EPAS1-mutated paragangliomas associated with haemoglobin disorders.

We report a large series of 40 patients presenting EPAS1-mutated paraganglioma (PGL) in whom we investigated a cause underlying chronic hypoxia. Four patients suffered from hypoxaemic heart disease. In patients with available haemoglobin electrophoresis results, 59% presented with a haemoglobin disorder, including six with sickle cell disease, five with sickle cell trait and two with heterozygous haemoglobin C disease. Histological and transcriptomic characterization of EPAS1 tumours revealed increased angiogenesis and high similarities with pseudohypoxic PGLs caused by VHL gene mutations. Sickle haemoglobinopathy carriers could thus be at increased risk for developing EPAS1-PGLs, which should be taken into account in their management and surveillance.

Progress and challenges in experimental models for pheochromocytoma and paraganglioma.

Experimental models for pheochromocytoma and paraganglioma are needed for basic pathobiology research and for preclinical testing of drugs to improve treatment of patients with these tumors, especially patients with metastatic disease. The paucity of models reflects the rarity of the tumors, their slow growth, and their genetic complexity. While there are no human cell line or xenograft models that faithfully recapitulate the genotype or phenotype of these tumors, the past decade has shown progress in development and utilization of animal models, including a mouse and a rat model for SDH-deficient pheochromocytoma associated with germline Sdhb mutations. There are also innovative approaches to preclinical testing of potential treatments in primary cultures of human tumors. Challenges with these primary cultures include how to account for heterogeneous cell populations that will vary depending on the initial tumor dissociation and how to distinguish drug effects on neoplastic vs normal cells. The feasible duration for maintaining cultures must also be balanced against time required to reliably assess drug efficacy. Considerations potentially important for all in vitro studies include species differences, phenotype drift, changes that occur in the transition from tissue to cell culture, and the O2 concentration in which cultures are maintained.

New advances in endocrine hypertension: from genes to biomarkers.

Hypertension (HT) is a major cardiovascular risk factor that affects 10% to 40% of the general population in an age-dependent manner. Detection of secondary forms of HT is particularly important because it allows the targeted management of the underlying disease. Among hypertensive patients, the prevalence of endocrine HT reaches up to 10%. Adrenal diseases are the most frequent cause of endocrine HT and are associated with excess production of mineralocorticoids (mainly primary aldosteronism), glucocorticoids (Cushing syndrome), and catecholamines (pheochromocytoma). In addition, a few rare diseases directly affecting the action of mineralocorticoids and glucocorticoids in the kidney also lead to endocrine HT. Over the past years, genomic and genetic studies have allowed improving our knowledge on the molecular mechanisms of endocrine HT. Those discoveries have opened new opportunities to transfer knowledge to clinical practice for better diagnosis and specific treatment of affected subjects. In this review, we describe the physiology of adrenal hormone biosynthesis and action, the clinical and biochemical characteristics of different forms of endocrine HT, and their underlying genetic defects. We discuss the impact of these discoveries on diagnosis and management of patients, as well as new perspectives related to the use of new biomarkers for improved patient care.

SDHx mutation and pituitary adenoma: can in vivo 1H-MR spectroscopy unravel the link?

