The benefits of mild ketosis
The study of Breitkreuz et al. shows that ketosis might not be necessary to improve the cachectic state of cancer patients. In recent years, however, more evidence has emerged from both animal and laboratory studies indicating that cancer patients could benefit further from a very low CHO KD. In their mouse models, Tisdale et al. already noted that the KD not only attenuated the cachectic effects of the tumor, but also that the tumors grew more slowly (although they did not attribute this to a direct anti-tumor effect of β-hydroxybutyrate). Tumor growth inhibition through a KD has now been established in many animal models, is supported by a few clinical case reports, and laboratory studies have begun to reveal the underlying molecular mechanisms.
In vitro studies
More than 30 years ago, Magee et al. were the first to show that treating transformed cells with various, albeit supra-physiological, concentrations of β-hydroxybutyrate causes a dose-dependent and reversible inhibition of cell proliferation [116]. Their interpretation of the results that ''...ketone bodies interfere with either glucose entry or glucose metabolism...'' has been confirmed and further specified by Fine et al., who connected the inhibition of glycolysis in the presence of abundant ketone bodies to the overexpression of uncoupling protein-2 (UCP-2), a mitochondrial defect occurring in many tumor cells [127]. In normal cells, abundant acetyl-CoA and citrate from the breakdown of fatty acids and ketone bodies would inhibit key enzymes of glycolysis to ensure stable ATP levels; in tumor cells, however, the same phenomenon would imply a decrease in ATP production if the compensatory ATP production in the mitochondria was impaired. For several colon and breast cancer cell lines, Fine et al. showed that the amount of ATP loss under treatment with acetoacetate was related to the level of UCP-2 expression.
Very recently, Maurer et al. demonstrated that glioma cells - although not negatively influenced by β-hydroxybutyrate - are not able to use this ketone body as a substitute for glucose when starved of the latter, contrary to benign neuronal cells [128]. This supports the hypothesis that under low glucose concentrations, ketone bodies could serve benign cells as a substitute for metabolic demands while offering no such benefit to malign cells.
Animal studies
To our knowledge, the first and - with a total of 303 rats and nine experiments - most extensive study of a KD in animals was conducted by van Ness van Alstyne and Beebe in 1913 [129]. Experiments were divided into two classes: in the first class, rats in the treatment arm were fed a CHO-free diet consisting of casein and lard for several weeks before plantation of a Buffalo sarcoma, while the control arm received either bread only or casein, lard and lactose. Rats on the CHO-free diet not only gained more weight than the controls, but also exhibited much less tumor growth and mortality rates, the differences being "... so striking as to leave no room for doubt that the diet was an important factor in enabling the rats to resist the tumor after growth had started." [and they used casein!] In a second class of experiments using either the slow-growing Jensen sarcoma or the aggressive Buffalo sarcoma, the rats were put on the CHO-free diet on the same day that the tumor was planted. This time, differences between the treatment and control groups were "... so slight that ... one is left in no doubt of the ineffectiveness of non-carbohydrate feeding begun at the time of tumor implantation." Interestingly, this parallels the observation of Fearon et al. that rats who started to receive a KD at the same day as tumor transplantation did not differ from controls in either body or tumor weight after 14 d [120]. In these rats, it was noted that despite persistent ketosis, blood glucose levels were not significantly lower than in controls which were also fed ad libitum. This stability of blood glucose, independent of ketosis, was subsequently confirmed in studies in which mice were fed ad libitum on a KD [84,114,121-123,130] although two studies reported a drop in blood glucose concentrations compared with the control group [116,131]. In the study of Magee et al., however, diet was presented as a liquid vegetable oil and energy intake was not monitored, allowing for the possibility that the animals underate voluntarily, in this way consuming a "caloric restricted KD" used in several experimental settings from the Seyfried lab [84,114,132], which was shown therein to be superior to the unrestricted KD in tumor growth control. That "caloric restriction" per se can hamper tumor growth has been impressively demonstrated already in 1942 by A. Tannenbaum in a series of comprehensive mouse models with different mouse strains and tumor induction types [133]. Throughout all experimental series, a strict restriction of food intake (impeding weight gain) several weeks before inducing tumorigenesis by application of 3,4 benzpyrene decreased the appearance rate and appearance time of tumors in the diet mice compared to the ad libitum controls. Notably, the calorie-restricted diet was composed of 53% CHOs compared to 69% in the control group. Despite a lack of data on blood glucose and ketone body levels, it could be speculated that the strict restriction of food per se (to 50-60% of the control group) induced a ketotic state and thus the ketones were - at least to some extend - responsible for the effects observed.
