Molecular imaging in oncology: Current impact and future directions.

The authors define molecular imaging, according to the Society of Nuclear Medicine and Molecular Imaging, as the visualization, characterization, and measurement of biological processes at the molecular and cellular levels in humans and other living systems. Although practiced for many years clinically in nuclear medicine, expansion to other imaging modalities began roughly 25 years ago and has accelerated since. That acceleration derives from the continual appearance of new and highly relevant animal models of human disease, increasingly sensitive imaging devices, high-throughput methods to discover and optimize affinity agents to key cellular targets, new ways to manipulate genetic material, and expanded use of cloud computing. Greater interest by scientists in allied fields, such as chemistry, biomedical engineering, and immunology, as well as increased attention by the pharmaceutical industry, have likewise contributed to the boom in activity in recent years. Whereas researchers and clinicians have applied molecular imaging to a variety of physiologic processes and disease states, here, the authors focus on oncology, arguably where it has made its greatest impact. The main purpose of imaging in oncology is early detection to enable interception if not prevention of full-blown disease, such as the appearance of metastases. Because biochemical changes occur before changes in anatomy, molecular imaging-particularly when combined with liquid biopsy for screening purposes-promises especially early localization of disease for optimum management. Here, the authors introduce the ways and indications in which molecular imaging can be undertaken, the tools used and under development, and near-term challenges and opportunities in oncology.

Engineered tRNAs suppress nonsense mutations in cells and in vivo.

Nonsense mutations are the underlying cause of approximately 11% of all inherited genetic diseases1. Nonsense mutations convert a sense codon that is decoded by tRNA into a premature termination codon (PTC), resulting in an abrupt termination of translation. One strategy to suppress nonsense mutations is to use natural tRNAs with altered anticodons to base-pair to the newly emerged PTC and promote translation2-7. However, tRNA-based gene therapy has not yielded an optimal combination of clinical efficacy and safety and there is presently no treatment for individuals with nonsense mutations. Here we introduce a strategy based on altering native tRNAs into  efficient suppressor tRNAs (sup-tRNAs) by individually fine-tuning their sequence to the physico-chemical properties of the amino acid that they carry. Intravenous and intratracheal lipid nanoparticle (LNP) administration of sup-tRNA in mice restored the production of functional proteins with nonsense mutations. LNP-sup-tRNA formulations caused no discernible readthrough at endogenous native stop codons, as determined by ribosome profiling. At clinically important PTCs in the cystic fibrosis transmembrane conductance regulator gene (CFTR), the sup-tRNAs re-established expression and function in cell systems and patient-derived nasal epithelia and restored airway volume homeostasis. These results provide a framework for the development of tRNA-based therapies with a high molecular safety profile and high efficacy in targeted PTC suppression.

Transgenic ferret models define pulmonary ionocyte diversity and function.

Speciation leads to adaptive changes in organ cellular physiology and creates challenges for studying rare cell-type functions that diverge between humans and mice. Rare cystic fibrosis transmembrane conductance regulator (CFTR)-rich pulmonary ionocytes exist throughout the cartilaginous airways of humans1,2, but limited presence and divergent biology in the proximal trachea of mice has prevented the use of traditional transgenic models to elucidate ionocyte functions in the airway. Here we describe the creation and use of conditional genetic ferret models to dissect pulmonary ionocyte biology and function by enabling ionocyte lineage tracing (FOXI1-CreERT2::ROSA-TG), ionocyte ablation (FOXI1-KO) and ionocyte-specific deletion of CFTR (FOXI1-CreERT2::CFTRL/L). By comparing these models with cystic fibrosis ferrets3,4, we demonstrate that ionocytes control airway surface liquid absorption, secretion, pH and mucus viscosity-leading to reduced airway surface liquid volume and impaired mucociliary clearance in cystic fibrosis, FOXI1-KO and FOXI1-CreERT2::CFTRL/L ferrets. These processes are regulated by CFTR-dependent ionocyte transport of Cl- and HCO3-. Single-cell transcriptomics and in vivo lineage tracing revealed three subtypes of pulmonary ionocytes and a FOXI1-lineage common rare cell progenitor for ionocytes, tuft cells and neuroendocrine cells during airway development. Thus, rare pulmonary ionocytes perform critical CFTR-dependent functions in the proximal airway that are hallmark features of cystic fibrosis airway disease. These studies provide a road map for using conditional genetics in the first non-rodent mammal to address gene function, cell biology and disease processes that have greater evolutionary conservation between humans and ferrets.

