Advances and challenges in measuring hepatic glucose uptake with FDG PET: implications for diabetes research.

The liver plays a crucial role in the control of glucose homeostasis and is therefore of great interest in the investigation of the development of type 2 diabetes. Hepatic glucose uptake (HGU) can be measured through positron emission tomography (PET) imaging with the tracer [18F]-2-fluoro-2-deoxy-D-glucose (FDG). HGU is dependent on many variables (e.g. plasma glucose, insulin and glucagon concentrations), and the metabolic state for HGU assessment should be chosen with care and coherence with the study question. In addition, as HGU is influenced by many factors, protocols and measurement conditions need to be standardised for reproducible results. This review provides insights into the protocols that are available for the measurement of HGU by FDG PET and discusses the current state of knowledge of HGU and its impairment in type 2 diabetes. Overall, a scanning modality that allows for the measurement of detailed kinetic information and influx rates (dynamic imaging) may be preferable to static imaging. The combination of FDG PET and insulin stimulation is crucial to measure tissue-specific insulin sensitivity. While the hyperinsulinaemic-euglycaemic clamp allows for standardised measurements under controlled blood glucose levels, some research questions might require a more physiological approach, such as oral glucose loading, with both advantages and complexities relating to fluctuations in blood glucose and insulin levels. The available approaches to address HGU hold great potential but await more systematic exploitation to improve our understanding of the mechanisms underlying metabolic diseases. Current findings from the investigation of HGU by FDG PET highlight the complex interplay between insulin resistance, hepatic glucose metabolism, NEFA levels and intrahepatic lipid accumulation in type 2 diabetes and obesity. Further research is needed to fully understand the underlying mechanisms and potential therapeutic targets for improving HGU in these conditions.

Dynamic total-body PET/CT imaging with reduced acquisition time shows acceptable performance in quantification of [18F]FDG tumor kinetic metrics.

PURPOSE: To investigate the feasibility of reducing the acquisition time for continuous dynamic positron emission tomography (PET) while retaining acceptable performance in quantifying kinetic metrics of 2-[18F]-fluoro-2-deoxy-D-glucose ([18F]FDG) in tumors. METHODS: In total, 78 oncological patients underwent total-body dynamic PET imaging for ≥ 60 min, with 8, 20, and 50 patients receiving full activity (3.7 MBq/kg), half activity (1.85 MBq/kg), and ultra-low activity (0.37 MBq/kg) of [18F]FDG, respectively. The dynamic data were divided into 21-, 30-, 45- and ≥ 60-min groups. The kinetic analysis involved model fitting to derive constant rates (VB, K1 to k3, and Ki) for both tumors and normal tissues, using both reversible and irreversible two-tissue-compartment models. One-way ANOVA with repeated measures or the Freidman test compared the kinetic metrics among groups, while the Deming regression assessed the correlation of kinetic metrics among groups. RESULTS: All kinetic metrics in the 30-min and 45-min groups were statistically comparable to those in the ≥ 60-min group. The relative differences between the 30-min and ≥ 60-min groups ranged from 12.3% ± 15.1% for K1 to 29.8% ± 30.0% for VB, and those between the 45-min and ≥ 60-min groups ranged from 7.5% ± 8.7% for Ki to 24.0% ± 24.3% for VB. However, this comparability was not observed between the 21-min and ≥ 60-min groups. The significance trend of these comparisons remained consistent across different models (reversible or irreversible), administrated activity levels, and partial volume corrections for lesions. Significant correlations in tumor kinetic metrics were identified between the 30-/45-min and ≥ 60-min groups, with Deming regression slopes > 0.813. In addition, the comparability of kinetic metrics between the 30-min and ≥ 60-min groups were established for normal tissues. CONCLUSION: The acquisition time for dynamic PET imaging can be reduced to 30 min without compromising the ability to reveal tumor kinetic metrics of [18F]FDG, using the total-body PET/CT system.

Exploration the feasibility and additional value of [18F]FDG/[68Ga]Ga-FAPI-04 dual-low-activity-tracer one-stop total-body PET imaging at 34 min post-injection of [68Ga]Ga-FAPI-04.