Germline mutations in genes encoding succinate dehydrogenase (SDH) are frequently involved in pheochromocytoma/paraganglioma (PPGL) development and were implicated in patients with the '3PAs' syndrome (associating pituitary adenoma (PA) and PPGL) or isolated PA. However, the causality link between SDHx mutation and PA remains difficult to establish, and in vivo tools for detecting hallmarks of SDH deficiency are scarce. Proton magnetic resonance spectroscopy (1H-MRS) can detect succinate in vivo as a biomarker of SDHx mutations in PGL. The objective of this study was to demonstrate the causality link between PA and SDH deficiency in vivo using 1H-MRS as a novel noninvasive tool for succinate detection in PA. Three SDHx-mutated patients suffering from a PPGL and a macroprolactinoma and one patient with an apparently sporadic non-functioning pituitary macroadenoma underwent MRI examination at 3 T. An optimized 1H-MRS semi-LASER sequence (TR = 2500 ms, TE = 144 ms) was employed for the detection of succinate in vivo. Succinate and choline-containing compounds were identified in the MR spectra as single resonances at 2.44 and 3.2 ppm, respectively. Choline compounds were detected in all the tumors (three PGL and four PAs), while a succinate peak was only observed in the three macroprolactinomas and the three PGL of SDHx-mutated patients, demonstrating SDH deficiency in these tumors. In conclusion, the detection of succinate by 1H-MRS as a hallmark of SDH deficiency in vivo is feasible in PA, laying the groundwork for a better understanding of the biological link between SDHx mutations and the development of these tumors.

Succinate Dehydrogenase, Succinate, and Superoxides: A Genetic, Epigenetic, Metabolic, Environmental Explosive Crossroad.

Research focused on succinate dehydrogenase (SDH) and its substrate, succinate, culminated in the 1950s accompanying the rapid development of research dedicated to bioenergetics and intermediary metabolism. This allowed researchers to uncover the implication of SDH in both the mitochondrial respiratory chain and the Krebs cycle. Nowadays, this theme is experiencing a real revival following the discovery of the role of SDH and succinate in a subset of tumors and cancers in humans. The aim of this review is to enlighten the many questions yet unanswered, ranging from fundamental to clinically oriented aspects, up to the danger of the current use of SDH as a target for a subclass of pesticides.

Persistent Properties of a Subpopulation of Cancer Cells Overexpressing the Hedgehog Receptor Patched.

Despite the development of new therapeutic strategies, cancer remains one of the leading causes of mortality worldwide. One of the current major challenges is the resistance of cancers to chemotherapy treatments inducing metastases and relapse of the tumor. The Hedgehog receptor Patched (Ptch1) is overexpressed in many types of cancers. We showed that Ptch1 contributes to the efflux of doxorubicin and plays an important role in the resistance to chemotherapy in adrenocortical carcinoma (ACC), a rare cancer which presents strong resistance to the standard of care chemotherapy treatment. In the present study, we isolated and characterized a subpopulation of the ACC cell line H295R in which Ptch1 is overexpressed and more present at the cell surface. This cell subpopulation is more resistant to doxorubicin, grows as spheroids, and has a greater capability of clonogenicity, migration, and invasion than the parental cells. Xenograft experiments performed in mice and in ovo showed that this cell subpopulation is more tumorigenic and metastatic than the parental cells. These results suggest that this cell subpopulation has cancer stem-like or persistent cell properties which were strengthened by RNA-seq. If present in tumors from ACC patients, these cells could be responsible for therapy resistance, relapse, and metastases.

[Succinate dehydrogenase in cancer]. / Rôle de la succinate déshydrogénase dans le cancer.

Succinate dehydrogenase (SDH) is a mitochondrial enzyme that participates in both the tricarboxylic acid cycle and the electron transport chain. Mutations in genes encoding SDH are responsible for a predisposition to pheochromocytomas and paragangliomas, and more rarely, to gastrointestinal stromal tumors or renal cell carcinomas. A decrease in SDH activity, not explained by genetics, has also been observed in more common cancers. One of the consequences of the inactivation of SDH is the excessive production of its substrate, succinate, which acts as an oncometabolite by promoting a pseudohypoxic status and an extensive epigenetic rearrangement. Understanding SDH-related oncogenesis now makes it possible to develop innovative diagnostic methods and to consider targeted therapies for the management of affected patients.