Human distal airways contain a multipotent secretory cell that can regenerate alveoli.

The human lung differs substantially from its mouse counterpart, resulting in a distinct distal airway architecture affected by disease pathology in chronic obstructive pulmonary disease. In humans, the distal branches of the airway interweave with the alveolar gas-exchange niche, forming an anatomical structure known as the respiratory bronchioles. Owing to the lack of a counterpart in mouse, the cellular and molecular mechanisms that govern respiratory bronchioles in the human lung remain uncharacterized. Here we show that human respiratory bronchioles contain a unique secretory cell population that is distinct from cells in larger proximal airways. Organoid modelling reveals that these respiratory airway secretory (RAS) cells act as unidirectional progenitors for alveolar type 2 cells, which are essential for maintaining and regenerating the alveolar niche. RAS cell lineage differentiation into alveolar type 2 cells is regulated by Notch and Wnt signalling. In chronic obstructive pulmonary disease, RAS cells are altered transcriptionally, corresponding to abnormal alveolar type 2 cell states, which are associated with smoking exposure in both humans and ferrets. These data identify a distinct progenitor in a region of the human lung that is not found in mouse that has a critical role in maintaining the gas-exchange compartment and is altered in chronic lung disease.

Axicabtagene ciloleucel vs standard of care in second-line large B-cell lymphoma: outcomes by metabolic tumor volume.

ABSTRACT: Metabolic tumor volume (MTV) assessed using 2-deoxy-2-[18F]fluoro-d-glucose positron emission tomography, a measure of tumor burden, is a promising prognostic indicator in large B-cell lymphoma (LBCL). This exploratory analysis evaluated relationships between baseline MTV (categorized as low [median or less] vs high [greater than median]) and clinical outcomes in the phase 3 ZUMA-7 study (NCT03391466). Patients with LBCL relapsed within 12 months of or refractory to first-line chemoimmunotherapy were randomized 1:1 to axicabtagene ciloleucel (axi-cel; autologous anti-CD19 chimeric antigen receptor T-cell therapy) or standard care (2-3 cycles of chemoimmunotherapy followed by high-dose chemotherapy with autologous stem cell transplantation in patients who had a response). All P values are descriptive. Within high- and low-MTV subgroups, event-free survival (EFS) and progression-free survival (PFS) were superior with axi-cel vs standard care. EFS in patients with high MTV (vs low MTV) was numerically shorter with axi-cel and was significantly shorter with standard care. PFS was shorter in patients with high MTV vs low MTV in both the axi-cel and standard-care arms, and median MTV was lower in patients in ongoing response at data cutoff vs others. Median MTV was higher in patients treated with axi-cel who experienced grade ≥3 neurologic events or cytokine release syndrome (CRS) than in patients with grade 1/2 or no neurologic events or CRS, respectively. Baseline MTV less than or equal to median was associated with better clinical outcomes in patients receiving axi-cel or standard care for second-line LBCL. The trial was registered at www.clinicaltrials.gov as #NCT03391466.

Ivacaftor for Chronic Obstructive Pulmonary Disease - Results from a Phase 2, Randomized Controlled Trial.