OBJECTIVES: To validate the feasibility of one-stop 2-[18F]fluoro-2-deoxy-D-glucose ([18F]FDG) and [68Ga]Ga-fibroblast activation protein inhibitor-04 ([68Ga]Ga-FAPI-04) dual-low-activity-tracer positron emission tomography/computed tomography (PET/CT) at 34 min post-injection of [68Ga]Ga-FAPI-04 and explore its additional value. METHODS: Thirty pairs of patients with suspected malignancies who underwent dual-tracer imaging were enrolled in this retrospective study. The images were reconstructed at 34-39 and 50-60 min after additional injection of [68Ga]Ga-FAPI-04 (in one-stop FDG-FAPI PET/CT, named PETFDG, PETD34-39, and PETD50-60; in the 2-day protocol, named PETFDG', PETF34-39, and PETF50-60, respectively). Tumour-to-normal ratios (TNR) of lesions in PETFDG, PETD34-39, and PETD50-60 and TNR of lesions in PETF34-39 and PETF50-60 were evaluated separately. To evaluate the potential added value of one-stop FDG-FAPI PET/CT over the 2-day protocol, TNRs of PETFDG, PETD34-39, and PETD50-60 were compared with PETF34-39. The lesion detectability of the two imaging protocols was evaluated by chi-square test. RESULTS: Comparing FAPI-weighted PET (PETD34-39 and PETD50-60) and single-tracer imaging (PETFDG) in one-stop FDG-FAPI PET/CT, TNRs of FAPI-weighted PET were higher than those of PETFDG. PETD34-39 and PETD50-60 showed similar performance in lesion detectability and TNRs (all P > 0.05). In the 2-day protocol, there are no statistically significant differences in TNRs of all lesions at PETF34-39 and PETF50-60. Comparing one-stop FDG-FAPI PET/CT with the 2-day protocol, TNRs of PETF34-39 were significantly higher than those of PETFDG but lower than those of PETD34-39 and PETD50-60. Lesion detectability in the one-stop FDG-FAPI PET/CT was higher than that in the 2-day protocol. The average radiation dose in one-stop FDG-FAPI PET/CT was significantly lower than that in the 2-day protocol (P<0.001). CONCLUSION: One-stop FDG-FAPI PET/CT at 34 min could provide sufficient information to meet clinical diagnosis and showed better lesion detectability than that in the 2-day protocol.

Validation of cardiac image-derived input functions for functional PET quantification.

PURPOSE: Functional PET (fPET) is a novel technique for studying dynamic changes in brain metabolism and neurotransmitter signaling. Accurate quantification of fPET relies on measuring the arterial input function (AIF), traditionally achieved through invasive arterial blood sampling. While non-invasive image-derived input functions (IDIF) offer an alternative, they suffer from limited spatial resolution and field of view. To overcome these issues, we developed and validated a scan protocol for brain fPET utilizing cardiac IDIF, aiming to mitigate known IDIF limitations. METHODS: Twenty healthy individuals underwent fPET/MR scans using [18F]FDG or 6-[18F]FDOPA, utilizing bed motion shuttling to capture cardiac IDIF and brain task-induced changes. Arterial and venous blood sampling was used to validate IDIFs. Participants performed a monetary incentive delay task. IDIFs from various blood pools and composites estimated from a linear fit over all IDIF blood pools (3VOI) and further supplemented with venous blood samples (3VOIVB) were compared to the AIF. Quantitative task-specific images from both tracers were compared to assess the performance of each input function to the gold standard. RESULTS: For both radiotracer cohorts, moderate to high agreement (r: 0.60-0.89) between IDIFs and AIF for both radiotracer cohorts was observed, with further improvement (r: 0.87-0.93) for composite IDIFs (3VOI and 3VOIVB). Both methods showed equivalent quantitative values and high agreement (r: 0.975-0.998) with AIF-derived measurements. CONCLUSION: Our proposed protocol enables accurate non-invasive estimation of the input function with full quantification of task-specific changes, addressing the limitations of IDIF for brain imaging by sampling larger blood pools over the thorax. These advancements increase applicability to any PET scanner and clinical research setting by reducing experimental complexity and increasing patient comfort.

Preclinical toxicity studies supporting 2DG for treatment of status epilepticus.

2-deoxy-D-glucose (2DG) has been proposed as a potential antiseizure treatment based on seizure suppressive actions in multiple acute and chronic seizure models, including models of status epilepticus (SE). Here we summarize recently completed preclinical toxicological studies of single doses of an intravenous formulation of 2DG supporting potential safety of 2DG for acute treatment of SE and acute repetitive seizures (ARS).

Physiologically Achievable Concentration of 2-Deoxy-D-Glucose Stimulates IFN-γ Secretion in Activated T Cells In Vitro.