Title: Rôle de la succinate déshydrogénase dans le cancer. Abstract: La succinate déshydrogénase (SDH) est une enzyme mitochondriale qui participe au cycle de Krebs et à la chaîne respiratoire. Quand elles sont à l'origine de cancers, les mutations des gènes codant les différentes sous-unités de la SDH sont responsables d'une prédisposition aux phéochromocytomes et aux paragangliomes, et, plus rarement, aux tumeurs stromales gastro-intestinales ou au cancer du rein. Une diminution de l'activité de la SDH, non expliquée par la génétique, s'observe aussi dans certains cancers plus fréquents. Une des conséquences de l'inactivation de la SDH est la production excessive de son substrat, le succinate, qui joue un rôle d'oncométabolite en promouvant un statut pseudohypoxique et d'importants remaniements épigénétiques. La compréhension de l'oncogenèse liée à la succinate déshydrogénase permet aujourd'hui de développer des méthodes diagnostiques innovantes et d'envisager des thérapies ciblées pour la prise en charge des patients atteints.

Preclinical evaluation of targeted therapies in Sdhb-mutated tumors.

Therapies for metastatic SDHB-dependent pheochromocytoma and paraganglioma (PPGL) are limited and poorly efficient. New targeted therapies and identification of early non-invasive biomarkers of response are thus urgently needed for these patients. We characterized an in vivo allograft model of spontaneously immortalized murine chromaffin cells (imCC) with inactivation of the Sdhb gene by dynamic contrast-enhanced MRI (DCE-MRI) and 18FDG-PET. We evaluated the response to several therapies: IACS-010759 (mitochondrial respiratory chain complex I inhibitor), sunitinib (tyrosine kinase inhibitor with anti-angiogenic activity), talazoparib (poly ADP ribose polymerase (PARP) inhibitor) combined or not to temozolomide (alkylating agent), pharmacological inhibitors of HIF2a (PT2385 and PT2977 (belzutifan)) and molecular inactivation of HIF2a (imCC Sdhb-/- shHIF2a). Multimodal imaging was performed, including magnetic resonance spectroscopy (1H-MRS) to monitor the level of succinate in vivo. The allografted model of Sdhb-/- imCC reflected SDHB-deficient tumors, with increased angiogenesis and a particular avidity for 18FDG. After 14 days of treatment, IACS-010759, sunitinib and talazoparib at high doses allowed a significant reduction of the tumor volumes. In contrast to the tumor growth inhibition observed in Sdhb-/- shHIF2a imCC tumors, pharmacological inhibitors of HIF2a (PT2385 and belzutifan) showed no antitumor action in this model, alone or in combination with sunitinib. 1H-MRS, but not DCE-MRI, enabled the monitoring response to sunitinib, which was the best treatment in this study, promoting a decrease in succinate levels detected in vivo. This study paves the way for new therapeutic options and reveals a potential new early biomarker of response to treatment in SDHB-dependent PPGL.

Low-grade oncocytic renal tumor (LOT): mutations in mTOR pathway genes and low expression of FOXI1.

Low-grade oncocytic renal tumor (LOT) is an emerging provisional entity, described as rare solid renal oncocytic/eosinophilic tumor sharing diffuse CK7 and negative CD117 immunoprofile. The links between LOT and other eosinophilic chromophobe like-renal cell carcinomas (RCC) are currently discussed. We sequenced tumoral DNA with a next generation sequencing panel for kidney cancer and carried out immunohistochemical analyses with CK7, CD117, SDHB, 4EBP1-P, S6K-P, and FOXI1 antibodies in a series of ten cases of LOT (9 females, 1 male; mean age at surgery: 66 years, 42.3 to 83.4) retrospectively diagnosed from a cohort of 272 tumors initially classified as chromophobe RCC (CHRCC). All LOT were single, without known hereditary predisposition, classified stage pT1 (70%), pT2 (20%) or pT3a (10%). Morphological features were similar to previous descriptions and clinical behavior was indolent for the six cases with available follow-up. We identified genetic variations in mTOR pathway related genes in 80% of cases, MTOR (7 cases) or TSC1 (1 case). Expression of FOXI1 was absent in all cases. In 9 LOT, 4EBP1-P and S6K-P were overexpressed, suggesting mTOR pathway activation.Our data highlights the major role of mTOR pathway in tumorigenesis of LOT mostly due to activating MTOR gene variations. Absence of FOXI1 expression is a strong argument to distinguish LOT from eosinophilic CHRCC and to bring them closer to other recently described FOXI1 negative eosinophilic-CHRCC like with MTOR/TSC mutations. Altogether, our data argue to consider LOT as a distinct entity with a favorable clinical outcome. However, in case of metastasis, an accurate diagnosis of LOT would be essential for the patient's management and could allow targeted therapy.

