Ketogenic diets and cancer in preclinical mouse models: more hype (and protein restriction) than reality?

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This represents an early draft of a section of an upcoming scientific review.

One of the most exciting series of findings involves reduction of xenograft growth in mouse cancer models among animals given the ketogenic diet. A much-celebrated meta-analysis of mouse cancer models by Klement et al., 2016 (using at least a 2:1 ketogenic ratio and included controls with 50% of energy without additional treatment and reported tumor growth and survival endpoints) showed a robust reduction in tumor growth was found in mice fed the ketogenic diet compared to mice fed standard rodent chow [1]. However, severe methodological problems may have caused this meta-analysis and the studies it includes to poorly represent the preclinical efficacy of the ketogenic diet. Among the twelve studies included in this meta-analysis, nine used ketogenic diets with protein content (as a percent of total kilocalories) ranging from 61% to as low as 22% of that in the control diets, or in other words, involving between a half and a four-fold protein restriction (with a mean protein intake of 42% that of the control diet) [2–11]. This is important because it has long been known that protein restriction causes a robust delay of chemically induced and xenograft tumor growth [12–16]. Methionine restriction, which also occurs during protein restriction, achieves the same effect [17–19], including among several types of glioma xenografts [20], the most prominent and much-celebrated cancer type to ostensibly respond to the ketogenic diet in the ketogenic diet literature [1]. Therefore, the positive results of these nine ketogenic diet studies are confounded by protein restriction, and it is unclear what role the ketogenic component of the diet played in the results reported.

Furthermore, while Klement et al., 2016 reported positive findings for the remaining three studies, two of these findings occurred in comparison to a high-fat diet [3,5], which has been shown in several studies to increase the rate of xenograft tumor growth [21,22]. In fact, in these same two papers, a comparison with a third group of mice fed standard rodent chow was reported, and this showed no significant difference with the ketogenic diet-fed mice (except for a small difference at day 39 in one paper, which favored the standard chow mice) [3,5]. Surprisingly, the comparison with this control group of mice, which would suggest no efficacy of the ketogenic diet beyond simply being better than a high-fat diet, was for some reason not reported by Klement et al., 2016, further biasing the meta-analysis.

The third of the three diets that matched for protein did see a beneficial effect for the ketogenic diet, but this version of the ketogenic diet was highly unusual, with non-ketogenic carbohydrate kilocalorie % and ketogenesis driven by supplemental medium-chain triglycerides (Martuscello et al., 2016). Interestingly, Martuscello’s MCT group had a 5% higher protein as % of macronutrients than control (21% vs 26%) and may have consumed more food: they gained more weight than the other groups.

This finding is promising, but future studies will need to confirm these results with less confounded feeding designs. Of note, one high-profile mouse study published last year in Nature also used a highly restricted protein intake in the ketogenic group—a mere 25% of the intake of the control group (Hopkins et al., 2018), perhaps exemplifying the prevalence of this design flaw in the ketogenic diet literature. Other recent papers, such as one that showed normalization of the metabolome in a breast cancer xenograft model with the administration of a ketogenic diet, are also confounded by profound protein restriction (Licha et al., 2019).

A list of studies discussed in this article can be found below:

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Note: the 2013 Poff article listed in the spreadsheet above refers to a different study (but published in the same year) than that examined by Klement et al., 2016, with a slightly different macronutrient composition than that listed. This difference does not substantially change the analysis or conclusion.

1.        Klement, R.J.; Champ, C.E.; Otto, C.; Kämmerer, U. Anti-tumor effects of ketogenic diets in mice: A meta-analysis. PLoS One 2016, 11, 1–16.

2.        Zhou, W.; Mukherjee, P.; Kiebish, M.A.; Markis, W.T.; Mantis, J.G.; Seyfried, T.N. The calorically restricted ketogenic diet, an effective alternative therapy for malignant brain cancer. Nutr. Metab. (Lond). 2007, 4, 5.

