Antimetabolites in the Treatment of Solid Tumors: Competitive Inhibition by Structural Analogs

Enzymatic Inhibition with Structural Analogs

Antimetabolites such as methotrexate -a folate analog- have been used to halt tumor progression or disrupt embryogenic growth since the 1950s. 2-deoxy-D-glucose (2DG) -an undegradable pseudosugar with antiproliferative effects- has served routinely in cancer diagnosis through Positron Emission Tomography, combined with ¹⁸F as radiotracer, and is both safe and effective in humans when used on its own [1]. Sodium ascorbate (which happens to be a hexose or six-carbon glucose analog) is widely recognized for its strong antimicrobial and antineoplastic properties, when properly applied in pharmacological doses [2]. Intravenous D-Glucosamine has been shown to induce tumor hemorrhage, halt cell proliferation and prolong survival since 1970 [3].

In one form or another, the principle of Enzymatic Inhibition with Structural Analogs (EISA) for the treatment of solid tumors is being deployed in many clinics to induce an acute energy constriction in neoplastic tissues, usually as an adjunct to known chemotherapeutic regimes such as R-CHOP-21. Such an approach requires the implementation of carefully engineered dietary restrictions, as suitable to the host, coupled with metabolic disruptors. This is routinely accomplished through intravenous injections of structural analogs of glutamine, glucose and pyruvate, administered under deep physiological ketosis.

In the clinical setting, physiological ketosis is defined as: ketonemia ≥4 mM/L, glycemia ≤3 mM/L, and pH 7.4 (±0.1). This state is arrived at by way of a low-calorie ketogenic diet, or short-term water-only fasting (usually 72 hours), and has been extensively proven to optimize chemotherapy regimens of any type [4-7].

In the last two decades, this author and many other researchers and clinicians have acquired deep functional knowledge on the use of the glucose analog 2-deoxy-D-glucose as an adjuvant in the treatment of highly glycolytic tumors [1, 8]. As stated, the intravenous use of pharmacological doses of sodium ascorbate has proven to be selectively cytotoxic for cancer cells of multiple tumor types, both in vitro [9] and in vivo [10], as well as in Homo, including pancreatic adenocarcinoma [11-13].

Safe and effective metabolic interventions intended to optimize standard therapeutics such as tumor-reducing surgery, cytotoxic chemotherapy, immunotherapy and radiotherapy are readily available. The intended purpose of these adjuvant metabolic interventions is to increase the therapeutic index (TI) of orthodox treatments. However, there are many reports of tumor remissions and substantial improvements in five-year survival rates when employed on their own as a sole form of treatment in individuals deemed non suitable for cytotoxic chemotherapy [14-17].

Rationale: Exploiting the functional asymmetry of cancer cells with structural analogs

It has long been established that virtually all solid tumors that constitute metastasis have a high Standardized Uptake Value or SUVmax in Positron Emission Tomography scans [18]. All manner of cancers exhibit a hypermetabolic phenotype, proven to be quantitatively predictive of invasiveness and survival [10-21]. Adding labeled nutritional substrates to cell cultures and experimental animals in countless studies has shown that substantial amounts of carbon donors metabolized by neoplastic cells require transmembrane carriers, as well as an oversized enzymatic machinery. There is, for instance, unmistakable evidence of a proportional, direct correlation (r 0.83) between GLUT1 and cancer invasiveness [22-24]. Key rate-limiting enzymes such as glutamate dehydrogenase (GLUD), hexokinase II (HK2), and isoenzyme 5 in the lactate dehydrogenase family (LDHA) are intensely overexpressed in neoplastic cells, dwarfing its enzymatic expression in normal cells by an order of magnitude [25-29]. Tumor avidity for glutamine, glucose and pyruvate, as well as the overexpression of their catalyzing enzymes and transporters is already being exploited therapeutically by hundreds of doctors from multiple countries in a systematic manner [30-33].

Such approach takes advantage of the compensatory increase of fermentative glycolysis and glutaminolysis in neoplastic cells. This aberrant form of fermentative metabolism takes place even under high partial pressure of oxygen in the tissue (ptiO₂), hence named the “facultative” anaerobiosis of cancer cells [34]. Although in vivo, the actual yield of oxidative phosphorylation (~28 molecules of ATP) is somewhat lower than the theoretical yield (~36 molecules), this functional asymmetry has profound implications. It is deemed that the relative inefficiency in the energetic yield of glucose fermentation is approximately 14-fold lower than that of respiration (2 moles and 28 moles of ATP per mole of glucose, respectively). Poor ATP output induces cancer cells to an overexpression of GLUT transporters [23], hexokinase-II [26], and isoenzyme “A” (LDHA) also named LDH-5 [29]. Therefore, blocking these metabolic pathways has strong disruptive effects on neoplastic tissues (sometimes inducing acute tumor necrosis) while sparing the host.