2-deoxy-D-glucose (2DG) is a glycolysis and protein N-glycosylation inhibitor with promising anti-tumor and immunomodulatory effects. However, 2DG can also suppress T cell function, including IFN-γ secretion. Few human T cell studies have studied low-dose 2DG, which can increase IFN-γ in a Jurkat clone. We therefore investigated 2DG's effect on IFN-γ in activated human T cells from PBMCs, with 2DG treatment commenced either concurrently with activation or 48 h after activation. Concurrent 2DG treatment decreased IFN-γ secretion in a dose-dependent manner. However, 2DG treatment of pre-activated T cells had a hormetic effect on IFN-γ, with 0.15-0.6 mM 2DG (achievable in vivo) increasing and >2.4 mM 2DG reducing its secretion. In contrast, IL-2 levels declined monotonously with increasing 2DG concentration. Lower 2DG concentrations reduced PD-1 and increased CD69 expression regardless of treatment timing. The absence of increased T-bet or Eomes expression or IFNG transcription suggests another downstream mechanism. 2DG dose-dependently induced the unfolded protein response, suggesting a possible role in increased IFN-γ secretion, possibly by increasing the ER folding capacity for IFN-γ via increased chaperone expression. Overall, low-dose, short-term 2DG exposure could potentially improve the T cell anti-tumor response.

The Impact of Glycolysis and Its Inhibitors on the Immune Response to Inflammation and Autoimmunity.

Glucose metabolism is a crucial biological pathway maintaining the activation of extra- and intracellular signaling pathways involved in the immune response. Immune cell stimulation via various environmental factors results in their activation and metabolic reprogramming to aerobic glycolysis. Different immune cells exhibit cell-type-specific metabolic patterns when performing their biological functions. Numerous published studies have shed more light on the importance of metabolic reprogramming in the immune system. Moreover, this knowledge is crucial for revealing new ways to target inflammatory pathologic states, such as autoimmunity and hyperinflammation. Here, we discuss the role of glycolysis in immune cell activity in physiological and pathological conditions, and the potential use of inhibitors of glycolysis for disease treatment.

Differential role of oxidative stress in synaptic and nonsynaptic in vitro ictogenesis.

It is well established that dysfunctional glucose metabolism and in particular hypoglycemia can lead to hyperexcitability and exacerbate epileptic seizures. The precise mechanisms behind this form of hyperexcitability are still unresolved. The present study investigates to what extent oxidative stress can account for the acute proconvulsant effect of hypoglycemia. We used the glucose derivative 2-deoxy-d-glucose (2-DG) to mimic glucose deprivation in hippocampal slices during the extracellular recording of interictal-like (IED) and seizure-like (SLE) epileptic discharge in areas CA3 and CA1. After induction of IED in area CA3 by perfusion of Cs+ (3 mM), MK801 (10 µM), and bicuculline (10 µM), subsequent application of 2-DG (10 mM) resulted in the appearance of SLE in 78.3% of experiments. This effect was only observed in area CA3 and was reversibly blocked by tempol (2 mM), a scavenger of reactive oxygen species, in 60% of experiments. Preincubation with tempol reduced the incidence of 2-DG-induced SLE to 40%. Low-Mg2+-induced SLE in area CA3 and in the entorhinal cortex (EC) was also reduced by tempol. In contrast, to the above models, which depend on synaptic transmission, nonsynaptic epileptiform field bursts induced in area CA3 by a combination of Cs+ (5 mM) and Cd2+ (200 µM), or in area CA1 using the "low-Ca2+ model," was unaffected or even enhanced by tempol. These results indicate that oxidative stress significantly contributes to 2-DG-induced seizures in area CA3 and that the impact of oxidative stress differs between synaptic and nonsynaptic ictogenesis.NEW & NOTEWORTHY The main findings of the current study are that area CA3 but not area CA1 can support 2-DG-induced seizure activity, that oxidative stress significantly contributes to 2-DG-induced seizure activity in area CA3, and that the impact of oxidative stress differs between synaptic and nonsynaptic epileptiform activity. In in vitro models where ictogenesis depends on synaptic interactions, oxidative stress lowers the seizure threshold, whereas in nonsynaptic models seizure threshold is unchanged or even increased.

Controlling Herpes Simplex Virus-Induced Immunoinflammatory Lesions Using Metabolic Therapy: a Comparison of 2-Deoxy-d-Glucose with Metformin.