International initiative for a curated SDHB variant database improving the diagnosis of hereditary paraganglioma and pheochromocytoma.

BACKGROUND: SDHB is one of the major genes predisposing to paraganglioma/pheochromocytoma (PPGL). Identifying pathogenic SDHB variants in patients with PPGL is essential to the management of patients and relatives due to the increased risk of recurrences, metastases and the emergence of non-PPGL tumours. In this context, the 'NGS and PPGL (NGSnPPGL) Study Group' initiated an international effort to collect, annotate and classify SDHB variants and to provide an accurate, expert-curated and freely available SDHB variant database. METHODS: A total of 223 distinct SDHB variants from 737 patients were collected worldwide. Using multiple criteria, each variant was first classified according to a 5-tier grouping based on American College of Medical Genetics and NGSnPPGL standardised recommendations and was then manually reviewed by a panel of experts in the field. RESULTS: This multistep process resulted in 23 benign/likely benign, 149 pathogenic/likely pathogenic variants and 51 variants of unknown significance (VUS). Expert curation reduced by half the number of variants initially classified as VUS. Variant classifications are publicly accessible via the Leiden Open Variation Database system (https://databases.lovd.nl/shared/genes/SDHB). CONCLUSION: This international initiative by a panel of experts allowed us to establish a consensus classification for 223 SDHB variants that should be used as a routine tool by geneticists in charge of PPGL laboratory diagnosis. This accurate classification of SDHB genetic variants will help to clarify the diagnosis of hereditary PPGL and to improve the clinical care of patients and relatives with PPGL.

Loss of SDHB Promotes Dysregulated Iron Homeostasis, Oxidative Stress, and Sensitivity to Ascorbate.

Succinate dehydrogenase is a key enzyme in the tricarboxylic acid cycle and the electron transport chain. All four subunits of succinate dehydrogenase are tumor suppressor genes predisposing to paraganglioma, but only mutations in the SDHB subunit are associated with increased risk of metastasis. Here we generated an Sdhd knockout chromaffin cell line and compared it with Sdhb-deficient cells. Both cell types exhibited similar SDH loss of function, metabolic adaptation, and succinate accumulation. In contrast, Sdhb-/- cells showed hallmarks of mesenchymal transition associated with increased DNA hypermethylation and a stronger pseudo-hypoxic phenotype compared with Sdhd-/- cells. Loss of SDHB specifically led to increased oxidative stress associated with dysregulated iron and copper homeostasis in the absence of NRF2 activation. High-dose ascorbate exacerbated the increase in mitochondrial reactive oxygen species, leading to cell death in Sdhb-/- cells. These data establish a mechanism linking oxidative stress to iron homeostasis that specifically occurs in Sdhb-deficient cells and may promote metastasis. They also highlight high-dose ascorbate as a promising therapeutic strategy for SDHB-related cancers. SIGNIFICANCE: Loss of different succinate dehydrogenase subunits can lead to different cell and tumor phenotypes, linking stronger 2-OG-dependent dioxygenases inhibition, iron overload, and ROS accumulation following SDHB mutation.

Sunitinib-induced cardiac hypertrophy and the endothelin axis.

