Unexpected chronic lymphocytic leukemia B cell activation by bisphosphonates.

Chronic lymphocytic leukemia (CLL) is a disease of the elderly, often presenting comorbidities like osteoporosis and requiring, in a relevant proportion of cases, treatment with bisphosphonates (BPs). This class of drugs was shown in preclinical investigations to also possess anticancer properties. We started an in vitro study of the effects of BPs on CLL B cells activated by microenvironment-mimicking stimuli and observed that, depending on drug concentration, hormetic effects were induced on the leukemic cells. Higher doses induced cytotoxicity whereas at lower concentrations, more likely occurring in vivo, the drugs generated a protective effect from spontaneous and chemotherapy-induced apoptosis, and augmented CLL B cell activation/proliferation. This CLL-activation effect promoted by the BPs was associated with markers of poor CLL prognosis and required the presence of bystander stromal cells. Functional experiments suggested that this phenomenon involves the release of soluble factors and is increased by cellular contact between stroma and CLL B cells. Since CLL patients often present comorbidities such as osteoporosis and considering the diverse outcomes in both CLL disease progression and CLL response to treatment among patients, illustrating this phenomenon holds potential significance in driving additional investigations.

Mandatory role of endoplasmic reticulum and its pentose phosphate shunt in the myocardial defense mechanisms against the redox stress induced by anthracyclines.

Anthracyclines' cardiotoxicity involves an accelerated generation of reactive oxygen species. This oxidative damage has been found to accelerate the expression of hexose-6P-dehydrogenase (H6PD), that channels glucose-6-phosphate (G6P) through the pentose phosphate pathway (PPP) confined within the endoplasmic/sarcoplasmic reticulum (SR). To verify the role of SR-PPP in the defense mechanisms activated by doxorubicin (DXR) in cardiomyocytes, we tested the effect of this drug in H6PD knockout mice (H6PD-/-). Twenty-eight wildtype (WT) and 32 H6PD-/- mice were divided into four groups to be treated with intraperitoneal administration of saline (untreated) or DXR (8 mg/Kg once a week for 3 weeks). One week thereafter, survivors underwent imaging of 18F-deoxyglucose (FDG) uptake and were sacrificed to evaluate the levels of H6PD, glucose-6P-dehydrogenase (G6PD), G6P transporter (G6PT), and malondialdehyde. The mRNA levels of SR Ca2+-ATPase 2 (Serca2) and ryanodine receptors 2 (RyR2) were evaluated and complemented with Hematoxylin/Eosin staining and transmission electron microscopy. During the treatment period, 1/14 DXR-WT and 12/18 DXR-H6PD-/- died. At microPET, DXR-H6PD-/- survivors displayed an increase in left ventricular size (p < 0.001) coupled with a decreased urinary output, suggesting a severe hemodynamic impairment. At ex vivo analysis, H6PD-/- condition was associated with an oxidative damage independent of treatment type. DXR increased H6PD expression only in WT mice, while G6PT abundance increased in both groups, mismatching a generalized decrease of G6PD levels. Switching-off SR-PPP impaired reticular accumulation of Ca2+ decelerating Serca2 expression and upregulating RyR2 mRNA level. It thus altered mitochondrial ultrastructure eventually resulting in a cardiomyocyte loss. The recognized vulnerability of SR to the anthracycline oxidative damage is counterbalanced by an acceleration of G6P flux through a PPP confined within the reticular lumen. The interplay of SR-PPP with the intracellular Ca2+ exchanges regulators in cardiomyocytes configure the reticular PPP as a potential new target for strategies aimed to decrease anthracycline toxicity.

Divergent Oxidative Stress in Normal Tissues and Inflammatory Cells in Hodgkin and Non-Hodgkin Lymphoma.