Herpes simplex virus (HSV) infection of the eye can result in a blinding immunoinflammatory lesion in the cornea called herpetic stromal keratitis (HSK). This lesion is orchestrated by T cells and can be reduced in magnitude by anti-inflammatory drugs and procedures that change the balance of cellular participants in lesions. This report evaluates the effect of drugs that cause metabolic reprogramming on lesion expression using two drugs that affect glucose metabolism: 2-deoxy-d-glucose (2DG) and metformin. Both drugs could limit HSK severity, but 2DG therapy could result in herpes encephalitis if used when replicating virus was still present. The reason metformin was a safer therapy was its lack of marked inhibitory effects on inflammatory cells particularly interferon-γ (IFN-γ)-producing Th1 and CD8 T cells in the trigeminal ganglion (TG), in which HSV latency is established and sustained. Additionally, whereas 2DG in TG cultures with established latency accelerated the termination of latency, this did not occur in the presence of metformin, likely because the inflammatory cells remained functional. Our results support the value of metabolic reprogramming to control viral immunoinflammatory lesions, but the approach used should be chosen with caution. IMPORTANCE Herpes simplex virus (HSV) infection of the eye is an example where damaging lesions are in part the consequence of a host response to the infection. Moreover, it was shown that changing the representation of cellular participants in the inflammatory reaction can minimize lesion severity. This report explores the value of metabolic reprogramming using two drugs that affect glucose metabolism to achieve cellular rebalancing. It showed that two drugs, 2-deoxy-d-glucose (2DG) and metformin, effectively diminished ocular lesion expression, but only metformin avoided the complication of HSV spreading to the central nervous system (CNS) and causing herpetic encephalitis. The report provides some mechanistic explanations for the findings.

2-Deoxy-D-glucose simultaneously targets glycolysis and Wnt/ß-catenin signaling to inhibit cervical cancer progression.

Cervical cancer is one of the most common female malignant tumors, with typical cancer metabolism characteristics of increased glycolysis flux and lactate accumulation. 2-Deoxy-D-glucose (2-DG) is a glycolysis inhibitor that acts on hexokinase, the first rate-limiting enzyme in the glycolysis pathway. In this research, we demonstrated that 2-DG effectively reduced glycolysis and impaired mitochondrial function in cervical cancer cell lines HeLa and SiHa. Cell function experiments revealed that 2-DG significantly inhibited cell growth, migration, and invasion, and induced G0/G1 phase arrest at non-cytotoxic concentrations. In addition, we found that 2-DG down-regulated Wingless-type (Wnt)/ß-catenin signaling. Mechanistically, 2-DG accelerated the degradation of ß-catenin protein, which resulted in the decrease of ß-catenin expression in both nucleus and cytoplasm. The Wnt agonist lithium chloride and ß-catenin overexpression vector could partially reverse the inhibition of malignant phenotype by 2-DG. These data suggested that 2-DG exerted its anti-cancer effects on cervical cancer by co-targeting glycolysis and Wnt/ß-catenin signaling. As expected, the combination of 2-DG and Wnt inhibitor synergistically inhibited cell growth. It is noteworthy that, down-regulation of Wnt/ß-catenin signaling also inhibited glycolysis, indicating a similar positive feedback regulation between glycolysis and Wnt/ß-catenin signaling. In conclusion, we investigated the molecular mechanism by which 2-DG inhibits the progression of cervical cancer in vitro, elucidated the interregulation between glycolysis and Wnt/ß-catenin signaling, and preliminarily explored the effect of combined targeting of glycolysis and Wnt/ß-catenin signaling on cell proliferation, which provides more possibilities for the formulation of subsequent clinical treatment strategies.

