamin, is derived from food and is converted in the body to tetrahydrofolate (THF), which plays a role in various biochemical reactions. Through folate-mediated one-carbon metabolism (FOCM), THF provides essential nucleotides or one-carbon units necessary for DNA replication and methylation, highlighting its crucial role in epigenetics.

The earliest reports connecting FOCM with cancer treatment date back to the 1940s. The antifolate drug methotrexate was discovered to alleviate symptoms of acute lymphoblastic leukemia (ALL) and has been significant in treating acute leukemia, choriocarcinoma, and lung adenocarcinoma, indicating that targeting FOCM is an effective method for cancer treatment.

This article focuses on key enzymes in FOCM, particularly SHMT2 and MTHFD2, as potential targets for cancer therapy. It discusses their expression in cancer, potential functions, regulatory mechanisms, and some inhibitors, providing references for the future development of corresponding drugs.

Folate-mediated one-carbon metabolism (Xie et al.,2024).

Folate-Mediated One-Carbon Metabolism in Cancer

Folate-mediated one-carbon metabolism is a complex network of metabolic pathways that are intricately linked to folate metabolism. These pathways are crucial for the interconversion of serine and glycine, de novo synthesis of purines and thymidylate, and the methylation of homocysteine to methionine. FOCM is compartmentalized within cells, with different components localized in the cytoplasm, nucleus, or mitochondria. This spatial organization allows FOCM to act as a regulator and sensor of the cellular nutritional state, controlling the distribution of one-carbon units among various metabolic receptors. This, in turn, influences the synthesis of nucleotides, specific amino acids, S-adenosylmethionine (SAM), and glutathione.

Moreover, FOCM plays a role in maintaining redox balance by contributing to the production of glutathione, which is vital for cellular antioxidant defenses. Thus, FOCM not only provides essential nutrients necessary for cellular growth and proliferation but also regulates the nutritional status of cells through its epigenetic and redox functions. Abnormalities in folate metabolism have been observed in various cancers, including breast, liver, lung, colorectal, and bladder cancers, highlighting the universality of research on folate metabolism in cancer. This indicates that FOCM is a promising target for cancer treatment due to its dual role in supplying necessary nutrients for cell proliferation and influencing the stability of redox states within the body.

Antifolate Drugs in Cancer Treatment

Currently, the U.S. Food and Drug Administration (FDA) recognizes several antifolate antitumor drugs, primarily inhibitors of thymidylate synthase (TYMS) and dihydrofolate reductase (DHFR). These drugs function by interfering with the synthesis of purines and thymidine, thereby inhibiting DNA replication in cancer cells. They have been widely used in treating conditions like acute lymphoblastic leukemia, colorectal cancer, advanced non-small cell lung cancer, and peripheral lymphocyte lymphoma.

Recently, there has been increasing interest in mitochondrial enzymes SHMT2 and MTHFD2 in cancer therapy. Mitochondria are not only energy factories for cells but also provide a range of nutrients essential for cellular growth. Analyses of mRNA in cancer cells have revealed that SHMT2 and MTHFD2 are among the top five genes with the highest expression levels in various tumors. Abnormal expression of these enzymes can disrupt DNA synthesis and oxidative balance, further establishing FOCM as an attractive target for cancer treatment.

Given the significant relationship between mitochondrial metabolism and cancer, SHMT2 and MTHFD2 are highlighted as potential therapeutic targets. Their role in FOCM underscores the importance of investigating these enzymes further to develop effective cancer therapies that exploit metabolic vulnerabilities within tumor cells.

SHMT2 in Cancer Metabolism

Large-scale genomic studies of human tumors have revealed that serine hydroxymethyltransferase 2 (SHMT2) is essential for the survival of cancer cells. This enzyme is highly expressed in various cancers, including gliomas, colorectal cancer, and breast cancer. Notably, the knockout of SHMT2 has been shown to inhibit the proliferation of cancer cells, suggesting that targeting SHMT2 could provide a promising avenue for cancer therapy.

Impact on Amino Acid Metabolism

Serine plays a crucial role as a one-carbon unit donor in the folate cycle, contributing to nucleotide synthesis, methylation reactions, and the formation of NADPH, which is vital for maintaining redox homeostasis. SHMT2 catalyzes the conversion of serine into glycine and CH2-THF in the mitochondria. Glycine is a precursor for downstream metabolites such as glutathione and purines, while CH2-THF facilitates the transfer of one-carbon units to the cytoplasm, supporting the synthesis of purines and thymidylate. The ability of SHMT2 to influence amino acid metabolism underscores its role in maintaining cellular redox balance and nucleotide synthesis.