RATIONALE: Patients with chronic obstructive pulmonary disease (COPD) exhibit acquired cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction. CFTR modulators may improve outcomes in patients with COPD, although recent data regarding magnitude of benefit have been inconclusive and effects on mucociliary clearance are unknown. We conducted a phase 2, randomized, double blind placebo control trial to determine safety and tolerability, and explore the potential mechanism of ivacaftor for the treatment of COPD. METHODS: We randomized 40 patients with moderate to severe COPD and symptoms of chronic bronchitis to ivacaftor (N=30) or placebo (N=10) 150mg BID for 12 weeks. Primary endpoints included evaluation of safety of ivacaftor and pharmacokinetics (PK). Secondary endpoints included measures of CFTR activity and clinical outcomes. RESULTS: Ivacaftor was safe and tolerable with similar rates of adverse events rates between groups. Most common adverse event was diarrhea in the ivacaftor group and acute COPD exacerbation in the placebo group. PK analysis found the mean area under the curve over 12 hours (AUC12) to be 72% of the previously reported AUC12 in cystic fibrosis (CF). Treatment with ivacaftor did not improve sweat chloride, whole lung mucociliary clearance, lung function or respiratory symptoms. CONCLUSION: Ivacaftor was safe and well tolerated, but did not improve measures of CFTR activity or mucus clearance. As serum concentrations achieved were lower than observed in CF at the same dose, and modulation of wild type CFTR differs from G551D, further dose determination studies are needed to better understand treatment efficacy of CFTR potentiators in COPD. Clinical trial registration available at www. CLINICALTRIALS: gov, ID: NCT03085485.

Pulmonary Fibrosis Stakeholder Summit: A Joint NHLBI, Three Lakes Foundation, and Pulmonary Fibrosis Foundation Workshop Report.

Despite progress in elucidation of disease mechanisms, identification of risk factors, biomarker discovery, and the approval of two medications to slow lung function decline in idiopathic pulmonary fibrosis and one medication to slow lung function decline in progressive pulmonary fibrosis, pulmonary fibrosis remains a disease with a high morbidity and mortality. In recognition of the need to catalyze ongoing advances and collaboration in the field of pulmonary fibrosis, the NHLBI, the Three Lakes Foundation, and the Pulmonary Fibrosis Foundation hosted the Pulmonary Fibrosis Stakeholder Summit on November 8-9, 2022. This workshop was held virtually and was organized into three topic areas: 1) novel models and research tools to better study pulmonary fibrosis and uncover new therapies, 2) early disease risk factors and methods to improve diagnosis, and 3) innovative approaches toward clinical trial design for pulmonary fibrosis. In this workshop report, we summarize the content of the presentations and discussions, enumerating research opportunities for advancing our understanding of the pathogenesis, treatment, and outcomes of pulmonary fibrosis.

Metachrony drives effective mucociliary transport via a calcium-dependent mechanism.

The mucociliary transport apparatus is critical for maintaining lung health via the coordinated movement of cilia to clear mucus and particulates. A metachronal wave propagates across the epithelium when cilia on adjacent multiciliated cells beat slightly out of phase along the proximal-distal axis of the airways in alignment with anatomically directed mucociliary clearance. We hypothesized that metachrony optimizes mucociliary transport (MCT) and that disruptions of calcium signaling would abolish metachrony and decrease MCT. We imaged bronchi from human explants and ferret tracheae using micro-optical coherence tomography (µOCT) to evaluate airway surface liquid depth (ASL), periciliary liquid depth (PCL), cilia beat frequency (CBF), MCT, and metachrony in situ. We developed statistical models that included covariates of MCT. Ferret tracheae were treated with BAPTA-AM (chelator of intracellular Ca2+), lanthanum chloride (nonpermeable Ca2+ channel competitive antagonist), and repaglinide (inhibitor of calaxin) to test calcium dependence of metachrony. We demonstrated that metachrony contributes to mucociliary transport of human and ferret airways. MCT was augmented in regions of metachrony compared with nonmetachronous regions by 48.1%, P = 0.0009 or 47.5%, P < 0.0020 in humans and ferrets, respectively. PCL and metachrony were independent contributors to MCT rate in humans; ASL, CBF, and metachrony contribute to ferret MCT rates. Metachrony can be disrupted by interference with calcium signaling including intracellular, mechanosensitive channels, and calaxin. Our results support that the presence of metachrony augments MCT in a calcium-dependent mechanism.NEW & NOTEWORTHY We developed a novel imaging-based analysis to detect coordination of ciliary motion and optimal coordination, a process called metachrony. We found that metachrony is key to the optimization of ciliary-mediated mucus transport in both ferret and human tracheal tissue. This process appears to be regulated through calcium-dependent mechanisms. This study demonstrates the capacity to measure a key feature of ciliary coordination that may be important in genetic and acquired disorders of ciliary function.

