A subsequent study published in 1974 focused on methionine specifically [ 71 ]. This study was conducted on tissue cultures including W-256 (a rat breast cancer cell line), L1210 (a mouse lymphatic leukemia cell line), J111 (a human leukemia cell line), liver epithelial and liver fibroblasts of rats, skin fibroblasts of mice, and human breast and prostate cells that were normal or malignant [ 71 ]. The cells were cultured in folic acid- and cyanocobalamin-rich medium that either contained methionine or was methionine-free with a homocysteine supplement. Despite the media containing other methyl donors, the growth of the malignant cells was significantly impaired in the methionine-depleted media, while the normal cell growth was unchanged. These effects were attributed to the ability of normal cells to recycle homocysteine through methionine synthase to supply methionine endogenously. While this is true for normal cells, malignant cells lack the enzyme required to recycle homocysteine therefore giving methionine restriction the capacity to alter cancer cells while maintaining normal, healthy cells [ 72 73 ]. This enables the possibility that methionine restriction, as a therapeutic, may be able to specifically target cancer cells, preventing off-target effects on normal cellular processes. The following sections of this review provide an overview of the literature regarding methionine restriction and specific cancer types, including prostate, breast, and colorectal cancers ( Table 1 ).

In 1959, one of the early studies conducted in methionine restriction evaluated several outcomes produced from diets lacking specific amino acids. The study was conducted on rats fed isocaloric diets that were complete in all amino acids or devoid of one essential amino acid [ 70 ]. After transplantation of the Walker tumor, and 10-day preparative diet, rats were divided into different groups. Each group was fed a specific diet with different amino acid compositions for 5 days. While the initial aim of this study was to distinguish between two opposing views on nitrogen balance and amino acid restriction, the results showed a significant reduction in tumor growth in the rats fed diets lacking either methionine, valine, or isoleucine [ 70 ].

In another study, PC-3, DU-145, and LNCaP human prostate cancer cell lines were cultured in complete- or methionine-free media and methionine dependency was evaluated [ 77 ]. The results showed that PC-3 is completely methionine-dependent, while DU-145 cells were mildly dependent, and LNCaP cells were almost completely methionine-independent. These data indicate that the responses to methionine restriction vary across different cancers, although MR inhibited growth of all three cancer cell lines [ 77 ]. The mechanisms by which MR reduced cancer growth also differed between the cell lines, with MR upregulating p21 and p27 (cell cycle inhibitors that halt cell cycle progression) in LNCaP cells, but only increasing p27 in PC-3 cells [ 77 ]. Further, the PC-3 cells began to undergo apoptosis within six days of MR, whereas the LNCaP cells were relatively resistant to MR-induced apoptosis [ 77 ]. Together, these data indicate a precision diet such as MR may benefit a subpopulation of patients with prostate cancer.

Methionine restriction has been shown to induce apoptosis in the human prostate cancer cell lines, PC3 and DU145 [ 75 77 ]. MR inhibits Raf and Akt oncogenic pathways, while increasing caspase-9 and the mitochondrial pro-apoptotic protein, Bak [ 75 76 ]. Restricting media methionine concentrations damages mitochondrial integrity, leading to apoptosis in both prostate cancer cell lines [ 75 76 ]. Additionally, energy production was impaired and ROS production was decreased. Caspase-dependent and -independent apoptosis was observed in response to MR [ 75 76 ]. Other studies have identified that c-Jun N-terminal kinases (JNK1) is a critical regulator of MR-induced apoptosis in prostate cancer cells [ 78 ].

Another target of MR in prostate cancer cells is thymidylate synthase (TS). Thymidylate synthase is the enzyme that catalyzes the methylation of deoxyuridylic acid during nucleotide biosynthesis and is thus an important target for cancer treatment. The chemotherapy drug, 5-fluorouracil (5-fu), inhibits TS activity by disrupting action of TS, causing DNA and RNA damage, making 5-fu an effective and commonly used cancer treatment [ 74 ]. However, 5-fu has also been reported to increase TS protein expression, resulting in 5-fu drug resistance [ 92 ]. Interestingly, several studies have shown that MR and 5-fu have synergistic anti-cancer effects [ 12 87 ]. MR selectively reduces TS activity in prostate cancer cells by ~80% within 48 h, but does not affect TS activity in normal prostate epithelial cells [ 74 ]. Importantly, MR also reduces TS protein expression, potentially explaining the synergy between MR and 5-fu [ 74 ]. That MR also reduces TS protein expression may make MR an attractive treatment alongside 5-fu to help combat resistance to 5-fluoruracil.