The extensive reprogramming of energy metabolism undergone by cancer cells explains the intense glucose and/or glutamine uptake shown by solid tumors and is, as mentioned, the basis for the Positron Emission Tomography (PET). Using ¹⁸Fluoro-DeoxyD-Glucose or ¹⁸Fluoroglutamine as a radiotracer, the absorption of these substrates reveals hypermetabolic tissues. In PET-positive tumors, with a Standardized Uptake Value equal to or higher than 3 (SUVmax ≥ 3), glycolysis and glutaminolysis are known to be overexpressed by a factor of 10, 30, or higher, even under a ptiO2 high enough to sustain oxidative phosphorylation [35]. Despite their clinical heterogeneity, this central feature of cancer, the Warburg effect, is a universal phenotypical hallmark of all malignant tumors [36].

Ongoing research and challenges

To date, several thousand technical papers on a vast array of metabolic approaches have been published [37]. As per the peer-reviewed literature, over the last two decades multiple medical centers and clinical research groups in several countries have acquired experience and functional knowledge on the use of structural analogs of glutamine, glucose, and pyruvate as non-toxic metabolic disruptors [38, 39]. A summarized list of these antimetabolites includes 2DG, ascorbate, D-glucosamine, telaglenastat, piperazine erastin, L-asparaginase, DON, 3BP, EGCG and oxamate. Several other compounds are being tested preclinically.

Applying the principle of competitive inhibition with structural analogs as an adjunct -or even as a sole form of treatment, with no other concomitant therapeutic interventions- has proven effective in at least 12 types of malignancies, including kidney, bladder, breast, stomach, colon, lung, prostate, and pancreatic cancer, as well as lymphoma, sarcoma, and melanoma [40-46].

Unexpected findings in the clinical setting following adjuvant or palliative interventions, i.e. tumor remissions and doble- or even triple-digit percentual increases in overall survival time (OS), have led clinicians to conclude that functional constrictions induced by antimetabolites are an effective, non-toxic, economical method to improve the therapeutic index of standard cancer treatments. By our calculations, as of March 2026, in just a few medical centers across Latin America and Spain, well over 1.007.000 metabolic treatment sessions have been administered on ≈7.900 cancer patients over the course of 10 years.

As of this writing, 13 prospective, phase three clinical studies concerning some form of metabolic therapy are registered in http://www.clinicaltrials.gov . Although hundreds of doctors already employ different modalities of metabolic therapy due to their safety and simplicity, more clinical trials are necessary to test emerging molecular candidates and to assess the efficacy of combinatorial metabolic approaches to cancer.

Conclusion

Standard treatments of cancer can be substantially improved by the adjunct introduction of antimetabolites of glutamine, glucose, pyruvate and possibly other nutritional substrates. The competitive inhibition of several rate-limiting enzymes within neoplastic cells using already available structural analogs may improve therapeutic outcomes in a non-toxic, cost-effective way.

Funding: All funds required for the research, writing and publication of this paper have been provided entirely by the author.

Conflicts of Interests: As of this writing, the author has no competing interests (directly or indirectly, by ownership or by affiliation) with any brand, company, or institution. No outside contributors had any role in the design, implementation, or analysis of the study or the write-up of this paper.