Evaluation of online teaching modules for PSMA PET interpretation.

PURPOSE: The proliferation of US FDA-approved prostate-specific membrane antigen (PSMA)-targeted positron emission tomography (PET) imaging agents as a means to evaluate prostate cancer patients, and the expanding knowledge of interpretive pitfalls, has led to the generation of multiple online training modules geared toward the reading of each individual agent, each taking different approaches to criteria for interpretation, which may contribute to the variability of reporting in clinical practice. MATERIALS AND METHODS: The websites of the marketers of each FDA-approved agent [68Ga-PSMA-11 (Illuccix; Telix Pharmaceuticals), 68Ga-PSMA-11 (Locametz; Novartis Pharmaceuticals), 18F-rh-PSMA-7.3 (Posluma; Blue Earth Diagnostics)], and the website of the Society of Nuclear Medicine and Molecular Imaging [18F-DCFPyL (Pylarify)] were examined. All information pertaining to reader training, including videos, PDFs, and PowerPoint presentations, were reviewed. RESULTS: Videos from each module covered interpretive approach and pitfalls and ranged in length from a total of 20 min up to 315 min. Each module provided a different approach to PSMA PET scan findings, and on a different number and breadth of interpretive tips and pitfalls (a total of approximately 12-30 in all). CONCLUSIONS: Each of the four PSMA PET reader training modules covered important interpretive pitfalls. The lengths of the video portions of each module varied considerably, suggesting variable investments in time necessary to complete each module. The differences in the modules could contribute to inconsistency among readers depending on which module(s) they may have completed and which radiotracer(s) they are using.

Implementation of PSMA PET/CT and alignment of ordering to SNMMI appropriate use criteria in a large network system.

INTRODUCTION: The Society of Nuclear Medicine and Molecular Imaging (SNMMI) provides appropriate use criteria (AUC) for prostate-specific membrane antigen positron emission tomography/computed tomography (PSMA PET/CT) which include guidance on imaging in newly diagnosed prostate cancer and in patients with biochemically recurrent (BCR) disease. This study aims to examine trends in PSMA implementation and the prevalence and outcomes of scans ordered in scenarios deemed rarely appropriate or not meeting SNMMI AUC. METHODS: We retrospectively identified patients who were diagnosed with presumptive National Comprehensive Cancer Network unfavorable intermediate, high, or very high risk prostate cancer, patients who underwent staging for BCR, and all patients staged with PSMA between July 2021 and March 2023. Positivity was validated by adherence to a predetermined reference standard. RESULTS: The frequency of PSMA use increased in initial staging from 24% to 80% and work-up of BCR from 91% to 99% over our study period. In addition, 5% (17/340) of PSMA scans ordered for initial staging did not meet AUC and 3% (15/557) of posttreatment scans were deemed rarely appropriate. Initial staging orders not meeting SNMMI AUC resulted in no positivity (0/17), while rarely appropriate posttreatment scans were falsely positive in 75% (3/4) of cases. Urologists (53%, 17/32) comprised the largest ordering specialty in rarely appropriate use. CONCLUSION: The frequency of PSMA use rose across the study period. A significant minority of patients received PSMA PET/CT in rarely appropriate scenarios yielding no positivity in initial staging and significant false positivity post-therapy. Further education of providers and electronic medical record-based interventions could help limit the rarely appropriate use of PET imaging.