Glucose and mannose analogs inhibit KSHV replication by blocking N-glycosylation and inducing the unfolded protein response.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent for Kaposi's sarcoma (KS), an HIV/AIDS-associated malignancy. Effective treatments against KS remain to be developed. The sugar analog 2-deoxy- d-glucose (2-DG) is an anticancer agent that is well-tolerated and safe in patients and was recently demonstrated to be a potent antiviral, including KSHV and severe acute respiratory syndrome coronavirus 2. Because 2-DG inhibits glycolysis and N-glycosylation, identifying its molecular targets is challenging. Here we compare the antiviral effect of 2-DG with 2-fluoro-deoxy- d-glucose, a glycolysis inhibitor, and 2-deoxy-fluoro- d-mannose (2-DFM), a specific N-glycosylation inhibitor. At doses similar to those clinically achievable with 2-DG, the three drugs impair KSHV replication and virion production in iSLK.219 cells via downregulation of viral structural glycoprotein expression (K8.1 and gB), being 2-DFM the most potent KSHV inhibitor. Consistently with the higher potency of 2-DFM, we found that d-mannose rescues KSHV glycoprotein synthesis and virus production, indicating that inhibition of N-glycosylation is the main antiviral target using d-mannose competition experiments. Suppression of N-glycosylation by the sugar drugs triggers ER stress. It activates the host unfolded protein response (UPR), counteracting KSHV-induced inhibition of the protein kinase R-like endoplasmic reticulum kinase branch, particularly activating transcription factor 4 and C/EBP homologous protein expression. Finally, we demonstrate that sugar analogs induce autophagy (a prosurvival mechanism) and, thus, inhibit viral replication playing a protective role against KSHV-induced cell death, further supporting their direct antiviral effect and potential therapeutic use. Our work identifies inhibition of N-glycosylation leading to ER stress and UPR as an antienveloped virus target and sugar analogs such as 2-DG and the newly identified 2-DFM as antiviral drugs.

One-stop [18F]FDG and [68Ga]Ga-DOTA-FAPI-04 total-body PET/CT examination with dual-low activity: a feasibility study.

PURPOSE: Positron emission tomography/computed tomography (PET/CT) based on fibroblast activation protein inhibitors (FAPI) has shown complementary values to 2-[18F]-fluoro-2-deoxy-D-glucose ([18F]FDG) in cancer imaging. This study aimed to investigate the feasibility of a one-stop FDG-FAPI dual-tracer imaging protocol with dual-low activity for oncological imaging. METHODS: Nineteen patients with malignancies underwent one-stop [18F]FDG (0.37 MBq/kg) PET (PETFDG) and dual-tracer PET 30-40 and 50-60 min (hereafter, PETD30-40 and PETD50-60, respectively) after additional injection of [68Ga]Ga-DOTA-FAPI-04 (0.925 MBq/kg), with a single diagnostic CT to generate the PET/CT. The lesion detection rate and tumor-to-normal ratios (TNRs) of tracer uptake were compared between PETFDG/CT and PETD50-60/CT and between PETD50-60/CT and PETD30-40/CT. In addition, a visual scoring system was established to compare the lesion detectability. RESULTS: The dual-tracer PETD50-60 and PETD30-40/CT showed similar performance in detecting primary tumors but presented significantly higher lesion TNRs than PETFDG. Significantly, more metastases with higher TNRs were identified on PETD50-60 than PETFDG (491 vs. 261, P < 0.001). The dual-tracer PETD50-60 received significantly higher visual scores than single PETFDG (111 vs. 10) in demonstrating both primary tumors (12 vs. 2) and metastases (99 vs. 8). However, these differences were not significant between PETD50-60 and PETD30-40. These resulted in tumor upstaging in 44.4% patients taking PET/CT for initial assessment, and more recurrences (68 vs. 7) were identified in patients taking PET/CT for restaging, both on PETD50-60 and PETD30-40, compared to PETFDG. The reduced effective dosimetry per patient (26.2 ± 2.57 mSv) was equal to that of a single standard whole-body PET/CT. CONCLUSION: The one-stop dual-tracer dual-low-activity PET imaging protocol combines the strengths of [18F]FDG and [68Ga]Ga-DOTA-FAPI-04 with shorter duration and lesser radiation and is thus clinically applicable.

Reduced D2 /D3 Receptor Binding and Glucose Metabolism in a Macaque Model of Huntington's Disease.

BACKGROUND: Dopamine system dysfunction and altered glucose metabolism are implicated in Huntington's disease (HD), a neurological disease caused by mutant huntingtin (mHTT) expression. OBJECTIVE: The aim was to characterize alterations in cerebral dopamine D2 /D3 receptor density and glucose utilization in a newly developed AAV-mediated NHP model of HD that expresses mHTT throughout numerous brain regions. METHODS: Positron emission tomography (PET) imaging was performed using [18 F]fallypride to quantify D2 /D3 receptor density and 2-[18 F]fluoro-2-deoxy-d-glucose ([18 F]FDG) to measure cerebral glucose utilization in these HD macaques. RESULTS: Compared to controls, HD macaques showed significantly reduced dopamine D2 /D3 receptor densities in basal ganglia (P < 0.05). In addition, HD macaques displayed significant glucose hypometabolism throughout the cortico-basal ganglia network (P < 0.05). CONCLUSIONS: [18 F]Fallypride and [18 F]FDG are PET imaging biomarkers of mHTT-mediated disease progression that can be used as noninvasive outcome measures in future therapeutic studies with this AAV-mediated HD macaque model. © 2022 International Parkinson and Movement Disorder Society.

