Hyperpolarized Magnetic Resonance Spectroscopic Imaging Research Articles

Abstract 4156: Monitoring early chemotherapy response in murine pancreatic ductal adenocarcinoma using non-invasive hyperpolarized magnetic resonance spectroscopic imaging with 13C-pyruvate

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) is a rarely operable cancer with a very poor prognosis. Nearly 100% of affected patients receive one of two alternative first-line chemotherapies during the course of their disease: 1.) Gemcitabine-Abraxane (G/A) or 2.) FOLFIRINOX. Treatment success varies widely between the patients due to individual molecular, structural tumor and metabolic heterogeneity. In this project, we use hyperpolarized 13C-pyruvate magnetic resonance spectroscopic imaging (HP-MRSI) to non-invasively characterize early metabolic changes in murine endogenous PDAC in response to the mentioned chemotherapies. Methods: 18 KPC (Ptf1awt/cre(C)Kraswt/G12D(K)Trp53fl/fl(P)) mice with 74 distinct tumour nodules, visually identified based on T2-weighted anatomical imaging, were used in this study. The conversion of pyruvate to lactate was detected by HP-MRSI, performed on 7 T preclinical system using modified 3D balanced steady-state free precession (bSSFP) sequence on day 0 pre-therapy and on day 3 post-therapy. 100μg/g G (intraperitoneally) and 30μg/g A (intravenously) were applied twice on day 0 post-scan and day 2 (n=6). FOLFIRINOX (60μg/g 5-Fluoruracil, 27μg/g Irinotecan, 12.8μg/g Oxaliplatin, 60μg/g Leucovorin) was injected intravenously once on day 0 post-scan (n=6). Vehicle mice (n=6) were left untreated. Area under the curves (AUC) of lactate to pyruvate time-courses (AUCl/AUCp) were calculated. Primary murine PDAC cell lines were established from biopsies of 58 imaging region, sequenced using 3’RNA-Seq (Poly-ASeq) method and analysed using Dseq2 R package. Results: G/A and FOLFIRINOX-treated tumors exhibited heterogeneous metabolic responses in distinct nodules, while non-treated group tumors remained stable. A significant difference in metabolic changes (p = 0.01, ANOVA) from day 0 to day 3 was observed between FOLFIRINOX, G/A, and non-treated groups. Low glycolytic tumors showed a positive trend, more pronounced with FOLFIRINOX than G/A treatment. Metabolically active nodules with high AUCl/AUCp on day 0 reduced lactate turnover under therapy. These findings suggest that metabolic HP-MRSI can detect treatment-induced changes in tumor composition and subtype-specific epithelial/mesenchymal plasticity, correlating with the metabolic phenotype. Further investigation will include histology, subtype-specific biomarkers, transcriptional signatures, and regional transcriptome analysis to explain molecular changes in metabolism. Conclusions: We have applied a non-invasive approach for detection of metabolic changes under chemotherapy in murine PDAC. These findings will contribute in the long term to the establishment of predictive imaging biomarkers for PDAC. Citation Format: Irina Heid, Anna-Maria Schmidmüller, Simon Baller, Jason G. Skinner, Geoffrey J. Topping, Wolfgang Gottwald, Hussein Trabulssi, Rupert Öllinger, Katja Steiger, Roland Rad, Franz Schilling, Rickmer F. Braren. Monitoring early chemotherapy response in murine pancreatic ductal adenocarcinoma using non-invasive hyperpolarized magnetic resonance spectroscopic imaging with 13C-pyruvate [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4156.

