Molecular Imaging with PET-CT and PET-MRI in Pediatric Musculoskeletal Diseases.

Molecular imaging has emerged as an integral part of oncologic imaging. Given the physiologic changes that precede anatomic changes, molecular imaging can enable early detection of disease and monitoring of response. [18F] Fluorodeoxyglucose (FDG) Positron emission tomography (PET) is the predominant molecular imaging modality used in oncologic assessment and can be performed using PET/CT or PET/MR. In pediatric patients, PET/MRI imaging is generally preferred due to low radiation exposure and PET/MRI is particularly advantageous for imaging musculoskeletal (MSK) diseases, as MRI provides superior characterization of tissue changes as compared to CT. In this article, we provide an overview of the typical role of PET CT/MRI in assessment of some common pediatric malignancies and benign MSK diseases with case examples. We also discuss the relative advantages of PET/MRI compared to PET/CT, and review published data with a primary focus on the use of PET/MR.

Molecular Imaging in Soft-tissue Sarcoma: Evolving Role of FDG PET.

Soft tissue sarcomas are a rare and heterogenous group of tumors that account for 2% of all cancer-related deaths. Molecular imaging with FDG PET can offer valuable metabolic information to help inform clinical management of soft tissue sarcomas that is unique and complementary to conventional diagnostic imaging techniques. FDG PET imaging often correlates with tumor grade, can help guide biopsy, and frequently detects additional sites of disease compared to conventional imaging in patients being considered for definitive or salvage therapy. Traditional size-based evaluation of treatment response is often inadequate in soft tissue sarcoma and changes in metabolic activity can add significant value to interim and end of treatment imaging for high-grade sarcomas. FDG PET can be used for detection of recurrence or malignant transformation and thus play a vital role in surveillance. This article reviews the evolving role of FDG PET in initial diagnosis, staging, treatment response assessment, and restaging. Further studies on the use of FDG PET in soft sarcoma are needed, particularly for rare histopathologic subtypes.

64Cu-DOTATATE Uptake in a Pulmonary Hamartoma.

ABSTRACT: DOTATATE PET/CT is frequently used to evaluate indeterminant pulmonary nodules suspected to be pulmonary carcinoid. We report an unexpected case of pulmonary hamartoma demonstrating 64Cu-DOTATATE uptake in a 43-year-old woman with a slowly enlarging pulmonary nodule. Histopathological staining showed somatostatin receptor 2 expression on vascular endothelial cells and a proportion of cartilage and smooth muscle cells within the hamartoma.

PSMA theragnostics for metastatic castration resistant prostate cancer.

There has been tremendous growth in the development of theragnostics for personalized cancer diagnosis and treatment over the past two decades. In prostate cancer, the new generation of prostate specific membrane antigen (PSMA) small molecular inhibitor-based imaging agents achieve extraordinary tumor to background ratios and allow their therapeutic counterparts to deliver effective tumor doses while minimizing normal tissue toxicity. The PSMA targeted small molecule positron emission tomography (PET) agents 18F-DCFPyL (2-(3-{1-carboxy-5-((6-(18)F-fluoro-pyridine-3-carbonyl)-amino)-pentyl}-ureido)-pentanedioic acid) and Gallium-68 (68Ga)-PSMA-11 have been approved by the United States Food and Drug Administration (FDA) for newly diagnosed high risk prostate cancer patients and for patients with biochemical recurrence. More recently, the Phase III VISION trial showed that Lutetium-177 (177Lu)-PSMA-617 treatment increases progression-free survival and overall survival in patients with heavily pre-treated advanced PSMA-positive metastatic castration-resistant prostate cancer (mCRPC). Here, we review the PSMA targeted theragnostic pairs under clinical investigation for detection and treatment of metastatic prostate cancer.

Phantom study of SPECT/CT augmented reality for intraoperative localization of sentinel lymph nodes in head and neck melanoma.