Directly Bound Deuterons Increase X-Nuclei Hyperpolarization using Dynamic Nuclear Polarization.

Deuterated 13 C sites in sugars (D-glucose and 2-deoxy-D-glucose) showed 6.3-to-17.5-fold higher solid-state dynamic nuclear polarization (DNP) levels than their respective protonated sites at 3.35T. This effect was found to be unrelated to the protonation of the bath. Deuterated 15 N in sites bound to exchangeable protons ([15 N2 ]urea) showed a 1.3-fold higher polarization than their respective protonated sites at the same magnetic field. This relatively smaller effect was attributed to incomplete deuteration of the 15 N sites due to the solvent mixture. For a 15 N site that is not bound to protons or deuterons ([15 N]nitrate), deuteration of the bath did not affect the polarization level. These findings suggest a phenomenon related to DNP of X-nuclei directly bound to deuteron(s) as opposed to proton(s). It appears that direct binding to deuterons increases the solid-state DNP polarization level of X-nuclei which are otherwise bound to protons.

2-Deoxy-D-glucose inhibits lymphocytic choriomeningitis virus propagation by targeting glycoprotein N-glycosylation.

BACKGROUND: Increased glucose uptake and utilization via aerobic glycolysis are among the most prominent hallmarks of tumor cell metabolism. Accumulating evidence suggests that similar metabolic changes are also triggered in many virus-infected cells. Viral propagation, like highly proliferative tumor cells, increases the demand for energy and macromolecular synthesis, leading to high bioenergetic and biosynthetic requirements. Although significant progress has been made in understanding the metabolic changes induced by viruses, the interaction between host cell metabolism and arenavirus infection remains unclear. Our study sheds light on these processes during lymphocytic choriomeningitis virus (LCMV) infection, a model representative of the Arenaviridae family. METHODS: The impact of LCMV on glucose metabolism in MRC-5 cells was studied using reverse transcription-quantitative PCR and biochemical assays. A focus-forming assay and western blot analysis were used to determine the effects of glucose deficiency and glycolysis inhibition on the production of infectious LCMV particles. RESULTS: Despite changes in the expression of glucose transporters and glycolytic enzymes, LCMV infection did not result in increased glucose uptake or lactate excretion. Accordingly, depriving LCMV-infected cells of extracellular glucose or inhibiting lactate production had no impact on viral propagation. However, treatment with the commonly used glycolytic inhibitor 2-deoxy-D-glucose (2-DG) profoundly reduced the production of infectious LCMV particles. This effect of 2-DG was further shown to be the result of suppressed N-linked glycosylation of the viral glycoprotein. CONCLUSIONS: Although our results showed that the LCMV life cycle is not dependent on glucose supply or utilization, they did confirm the importance of N-glycosylation of LCMV GP-C. 2-DG potently reduces LCMV propagation not by disrupting glycolytic flux but by inhibiting N-linked protein glycosylation. These findings highlight the potential for developing new, targeted antiviral therapies that could be relevant to a wider range of arenaviruses.

A closed, autologous bioprocess optimized for TCR-T cell therapies.

Autologous cell therapy has proven to be an effective treatment for hematological malignancies. Cell therapies for solid tumors are on the horizon, however the high cost and complexity of manufacturing these therapies remain a challenge. Routinely used open steps to transfer cells and reagents through unit operations further burden the workflow reducing efficiency and increasing the chance for human error. Here we describe a fully closed, autologous bioprocess generating engineered TCR-T cells. This bioprocess yielded 5-12 × 10e9 TCR-expressing T cells, transduced at low multiplicity of infections, within 7-10 days, and cells exhibited an enriched memory T-cell phenotype and enhanced metabolic fitness. It was demonstrated that activating, transducing, and expanding leukapheresed cells in a bioreactor without any T-cell or peripheral blood mononuclear cell enrichment steps had a high level of T-cell purity (~97%). Several critical process parameters of the bioreactor, including culturing at a high cell density (7e6 cells/mL), adjusting rocking agitations during phases of scale-up, lowering glycolysis through the addition of 2-deoxy- d-glucose, and modulating interleukin-2 levels, were investigated on their roles in regulating transduction efficiency, cell growth, and T-cell fitness such as T-cell memory phenotype and resistance to activation-induced cell death. The bioprocess described herein supports scale-out feasibility by enabling the processing of multiple patients' batches in parallel within a Grade C cleanroom.