Influence on Mitochondrial Respiratory Chain

SHMT2 is also linked to the mitochondrial respiratory chain and redox state. Studies indicate that the knockout of SHMT2 leads to downregulation of mitochondrial respiratory chain complexes, specifically Complexes I and IV. In contrast, overexpression of SHMT2 appears to enhance their function. This suggests that SHMT2 is beneficial for the normal functioning of the mitochondrial respiratory chain. The downregulation of these complexes following SHMT2 inhibition seems to be primarily translational rather than transcriptional, highlighting the need for further investigation into the mechanisms of how SHMT2 affects the assembly of these complexes.

Regulatory Mechanisms

The expression of SHMT2 is regulated by several known signaling pathways and transcription factors. For instance, mTOR, a serine/threonine kinase, plays a crucial role in regulating the synthesis of nucleotides, lipids, and proteins. The activation of mTORC1 inhibits glycogen synthase kinase-3 (GSK-3), which in turn promotes the upregulation of SHMT2 expression. Additionally, the JAK/STAT signaling pathway is involved in various cellular processes, including proliferation and apoptosis. In LNCaP cells, IL-6 activates JAK2/STAT3, leading to increased SHMT2 expression in mitochondria. Other pathways, such as those involving NRF2, also regulate SHMT2 expression, particularly under oxidative stress conditions. Given its association with redox homeostasis and metabolic reprogramming, targeting SHMT2 could be a promising strategy for cancer therapy.

Development of Inhibitors

The expression patterns of SHMT2 and its impact on cancer cell behavior suggest that developing selective inhibitors of this enzyme could be beneficial for cancer treatment. While NSC127755 was the first identified SHMT inhibitor, its development was limited due to adverse effects. Leukopsin has also shown inhibitory effects on both SHMT isoforms but has not been applied clinically due to its conversion into other folate analogs in the body.

In 2015, researchers discovered a plant-derived SHMT inhibitor, compound 2.12, which induced apoptosis in lung cancer cells. Following this, another plant-derived compound, SHIN1, was optimized and demonstrated enhanced SHMT activity in vitro. However, its rapid clearance hindered further in vivo studies. An optimized version, SHIN2, exhibited significant inhibitory effects on T-cell acute lymphoblastic leukemia (T-ALL) cell proliferation and showed anti-leukemia activity in mouse models. Additionally, virtual screening identified pyrrolidine structures as potential SHMT2 inhibitors, while studies on pyrrolo[2,3-b]pyridine folate analogs revealed that AGF347 has broad anticancer effects across multiple cancer types.

Recent research has provided structural insights into antifolate drugs, with compounds like lometrexol (LTX) and pemetrexed (PTX) demonstrating inhibitory activity against SHMT2. Their efficacy as SHMT2 inhibitors suggests that they could serve as excellent starting points for developing effective SHMT2-targeted therapies.

MTHFD2 in Cancer Metabolism and Treatment

MTHFD2 (methylenetetrahydrofolate dehydrogenase 2) is expressed in the mitochondria of embryonic and transformed cells. It catalyzes two consecutive steps in the folate pathway, leading to the production of formate, which serves as a one-carbon unit essential for various metabolic processes. Research indicates that MTHFD2 is significantly upregulated in many rapidly proliferating tumors, such as breast and colorectal cancers, and its high expression correlates with poor patient survival. Notably, MTHFD2 is rarely expressed in healthy adult tissues, which highlights its potential as a therapeutic target in cancer.

Impact on Purine Synthesis

Recent studies have elucidated MTHFD2's role in cancer metabolism, particularly in purine synthesis and NADPH production. The activation of the mTORC1 pathway can induce MTHFD2 expression in breast cancer cells, and MTHFD2 gene knockout leads to the accumulation of key intermediates in purine synthesis, such as aminoimidazole-4-carboxamide ribonucleoside (AICAR). Moreover, the MYCN oncogene enhances purine synthesis via MTHFD2, influencing cancer progression. The knockdown of MTHFD2 impairs cell proliferation, colony formation, migration, and DNA synthesis, underscoring its critical role in supporting rapid cell growth.

Influence on Redox Homeostasis

MTHFD2 also contributes to maintaining redox balance as an NAD(P)-dependent enzyme. In colorectal cancer, the MTHFD2 inhibitor LY345899 was shown to reduce NADPH production and disrupt redox homeostasis, leading to decreased tumor proliferation and metastasis. Mechanistically, MTHFD2 expression is upregulated by c-Myc through the KRAS-activated AKT and ERK pathways, further emphasizing its importance in metabolic regulation within cancer cells.