Animal studies that have investigated the effects of a KD on tumor progression and host survival
Concerning fat quality, Freedland et al. observed that a diet rich in corn oil might stimulate prostate cancer growth to a greater extent than one rich in saturated fat [134].A recent study suggests, however, that tumor growth inhibition neither depends on fat quality nor ketone body levels[131]. In this case, mice injected with either murine squamous cell carcinoma or human colorectal carcinoma cells received a low CHO, high-protein diet in which ~ 60 E% was derived from protein, 10-15 E% from CHO and ~ 25 E% from fat. No systemic ketosis was measured, yet tumors grew significantly less compared with a standard diet containing 55 E% from CHO and 22 E% from the same fat source. IGF1 levels and body weight remained stable, so these findings could not be attributed to one of these factors. There was, however, a significant drop in blood glucose, insulin and lactate levels, and a positive correlation between blood lactate as well as insulin levels and tumor growth was found. The study of Venkateskwaran et al. indicates that in prostate cancer insulin and/or IGF1 play major roles in driving tumor cell proliferation [97].
The diversity of these findings should not be surprising, given the variety of mice strains, tumor cell lines, diet composition and time of diet initiation relative to tumor planting. Instead, it seems remarkable that the same basic treatment, namely drastic restriction of CHOs, apparently induces anti-tumoral effects via different pathways. Thus, it may depend on the circumstances which variables - including blood glucose, insulin, lactate, IGF1, fat quality and ketone bodies - are the best predictors of and responsible for the anti-tumor effects of very low CHO diets.
Human studies
Until now, no randomized controlled trials have been conducted to evaluate the effects of a KD on tumor growth and patient survival. It has to be noted in general, however, that any dietary intervention requiring a dramatic change of life style makes randomized studies nearly impossible - however, even prospective cohort studies are missing. There is only anecdotal evidence that such a diet might be effective as a supportive treatment. One study investigated whether a high-fat diet (80% non-nitrogenous calories from fat) would inhibit tumor cell replication compared to a high-dextrose diet (100% non-nitrogenous calories from dextrose) in 27 patients with gastro-intestinal cancers [137]. Diets were administered parenterally and cell proliferation assessed using thymidine labeling index on tumor samples. After 14 days, the authors found a non-significant trend for impaired proliferation in the high-fat group. Whether ketosis was achieved with this regime was not evaluated, but blood glucose levels were comparable in both trial groups. A very recent pilot trial demonstrated the feasibility of a low CHO up to a ketogenic regimen implemented for 12 weeks in very advanced outpatient cancer patients. Notably, severe side effects were not observed, nearly all standard blood parameters improved and some measures of quality of life changed for the better [138]. The first attempt to treat cancer patients with a long-term controlled KD was reported by L. Nebeling in 1995 for two pediatric patients with astrocytoma [139]. The results of those two cases were very encouraging and the diet was described in detail in another publication [140]. Implementing a KD with additional calorie restriction in a female patient with glioblastoma multiforme clearly demonstrated that this intervention was able to stop tumor growth [132]. This was achieved, however, on the expense of a dramatic weigh loss of 20% over the intervention period, which is no option for the majority of metastatic cancer patients being in a catabolic state. A first clinical study applying a non-restricted KD for patients with glioblastoma (ERGO-study, NHI registration number NCT00575146), which was presented at the 2010 ASCO meeting [141], showed good feasibility and suggested some anti-tumor activity. The protocol of another clinical interventional trial (RECHARGE trial, NCT00444054) treating patients with metastatic cancer by a very low CHO diet was published in 2008 [142], and preliminary data from this study presented at the 2011 ASCO-meeting showed a clear correlation between disease stability or partial remission and high ketosis, independent of weight loss and unconscious caloric restriction of the patients [136]. While a randomized study for the treatment of prostate cancer patents applying the Atkins diet (NCT00932672) is currently recruiting patients at the Duke University, another trial posted at the clinical trials database (ClinicalTrials.gov) is not yet open for recruitment (NCT01092247). Very recently, two Phase I studies applying a ketogenic diet based on KetoCal® 4:1 started recruitment at the University of Iowa, intended to treat prostate cancer patients (KETOPAN, NCT01419483) and non-small cell lung cancer (KETOLUNG, NCT01419587). Thus, in the future, several data should be available to judge whether this kind of nutrition is useful as either a supportive or even therapeutic treatment option for cancer patients.