BACKGROUND: Previous studies reported mitochondrial and endoplasmic reticulum redox stress in peripheral blood mononucleated cells (PBMCs) of treatment-naïve Hodgkin lymphoma (HL) patients. Here, we assessed whether this response also applies to non-HL (NHL) patients, and whether the oxidative damage is a selective feature of PBMCs or, rather, also affects tissues not directly involved in the inflammatory response. METHODS: Isolated PBMCs of 28 HL, 9 diffuse large B cell lymphoma, 8 less aggressive-NHL, and 45 controls underwent flow cytometry to evaluate redox stress and uptake of the glucose analogue 2-NBDG. This analysis was complemented with the assay of malondialdehyde (MDA) levels and enzymatic activity of glucose-6P-dehydrogenase and hexose-6P-dehydrogenase (H6PD). In all lymphoma patients, 18F-fluoro-deoxyglucose uptake was estimated in the myocardium and skeletal muscles. RESULTS: Mitochondrial reactive oxygen species generation and MDA levels were increased only in HL patients as well as H6PD activity and 2-NBDG uptake. Similarly, myocardial FDG retention was higher in HL than in other groups as opposed to a similar tracer uptake in the skeletal muscle. CONCLUSIONS: Redox stress of PBMCs is more pronounced in HL with respect to both NHL groups. This phenomenon is coherent with an increased activity of H6PD that also extends to the myocardium.

Gene's expression underpinning the divergent predictive value of [18F]F-fluorodeoxyglucose and prostate-specific membrane antigen positron emission tomography in primary prostate cancer: a bioinformatic and experimental study.

BACKGROUND: Positron Emission Tomography (PET) imaging with Prostate-Specific Membrane Antigen (PSMA) and Fluorodeoxyglucose (FDG) represent promising biomarkers for risk-stratification of Prostate Cancer (PCa). We verified whether the expression of genes encoding for PSMA and enzymes regulating FDG cellular uptake are independent and additive prognosticators in PCa. METHODS: mRNA expression of genes involved in glucose metabolism and PSMA regulation obtained from primary PCa specimens were retrieved from open-source databases and analyzed using an integrative bioinformatics approach. Machine Learning (ML) techniques were used to create predictive Progression-Free Survival (PFS) models. Cellular models of primary PCa with different aggressiveness were used to compare [18F]F-PSMA-1007 and [18F]F-FDG uptake kinetics in vitro. Confocal microscopy, immunofluorescence staining, and quantification analyses were performed to assess the intracellular and cellular membrane PSMA expression. RESULTS: ML analyses identified a predictive functional network involving four glucose metabolism-related genes: ALDOB, CTH, PARP2, and SLC2A4. By contrast, FOLH1 expression (encoding for PSMA) did not provide any additive predictive value to the model. At a cellular level, the increase in proliferation rate and migratory potential by primary PCa cells was associated with enhanced FDG uptake and decreased PSMA retention (paralleled by the preferential intracellular localization). CONCLUSIONS: The overexpression of a functional network involving four glucose metabolism-related genes identifies a higher risk of disease progression since the earliest phases of PCa, in agreement with the acknowledged prognostic value of FDG PET imaging. By contrast, the prognostic value of PSMA PET imaging is independent of the expression of its encoding gene FOLH1. Instead, it is influenced by the protein docking to the cell membrane, regulating its accessibility to tracer binding.

Mitochondrial Generated Redox Stress Differently Affects the Endoplasmic Reticulum of Circulating Lymphocytes and Monocytes in Treatment-Naïve Hodgkin's Lymphoma.

BACKGROUND: The redox stress caused by Hodgkin's lymphoma (HL) also involves the peripheral blood mononucleated cells (PBMCs) even before chemotherapy. Here, we tested whether lymphocytes and monocytes show a different response to the increased mitochondrial generation of reactive oxygen species (ROS). METHODS: PBMCs, isolated from the blood of treatment-naïve HL patients and control subjects, underwent assessment of malondialdehyde content and enzymatic activity of both hexose- and glucose-6P dehydrogenase (H6PD and G6PD) as well as flow cytometric analysis of mitochondrial ROS content. These data were complemented by evaluating the uptake of the fluorescent glucose analogue 2-NBDG that is selectively stored within the endoplasmic reticulum (ER). RESULTS: Malondialdehyde content was increased in the whole population of HL PBMCs. The oxidative damage matched an increased activity of G6PD, and even more of H6PD, that trigger the cytosolic and ER pentose phosphate pathways, respectively. At flow cytometry, the number of recovered viable cells was selectively decreased in HL lymphocytes that also showed a more pronounced increase in mitochondrial ROS generation and 2-NBDG uptake, with respect to monocytes. CONCLUSIONS: PBMCs of HL patients display a selective mitochondrial and ER redox stress most evident in lymphocytes already before the exposure to chemotherapy toxicity.