Enhancing Cancer Diagnosis with Real-Time Feedback: Tumor Metabolism through Hyperpolarized 1-13C Pyruvate MRSI

This review article discusses the potential of hyperpolarized (HP) 13C magnetic resonance spectroscopic imaging (MRSI) as a noninvasive technique for identifying altered metabolism in various cancer types. Hyperpolarization significantly improves the signal-to-noise ratio for the identification of 13C-labeled metabolites, enabling dynamic and real-time imaging of the conversion of [1-13C] pyruvate to [1-13C] lactate and/or [1-13C] alanine. The technique has shown promise in identifying upregulated glycolysis in most cancers, as compared to normal cells, and detecting successful treatment responses at an earlier stage than multiparametric MRI in breast and prostate cancer patients. The review provides a concise overview of the applications of HP [1-13C] pyruvate MRSI in various cancer systems, highlighting its potential for use in preclinical and clinical investigations, precision medicine, and long-term studies of therapeutic response. The article also discusses emerging frontiers in the field, such as combining multiple metabolic imaging techniques with HP MRSI for a more comprehensive view of cancer metabolism, and leveraging artificial intelligence to develop real-time, actionable biomarkers for early detection, assessing aggressiveness, and interrogating the early efficacy of therapies.

Excess exogenous pyruvate inhibits lactate dehydrogenase activity in live cells in an MCT1-dependent manner

Cellular pyruvate is an essential metabolite at the crossroads of glycolysis and oxidative phosphorylation, capable of supporting fermentative glycolysis by reduction to lactate mediated by lactate dehydrogenase (LDH) among other functions. Several inherited diseases of mitochondrial metabolism impact extracellular (plasma) pyruvate concentrations, and [1-13C]pyruvate infusion is used in isotope-labeled metabolic tracing studies, including hyperpolarized magnetic resonance spectroscopic imaging. However, how these extracellular pyruvate sources impact intracellular metabolism is not clear. Herein, we examined the effects of excess exogenous pyruvate on intracellular LDH activity, extracellular acidification rates (ECARs) as a measure of lactate production, and hyperpolarized [1-13C]pyruvate-to-[1-13C]lactate conversion rates across a panel of tumor and normal cells. Combined LDH activity and LDHB/LDHA expression analysis intimated various heterotetrameric isoforms comprising LDHA and LDHB in tumor cells, not only canonical LDHA. Millimolar concentrations of exogenous pyruvate induced substrate inhibition of LDH activity in both enzymatic assays ex vivo and in live cells, abrogated glycolytic ECAR, and inhibited hyperpolarized [1-13C]pyruvate-to-[1-13C]lactate conversion rates in cellulo. Of importance, the extent of exogenous pyruvate-induced inhibition of LDH and glycolytic ECAR in live cells was highly dependent on pyruvate influx, functionally mediated by monocarboxylate transporter-1 localized to the plasma membrane. These data provided evidence that highly concentrated bolus injections of pyruvate in vivo may transiently inhibit LDH activity in a tissue type- and monocarboxylate transporter-1–dependent manner. Maintaining plasma pyruvate at submillimolar concentrations could potentially minimize transient metabolic perturbations, improve pyruvate therapy, and enhance quantification of metabolic studies, including hyperpolarized [1-13C]pyruvate magnetic resonance spectroscopic imaging and stable isotope tracer experiments.

Hyperpolarized 13C pyruvate magnetic resonance spectroscopy for in vivo metabolic phenotyping of rat HCC

The in vivo assessment of tissue metabolism represents a novel strategy for the evaluation of oncologic disease. Hepatocellular carcinoma (HCC) is a high-prevalence, high-mortality tumor entity often discovered at a late stage. Recent evidence indicates that survival differences depend on metabolic alterations in tumor tissue, with particular focus on glucose metabolism and lactate production. Here, we present an in vivo imaging technique for metabolic tumor phenotyping in rat models of HCC. Endogenous HCC was induced in Wistar rats by oral diethyl-nitrosamine administration. Peak lactate-to-alanine signal ratios (L/A) were assessed with hyperpolarized magnetic resonance spectroscopic imaging (HPMRSI) after [1-13C]pyruvate injection. Cell lines were derived from a subset of primary tumors, re-implanted in nude rats, and assessed in vivo with dynamic hyperpolarized magnetic resonance spectroscopy (HPMRS) after [1-13C]pyruvate injection and kinetic modelling of pyruvate metabolism, taking into account systemic lactate production and recirculation. For ex vivo validation, enzyme activity and metabolite concentrations were spectroscopically quantified in cell and tumor tissue extracts. Mean peak L/A was higher in endogenous HCC compared to non-tumorous tissue. Dynamic HPMRS revealed higher pyruvate-to-lactate conversion rates (kpl) and lactate signal in subcutaneous tumors derived from high L/A tumor cells, consistent with ex vivo measurements of higher lactate dehydrogenase (LDH) levels in these cells. In conclusion, HPMRS and HPMRSI reveal distinct tumor phenotypes corresponding to differences in glycolytic metabolism in HCC tumor tissue.