Is there a role for carbohydrate restriction in the prevention of cancer?
"Prevention of cancer" can refer to either the inhibition of carcinogenesis per se or - once that cells made the transition to malignancy - the sufficient delay of tumor growth, so that it remains undetected and asymptomatic during a subject's lifespan. There is evidence that even modest CHO restriction may influence both of these mechanisms positively through various pathways. The IGF1R-IR pathway has already been discussed: once a potentially carcinogenic somatic mutation has occurred, the probability for carcinogenesis of a cell that is borderline between apoptosis and malignancy might be raised by high levels of insulin and IGF1 in the micro-environment. Once a cell became malignant, high insulin and IGF1 levels might accelerate proliferation and progression towards a more aggressive, glycolytic phenotype. In rats treated with the carcinogen N-methyl-N-nitrosourea, it has been shown that lowering the CHO content of the diet from 60 E% to 40 E% with a simultaneous increase in protein was sufficient to lower postprandial insulin levels as well as decrease the appearance rate of tumors from (18.2 ± 1.3)%/wk to (12.9 ± 1.4)%/wk (p < 0.05), however with no statistically significant effect on tumor latency and weight measured after 10 wk [143]. Similarly, a recent study reported that NOP mice, which normally have a 70 - 80% chance of developing breast cancer over their lifetime due to genetic mutations, stayed tumor-free at 1 year of age when their calories from CHO were limited to 15%, while almost half of those on a 55% CHO diet developed tumors [131]. Notably, only 3 out of 11 mice in the 15% CHO group died with having a tumor compared to 7 out of 10 in the 55% CHO group; at death, significantly lower plasma insulin levels had been measured for the low CHO group. These results support the epidemiological [25,29,31,32] and in vitro [81,144] findings that high CHO diets, in particular those including high GI foods, promote mammary tumorigenesis via the sustained action of insulin.
Lower insulin levels may further increase the chance of intermittent ketosis, in particular if CHO restriction is combined with exercise, calorie restriction or intermittent fasting. Seyfried and Shelton [40] pointed out the possibility of ketone bodies to help in cancer prevention through their ability to protect the mitochondria from inflammation and ROS. Being more satiating than low-fat diets [145,146], a low CHO diet would make it easier to avoid caloric overconsumption or to implement intermittent fasting as an additional lifestyle change [147].