Therapeutic efficacy of proton transport inhibitors alone or in combination with cisplatin in triple negative and hormone sensitive breast cancer models.

Triple negative breast cancers (TNBCs) are very aggressive and have a poor prognosis due to lack of efficacious therapies. The only effective treatment is chemotherapy that however is frequently hindered by the occurrence of drug resistance. We approached this problem in vitro and in vivo on a triple negative and a hormone sensitive breast cancer cell lines: 4T1 and TS/A. A main defense mechanism of tumors is the extrusion of intracellular protons derived from the metabolic shift to glycolysis, and necessary to maintain an intracellular pH compatible with life. The resulting acidic extracellular milieu bursts the malignant behavior of tumors and impairs chemotherapy. Therefore, we investigated the efficacy of combined therapies that associate cisplatin (Cis) with proton exchanger inhibitors, such as esomeprazole (ESO) and 5-(N-ethyl-N-isopropyl)amiloride (EIPA). Our results demonstrate that in the 4T1 triple negative model the combined therapy Cis plus EIPA is significantly more effective than the other treatments. Instead, in the TS/A tumor the best therapeutic result is obtained with ESO alone. Remarkably, in both 4T1 and TS/A tumors these treatments correlate with increase of CD8+  T lymphocytes and dendritic cells, and a dramatic reduction of M2 macrophages and other suppressor myeloid cells (MDSC) in the tumor infiltrates.

Fundamental Role of Pentose Phosphate Pathway within the Endoplasmic Reticulum in Glutamine Addiction of Triple-Negative Breast Cancer Cells.

Cancer utilization of large glutamine equivalents contributes to diverging glucose-6-P flux toward the pentose phosphate shunt (PPP) to feed the building blocks and the antioxidant responses of rapidly proliferating cells. In addition to the well-acknowledged cytosolic pathway, cancer cells also run a largely independent PPP, triggered by hexose-6P-dehydrogenase within the endoplasmic reticulum (ER), whose activity is mandatory for the integrity of ER-mitochondria networking. To verify whether this reticular metabolism is dependent on glutamine levels, we complemented the metabolomic characterization of intermediates of the glucose metabolism and tricarboxylic acid cycle with the estimation of proliferating activity, energy metabolism, redox damage, and mitochondrial function in two breast cancer cell lines. ER-PPP activity and its determinants were estimated by the ER accumulation of glucose analogs. Glutamine shortage decreased the proliferation rate despite increased ATP and NADH levels. It depleted NADPH reductive power and increased malondialdehyde content despite a marked increase in glucose-6P-dehydrogenase. This paradox was explained by the deceleration of ER-PPP favored by the decrease in hexose-6P-dehydrogenase expression coupled with the opposite response of its competitor enzyme glucose-6P-phosphatase. The decreased ER-PPP activity eventually hampered mitochondrial function and calcium exchanges. These data configure the ER-PPP as a powerful, unrecognized regulator of cancer cell metabolism and proliferation.

Metformin and Cancer Glucose Metabolism: At the Bench or at the Bedside?

Several studies reported that metformin, the most widely used drug for type 2 diabetes, might affect cancer aggressiveness. The biguanide seems to directly impair cancer energy asset, with the consequent phosphorylation of AMP-activated protein kinase (AMPK) inhibiting cell proliferation and tumor growth. This action is most often attributed to a well-documented blockage of oxidative phosphorylation (OXPHOS) caused by a direct interference of metformin on Complex I function. Nevertheless, several other pleiotropic actions seem to contribute to the anticancer potential of this biguanide. In particular, in vitro and in vivo experimental studies recently documented that metformin selectively inhibits the uptake of 2-[18F]-Fluoro-2-Deoxy-D-Glucose (FDG), via an impaired catalytic function of the enzyme hexose-6P-dehydrogenase (H6PD). H6PD triggers a still largely uncharacterized pentose-phosphate pathway (PPP) within the endoplasmic reticulum (ER) that has been found to play a pivotal role in feeding the NADPH reductive power for both cellular proliferation and antioxidant responses. Regardless of its exploitability in the clinical setting, this metformin action might configure the ER metabolism as a potential target for innovative therapeutic strategies in patients with solid cancers and potentially modifies the current interpretative model of FDG uptake, attributing PET/CT capability to predict cancer aggressiveness to the activation of H6PD catalytic function.