Anti-angiogenics drugs in clinical use for cancer treatment induce cardiotoxic side effects. The endothelin axis is involved in hypertension and cardiac remodelling, and addition of an endothelin receptor antagonist to the anti-angiogenic sunitinib was shown to reduce cardiotoxicity of sunitinib in mice. Here, we explored further the antidote effect of the endothelin receptor antagonist macitentan in sunitinib-treated animals on cardiac remodeling. Methods: Tumor-bearing mice treated per os daily by sunitinib or vehicle were imaged before and after 1, 3 and 6 weeks of treatment by positron emission tomography using [18F]fluorodeoxyglucose and by echocardiography. Non-tumor-bearing animals were randomly assigned to be treated per os daily by vehicle or sunitinib or macitentan or sunitinib+macitentan, and imaged by echocardiography after 5 weeks. Hearts were harvested for histology and molecular analysis at the end of in vivo exploration. Results: Sunitinib treatment increases left ventricular mass and ejection fraction and induces cardiac fibrosis. Sunitinib also induces an early increase in cardiac uptake of [18F]fluorodeoxyglucose, which is significantly correlated with increased left ventricular mass at the end of treatment. Co-administration of macitentan prevents sunitinib-induced hypertension, increase in ejection fraction and cardiac fibrosis, but fails to prevent increase of the left ventricular mass. Conclusion: Early metabolic changes predict sunitinib-induced cardiac remodeling. Endothelin blockade can prevent some but not all cardiotoxic side-effects of sunitinib, in particular left ventricle hypertrophy that appears to be induced by sunitinib through an endothelin-independent mechanism.

Germline DLST Variants Promote Epigenetic Modifications in Pheochromocytoma-Paraganglioma.

CONTEXT: Pheochromocytomas and paragangliomas (PPGLs) are neuroendocrine tumors in which altered central metabolism appears to be a major driver of tumorigenesis, and many PPGL genes encode proteins involved in the tricarboxylic acid (TCA) cycle. OBJECTIVE/DESIGN: While about 40% of PPGL cases carry a variant in a known gene, many cases remain unexplained. In patients with unexplained PPGL showing clear evidence of a familial burden or multiple tumors, we aimed to identify causative factors using genetic analysis of patient DNA and functional analyses of identified DNA variants in patient tumor material and engineered cell lines. PATIENTS AND SETTING: Patients with a likely familial cancer burden of pheochromocytomas and/or paragangliomas and under investigation in a clinical genetic and clinical research setting in university hospitals. RESULTS: While investigating unexplained PPGL cases, we identified a novel variant, c.1151C>T, p.(Pro384Leu), in exon 14 of the gene encoding dihydrolipoamide S-succinyltransferase (DLST), a component of the multi-enzyme complex 2-oxoglutarate dehydrogenase. Targeted sequence analysis of further unexplained cases identified a patient carrying a tumor with compound heterozygous variants in DLST, consisting of a germline variant, c.1121G>A, p.(Gly374Glu), together with a somatic missense variant identified in tumor DNA, c.1147A>G, p.(Thr383Ala), both located in exon 14. Using a range of in silico and functional assays we show that these variants are predicted to be pathogenic, profoundly impact enzyme activity, and result in DNA hypermethylation. CONCLUSIONS: The identification and functional analysis of these DLST variants further validates DLST as an additional PPGL gene involved in the TCA cycle.

Screening of a Large Cohort of Asymptomatic SDHx Mutation Carriers in Routine Practice.