References

1. Prieto Gratacós E, García Oliver P, Álvarez RP, Redal MA, Sosa I, et al. (2020) Safety of Antimetabolite 2-Deoxy-D-arabinohexose (2DG) as a Coadjuvant Metabolic Intervention in 268 Cancer Patients. Cancer Medicine Journal Review 3: S1.
2. Prieto Gratacós E1, Laguzzi M. (2020) Pharmacokinetics of SixCarbon Analogues of L-Glucose in Tumour-Bearing Humans (Series I: Ascorbate). Oncol Res Ther 5: 10100.
3. Bekesi JG, Richard J. (1970) Inhibitory Effects of d-Glucosamine on the Growth of Walker 256 Carcinosarcoma and on Protein, RNA, and DNA Synthesis. Winzler Cancer Res. 30: 2905-2912.
4. Brandhorst S. (2021) Fasting and fasting-mimicking diets for chemotherapy augmentation. Gero Science. 43:1201-1216.
5. Li N, Sun YJ,Huang LY, Li RR, Zhang JS, et al. (2025) Fastingmimicking diet potentiates anti-tumor effects of CDK4/6 inhibitors against breast cancer by suppressing NRAS- and IGF1-mediated mTORC1 signaling. Drug Resist Updat.78: 101161.
6. Lee C, Raffaghello L, Longo VD, (2012) Starvation, detoxification, and multidrug resistance in cancer therapy. Drug Resist Updat.15:114-22.
7. Groot SD, Lugtenberg RT, Cohen D, Welters MJP, Ehsan I, et al. (2020) Fasting mimicking diet as an adjunct to neoadjuvant chemotherapy for breast cancer in the multicentre randomized phase 2 DIRECT trial. Nat Commun. 11: 3083.
8. Mark Stein, Hongxia Lin, Chandrika Jeyamohan, Dmitri Dvorzhinski, Murugesan Gounder, et al. (2010) Targeting tumor metabolism with 2-deoxyglucose in patients with castrate-resistant prostate cancer and advanced malignancies. Prostate. 70:1388-1394.
9. Jiang X, Liu J, Mao J, Han W, Fan Y, et al. (2023) Pharmacological ascorbate potentiates combination nanomedicines and reduces cancer cell stemness to prevent post-surgery recurrence and systemic metastasis. Biomaterials. 295: 122037.
10. Chen Q, Espey MG, Sun AY, Pooput C, Kirk KL, et al. (2008) Pharmacologic doses of ascorbate act as a prooxidant and decrease growth of aggressive tumor xenografts in mice. Proc Natl Acad Sci 105:11105-9.
11. Ma Y, Chapman J, Levine M, Polireddy K, Drisko J, et al. (2014) High dose parenteral ascorbate enhanced chemosensitivity of ovarian cancer and reduced toxicity of chemotherapy Sci Transl Med. 6: 222ra18.
12. Gratacós PE, Redal AMA, Amador V, Sosa I, Laguzzi M, et al. (2018) Metabolic Therapy of Pancreatic Cancer. Clinics in Oncology 3: 1512.
13. Cameron E, Campbell A. (1974) The orthomolecular treatment of cancer. II. Clinical trial of high-dose ascorbic acid supplements in advanced human cancer. Chem Biol Interact. 9: 285-315.
14. DeVita VT, Lawrence TS, Rosenberg SA, (2008) DeVita, Hellman, and Rosenberg’s Cancer: Principles and Practice of Oncology. Lippincott Williams & Wilkins.
15. Moon SY, Joo KR, So YR, Lim JU, Cha JM, Et al. (2013) Predictive value of maximum standardized uptake value (SUVmax) on 18F-FDG PET/CT in patients with locally advanced or metastatic pancreatic cancer. Clin Nucl Med. 38: 778-83.
16. Krengli M, Ferrara E, Guaschino R, Puta E, Turri L, Et al. (2022) 18F-FDG PET/CT as predictive and prognostic factor in esophageal cancer treated with combined modality treatment. Ann Nucl Med. 36: 450-459.
17. Martiniova L, Kamel S, Kairemo K, Benjamin R, Somaiah N, et al. (2024) Predictive Value of Quantitative Parameters of (18)F-FDG PET/CT in Patients with Liposarcoma. Diagnostics 14: 2021.
18. Davis-Yadley AH, Abbott AM, Pimiento JM, Chen DT, Malafa MP, et al. (2016) Increased Expression of the GLUT-1 Gene is Associated with Worse Overall Survival in Resected Pancreatic Adenocarcinoma. Pancreas. 45: 974-979.
19. Yuan Q, Liu H, Song C, Tian Y, Hassan W, et al. (2025) Prevalence of GLUT1 overexpression in human cancers a systematic review and meta-analysis. Discov Oncol. 16: 1761.
20. Ito S, Fukusato T, Nemoto T, Sekihara H, Seyama Y, et al. (2002) Coexpression of Glucose Transporter 1 and Matrix Metalloproteinase-2 in Human Cancers. J Natl Cancer Inst. 94: 1080-1091.
21. Mathupala SP, Ko YH, Pedersen PL. (2006) Hexokinase II: cancer’s double- edged sword acting as both facilitator and gatekeeper of malignancy when bound to mitochondria. Oncogene. 25: 4777-86.