3.        Freedland, S.J.; Mavropoulos, J.; Wang, A.; Darshan, M.; Demark-Wahnefried, W.; Aronson, W.J.; Cohen, P.; Hwang, D.; Peterson, B.; Fields, T.; et al. Carbohydrate restriction, prostate cancer growth, and the insulin-like growth factor axis. Prostate 2008, 68, 11–9.

4.        Otto, C.; Kaemmerer, U.; Illert, B.; Muehling, B.; Pfetzer, N.; Wittig, R.; Voelker, H.U.; Thiede, A.; Coy, J.F. Growth of human gastric cancer cells in nude mice is delayed by a ketogenic diet supplemented with omega-3 fatty acids and medium-chain triglycerides. BMC Cancer 2008, 8.

5.        Mavropoulos, J.C.; Buschemeyer, W.C.; Tewari, A.K.; Rokhfeld, D.; Pollak, M.; Zhao, Y.; Febbo, P.G.; Cohen, P.; Hwang, D.; Devi, G.; et al. The effects of varying dietary carbohydrate and fat content on survival in a murine LNCaP prostate cancer xenograft model. Cancer Prev. Res. (Phila). 2009, 2, 557–65.

6.        Rieger, J.; Bähr, O.; Maurer, G.D.; Hattingen, E.; Franz, K.; Brucker, D.; Walenta, S.; Kämmerer, U.; Coy, J.F.; Weller, M.; et al. ERGO: a pilot study of ketogenic diet in recurrent glioblastoma. Int. J. Oncol. 2014, 44, 1843–52.

7.        Maurer, G.D.; Brucker, D.P.; Bähr, O.; Harter, P.N.; Hattingen, E.; Walenta, S.; Mueller-Klieser, W.; Steinbach, J.P.; Rieger, J. Differential utilization of ketone bodies by neurons and glioma cell lines: a rationale for ketogenic diet as experimental glioma therapy. BMC Cancer 2011, 11, 315.

8.        Abdelwahab, M.G.; Fenton, K.E.; Preul, M.C.; Rho, J.M.; Lynch, A.; Stafford, P.; Scheck, A.C. The ketogenic diet is an effective adjuvant to radiation therapy for the treatment of malignant glioma. PLoS One 2012, 7, e36197.

9.        Hao, G.-W.; Chen, Y.-S.; He, D.-M.; Wang, H.-Y.; Wu, G.-H.; Zhang, B. Growth of human colon cancer cells in nude mice is delayed by ketogenic diet with or without omega-3 fatty acids and medium-chain triglycerides. Asian Pac. J. Cancer Prev. 2015, 16, 2061–8.

10.      Martuscello, R.T.; Vedam-Mai, V.; McCarthy, D.J.; Schmoll, M.E.; Jundi, M.A.; Louviere, C.D.; Griffith, B.G.; Skinner, C.L.; Suslov, O.; Deleyrolle, L.P.; et al. A Supplemented High-Fat Low-Carbohydrate Diet for the Treatment of Glioblastoma. Clin. Cancer Res. 2016, 22, 2482–95.

11.      Dang, M.T.; Wehrli, S.; Dang, C. V.; Curran, T. The ketogenic diet does not affect growth of Hedgehog pathway medulloblastoma in mice. PLoS One 2015, 10.

12.      Fontana, L.; Adelaiye, R.M.; Rastelli, A.L.; Miles, K.M.; Ciamporcero, E.; Longo, V.D.; Nguyen, H.; Vessella, R.; Pili, R. Dietary protein restriction inhibits tumor growth in human xenograft models of prostate and breast cancer. Oncotarget 2013, 4, 2451–2461.

13.      Hawrylewicz, E.J.; Huang, H.H.; Liu, J.M. Dietary protein, enhancement of N-nitrosomethylurea-induced mammary carcinogenesis, and their effect on hormone regulation in rats. Cancer Res. 1986, 46, 4395–9.