OBJECTIVE: To show that augmented reality (AR) visualization of single-photon emission computed tomography (SPECT)/computed tomography (CT) data in 3D can be used to accurately localize targets in the head and neck region. MATERIALS AND METHODS: Eight head and neck styrofoam phantoms were painted with a mixture of radioactive solution (Tc-99m) detectable with a handheld gamma probe and fluorescent ink visible only under ultraviolet (UV) light to create 10-20 simulated lymph nodes on their surface. After obtaining SPECT/CT images of these phantoms, virtual renderings of the nodes were generated from the SPECT/CT data and displayed using a commercially available AR headset. For each of three physician evaluators, the time required to localize lymph node targets was recorded (1) using the gamma probe alone and (2) using the gamma probe while wearing the AR headset. In addition, the surface localization accuracy when using the AR headset was evaluated by measuring the misalignment between the locations visually marked by the evaluators and the ground truth locations identified using UV stimulation of the ink at the site of the nodes. RESULTS: For all three evaluators, using the AR headset significantly reduced the time to detect targets (P = 0.012, respectively) compared to using the gamma probe alone. The average misalignment between the location marked by the evaluators and the ground truth location was 8.6 mm. CONCLUSION: AR visualization of SPECT/CT data in 3D allows for accurate localization of targets in the head and neck region, and may reduce the localization time of targets.

18F-FDG PET/CT for Evaluation of Post-Transplant Lymphoproliferative Disorder (PTLD).

Post-transplant lymphoproliferative disorders (PTLD) are a spectrum of heterogeneous lymphoproliferative conditions that are serious and possibly fatal complications after solid organ or allogenic hematopoietic stem cell transplantation. Most PTLD are attributed to Epstein-Barr virus reactivation in B-cells in the setting of immunosuppression after transplantation. Early diagnosis, accurate staging, and timely treatment are of vital importance to reduce morbidity and mortality. Given the often nonspecific clinical presentation and disease heterogeneity of PTLD, tissue biopsy and histopathological analysis are essential to establish diagnosis and most importantly, determine the subtype of PTLD, which guides treatment options. Advanced imaging modalities such as 18F-FDG PET/CT have played an increasingly important role and have shown high sensitivity and specificity in detection, staging, and assessing treatment response in multiple clinical studies over the last two decades. However, larger multicenter prospective validation is still needed to further establish the clinical utility of PET imaging in the management of PTLD. Significantly, new hybrid imaging modalities such as PET/MR may help reduce radiation exposure, which is especially important in pediatric transplant patients.

The Clinical Utility of 18F-Fluciclovine PET/CT in Biochemically Recurrent Prostate Cancer: an Academic Center Experience Post FDA Approval.

PURPOSE: To evaluate the diagnostic performance and clinical utility of 18F-fluciclovine PET/CT in patients with biochemical recurrence (BCR) of prostate cancer (PC). METHODS: 18F-Fluciclovine scans of 165 consecutive men with BCR after primary definitive treatment with prostatectomy (n = 102) or radiotherapy (n = 63) were retrospectively evaluated. Seventy patients had concurrent imaging with at least one other conventional modality (CT (n = 31), MRI (n = 31), or bone scan (n = 26)). Findings from 18F-fluciclovine PET were compared with those from conventional imaging modalities. The positivity rate and impact of 18F-fluciclovine PET on patient management were recorded. In 33 patients who underwent at least one other PET imaging (18F-NaF PET/CT (n = 12), 68Ga-PSMA11 PET/CT (n = 5), 18F-DCFPyL PET/CT (n = 20), and 68Ga-RM2 PET/MRI (n = 5)), additional findings were evaluated. RESULTS: The overall positivity rate of 18F-fluciclovine PET was 67 %, which, as expected, increased with higher prostate-specific antigen (PSA) levels (ng/ml): 15 % (PSA < 0.5), 50 % (0.5 ≤ PSA < 1), 56 % (1 ≤ PSA < 2), 68 % (2 ≤ PSA < 5), and 94 % (PSA ≥ 5), respectively. One hundred and two patients (62 %) had changes in clinical management based on 18F-fluciclovine PET findings. Twelve of these patients (12 %) had lesion localization on 18F-fluciclovine PET, despite negative conventional imaging. Treatment plans of 14 patients with negative 18F-fluciclovine PET were changed based on additional PET imaging with a different radiopharmaceutical. CONCLUSION: 18F-Fluciclovine PET/CT remains a useful diagnostic tool in the workup of patients with BCR PC, changing clinical management in 62 % of participants in our cohort.

Fungal endocarditis resembling primary cardiac malignancy in a patient with B-cell ALL with culture confirmation.

Fungal endocarditis is a rare subtype of infective endocarditis that often presents with nonspecific symptoms in patients with complex medical histories, making diagnosis challenging. Patients with a history of ALL may present with congestive heart failure, chemo-induced cardiomyopathy, acute coronary syndrome, cardiac lymphomatous metastasis, or infections. We present the case of a patient with a history of ALL who presented with acute coronary syndrome and imaging concerning for primary cardiac lymphoma, when in fact the patient ended up suffering from culture proven fungal endocarditis.