22. Mathupala SP, Ko YH, Pedersen PL. (2008) Hexokinase-2 bound to mitochondria: Cancer’s stygian link to the “Warburg effect” and a pivotal target for effective therapy. Semin Cancer Biol. 19: 17-24.
23. Jin L, Li D, Alesi GN, Fan J, Kang HB, et al. (2015) Glutamate dehydrogenase 1 signals through antioxidant glutathione and can be regulated by glutamine metabolism in cancer cells. Cancer Cell. 27: 257-70.
24. Fang Huang, Qiuyue Zhang, Hong Ma, Qing Lv, Tao Zhang, et al. (2014) Expression of glutaminase is upregulated in colorectal cancer and of clinical significance. Int J Clin Exp Pathol. 7:1093-1100.
25. Prieto Gratacós, E., Aguirre Gauto P (2020) Isoform A of Lactate Dehydrogenase (LDH-A), a potential ultra-early, high sensitivity biomarker of tumorigenesis. Br J Med Health Res 7: 2394-2967.
26. ISOM
27. ACAM
28. IFM
29. AIHM
30. Koppenol WH, Bounds PL, Dang CV () Otto Warburg’s contributions to current concepts of cancer metabolism. Nat Rev Cancer 11: 325-337.
31. Kang S, Kim EH, Hwang JE, Shin JH, Jeong YS, et al. (2019) Prognostic significance of high metabolic activity in breast cancer: PET signature in breast cancer. Biochem Biophys Res Commun 511: 185-191.
32. Seyfried TN, Flores RE, Poff AM, D’Agostino DP (2014) Cancer as a metabolic disease: implications for novel therapeutics. Carcinogenesis 35: 515-527.
33. https://pubmed.ncbi.nlm.nih.gov/?term=metabolic+treatment+of+canc er+clinical+studies
34. Stein M, Lin H, Jeyamohan C, Dvorzhinski D, Gounder M, et al. (2010) Targeting tumor metabolism with 2-deoxyglucose in patients with castrate-resistant prostate cancer and advanced malignancies. Prostate 70: 1388-1394.
35. Prieto Gratacós E, García Oliver P, Alvarez RP, Redal MA, Sosa I, et al. (2020) Nuliglucaemia Lucidae: Extreme Deprivation of Blood Glucose as Organic Context for Cancer Treatment with Antimetabolites.
36. Tripathi SC, Fahrmann JF, Vykoukal JV, Dennison JB, Hanash SM, et al. (2018) Targeting metabolic vulnerabilities of cancer: Small molecule inhibitors in clinic. Cancer Rep (Hoboken) 2: e1131.
37. Jackson JA, Riordan HD, Hunninghake RD, Riordan N (1995) High Dose Intravenous Vitamin C and Long Time Survival of a Patient With Cancer of Head of the Pancreas.
38. Jayathilake PG, Victori P, Pavillet CE, Lee CH, Voukantsis D, et al. (2024) Development of cancer metabolism as a therapeutic target: new pathways, patient studies, stratification and combination therapy. PLoS Comput Biol 20: e1011944.
39. Yang WH, Qiu Y, Stamatatos O, Janowitz T, Lukey MJ, et al. (2021) Enhancing the Efficacy of Glutamine Metabolism Inhibitors in Cancer Therapy. Trends Cancer 7: 790-804.
40. Khodabakhshi A, Akbari ME, Mirzaei HR, Seyfried TN, Kalamian M, et al. (2021) Effects of Ketogenic metabolic therapy on patients with breast cancer: A randomized controlled clinical trial. Clin Nutr 40: 751758.
41. Fritz H, Flower G, Weeks L, Cooley K, Callachan M, et al. (2014) Intravenous Vitamin C and Cancer: A Systematic Review. Integr Cancer Ther 13: 280-300.
42. Ou J, Zhu X, Chen P, Du Y, Lu Y, et al. (2020) A randomized phase II trial of best supportive care with or without hyperthermia and vitamin C for heavily pretreated, advanced, refractory non-small-cell lung cancer. J Adv Res 24: 175-182.
43. Prieto Gratacós E, García Oliver P, Álvarez R, Redal MA, Sosa I, et al. (2020) Safety of Antimetabolite 2-Deoxy-D-arabinohexose (2DG) as a Coadjuvant Metabolic Intervention in 268 Cancer Patients.
44. Stein M, Lin H, Jeyamohan C, Dvorzhinski D, Gounder M, et al. (2010) Targeting Tumor Metabolism With 2-Deoxyglucose in Patients With Castrate-Resistant Prostate Cancer and Advanced Malignancies. Prostate 70: 1388-1394.
45. Prieto Gratacós E, Alvarez, Redal MA, Amador V, Sosa I, et al. (2018) Metabolic Therapy of Pancreatic Cancer. Clin Oncol 3: 1512.
46. Prieto Gratacós E, Redal MA, Alvarez R (2019) Impact of Non-Toxic Metabolic Disruptors on Overall Survival and One Year Survival Rate in Exocrine Pancreatic Cancer.