CONTEXT: When an SDHx mutation is identified in a patient with a pheochromocytoma (PCC) or a paraganglioma (PGL), predictive genetic testing can detect mutation carriers that would benefit from screening protocols. OBJECTIVE: To define the tumor detection rate in a large cohort of asymptomatic SDHX mutation carriers. DESIGN AND SETTING: Retrospective multicentric study in 6 referral centers. PATIENTS: Between 2005 and 2019, 249 asymptomatic SDHx (171 SDHB, 31 SDHC, 47 SDHD) mutation carriers, with at least 1 imaging work-up were enrolled. RESULTS: Initial work-up, including anatomical (98% of subjects [97-100% according to center]) and/or functional imaging (67% [14-90%]) detected 48 tumors in 40 patients. After a negative initial work-up, 124 patients benefited from 1 to 9 subsequent follow-up assessments (mean: 1.9 per patient), with a median follow-up time of 5 (1-13) years. Anatomical (86% [49-100 %]) and/or functional imaging (36% [7-60 %]) identified 10 new tumors (mean size: 16 mm [4-50]) in 10 patients. Altogether, 58 tumors (55 paraganglioma [PGL], including 45 head and neck PGL, 2 pheochromocytoma [PCC], 1 gastrointestinal stromal tumor [GIST]), were detected in 50 patients (22 [13%] SDHB, 1 [3.2%] SDHC, and 27 [57%] SDHD), with a median age of 41 years old [11-86], 76% without catecholamine secretion and 80% during initial imaging work-up. CONCLUSIONS: Imaging screening enabled detection of tumors in 20% of asymptomatic SDHx mutation carriers, with a higher detection rate in SDHD (57%) than in SDHB (13%) and SDHC (3%) mutation carriers, arguing for a gene-by-gene approach. Prospective studies using well-defined protocols are needed to obtain strong and useful data.

An update on adult forms of hereditary pheochromocytomas and paragangliomas.

PURPOSE OF REVIEW: Pheochromocytomas and paragangliomas (PPGL) display a strong genetic determinism with 40% of inherited forms. The purpose of this review is to provide an update on current knowledge on adult forms of hereditary PPGL and their management. RECENT FINDINGS: PPGL are genetically-driven in 70% of cases, with germline and/or somatic mutations identified in more than 20 genes. Although eight new susceptibility genes have recently emerged, mutations on SDHx genes remain the most frequent. In addition to SDHB, mutations in SLC25A11, FH and MDH2 may predispose to a metastatic disease and somatic alterations including TERT and ATRX mutations, and the differential expression on noncoding RNAs are also associated with the occurrence of metastases.The biochemical diagnosis remains the mainstay of functional PPGL and does not differ between hereditary PPGL while the choice of the best nuclear imaging approach is dictated by the tumor type and can be influenced by the presence of a germline mutation (18F-DOPA PET/CT for cluster 2 mutation and Ga-DOTATATE PET/CT for cluster 1 mutation). SUMMARY: A systematic genetic testing and counselling is recommended for all PPGL patients and should lead to conservative surgery and an adapted follow up, in case of hereditary form.

Epigenetic and metabolic reprogramming of SDH-deficient paragangliomas.

Pheochromocytomas and paragangliomas (PPGL) are rare neuroendocrine tumors arising from the adrenal medulla or extra-adrenal paraganglia. Around 40% of all cases are caused by a germline mutation in a susceptibility gene, half of which being found in an SDHx gene (SDHA, SDHB, SDHC, SDHD or SDHAF2). They encode the four subunits and assembly factor of succinate dehydrogenase (SDH), a mitochondrial enzyme involved both in the tricarboxylic acid cycle and electron transport chain. SDHx mutations lead to the accumulation of succinate, which acts as an oncometabolite by inhibiting iron(II) and alpha-ketoglutarate-dependent dioxygenases thereby regulating the cell's hypoxic response and epigenetic processes. Moreover, SDHx mutations induce cell metabolic reprogramming and redox imbalance. Major discoveries in PPGL pathophysiology have been made since the initial discovery of SDHD gene mutations in 2000, improving the understanding of their biology and patient management. It indeed provides new opportunities for diagnostic tools and innovative therapeutic targets in order to improve the prognosis of patients affected by these rare tumors, in particular in the context of metastatic diseases associated with SDHB mutations. This review first describes an overview of the pathophysiology and then focuses on clinical implications of the epigenetic and metabolic reprogramming of SDH-deficient PPGL.

An overview of 20 years of genetic studies in pheochromocytoma and paraganglioma.