Cost-effectiveness of magnetic resonance imaging versus ultrasound for the detection of symptomatic full-thickness supraspinatus tendon tears.

BACKGROUND: The purpose of this study was to determine the value of magnetic resonance imaging (MRI) and ultrasound-based imaging strategies in the evaluation of a hypothetical population with a symptomatic full-thickness supraspinatus tendon (FTST) tear using formal cost-effectiveness analysis. METHODS: A decision analytic model from the health care system perspective for 60-year-old patients with symptoms secondary to a suspected FTST tear was used to evaluate the incremental cost-effectiveness of 3 imaging strategies during a 2-year time horizon: MRI, ultrasound, and ultrasound followed by MRI. Comprehensive literature search and expert opinion provided data on cost, probability, and quality of life estimates. The primary effectiveness outcome was quality-adjusted life-years (QALYs) through 2 years, with a willingness-to-pay threshold set to $100,000/QALY gained (2016 U.S. dollars). Costs and health benefits were discounted at 3%. RESULTS: Ultrasound was the least costly strategy ($1385). MRI was the most effective (1.332 QALYs). Ultrasound was the most cost-effective strategy but was not dominant. The incremental cost-effectiveness ratio for MRI was $22,756/QALY gained, below the willingness-to-pay threshold. Two-way sensitivity analysis demonstrated that MRI was favored over the other imaging strategies over a wide range of reasonable costs. In probabilistic sensitivity analysis, MRI was the preferred imaging strategy in 78% of the simulations. CONCLUSION: MRI and ultrasound represent cost-effective imaging options for evaluation of the patient thought to have a symptomatic FTST tear. The results indicate that MRI is the preferred strategy based on cost-effectiveness criteria, although the decision between MRI and ultrasound for an imaging center is likely to be dependent on additional factors, such as available resources and workflow.

Structure of human nSMase2 reveals an interdomain allosteric activation mechanism for ceramide generation.

Neutral sphingomyelinase 2 (nSMase2, product of the SMPD3 gene) is a key enzyme for ceramide generation that is involved in regulating cellular stress responses and exosome-mediated intercellular communication. nSMase2 is activated by diverse stimuli, including the anionic phospholipid phosphatidylserine. Phosphatidylserine binds to an integral-membrane N-terminal domain (NTD); however, how the NTD activates the C-terminal catalytic domain is unclear. Here, we identify the complete catalytic domain of nSMase2, which was misannotated because of a large insertion. We find the soluble catalytic domain interacts directly with the membrane-associated NTD, which serves as both a membrane anchor and an allosteric activator. The juxtamembrane region, which links the NTD and the catalytic domain, is necessary and sufficient for activation. Furthermore, we provide a mechanistic basis for this phenomenon using the crystal structure of the human nSMase2 catalytic domain determined at 1.85-Å resolution. The structure reveals a DNase-I-type fold with a hydrophobic track leading to the active site that is blocked by an evolutionarily conserved motif which we term the "DK switch." Structural analysis of nSMase2 and the extended N-SMase family shows that the DK switch can adopt different conformations to reposition a universally conserved Asp (D) residue involved in catalysis. Mutation of this Asp residue in nSMase2 disrupts catalysis, allosteric activation, stimulation by phosphatidylserine, and pharmacological inhibition by the lipid-competitive inhibitor GW4869. Taken together, these results demonstrate that the DK switch regulates ceramide generation by nSMase2 and is governed by an allosteric interdomain interaction at the membrane interface.

A fidelity mechanism in DNA polymerase lambda promotes error-free bypass of 8-oxo-dG.

8-oxo-7,8-dihydroxy-2'-deoxyguanosine (8-oxo-dG) has high mutagenic potential as it is prone to mispair with deoxyadenine (dA). In order to maintain genomic integrity, post-replicative 8-oxo-dG:dA mispairs are removed through DNA polymerase lambda (Pol λ)-dependent MUTYH-initiated base excision repair (BER). Here, we describe seven novel crystal structures and kinetic data that fully characterize 8-oxo-dG bypass by Pol λ. We demonstrate that Pol λ has a flexible active site that can tolerate 8-oxo-dG in either the anti- or syn-conformation. Importantly, we show that discrimination against the pro-mutagenic syn-conformation occurs at the extension step and identify the residue responsible for this selectivity. This residue acts as a kinetic switch, shunting repair toward long-patch BER upon correct dCMP incorporation, thus enhancing repair efficiency. Moreover, this switch also provides a potential mechanism to increase repair fidelity of MUTYH-initiated BER.