Magnetic resonance spectroscopy for the study of cns malignancies

Despite intensive research, brain tumors are amongst the malignancies with the worst prognosis; therefore, a prompt diagnosis and thoughtful assessment of the disease is required. The resistance of brain tumors to most forms of conventional therapy has led researchers to explore the underlying biology in search of new vulnerabilities and biomarkers. The unique metabolism of brain tumors represents one potential vulnerability and the basis for a system of classification. Profiling this aberrant metabolism requires a method to accurately measure and report differences in metabolite concentrations. Magnetic resonance-based techniques provide a framework for examining tumor tissue and the evolution of disease. Nuclear Magnetic Resonance (NMR) analysis of biofluids collected from patients suffering from brain cancer can provide biological information about disease status. In particular, urine and plasma can serve to monitor the evolution of disease through the changes observed in the metabolic profiles. Moreover, cerebrospinal fluid can be utilized as a direct reporter of cerebral activity since it carries the chemicals exchanged with the brain tissue and the tumor mass. Metabolic reprogramming has recently been included as one of the hallmarks of cancer. Accordingly, the metabolic rewiring experienced by these tumors to sustain rapid growth and proliferation can also serve as a potential therapeutic target. The combination of 13C tracing approaches with the utilization of different NMR spectral modalities has allowed investigations of the upregulation of glycolysis in the aggressive forms of brain tumors, including glioblastomas, and the discovery of the utilization of acetate as an alternative cellular fuel in brain metastasis and gliomas. One of the major contributions of magnetic resonance to the assessment of brain tumors has been the non-invasive determination of 2-hydroxyglutarate (2HG) in tumors harboring a mutation in isocitrate dehydrogenase 1 (IDH1). The mutational status of this enzyme already serves as a key feature in the clinical classification of brain neoplasia in routine clinical practice and pilot studies have established the use of in vivo magnetic resonance spectroscopy (MRS) for monitoring disease progression and treatment response in IDH mutant gliomas. However, the development of bespoke methods for 2HG detection by MRS has been required, and this has prevented the wider implementation of MRS methodology into the clinic. One of the main challenges for improving the management of the disease is to obtain an accurate insight into the response to treatment, so that the patient can be promptly diverted into a new therapy if resistant or maintained on the original therapy if responsive. The implementation of 13C hyperpolarized magnetic resonance spectroscopic imaging (MRSI) has allowed detection of changes in tumor metabolism associated with a treatment, and as such has been revealed as a remarkable tool for monitoring response to therapeutic strategies. In summary, the application of magnetic resonance-based methodologies to the diagnosis and management of brain tumor patients, in addition to its utilization in the investigation of its tumor-associated metabolic rewiring, is helping to unravel the biological basis of malignancies of the central nervous system.

Role of metabolic imaging in diagnosis of primary, metastatic, and recurrent prostate cancer.