MTHFD2 Regulatory Mechanisms

The regulation of MTHFD2 expression involves several signaling pathways. Similar to SHMT2, MTHFD2 is influenced by the mTOR pathway. In non-small cell lung cancer, KRAS activation stimulates AKT and ERK1/2 pathways, resulting in the transcriptional upregulation of MTHFD2 through c-Myc binding to its promoter. Additionally, various transcription factors, such as SOX7, act as tumor suppressors by binding to the MTHFD2 promoter and inhibiting its expression. The EWS-FLI1 fusion protein in Ewing sarcoma also positively regulates MTHFD2 expression, further implicating it in cancer metabolism and redox balance. Given its abnormal expression across various cancers, targeting MTHFD2 presents a promising strategy for cancer treatment.

Development of Inhibitors

The widespread expression of MTHFD2 in multiple cancers highlights its potential as a therapeutic target. Initial studies have identified MTHFD2 inhibitors, including LY345889, although its selectivity is limited. More selective inhibitors have been developed, such as DS44960156, which demonstrated good selectivity against MTHFD2 compared to MTHFD1. The subsequent optimization of this compound led to DS18561882, which exhibited potent inhibitory effects on MTHFD2 and significantly suppressed tumor proliferation in mouse xenograft models. This compound shows promise as a potential treatment for breast cancer, although further research is needed to explore its efficacy in other cancer types.

ALDH1L2 in NADPH Production

ALDH1L2 (Aldehyde Dehydrogenase 1 Family Member L2) is an important mitochondrial enzyme that plays a vital role in producing NADPH, which is crucial for cellular redox balance. Research has shown that the NADPH generated by ALDH1L2 can help mitigate oxidative stress in melanoma cells, thereby promoting tumor metastasis. In studies involving NSG mice, the knockout of ALDH1L2 significantly inhibited distant metastases, indicating its role in tumor spread. Conversely, the use of antioxidants was found to enhance metastasis, suggesting that ALDH1L2 may be essential for maintaining the balance of oxidative stress during cancer progression.

Interestingly, the expression of ALDH1L2 is significantly elevated in human colorectal tumor tissues compared to normal tissues. This upregulation is associated with poorer survival rates in patients with colorectal and lung adenocarcinomas. Higher levels of ALDH1L2 correlate with decreased survival, particularly when compared to patients with low expression of mitochondrial enzymes like SHMT2 and MTHFD2. This finding highlights the potential of ALDH1L2 as a biomarker for prognosis in various cancers.

SHMT1: The Cytoplasmic Isoform of SHMT2

SHMT1 (Serine Hydroxymethyltransferase 1) serves as the cytoplasmic isoform of SHMT2 and is involved in critical metabolic pathways related to nucleotide synthesis. It is known to play a role in the synthesis of deoxythymidine monophosphate (dTMP) and vitamin B6, both of which are essential for DNA stability and cellular function. SHMT1 prevents the integration of deoxyuridine monophosphate (dUMP) into double-stranded DNA, thereby safeguarding the integrity of the DNA structure.

Moreover, SHMT1 is implicated in the regulation of oncogenic factors such as interleukin-6 (IL-6) and interleukin-8 (IL-8) through its role in sialic acid metabolism. Its downregulation has been associated with decreased levels of Neu5Ac, a key metabolite, ultimately leading to inhibited growth and migration of ovarian cancer cells. In liver cancer, SHMT1's ability to inhibit metastasis may involve the suppression of reactive oxygen species (ROS) production via the nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1) pathway.

Additionally, specific compounds that target SHMT1, including pyran derivatives, have demonstrated the ability to induce apoptosis in lung cancer cells. This suggests that SHMT1 not only plays a crucial role in cancer metabolism but also presents potential therapeutic avenues for targeting cancer cell proliferation and survival.

References

1. Xie, Hong, et al. "The association between serum folate and elderly diastolic hypertension: results from the NHANES (2007–2018)." Blood Pressure 33.1 (2024): 2380002.
2. Yang, Chengcan, et al. "Folate-mediated one-carbon metabolism: a targeting strategy in cancer therapy." Drug Discovery Today 26.3 (2021): 817-825.
3. Lan, Xu, Martha S. Field, and Patrick J. Stover. "Cell cycle regulation of folate‐mediated one‐carbon metabolism." Wiley Interdisciplinary Reviews: Systems Biology and Medicine 10.6 (2018): e1426.