Nucleotide binding interactions modulate dNTP selectivity and facilitate 8-oxo-dGTP incorporation by DNA polymerase lambda.

8-Oxo-7,8,-dihydro-2'-deoxyguanosine triphosphate (8-oxo-dGTP) is a major product of oxidative damage in the nucleotide pool. It is capable of mispairing with adenosine (dA), resulting in futile, mutagenic cycles of base excision repair. Therefore, it is critical that DNA polymerases discriminate against 8-oxo-dGTP at the insertion step. Because of its roles in oxidative DNA damage repair and non-homologous end joining, DNA polymerase lambda (Pol λ) may frequently encounter 8-oxo-dGTP. Here, we have studied the mechanisms of 8-oxo-dGMP incorporation and discrimination by Pol λ. We have solved high resolution crystal structures showing how Pol λ accommodates 8-oxo-dGTP in its active site. The structures indicate that when mispaired with dA, the oxidized nucleotide assumes the mutagenic syn-conformation, and is stabilized by multiple interactions. Steady-state kinetics reveal that two residues lining the dNTP binding pocket, Ala(510) and Asn(513), play differential roles in dNTP selectivity. Specifically, Ala(510) and Asn(513) facilitate incorporation of 8-oxo-dGMP opposite dA and dC, respectively. These residues also modulate the balance between purine and pyrimidine incorporation. Our results shed light on the mechanisms controlling 8-oxo-dGMP incorporation in Pol λ and on the importance of interactions with the incoming dNTP to determine selectivity in family X DNA polymerases.

Unraveling Cholesterol Catabolism in Mycobacterium tuberculosis: ChsE4-ChsE5 α2ß2 Acyl-CoA Dehydrogenase Initiates ß-Oxidation of 3-Oxo-cholest-4-en-26-oyl CoA.

The metabolism of host cholesterol by Mycobacterium tuberculosis (Mtb) is an important factor for both its virulence and pathogenesis, although how and why cholesterol metabolism is required is not fully understood. Mtb uses a unique set of catabolic enzymes that are homologous to those required for classical ß-oxidation of fatty acids but are specific for steroid-derived substrates. Here, we identify and assign the substrate specificities of two of these enzymes, ChsE4-ChsE5 (Rv3504-Rv3505) and ChsE3 (Rv3573c), that carry out cholesterol side chain oxidation in Mtb. Steady-state assays demonstrate that ChsE4-ChsE5 preferentially catalyzes the oxidation of 3-oxo-cholest-4-en-26-oyl CoA in the first cycle of cholesterol side chain ß-oxidation that ultimately yields propionyl-CoA, whereas ChsE3 specifically catalyzes the oxidation of 3-oxo-chol-4-en-24-oyl CoA in the second cycle of ß-oxidation that generates acetyl-CoA. However, ChsE4-ChsE5 can catalyze the oxidation of 3-oxo-chol-4-en-24-oyl CoA as well as 3-oxo-4-pregnene-20-carboxyl-CoA. The functional redundancy of ChsE4-ChsE5 explains the in vivo phenotype of the igr knockout strain of Mycobacterium tuberculosis; the loss of ChsE1-ChsE2 can be compensated for by ChsE4-ChsE5 during the chronic phase of infection. The X-ray crystallographic structure of ChsE4-ChsE5 was determined to a resolution of 2.0 Å and represents the first high-resolution structure of a heterotetrameric acyl-CoA dehydrogenase (ACAD). Unlike typical homotetrameric ACADs that bind four flavin adenine dinucleotide (FAD) cofactors, ChsE4-ChsE5 binds one FAD at each dimer interface, resulting in only two substrate-binding sites rather than the classical four active sites. A comparison of the ChsE4-ChsE5 substrate-binding site to those of known mammalian ACADs reveals an enlarged binding cavity that accommodates steroid substrates and highlights novel prospects for designing inhibitors against the committed ß-oxidation step in the first cycle of cholesterol side chain degradation by Mtb.

Completing the specificity swap: Single-stranded DNA recognition by F and R100 TraI relaxase domains.

