In addition to their abnormally high proliferative rates, cancer cells universally demonstrate an abnormal metabolism that is characterized by an overutilization of glycolysis (GLY) relative to the more energy efficient mitochondrial oxidative phosphorylation (OXPHOS). Originally described over 80 years ago by Warburg, this altered metabolism, more recently termed metabolic reprogramming, is now viewed as a hallmark of cancer [1], and appears intimately tied to the proliferative state. The novel clinical implicationof these observations on tumor metabolism is that they suggest that a potential anticancer strategy lies in reverting this metabolism towards normal levels, i.e., forcing OXPHOS. Although this would not result in cell kill per se, it should result in stabilization of growth with minimal toxiity. However, although a number of targets exist for which active drugs can be directed, we contend that the major obstacle towards taking this to the clinic is the inability to measure cancer metabolism in the intact organism. The recent development of hyperpolarized 13C magnetic resonance spectroscopy (MRS) enables for the first time the real-time investigation of in vivo metabolism with more than a 10,000-fold signal increase over conventional 13C methods. Using 13C-labeled pyruvate (Pyr) as a substrate allows us to quantitatively follow the in vivo fate of pyruvate, which occupies a key nodal point in the metabolic pathway in which glucose is either converted to lactate (Lac;reflecting GLY) or acetyl CoA (generating bicarbonate [Bic] in the process;reflecting OXPHOS). With this technology, it is therefore possible to measure the 13C labeling of lactate and bicarbonate following the bolus injection of hyperpolarized [1-13C]-Pyr, thus permitting a Lac/Bic ratio to be calculated, which we propose to study as a marker of therapeutic response. Bevacizumab (BEV;Avastin(R)) is a monoclonal antibody (mab) that binds vascular endothelial growth factor (VEGF), thus inhibiting angiogenesis. It is widely used in a number of tumor types, including glioblastoma multiforme, the most malignant of the primary brain tumors. Although it can have dramatic initial effects, its duration tends to be relatively short-lived and associated with the development of refractory tumor progression. Although an intimate relationship between flow and metabolism is well documented, there has been little study of the impact of BEV on tumor metabolism. We have proposed that BEV acutely disrupts tumor metabolism at the tissue level, such as to force OXPHOS, and that this transient effect correlates with tumor stabilization [2]. By improving Bic detection to enable quantitation, we have observed a marked decrease in Lac/Bic ratio in transplanted glioblastoma tissue after anti-VEGF therapy, an effect that can be seen within three hours of administration, providing initial support for this counterintuitive hypothesis. Considering that despite extensive study into its antiangiogenic effects, neither a reliable early clinical marker of BEV effect nor o resistance development has been elucidated, our results offer a new and exciting direction for improving the impact of this very valuable oncotherapeutic. This project represents a collaboration between a group that has been on the cutting edge of this technology with the P.I., a clinician scientist who is familiar with clinical trials as well as laboratory models of brain cancer. Glioblastoma multiforme (GBM), the most commonly occurring primary brain tumor, is an excellent prototypical cancer with which to assess metabolic therapies because of its high rate of GLY and treatment refractoriness. However, it is important to note that the derived findings should apply to all cancer. Our experiments are designed in such a way that by the end of this funding period, we will have refined this technology so as to perform imaging with high resolution as well as to assess molecules such as glutamate which are "deeper" into the OXPHOS pathway (SA1), determine the time course and dose relationships of the BEV effect in transplanted glioblastoma (SA2), assess the impact of anti-VEGF therapy on "metabolic symbiosis" (SA3) and assess whether these findings extrapolate to a more clinically relevant model in which brain tumors develop "spontaneously" after exposure to a neurocarcinogen in utero (SA4). These experiments therefore should move us close to our ultimate goal of linking BEV's treatment impact with an optimal "lactate/bicarbonate ratio" that can be used clinically not only as a measure of therapeutic efficacy, but also as a therapeutic goal. The recent awarding of funds to purchase a clinical grade polarizer at Stanford should also us to accelerate the translation of findings from bench to bedside.

Public Health Relevance

Bevacizumab (Avastin(R)) is a widely used cancer therapeutic that, via binding vascular endothelial growth factor (VEGF), blocks tumor blood vessel formation (angiogenesis). This proposal is based on our hypothesis (which is supported by our Preliminary Data) that its acute, dramatic (albeit temporary) effects on cancer, specificall the most malignant primary brain tumor, results from a significant disruption of tumor metabolism in which the tissue is forced to utilize mitochondrial pathways for energy production, thus reversing the well-known Warburg effect. To assess this in the intact organism, we will use magnetic resonance spectroscopy (MRS) of hyperpolarized 13C-pyruvate, which enables measurement of lactate and bicarbonate production, reflecting glycolysis and oxidative phosphorylation, respectively. It is our ultimate goal to link bevacizumab's treatment effect with an optimal lactate/bicarbonate ratio that can be used clinically as a measure of effectiveness and therapeutic goal.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Medical Imaging Study Section (MEDI)
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Zhang, Huiming
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Stanford University
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United States
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