The present review describes the current role of metabolic imaging techniques such as multiparametric MRI (mpMRI), magnetic resonance spectroscopic imaging (MRSI), hyperpolarized MRSI, and positron emission tomography (PET) in the diagnosis of primary prostate cancer, surveillance of low-grade disease, detection of metastases, and evaluation of biochemical recurrence after therapy. The natural history of prostate cancer ranges from indolent disease that is optimally monitored by active surveillance, to highly aggressive disease that can be lethal. Current diagnostic methods remain imperfect in noninvasively distinguishing between silent versus aggressive tumors. Hence, there is a high demand for noninvasive imaging techniques that offer insight into biological behavior of prostate cancer cells. Characterization of prostate cancer metabolism is a promising area to provide such insights. Metabolic imaging may allow for greater detection and ultimately characterization of tumor based on aggressiveness and spread. Hence, it has the potential to monitor tumor activity, predict prognostic outcomes, and guide individualized therapies.

Dynamic Imaging of LDH Inhibition in Tumors Reveals Rapid In Vivo Metabolic Rewiring and Vulnerability to Combination Therapy

SUMMARYThe reliance of many cancers on aerobic glycolysis has stimulated efforts to develop lactate dehydrogenase (LDH) inhibitors. However, despite significant efforts, LDH inhibitors (LDHi) with sufficient specificity and in vivo activity to determine whether LDH is a feasible drug target are lacking. We describe an LDHi with potent, on-target, in vivo activity. Using hyperpolarized magnetic resonance spectroscopic imaging (HP-MRSI), we demonstrate in vivo LDH inhibition in two glycolytic cancer models, MIA PaCa-2 and HT29, and we correlate depth and duration of LDH inhibition with direct anti-tumor activity. HP-MRSI also reveals a metabolic rewiring that occurs in vivo within 30 min of LDH inhibition, wherein pyruvate in a tumor is redirected toward mitochondrial metabolism. Using HP-MRSI, we show that inhibition of mitochondrial complex 1 rapidly redirects tumor pyruvate toward lactate. Inhibition of both mitochondrial complex 1 and LDH suppresses metabolic plasticity, causing metabolic quiescence in vitro and tumor growth inhibition in vivo.

Dynamic Imaging of LDH Inhibition in Tumors Reveals Rapid in vivo Metabolic Rewiring and Vulnerability to Combination Therapy

The reliance of many cancers on aerobic glycolysis has stimulated substantial efforts to develop inhibitors of enzymes in this pathway, including lactate dehydrogenase (LDH). However, despite significant efforts, there have been no reports to date of an LDH inhibitor (LDHi) with sufficient specificity and in vivo activity to determine whether LDH is a feasible drug target and if LDH inhibition alone will suppress growth of glycolytic tumors. Here we describe an LDHi with potent, on-target in vivo activity that has allowed us to address these questions. Using hyperpolarized magnetic resonance spectroscopic imaging (HP-MRSI) of [13C]pyruvate metabolism, we demonstrate dynamic in vivo LDH inhibition in two glycolytic cancer models, MIA PaCa-2 and HT29, and we correlate LDH inhibition with direct anti-tumor activity. However, HP-MRSI also reveals a metabolic rewiring that occurs within 30 min of LDH inhibition in vivo, wherein pyruvate flux in tumor xenografts is redirected toward mitochondrial metabolism, consistent with perturbation of the NADH/NAD+ redox couple. HP-MRSI also shows that in vivo inhibition of mitochondrial complex 1 inhibition rapidly redirects tumor pyruvate flux toward lactate and, importantly, inhibition of both mitochondrial complex 1 and LDH suppresses metabolic plasticity and drives these cancer cells into metabolic quiescence in vivo while causeing tumor stasis in vivo. These data highlight the importance of pyruvate flux in tumor metabolic plasticity in vivo, even in glycolytic models, and they demonstrate the utility of HP-MRSI in monitoring drug-dependent changes in tumor metabolism.