Respiratory-gated PET imaging with reduced acquisition time for suspect malignancies: the first experience in application of total-body PET/CT.

OBJECTIVES: This study aimed to investigate the performance of respiratory-gating imaging with reduced acquisition time using the total-body positron emission tomography/computed tomography (PET/CT) scanner. METHODS: Imaging data of 71 patients with suspect malignancies who underwent total-body 2-[18F]-fluoro-2-deoxy-D-glucose PET/CT for 15 min with respiration recorded were analyzed. For each examination, four reconstructions were performed: Ungated-15, using all coincidences; Ungated-5, using data of the first 5 min; Gated-15 using all coincidences but with respiratory gating; and Gated-6 using data of the first 6 min with respiratory gating. Lesions were quantified and image quality was evaluated; both were compared between the four image sets. RESULTS: A total of 390 lesions were found in the thorax and upper abdomen. Lesion detectability was significantly higher in gated-15 (97.2%) than in ungated-15 (93.6%, p = 0.001) and ungated-5 (92.3%, p = 0.001), but comparable to Gated-6 (95.9%, p = 0.993). A total of 131 lesions were selected for quantitative analyses. Lesions in Gated-15 presented significantly larger standardized uptake values, tumor-to-liver ratio, and tumor-to-blood ratio, but smaller metabolic tumor volume, compared to those in Ungated-15 and Ungated-5 (all p < 0.001). These differences were more obvious in small lesions and in lesions from sites other than mediastinum/retroperitoneum. However, these indices were not significantly different between Gated-15 and Gated-6. Higher, but acceptable, image noise was identified in gated images than in ungated images. CONCLUSIONS: Respiratory-gating imaging with reduced scanning time using the total-body PET/CT scanner is superior to ungated imaging and can be used in the clinic. KEY POINTS: • In PET imaging, respiratory gating can improve lesion presentation and detectability but requires longer imaging time. • This single-center study showed that the total-body PET scanner allows respiratory-gated imaging with reduced and clinically acceptable scanning time.

Head-to-head intra-individual comparison of total-body 2-[18F]FDG PET/CT and digital PET/CT in patients with malignant tumor: how sensitive could it be?

OBJECTIVES: To comparatively evaluate the lesion-detecting ability of 2-[18F]FDG total-body PET/CT (TB PET/CT) and conventional digital PET/CT. METHODS: This study enrolled 67 patients (median age, 65 years; 24 female and 43 male patients) who underwent a TB PET/CT scan and a conventional digital PET/CT scan after a single 2-[18F]FDG injection (3.7 MBq/kg). Raw PET data for TB PET/CT were acquired over the course of 5 min, and images were reconstructed using data from the first 1, 2, 3, and 4 min and the entire 5 min (G1, G2, G3, G4, and G5, respectively). The conventional digital PET/CT scan acquired in 2-3 min per bed (G0). Two nuclear medicine physicians independently assessed subjective image quality using a 5-point Likert scale and recorded the number of 2-[18F]FDG-avid lesions. RESULTS: A total of 241 lesions (69 primary lesions; 32 liver, lung, and peritoneum metastases; and 140 regional lymph nodes) among 67 patients with various types of cancer were analyzed. The subjective image quality score and SNR (signal-to-noise ratio) increased gradually from G1 to G5, and these values were significantly higher than the values at G0 (all p < 0.05). Compared to conventional PET/CT, G4 and G5 of TB PET/CT detected an additional 15 lesions (2 primary lesions; 5 liver, lung, and peritoneum lesions; and 8 lymph node metastases). CONCLUSION: TB PET/CT was more sensitive than conventional whole-body PET/CT in detecting small (4.3 mm, maximum standardized uptake value (SUVmax) of 1.0) or low-uptake (tumor-to-liver ratio of 1.6, SUVmax of 4.1) lesions. CLINICAL RELEVANCE STATEMENT: This study explored the gain of the image quality and lesion detectability of TB PET/CT, compared to conventional PET/CT, and recommended the appropriate acquisition time for TB PET/CT in clinical practice with an ordinary 2-[18F] FDG dose. KEY POINTS: • TB PET/CT increases the effective sensitivity to approximately 40 times that of conventional PET scanners. • The subjective image quality score and signal-to-noise ratio of TB PET/CT from G1 to G5 were better than those of conventional PET/CT. • 2-[18F]FDG TB PET/CT with a 4-min acquisition time at a regular tracer dose detected an additional 15 lesions compared to conventional PET/CT.