Myocardial Metabolic Response Predicts Chemotherapy Curative Potential on Hodgkin Lymphoma: A Proof-of-Concept Study.

Genome sharing between cancer and normal tissues might imply a similar susceptibility to chemotherapy toxicity. The present study aimed to investigate whether curative potential of doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD) is predicted by the metabolic response of normal tissues in patients with Hodgkin lymphoma (HL). METHODS: According to current guidelines, 86 patients with advanced-stage (IIB-IVB) HL, prospectively enrolled in the HD0607 trial (NCT00795613), underwent 18 F-fluorodeoyglucose PET/CT imaging at diagnosis and, at interim, after two ABVD courses, to decide regimen maintenance or its escalation. In both scans, myocardial FDG uptake was binarized according to its median value. Death and disease relapse were recorded to estimate progression-free survival (PFS) during a follow-up with median duration of 43.8 months (range 6.97-60). RESULTS: Four patients (4.6%) died, while six experienced disease relapse (7%). Complete switch-off of cancer lesions and cardiac lighting predicted a favorable outcome at Kaplan-Mayer analyses. The independent nature and additive predictive value of their risk prediction were confirmed by the multivariate Cox regression analysis. CONCLUSION: Susceptibility of HL lesions to chemotherapy is at least partially determined by factors featuring the host who developed it.

The role of endoplasmic reticulum in in vivo cancer FDG kinetics.

A recent result obtained by means of an in vitro experiment with cancer cultured cells has configured the endoplasmic reticulum as the preferential site for the accumulation of 2-deoxy-2-[18F]fluoro-D-glucose (FDG). Such a result is coherent with cell biochemistry and is made more significant by the fact that the reticular accumulation rate of FDG is dependent upon extracellular glucose availability. The objective of the present paper is to confirm in vivo the result obtained in vitro concerning the crucial role played by the endoplasmic reticulum in FDG cancer metabolism. This study utilizes data acquired by means of a Positron Emission Tomography scanner for small animals in the case of CT26 models of cancer tissues. The recorded concentration images are interpreted within the framework of a three-compartment model for FDG kinetics, which explicitly assumes that the endoplasmic reticulum is the dephosphorylation site for FDG in cancer cells. The numerical reduction of the compartmental model is performed by means of a regularized Gauss-Newton algorithm for numerical optimization. This analysis shows that the proposed three-compartment model equals the performance of a standard Sokoloff's two-compartment system in fitting the data. However, it provides estimates of some of the parameters, such as the phosphorylation rate of FDG, more consistent with prior biochemical information. These results are made more solid from a computational viewpoint by proving the identifiability and by performing a sensitivity analysis of the proposed compartment model.

Genetic screening of children with marrow failure. The role of primary Immunodeficiencies.

The differential diagnosis of marrow failure (MF) is crucial in the diagnostic work-up, since genetic forms require specific care. We retrospectively studied all patients with single/multi-lineage MF evaluated in a single-center to identify the type and incidence of underlying molecular defects. The diepoxybutane test was used to screen Fanconi Anemia. Other congenital MFs have been searched using Sanger and/or Next Generation Sequencing analysis, depending on the available tools over the years. Between 2009-2019, 97 patients (aged 0-32 years-median 5) with single-lineage (29%) or multilineage (68%) MF were evaluated. Fifty-three (54%) and 28 (29%) were diagnosed with acquired and congenital MF, respectively. The remaining 16 (17%), with trilinear (n=9) and monolinear (n=7) MF, were found to have an underlying primary immunodeficiency (PID) and showed clinical and biochemical signs of immune-dysregulation in 10/16 (62%) and in 14/16 (87%) of cases, respectively. Clinical signs were also found in 22/53 (41%) and 8/28 (28%) patients with idiopathic and classical cMF, respectively. Eight out of 16 PIDs patients were successfully transplanted, four received immunosuppression, two did not require treatment, and the remaining two died. We show that patients with single/multi-lineage MF may have underlying PIDs in a considerable number of cases and that MF may represent a relevant clinical sign in patients with PIDs, thus widening their clinical phenotype. An accurate immunological work-up should be performed in all patients with MF, and PID-related genes should be considered when screening MF in order to identify disorders that may receive targeted treatments and/or appropriate conditioning regimens before transplant.