Paragangliomas and pheochromocytomas (PPGL) are rare neuroendocrine tumours characterized by a strong genetic determinism. Over the past 20 years, evolution of PPGL genetics has revealed that around 40% of PPGL are genetically determined, secondary to a germline mutation in one of more than twenty susceptibility genes reported so far. More than half of the mutations occur in one of the SDHx genes (SDHA, SDHB, SDHC, SDHD, SDHAF2), which encode the different subunits and assembly protein of a mitochondrial enzyme, succinate dehydrogenase. These susceptibility genes predispose to early forms (VHL, RET, SDHD, EPAS1, DLST), syndromic (RET, VHL, EPAS1, NF1, FH), multiple (SDHD, TMEM127, MAX, DLST, MDH2, GOT2) or malignant (SDHB, FH, SLC25A11) PPGL. The discovery of a germline mutation in one of these genes changes the patient's follow-up and allows genetic screening of affected families and the presymptomatic follow-up of relatives carrying a mutation.

TET-Mediated Hypermethylation Primes SDH-Deficient Cells for HIF2α-Driven Mesenchymal Transition.

Loss-of-function mutations in the SDHB subunit of succinate dehydrogenase predispose patients to aggressive tumors characterized by pseudohypoxic and hypermethylator phenotypes. The mechanisms leading to DNA hypermethylation and its contribution to SDH-deficient cancers remain undemonstrated. We examine the genome-wide distribution of 5-methylcytosine and 5-hydroxymethylcytosine and their correlation with RNA expression in SDHB-deficient tumors and murine Sdhb-/- cells. We report that DNA hypermethylation results from TET inhibition. Although it preferentially affects PRC2 targets and known developmental genes, PRC2 activity does not contribute to the DNA hypermethylator phenotype. We also prove, in vitro and in vivo, that TET silencing, although recapitulating the methylation profile of Sdhb-/- cells, is not sufficient to drive their EMT-like phenotype, which requires additional HIF2α activation. Altogether, our findings reveal synergistic roles of TET repression and pseudohypoxia in the acquisition of metastatic traits, providing a rationale for targeting HIF2α and DNA methylation in SDH-associated malignancies.

Concurrent imaging of vascularization and metabolism in a mouse model of paraganglioma under anti-angiogenic treatment.

Rationale: Deregulation of metabolism and induction of vascularization are major hallmarks of cancer. Using a new multimodal preclinical imaging instrument, we explored a sequence of events leading to sunitinib-induced resistance in a murine model of paraganglioma (PGL) invalidated for the expression of succinate dehydrogenase subunit B (Sdhb-/-). Methods: Two groups of Sdhb-/- tumors bearing mice were treated with sunitinib (6 weeks) or vehicle (3 weeks). Concurrent Positron Emission Tomography (PET) with 2' -deoxy-2'-[18F]fluoro-D-glucose (FDG), Computed Tomography (CT) and Ultrafast Ultrasound Imaging (UUI) imaging sessions were performed once a week and ex vivo samples were analyzed by western blots and histology. Results: PET-CT-UUI enabled to detect a rapid growth of Sdhb-/- tumors with increased glycolysis and vascular development. Sunitinib treatment prevented tumor growth, vessel development and reduced FDG uptake at week 1 and 2 (W1-2). Thereafter, imaging revealed tumor escape from sunitinib treatment: FDG uptake in tumors increased at W3, followed by tumor growth and vessel development at W4-5. Perfused vessels were preferentially distributed in the hypermetabolic regions of the tumors and the perfused volume increased during escape from sunitinib treatment. Finally, initial changes in total lesion glycolysis and maximum vessel length at W1 were predictive of resistance to sunitinib. Conclusion: These results demonstrate an adaptive resistance of Sdhb-/- tumors to six weeks of sunitinib treatment. Early metabolic changes and delayed vessel architecture changes were detectable and predictable in vivo early during anti-angiogenic treatment. Simultaneous metabolic, anatomical and functional imaging can monitor precisely the effects of anti-angiogenic treatment of tumors.