Does high [18F]FDG uptake always mean poor prognosis? Colon cancer with high-level microsatellite instability is associated with high [18F]FDG uptake on PET/CT.

OBJECTIVES: High-level microsatellite instability (MSI-high) is generally associated with higher F-18 fluorodeoxyglucose ([18F]FDG) uptake than stable microsatellite (MSI-stable) tumors. However, MSI-high tumors have better prognosis, which is in contrast with general understanding that high [18F]FDG uptake correlates with poor prognosis. This study evaluated metastasis incidence with MSI status and [18F]FDG uptake. METHODS: We retrospectively reviewed 108 right-side colon cancer patients who underwent preoperative [18F]FDG PET/CT and postoperative MSI evaluations using a standard polymerase chain reaction at five Bethesda guidelines panel loci. The maximum standard uptake value (SUVmax), SUVmax tumor-to-liver ratio (TLR), metabolic tumor volume (MTV), and total lesion glycolysis (TLG) of the primary tumor were measured using SUV 2.5 cut-off threshold. Student's t-test or Mann-Whitney U test was performed for continuous variables, and χ2 test or Fisher's exact test was performed for categorical variables (p value of < 0.05 for statistical significance). Medical records were reviewed for metastasis incidence. RESULTS: Our study population had 66 MSI-stable and 42 MSI-high tumors. [18F]FDG uptake was higher in MSI-high tumors than MSI-stable tumors (TLR, median (Q1, Q3): 7.95 (6.06, 10.54) vs. 6.08 (4.09, 8.82), p = 0.021). Multivariable subgroup analysis demonstrated that higher [18F]FDG uptake was associated with higher risks of distant metastasis in MSI-stable tumors (SUVmax: p = 0.025, MTV: p = 0.008, TLG: p = 0.019) but not in MSI-high tumors. CONCLUSION: MSI-high colon cancer is associated with high [18F]FDG uptake, but unlike MSI-stable tumors, the degree of [18F]FDG uptake does not correlate with the rate of distant metastasis. CLINICAL RELEVANCE STATEMENT: MSI status should be considered during PET/CT assessment of colon cancer patients, as the degree of [18F]FDG uptake might not reflect metastatic potential in MSI-high tumors. KEY POINTS: • High-level microsatellite instability (MSI-high) tumor is a prognostic factor for distant metastasis. • MSI-high colon cancers had a tendency of demonstrating higher [18F]FDG uptake compared to MSI-stable tumors. • Although higher [18F]FDG uptake is known to represent higher risks of distant metastasis, the degree of [18F]FDG uptake in MSI-high tumors did not correlate with the rate at which distant metastasis occurred.

2-DG Re-Normalized IFN-γ Production in T Cells Excluding TEMRA Cells from Patients with Aplastic Anemia.

Aplastic anemia (AA) is a T cell immune mediated autoimmune disease in which cytokines, particularly IFN-γ are pathogenesis factors. Glucose metabolism is closely related to effector functions of activated T cells, such as IFN-γ production. The characteristics of glucose metabolism and whether interfering with glucose metabolism could affect T cells produce IFN-γ ability in AA patients remains unknown. In this study, we examined the characteristics of glucose metabolism in T cells from AA patients and the effects of the glucose metabolism inhibitor 2-deoxy-D-glucose (2-DG) on the ability of T cell production IFN-γ. Our data demonstrated abnormal glucose metabolism in stimulated T cells from AA patients, mainly reflected by increased glucose uptake and lactate secretion. In addition, EM and TEMRA cells exhibit higher glucose uptake in patients with AA compared with healthy individuals. Moreover, the frequency of IFN-γ+ was reduced by 2-DG in T cell from AA patients. Unexpectedly, 2-DG re-normalized the frequency of IFN-γ+ in other T cell subsets, except for in the TEMRA. In conclusion, our study reveals for the first time the existence of enhanced aerobic glycolysis in T cells from AA patients, and different T cell subsets exhibit different extent glucose uptake requirements. Aerobic glycolysis regulation may be a potential therapeutic strategy for aberrant T cell immunity. Moreover, TEMRA may have specific metabolic abnormalities, which should receive more attention in future targeted immune metabolism research.