During conjugative plasmid transfer, one plasmid strand is cleaved and transported to the recipient bacterium. For F and related plasmids, TraI contains the relaxase or nickase activity that cleaves the plasmid DNA strand. F TraI36, the F TraI relaxase domain, binds a single-stranded origin of transfer (oriT) DNA sequence with high affinity and sequence specificity. The TraI36 domain from plasmid R100 shares 91% amino acid sequence identity with F TraI36, but its oriT DNA binding site differs by two of eleven bases. Both proteins readily distinguish between F and R100 binding sites. In earlier work, two amino acid substitutions in the DNA binding cleft were shown to be sufficient to change the R100 TraI36 DNA-binding specificity to that of F TraI36. In contrast, three substitutions could make F TraI36 more "R100-like", but failed to completely alter the specificity. Here we identify one additional amino acid substitution that completes the specificity swap from F to R100. To our surprise, adding further substitutions from R100 to the F background were detrimental to binding instead of being neutral, indicating that their effects were influenced by their structural context. These results underscore the complex and subtle nature of DNA recognition by relaxases and have implications for the evolution of relaxase binding sites and oriT sequences.

A distinct MaoC-like enoyl-CoA hydratase architecture mediates cholesterol catabolism in Mycobacterium tuberculosis.

The Mycobacterium tuberculosis (Mtb) igr operon plays an essential role in Mtb cholesterol metabolism, which is critical for pathogenesis during the latent stage of Mtb infection. Here we report the first structure of a heterotetrameric MaoC-like enoyl-CoA hydratase, ChsH1-ChsH2, which is encoded by two adjacent genes from the igr operon. We demonstrate that ChsH1-ChsH2 catalyzes the hydration of a steroid enoyl-CoA, 3-oxo-4,17-pregnadiene-20-carboxyl-CoA, in the modified ß-oxidation pathway for cholesterol side chain degradation. The ligand-bound and apoenzyme structures of ChsH1-ChsH2(N) reveal an unusual, modified hot-dog fold with a severely truncated central α-helix that creates an expanded binding site to accommodate the bulkier steroid ring system. The structures show quaternary structure shifts that accommodate the four rings of the steroid substrate and offer an explanation for why the unusual heterotetrameric assembly is utilized for hydration of this steroid. The unique αß heterodimer architecture utilized by ChsH1-ChsH2 to bind its distinctive substrate highlights an opportunity for the development of new antimycobacterial drugs that target a pathway specific to Mtb.

Non-stop mRNA decay: a special attribute of trans-translation mediated ribosome rescue.

Decoding of aberrant mRNAs leads to unproductive ribosome stalling and sequestration of components of the translation machinery. Bacteria have evolved three seemingly independent pathways to resolve stalled translation complexes. The trans-translation process, orchestrated by the hybrid transfer-messenger RNA (tmRNA) and its essential protein co-factor, small protein B (SmpB), is the principal translation quality control system for rescuing unproductively stalled ribosomes. Two specialized alternative rescue pathways, coordinated by ArfA and ArfB, have been recently discovered. The SmpB-tmRNA mediated trans-translation pathway, in addition to re-mobilizing stalled translation complexes, co-translationally appends a degradation tag to the associated nascent polypeptides, marking them for proteolysis by various cellular proteases. Another unique feature of trans-translation, not shared by the alternative rescue pathways, is the facility to recruit ribonuclease R (RNase R) for targeted degradation of non-stop mRNAs, thus preventing further futile cycles of translation. The distinct C-terminal lysine-rich (K-rich) domain of RNase R is essential for its recruitment to stalled ribosomes. To gain new insights into the structure and function of RNase R, we investigated its global architecture, the spatial arrangement of its distinct domains, and the identities of key functional residues in its unique K-rich domain. Small-angle X-ray scattering models of RNase R reveal a tri-lobed structure with flexible N- and C-terminal domains, and suggest intimate contacts between the K-rich domain and the catalytic core of the enzyme. Alanine-scanning mutagenesis of the K-rich domain, in the region spanning residues 735 and 750, has uncovered the precise amino acid determinants required for the productive engagement of RNase R on tmRNA-rescued ribosomes. Theses analyses demonstrate that alanine substitution of conserved residues E740 and K741result in profound defects, not only in the recruitment of RNase R to rescued ribosomes but also in the targeted decay of non-stop mRNAs. Additionally, an RNase R variant with alanine substitution at residues K749 and K750 exhibits extensive defects in ribosome enrichment and non-stop mRNA decay. In contrast, alanine substitution of additional conserved residues in this region has no effect on the known functions of RNase R. In vitro RNA degradation assays demonstrate that the consequential substitutions (RNase R(E740A/K741A) and RNase R(K749A/K750A)) do not affect the ability of the enzyme to degrade structured RNAs, indicating that the observed defect is specific to the trans-translation related activities of RNase R. Taken together, these findings shed new light on the global architecture of RNase R and provide new details of how this versatile RNase effectuates non-stop mRNA decay on tmRNA-rescued ribosomes.