Abstract 4106: Hyperpolarized [1-13C]-pyruvate/lactate magnetic resonance spectroscopic imaging of prostate cancer in vivo predicts response to lactate dehydrogenase inhibition

Abstract Purpose: Non-invasive magnetic resonance spectroscopic imaging (MRSI) of hyperpolarized (HP) 13C-labeled pyruvate and its metabolite lactate is being used to monitor the metabolic flux in solid tumors. In this study, we evaluate the potential of MRSI of HP [1-13C]-pyruvate and [1-13C]-lactate, in prostate cancer as a predictive biomarker for targeting lactate dehydrogenase. Experimental Design: Two human prostate cancer cell lines (DU145 and PC3) were grown as xenografts. The conversion of pyruvate to lactate in xenografts was measured, after intravenous delivery of hyperpolarized [1-13C] pyruvic acid, by MRSI. Steady state metabolomic analysis of xenograft tumors was performed by mass spectrometry and steady state lactate levels were measured with proton (1H) MRS. Perfusion and oxygenation of xenografts were measured with electron paramagnetic resonance (EPR) imaging. Tumor growth was assessed after lactate dehydrogenase (LDH) inhibition with FX-11 (42 µg/mouse/day for 5 days x 2 weekly cycles). Lactate production, pyruvate uptake, extracellular acidification rates and oxygen consumption of the prostate cancer cell lines was analyzed in vitro. Protein levels of glycolysis regulators were assessed with immunoblotting. Results: DU145 tumors demonstrated an enhanced conversion of pyruvate to lactate with HP [1-13C] MRSI relative to PC3 tumors (21% higher 13C-lactate/pyruvate p<0.05). In addition, DU145 xenografts have a 42% (p<0.05) reduction in 13C-lactate/pyruvate after FX-11 treatment while PC3 xenografts demonstrate no sensitivity to FX-11 treatment. In correlation FX-11 significantly delayed DU145 tumor volume doubling time by 3.4 days (p<0.05). By comparison no difference was observed between DU145 and PC3 xenografts in steady state measures of lactate, oxygenation, or perfusion. The two cell lines also exhibited similar pyruvate uptake, lactate production, extracellular acidification and oxygen consumption rates and sensitivity to FX-11 in vitro. Difference in the expression of glycolysis regulators such as Hif-1α, LDHA, MCT1, MCT4, HK2 and PFKP were observed in vitro but did not correlate with HP [13C]-lactate/pyruvate MRSI results or FX-11 sensitivity. Conclusions: Hyperpolarized [1-13C]-pyruvate MRSI of prostate cancer xenografts predicted the efficacy of targeting LDH when steady state markers of glycolysis, in vivo and in vitro, did not. Citation Format: Bradley T. Scroggins, Masayuki Matsuo, Ayla O. White, Keita Saito, Jeeva P. Munasinghe, Carole Sourbier, Kazutoshi Yamamoto, Vivian Diaz, James B. Mitchell, Murali C. Krishna, Deborah E. Citrin. Hyperpolarized [1-13C]-pyruvate/lactate magnetic resonance spectroscopic imaging of prostate cancer in vivo predicts response to lactate dehydrogenase inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4106.

Hyperpolarized 13C MR metabolic imaging can detect neuroinflammation in vivo in a multiple sclerosis murine model

Proinflammatory mononuclear phagocytes (MPs) play a crucial role in the progression of multiple sclerosis (MS) and other neurodegenerative diseases. Despite advances in neuroimaging, there are currently limited available methods enabling noninvasive detection of MPs in vivo. Interestingly, upon activation and subsequent differentiation toward a proinflammatory phenotype MPs undergo metabolic reprogramming that results in increased glycolysis and production of lactate. Hyperpolarized (HP) 13C magnetic resonance spectroscopic imaging (MRSI) is a clinically translatable imaging method that allows noninvasive monitoring of metabolic pathways in real time. This method has proven highly useful to monitor the Warburg effect in cancer, through MR detection of increased HP [1-13C]pyruvate-to-lactate conversion. However, to date, this method has never been applied to the study of neuroinflammation. Here, we questioned the potential of 13C MRSI of HP [1-13C]pyruvate to monitor the presence of neuroinflammatory lesions in vivo in the cuprizone mouse model of MS. First, we demonstrated that 13C MRSI could detect a significant increase in HP [1-13C]pyruvate-to-lactate conversion, which was associated with a high density of proinflammatory MPs. We further demonstrated that the increase in HP [1-13C]lactate was likely mediated by pyruvate dehydrogenase kinase 1 up-regulation in activated MPs, resulting in regional pyruvate dehydrogenase inhibition. Altogether, our results demonstrate a potential for 13C MRSI of HP [1-13C]pyruvate as a neuroimaging method for assessment of inflammatory lesions. This approach could prove useful not only in MS but also in other neurological diseases presenting inflammatory components.