Prostate cancer is the second leading cause of cancer death among adult men in the US and current treatment options include hormonal therapy to reduce testosterone levels, radiation therapy, or surgical procedures [ 90 ]. While there are treatment options available for prostate cancer, there are no known interventions to prevent the development of prostate cancer. Using a well-characterized mouse model for prostate cancer (Transgenic Adenocarcinoma of the Mouse Prostate; TRAMP), it was shown that dietary MR inhibits prostate cancer development especially in the anterior and dorsal lobes of the prostate, where the most severe lesions are found [ 82 ]. While the mechanism by which MR inhibits prostate cancer development is not known, evidence suggests that MR may work by inhibiting prostate cancer cell proliferation, inhibiting the insulin/IGF-1 axis, or by reducing polyamine synthesis [ 82 ]. The cells of the prostate produce high levels of polyamines and inhibition of polyamine synthesis is effective at suppressing tumor growth in prostate cancer [ 91 ]. Given the dependence of polyamine synthesis on methionine, the polyamine biosynthetic pathway may be a primary target of MR in prevention and/or treatment of prostate cancer.

4.3. Methionine Restriction and Breast Cancer

Breast cancer is the second most common form of cancer diagnosed in women. Depending on the type of breast cancer, treatment options include surgery to remove the cancer or the entire breast, chemotherapy, hormone therapy, radiation therapy, and in some cases, targeted therapy drugs or immunotherapy. Breast cancer cells are hormone receptor-positive if they express either (or both) of the estrogen and/or progesterone receptors and are considered HER-2-positive if the breast cancer cells overexpress the protein, HER-2 (human epidermal growth factor receptor 2). If the cells meet none of these criteria, it is triple negative breast cancer (TNBC). TNBC makes up about 16% of all breast cancer diagnoses [ 93 ]. Few studies have examined the efficacy of MR in breast cancer models.

To investigate the effects of MR on breast cancer, a comprehensive study employed MR in a xenograft model for breast cancer, an immortalized human breast cell line, and an invasive breast cancer cell line [ 85 ]. In the animal model, athymic nude mice were injected with MCF10AT1 breast cancer cells. The control group was fed a diet containing 0.86% methionine while the MR group was fed a diet containing 0.12% methionine for 12 weeks [ 85 ]. Methionine restriction inhibited tumor progression in the mice by decreasing cell proliferation and increasing apoptosis [ 85 ]. MR increased expression of p21, but not p27, in the mouse mammary gland. Studies in the breast cancer cell line and immortalized breast cell line supported the involvement of p21 in the mechanism of action of MR. Additional mechanisms proposed involved the MR-induced reduction in circulating insulin and IGF1, which have both been linked to tumor growth, and the MR-induced depletion of polyamines [ 85 ].

TNBC has fewer treatment options than other forms of breast cancer due to the lack of a response to hormone therapy and drugs that target HER-2. Methionine restriction may provide a way to enhance efficacy of potential treatment options for TNBC. Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) receptor agonists are an exciting possibility for cancer treatment due to their ability to induce apoptosis in cancer cells while having little effect on normal cells. Despite their efficacy in preclinical studies, TRAIL receptor agonists have not been successful in human clinical trials. Methionine deprivation enhances expression of TRAIL-R2 in TNBC cells, but not in normal breast epithelial cells [ 81 ]. Further, a methionine-free diet suppresses breast cancer growth and enhances the efficacy of the TRAIL-receptor 2 monoclonal antibody, lexatumumab, in inhibiting breast cancer growth in mice [ 81 ]. Studies have identified that out of 10 essential amino acids tested, depletion of methionine elicited the greatest inhibition of migration and invasion of TNBC cells [ 79 ]. Together, these data suggest that a combination of TRAIL receptor agonists with a methionine-restricted diet may enhance efficacy of this treatment [ 81 ].