Novel PET Tracers in the Management of Cardiac Sarcoidosis.

Sarcoidosis is a systemic inflammatory disease of unknown etiology, pathologically characterized by non-caseating granulomas involving several organs and tissues. This pathological process can eventually affect the heart during his course leading to fibrosis associated with systolic dysfunction, conduction disturbance, and even sudden cardiac death. Due to this prognostic impact, diagnosis is crucial to optimize clinical management. The low sensitivity of endomyocardial biopsy and its invasive nature prevents its application as a first-line diagnostic approach. Thus, several efforts have been dedicated to the identification of advanced imaging tools for the diagnosis and monitoring of cardiac involvement in systemic sarcoidosis, including Positron Emission Tomography (PET). Starting from strengths and disadvantages of 18F-Fluorodeoxyglucose (18F-FDG) PET imaging, the present narrative review will summarize state of the art and future perspectives about radiotracers other than 18F-FDG of potential interest in the field of CS, including somatostatin receptor- ligands, proliferation markers and hypoxia displaying agents.

Two high-rate pentose-phosphate pathways in cancer cells.

The relevant role of pentose phosphate pathway (PPP) in cancer metabolic reprogramming has been usually outlined by studying glucose-6-phosphate dehydrogenase (G6PD). However, recent evidence suggests an unexpected role for a less characterized PPP, triggered by hexose-6-phosphate dehydrogenase (H6PD) within the endoplasmic reticulum (ER). Studying H6PD biological role in breast and lung cancer, here we show that gene silencing of this reticular enzyme decreases cell content of PPP intermediates and D-ribose, to a similar extent as G6PD silencing. Decrease in overall NADPH content and increase in cell oxidative status are also comparable. Finally, either gene silencing impairs at a similar degree cell proliferating activity. This unexpected response occurs despite the absence of any cross-interference between the expression of both G6PD and H6PD. Thus, overall cancer PPP reflects the contribution of two different pathways located in the cytosol and ER, respectively. Disregarding the reticular pathway might hamper our comprehension of PPP role in cancer cell biology.

The Elusive Link Between Cancer FDG Uptake and Glycolytic Flux Explains the Preserved Diagnostic Accuracy of PET/CT in Diabetes.

This study aims to verify in experimental models of hyperglycemia induced by streptozotocin (STZ-DM) to what degree the high competition between unlabeled glucose and metformin (MET) treatment might affect the accuracy of cancer FDG imaging. The study included 36 "control" and 36 "STZ-DM" Balb/c mice, undergoing intraperitoneal injection of saline or streptozotocin, respectively. Two-weeks later, mice were subcutaneously implanted with breast (4 T1) or colon (CT26) cancer cells and subdivided in three subgroups for treatment with water or with MET at 10 or 750 mg/Kg/day. Two weeks after, mice were submitted to micro-PET imaging. Enzymatic pathways and response to oxidative stress were evaluated in harvested tumors. Finally, competition by glucose, 2-deoxyglucose (2DG) and the fluorescent analog 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose (2-NBDG) on FDG uptake was studied in 4 T1 and CT26 cultured cells. STZ-DM slightly decreased cancer volume and FDG uptake rate (MRF). More importantly, it also abolished MET capability to decelerate lesion growth and MRF. This metabolic reprogramming closely agreed with the activity of hexose-6-phosphate dehydrogenase within the endoplasmic reticulum. Finally, co-incubation with 2DG virtually abolished FDG and 2-NBDG uptake within the endoplasmic reticulum in cultured cells. These data challenge the current dogma linking FDG uptake to glycolytic flux and introduce a new model to explain the relation between glucose analogue uptake and hexoses reticular metabolism. This selective fate of FDG contributes to the preserved sensitivity of PET imaging in oncology even in chronic moderate hyperglycemic conditions.