14.      Appleton, B.S.; Campbell, T.C. Inhibition of aflatoxin-initiated preneoplastic liver lesions by low dietary protein. Nutr. Cancer 1982, 3, 200–6.

15.      Appleton, B.S.; Campbell, T.C. Dietary protein intervention during the postdosing phase of aflatoxin B1-induced hepatic preneoplastic lesion development. J. Natl. Cancer Inst. 1983, 70, 547–9.

16.      Appleton, B.S.; Campbell, T.C. Effect of high and low dietary protein on the dosing and postdosing periods of aflatoxin B1-induced hepatic preneoplastic lesion development in the rat. Cancer Res. 1983, 43, 2150–4.

17.      Gao, X.; Sanderson, S.M.; Dai, Z.; Reid, M.A.; Cooper, D.E.; Lu, M.; Richie, J.P.; Ciccarella, A.; Calcagnotto, A.; Mikhael, P.G.; et al. Dietary methionine restriction targets one carbon metabolism in humans and produces broad therapeutic responses in cancer. bioRxiv 2019, 627364.

18.      Kokkinakis, D.M.; Schold, S.C.; Hori, H.; Nobori, T. Effect of long-term depletion of plasma methionine on the growth and survival of human brain tumor xenografts in athymic mice. Nutr. Cancer 1997, 29, 195–204.

19.      Latimer, M.N.; Freij, K.W.; Cleveland, B.M.; Biga, P.R. Physiological and molecular mechanisms of methionine restriction. Front. Endocrinol. (Lausanne). 2018, 9.

20.      Hoffman, R.M.; Kokkinakis, D.M.; Frenkel, E.P. Total Methionine Restriction Treatment of Cancer. Methods Mol. Biol. 2019, 1866, 163–171.

21.      O’Neill, A.M.; Burrington, C.M.; Gillaspie, E.A.; Lynch, D.T.; Horsman, M.J.; Greene, M.W. High-fat Western diet-induced obesity contributes to increased tumor growth in mouse models of human colon cancer. Nutr. Res. 2016, 36, 1325–1334.

22.      Lloyd, J.C.; Antonelli, J.A.; Phillips, T.E.; Masko, E.M.; Thomas, J.A.; Poulton, S.H.M.; Pollack, M.; Freedland, S.J. Effect of Isocaloric Low Fat Diet on Prostate Cancer Xenograft Progression in a Hormone Deprivation Model. J. Urol. 2010, 183, 1619–1624.

23.      Hopkins, B.D.; Pauli, C.; Xing, D.; Wang, D.G.; Li, X.; Wu, D.; Amadiume, S.C.; Goncalves, M.D.; Hodakoski, C.; Lundquist, M.R.; et al. Suppression of insulin feedback enhances the efficacy of PI3K inhibitors. Nature 2018, 560, 499–503.

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3 Comments

    1. No you are right.

      Martuscello’s MCT group had a 5% higher protein as % of macronutrients than control (21% vs 26%), if my calculations are correct. Then, they may have consumed more food, if the weight changes are any indication. This may have accentuated the higher protein intake in the MCT group, above what the macronutrient % would indicate. So the protein AND calorie intake was probably very substantially higher in the MCT group in Martuscello.

      But it’s the only study where protein was anywhere near as high or higher. That’s what I meant by “matched”, though I’m not using that definition very precisely. I’m focused on the protein match. I will edit the post to make this clear. Thank you for the feedback.

      1. Kevin, in fact to make a fair comparison between high carb and high fat, there should be more protein in the high fat diet, because high fat diet increases your protein needs: https://www.ncbi.nlm.nih.gov/pubmed/31083047.

        Btw, regarding current polemic on Twitter, i think those who have been physically injured by low carb charlatans should try to sue them. But at the same time, we’ve to preserve free speech, even if it’s abused by the charlatans.

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