Avoidance of chronic inflammation
Another potential benefit of low CHO diets might lie in their influence upon inflammatory processes that take place within various tissues. Inflammation is a well-established driver of early tumorigenesis and accompanies most, if not all cancers [148]. Chronic, 'smouldering' inflammation can both cause and develop along with neoplasia. There is evidence that chronic intake of easily digestible CHOs is able to promote such an inflammatory state in leukocytes and endothelial cells [94]. In obese individuals [149] and healthy subjects who underwent eccentric exercise training [150], the inflammatory state was further augmented postprandially through a high CHO intake, but not through high-fat, low CHO meals in the latter study. Maybe more importantly, even moderate CHO restriction has been shown to effectively target several important markers of atherosclerosis and type II diabetes, both of which are associated with chronic inflammation [151-157]. Forsythe et al. showed that in overweight individuals with dyslipidemia a very low CHO diet had a more favorable effect than a low fat diet in reducing several markers of inflammation [158]. Given these findings, it can be hypothesized that a diet with a low GL positively affects cancer risk through reducing postprandial hyperglycemia and the associated inflammatory response.
In this context, it is important to note that a low CHO diet offers further possibilities to target inflammation through omission or inclusion of certain foods. Usually, CHO restriction is not only limited to avoiding sugar and other high-GI foods, but also to a reduced intake of grains. Grains can induce inflammation in susceptible individuals due to their content of omega-6 fatty acids, lectins and gluten [159,160]. In particular gluten might play a key role in the pathogenesis of auto-immune and inflammatory disorders and some malignant diseases. In the small intestine, gluten triggers the release of zonulin, a protein that regulates the tight junctions between epithelial cells and therefore intestinal, but also blood-brain barrier function. Recent evidence suggests that overstimulation of zonulin in susceptible individuals could dysregulate intercellular communication promoting tumorigenesis at specific organ sites [161].
Paleolithic-type diets, that by definition exclude grain products, have been shown to improve glycemic control and cardiovascular risk factors more effectively than typically recommended low-fat diets rich in whole grains [162]. These diets are not necessarily very low CHO diets, but focus on replacing high-GI modern foods with fruits and vegetables, in this way reducing the total GL. This brings us back to our initial perception of cancer as a disease of civilization that has been rare among hunter-gatherer societies until they adopted the Western lifestyle. Although there are certainly many factors contributing to this phenomenon, the evidence presented in this review suggests that reduction of the high CHO intake that accounts for typically > 50 E% in the Western diet may play its own important role in cancer prevention and outcome.
Conclusions
We summarize our main findings from the literature regarding the role of dietary CHO restriction in cancer development and outcome.
(i) Most, if not all, tumor cells have a high demand on glucose compared to benign cells of the same tissue and conduct glycolysis even in the presence of oxygen (the Warburg effect). In addition, many cancer cells express insulin receptors (IRs) and show hyperactivation of the IGF1R-IR pathway. Evidence exists that chronically elevated blood glucose, insulin and IGF1 levels facilitate tumorigenesis and worsen the outcome in cancer patients.
(ii) The involvement of the glucose-insulin axis may also explain the association of the metabolic syndrome with an increased risk for several cancers. CHO restriction has already been shown to exert favorable effects in patients with the metabolic syndrome. Epidemiological and anthropological studies indicate that restricting dietary CHOs could be beneficial in decreasing cancer risk.
(iii) Many cancer patients, in particular those with advanced stages of the disease, exhibit altered whole-body metabolism marked by increased plasma levels of inflammatory molecules, impaired glycogen synthesis, increased proteolysis and increased fat utilization in muscle tissue, increased lipolysis in adipose tissue and increased gluconeogenesis by the liver. High fat, low CHO diets aim at accounting for these metabolic alterations. Studies conducted so far have shown that such diets are safe and likely beneficial, in particular for advanced stage cancer patients.
(iv) CHO restriction mimics the metabolic state of calorie restriction or - in the case of KDs - fasting. The beneficial effects of calorie restriction and fasting on cancer risk and progression are well established. CHO restriction thus opens the possibility to target the same underlying mechanisms without the side-effects of hunger and weight loss.
(v) Some laboratory studies indicate a direct anti-tumor potential of ketone bodies. During the past years, a multitude of mouse studies indeed proved anti-tumor effects of KDs for various tumor types, and a few case reports and pre-clinical studies obtained promising results in cancer patients as well. Several registered clinical trials are going to investigate the case for a KD as a supportive therapeutic option in oncology.