Increased myocardial 18F-FDG uptake as a marker of Doxorubicin-induced oxidative stress.

BACKGROUND: Oxidative stress and its interference on myocardial metabolism play a major role in Doxorubicin (DXR) cardiotoxic cascade. METHODS: Mice models of neuroblastoma (NB) were treated with 5 mg DXR/kg, either free (Free-DXR) or encapsulated in untargeted (SL[DXR]) or in NB-targeting Stealth Liposomes (pep-SL[DXR] and TP-pep-SL[DXR]). Control mice received saline. FDG-PET was performed at baseline (PET1) and 7 days after therapy (PET2). At PET2 Troponin-I and NT-proBNP were assessed. Explanted hearts underwent biochemical, histological, and immunohistochemical analyses. Finally, FDG uptake and glucose consumption were simultaneously measured in cultured H9c2 in the presence/absence of Free-DXR (1 µM). RESULTS: Free-DXR significantly enhanced the myocardial oxidative stress. Myocardial-SUV remained relatively stable in controls and mice treated with liposomal formulations, while it significantly increased at PET2 with respect to baseline in Free-DXR. At this timepoint, myocardial-SUV was directly correlated with both myocardial redox stress and hexose-6-phosphate-dehydrogenase (H6PD) enzymatic activity, which selectively sustain cellular anti-oxidant mechanisms. Intriguingly, in vitro, Free-DXR selectively increased FDG extraction fraction without altering the corresponding value for glucose. CONCLUSION: The direct correlation between cardiac FDG uptake and oxidative stress indexes supports the potential role of FDG-PET as an early biomarker of DXR oxidative damage.

G6Pase location in the endoplasmic reticulum: Implications on compartmental analysis of FDG uptake in cancer cells.

The favourable kinetics of 18F-fluoro-2-deoxyglucose (FDG) permits to depict cancer glucose consumption by a single evaluation of late tracer uptake. This standard procedure relies on the slow radioactivity loss, usually attributed to the limited tumour expression of G6P-phosphatase (G6Pase). However, this classical interpretation intrinsically represents an approximation since, as in all tissues, cancer G6Pase activity is remarkable and is confined to the endoplasmic reticulum (ER), whose lumen must be reached by phosphorylated FDG to explain its hydrolysis and radioactivity release. The present study tested the impact of G6Pase sequestration on the mathematical description of FDG trafficking and handling in cultured cancer cells. Our data show that accounting for tracer access to the ER configures this compartment as the preferential site of FDG accumulation. This is confirmed by the reticular localization of fluorescent FDG analogues. Remarkably enough, reticular accumulation rate of FDG is dependent upon extracellular glucose availability, thus configuring the same ER as a significant determinant of cancer glucose metabolism.

Concentration-dependent metabolic effects of metformin in healthy and Fanconi anemia lymphoblast cells.

Metformin (MET) is the drug of choice for patients with type 2 diabetes and has been proposed for use in cancer therapy and for treating other metabolic diseases. More than 14,000 studies have been published addressing the cellular mechanisms affected by MET. However, several in vitro studies have used concentrations of the drug 10-100-fold higher than the plasmatic concentration measured in patients. Here, we evaluated the biochemical, metabolic, and morphologic effects of various concentrations of MET. Moreover, we tested the effect of MET on Fanconi Anemia (FA) cells, a DNA repair genetic disease with defects in energetic and glucose metabolism, as well as on human promyelocytic leukemia (HL60) cell lines. We found that the response of wild-type cells to MET is concentration dependent. Low concentrations (15 and 150 µM) increase both oxidative phosphorylation and the oxidative stress response, acting on the AMPK/Sirt1 pathway, while the high concentration (1.5 mM) inhibits the respiratory chain, alters cell morphology, becoming toxic to the cells. In FA cells, MET was unable to correct the energetic/respiratory defect and did not improve the response to oxidative stress and DNA damage. By contrast, HL60 cells appear sensitive also at 150 µM. Our findings underline the importance of the MET concentration in evaluating the effect of this drug on cell metabolism and demonstrate that data obtained from in vitro experiments, that have used high concentrations of MET, cannot be readily translated into improving our understanding of the cellular effects of metformin when used in the clinical setting.