Abstract LB-184: Hyperpolarized 13C-pyruvate magnetic resonance spectroscopic imaging of prostate cancer tumor xenografts

Abstract Purpose: Non-invasive magnetic resonance spectroscopic imaging (MRSI) of hyperpolarized 13C-labeled pyruvate and its metabolite lactate is being used to profile the metabolism of solid tumors. During this imaging, the metabolism of a bolus of 13C-labeled pyruvate to lactate is observed in the imaged tumor. Thus, the concentrations of pyruvate may not represent the physiological steady state of the tumor. We hypothesize that in our model system hyperpolarized 13C-pyruvate/lactate MRSI detects the glycolytic potential of prostate tumor xenografts not the metabolic steady state and may be used to predict response of tumors to metabolic inhibitors. Materials/Methods: Two non-androgen responsive prostate cancer cell lines, PC3 and DU145, were characterized for steady state metabolic differences by in vitro cell culture. Metabolic profiling in vitro was conducted by analysis of the extracellular acidification rate (glycolysis) and the oxygen consumption rate (oxidative phosphorylation). MCT1 and LDHA expression was monitored by immunoblotting. Pyruvate uptake and lactate production were also assayed to compare common glycolytic markers between cell lines. In vivo analysis of PC3 and DU145 tumor xenografts was conducted by MRSI of hyperpolarized 13C-labeled pyruvate/lactate. The effects of LDHA inhibition (FX11) on tumor growth were monitored for both types of tumor xenografts. Results: In vitro metabolic analysis showed that PC3 and DU145 cells have similar extracellular acidification rates (109and 116 mpH/min, respectively, p>0.05) and oxygen consumption rates (233 and 203 pmol O2/min, respectively, p>0.05) suggesting that their metabolic profile is similar with regards to glycolysis and oxidative phosphorylation. In addition, MCT1 and LDHA expression was similar, pyruvate uptake was about twice as high in PC3 cells than in DU145 cells (p<0.05) and lactate production was similar under aerobic conditions (494 nmoles/mg and 506 nmoles/mg, respectively, p>0.05). In vivo analysis by hyperpolarized 13C-pyruvate MRSI revealed that the lactate/pyruvate ratio was 21% (p<0.05) higher in DU145 tumor xenografts than in PC3 tumors suggesting that in vivo DU145 tumors have higher glycolytic potential. Finally LDHA inhibition with FX11 delayed growth of DU145 tumor xenografts by 7 days compared to no growth delay seen in FX11 treated PC3 tumor xenografts (p<0.05) suggesting that DU145 tumors are dependent on glycolysis for growth. Conclusions: In vitro analysis of PC3 and DU145 cells suggests that there is little difference at steady state in many of the markers of glycolysis between these cell lines. However hyperpolarized MRSI data indicates that DU145 tumors have a higher glycolytic potential than PC3 tumors. Our data suggest that hyperpolarized 13C-pyruvate MRSI could be more useful in predicting the response of tumors to metabolic inhibitors of glycolysis than steady state analysis. Citation Format: Bradley T. Scroggins, Masayuki Matsuo, Jeeva P. Munasinghe, Ayla O. White, Carole Sourbier, Shingo Matsumoto, W. Marston Linehan, James B. Mitchell, Murali C. Krishna, Deborah E. Citrin. Hyperpolarized 13C-pyruvate magnetic resonance spectroscopic imaging of prostate cancer tumor xenografts. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-184.