Triple Therapy for Cystic Fibrosis Phe508del-Gating and -Residual Function Genotypes.

BACKGROUND: Elexacaftor-tezacaftor-ivacaftor is a small-molecule cystic fibrosis transmembrane conductance regulator (CFTR) modulator regimen shown to be efficacious in patients with at least one Phe508del allele, which indicates that this combination can modulate a single Phe508del allele. In patients whose other CFTR allele contains a gating or residual function mutation that is already effectively treated with previous CFTR modulators (ivacaftor or tezacaftor-ivacaftor), the potential for additional benefit from restoring Phe508del CFTR protein function is unclear. METHODS: We conducted a phase 3, double-blind, randomized, active-controlled trial involving patients 12 years of age or older with cystic fibrosis and Phe508del-gating or Phe508del-residual function genotypes. After a 4-week run-in period with ivacaftor or tezacaftor-ivacaftor, patients were randomly assigned to receive elexacaftor-tezacaftor-ivacaftor or active control for 8 weeks. The primary end point was the absolute change in the percentage of predicted forced expiratory volume in 1 second (FEV1) from baseline through week 8 in the elexacaftor-tezacaftor-ivacaftor group. RESULTS: After the run-in period, 132 patients received elexacaftor-tezacaftor-ivacaftor and 126 received active control. Elexacaftor-tezacaftor-ivacaftor resulted in a percentage of predicted FEV1 that was higher by 3.7 percentage points (95% confidence interval [CI], 2.8 to 4.6) relative to baseline and higher by 3.5 percentage points (95% CI, 2.2 to 4.7) relative to active control and a sweat chloride concentration that was lower by 22.3 mmol per liter (95% CI, 20.2 to 24.5) relative to baseline and lower by 23.1 mmol per liter (95% CI, 20.1 to 26.1) relative to active control (P<0.001 for all comparisons). The change from baseline in the Cystic Fibrosis Questionnaire-Revised respiratory domain score (range, 0 to 100, with higher scores indicating better quality of life) with elexacaftor-tezacaftor-ivacaftor was 10.3 points (95% CI, 8.0 to 12.7) and with active control was 1.6 points (95% CI, -0.8 to 4.1). The incidence of adverse events was similar in the two groups; adverse events led to treatment discontinuation in one patient (elevated aminotransferase level) in the elexacaftor-tezacaftor-ivacaftor group and in two patients (anxiety or depression and pulmonary exacerbation) in the active control group. CONCLUSIONS: Elexacaftor-tezacaftor-ivacaftor was efficacious and safe in patients with Phe508del-gating or Phe508del-residual function genotypes and conferred additional benefit relative to previous CFTR modulators. (Funded by Vertex Pharmaceuticals; VX18-445-104 ClinicalTrials.gov number, NCT04058353.).

Comparison of a novel potentiator of CFTR channel activity to ivacaftor in ameliorating mucostasis caused by cigarette smoke in primary human bronchial airway epithelial cells.

BACKGROUND: Cystic Fibrosis causing mutations in the gene CFTR, reduce the activity of the CFTR channel protein, and leads to mucus aggregation, airway obstruction and poor lung function. A role for CFTR in the pathogenesis of other muco-obstructive airway diseases such as Chronic Obstructive Pulmonary Disease (COPD) has been well established. The CFTR modulatory compound, Ivacaftor (VX-770), potentiates channel activity of CFTR and certain CF-causing mutations and has been shown to ameliorate mucus obstruction and improve lung function in people harbouring these CF-causing mutations. A pilot trial of Ivacaftor supported its potential efficacy for the treatment of mucus obstruction in COPD. These findings prompted the search for CFTR potentiators that are more effective in ameliorating cigarette-smoke (CS) induced mucostasis. METHODS: Small molecule potentiators, previously identified in CFTR binding studies, were tested for activity in augmenting CFTR channel activity using patch clamp electrophysiology in HEK-293 cells, a fluorescence-based assay of membrane potential in Calu-3 cells and in Ussing chamber studies of primary bronchial epithelial cultures. Addition of cigarette smoke extract (CSE) to the solutions bathing the apical surface of Calu-3 cells and primary bronchial airway cultures was used to model COPD. Confocal studies of the velocity of fluorescent microsphere movement on the apical surface of CSE exposed airway epithelial cultures, were used to assess the effect of potentiators on CFTR-mediated mucociliary movement. RESULTS: We showed that SK-POT1, like VX-770, was effective in augmenting the cyclic AMP-dependent channel activity of CFTR. SK-POT-1 enhanced CFTR channel activity in airway epithelial cells previously exposed to CSE and ameliorated mucostasis on the surface of primary airway cultures. CONCLUSION: Together, this evidence supports the further development of SK-POT1 as an intervention in the treatment of COPD.