Doxorubicin Effect on Myocardial Metabolism as a Prerequisite for Subsequent Development of Cardiac Toxicity: A Translational 18F-FDG PET/CT Observation.

The present translational study aimed to verify whether serial 18F-FDG PET/CT predicts doxorubicin cardiotoxicity. Methods: Fifteen athymic mice were treated intravenously with saline (n = 5) or with 5 or 7.5 mg of doxorubicin per kilogram (n = 5 each) and underwent dynamic small-animal PET beforehand and afterward to estimate left ventricular (LV) metabolic rate of glucose (MRGlu). Thereafter, we retrospectively identified 69 patients who had been successfully treated with a regimen of doxorubicin, bleomycin, vinblastine, and dacarbazine for Hodgkin disease (HD) and had undergone 4 consecutive 18F-FDG PET/CT scans. Volumes of interest were drawn on LV myocardium to quantify mean SUV. All patients were subsequently interviewed by telephone (median follow-up, 30 mo); 36 of them agreed to undergo electrocardiography and transthoracic echocardiography. Results: In mice, LV MRGlu was 17.9 ± 4.4 nmol × min-1 × g-1 at baseline. Doxorubicin selectively and dose-dependently increased this value in the standard-dose (27.9 ± 9 nmol × min-1 × g-1, P < 0.05 vs. controls) and high-dose subgroups (37.2 ± 7.8 nmol × min-1 × g-1, P < 0.01 vs. controls, P < 0.05 vs. standard-dose). In HD patients, LV SUV showed a progressive increase during doxorubicin treatment that persisted at follow-up. New-onset cardiac abnormalities appeared in 11 of 36 patients (31%). In these subjects, pretherapy LV SUV was markedly lower with respect to the remaining patients (1.53 ± 0.9 vs. 3.34 ± 2.54, respectively, P < 0.01). Multivariate analysis confirmed the predictive value of baseline LV SUV for subsequent cardiac abnormalities. Conclusion: Doxorubicin dose-dependently increases LV MRGlu, particularly in the presence of low baseline 18F-FDG uptake. These results imply that low myocardial 18F-FDG uptake before the initiation of doxorubicin chemotherapy in HD patients may predict the development of chemotherapy-induced cardiotoxicity, suggesting that prospective clinical trials are warranted to test this hypothesis.

Defects in mitochondrial energetic function compels Fanconi Anaemia cells to glycolytic metabolism.

Energetic metabolism plays an essential role in the differentiation of haematopoietic stem cells (HSC). In Fanconi Anaemia (FA), DNA damage is accumulated during HSC differentiation, an event that is likely associated with bone marrow failure (BMF). One of the sources of the DNA damage is altered mitochondrial metabolism and an associated increment of oxidative stress. Recently, altered mitochondrial morphology and a deficit in the energetic activity in FA cells have been reported. Considering that mitochondria are the principal site of aerobic ATP production, we investigated FA metabolism in order to understand what pathways are able to compensate for this energy deficiency. In this work, we report that the impairment in mitochondrial oxidative phosphorylation (OXPHOS) in FA cells is countered by an increase in glycolytic flux. By contrast, glutaminolysis appears lower with respect to controls. Therefore, it is possible to conclude that in FA cells glycolysis represents the main pathway for producing energy, balancing the NADH/NAD+ ratio by the conversion of pyruvate to lactate. Finally, we show that a forced switch from glycolytic to OXPHOS metabolism increases FA cell oxidative stress. This could be the cause of the impoverishment in bone marrow HSC during exit from the homeostatic quiescent state. This is the first work that systematically explores FA energy metabolism, highlighting its flaws, and discusses the possible relationships between these defects and BMF.