A revised airway epithelial hierarchy includes CFTR-expressing ionocytes.

The airways of the lung are the primary sites of disease in asthma and cystic fibrosis. Here we study the cellular composition and hierarchy of the mouse tracheal epithelium by single-cell RNA-sequencing (scRNA-seq) and in vivo lineage tracing. We identify a rare cell type, the Foxi1+ pulmonary ionocyte; functional variations in club cells based on their location; a distinct cell type in high turnover squamous epithelial structures that we term 'hillocks'; and disease-relevant subsets of tuft and goblet cells. We developed 'pulse-seq', combining scRNA-seq and lineage tracing, to show that tuft, neuroendocrine and ionocyte cells are continually and directly replenished by basal progenitor cells. Ionocytes are the major source of transcripts of the cystic fibrosis transmembrane conductance regulator in both mouse (Cftr) and human (CFTR). Knockout of Foxi1 in mouse ionocytes causes loss of Cftr expression and disrupts airway fluid and mucus physiology, phenotypes that are characteristic of cystic fibrosis. By associating cell-type-specific expression programs with key disease genes, we establish a new cellular narrative for airways disease.

Expanded experience with cardiovascular black blood cinematic rendering.

Black blood cinematic rendering (BBCR) is a newly described preset for cinematic rendering, which creates photorealistic displays from volumetric data sets with the contrast-enhanced blood pool displayed as dark and transparent. That set of features potentially provides for enhanced visualization of endomyocardial and intraluminal pathology, as well as cardiac devices. The similarity of the images to black-blood magnetic resonance imaging (MRI) may allow for expansion of the evaluation of certain types of pathology into patient populations unable to undergo MRI. In the emergency setting, the rapid acquisition time and reasonable post-processing time make this technique clinically feasible. In this expanded experience, we demonstrate an expanded clinical experience with the BBCR technique, highlighting the applications for intraluminal cardiovascular evaluation, especially focused on current and potential emergency radiology applications.

Molecular Imaging of Infections: Emerging Techniques for Pathogen-Specific Diagnosis and Guided Therapy.

Evaluation of patients that may be infected is challenging. Imaging to identify or localize a site of infection is often limited because of the nonspecific nature of the findings on conventional imaging modalities. Available imaging methods lack the ability to determine if antibiotics are reaching the site of infection and are not optimized to follow response to therapy. Positron emission tomography (PET) is a method by which radiolabeled molecules can be used to detect metabolic perturbations or levels of expression of specific targets. The most common PET agent is the glucose analog 2-deoxy-2-[18F]fluoro-D-glucose (18F-FDG). 18F-FDG has some applicability to localizing a site of infection, but its lack of specificity limits its usefulness. There is a need for the development of pathogen-specific PET radiotracers to address the imaging shortcomings noted above. Preclinical and clinical progress has been made, but significant challenges remain.

Mucociliary clearance augmenting drugs block SARS-CoV-2 replication in human airway epithelial cells.

The coronavirus disease (COVID-19) pandemic, caused by SARS-CoV-2 coronavirus, is devastatingly impacting human health. A prominent component of COVID-19 is the infection and destruction of the ciliated respiratory cells, which perpetuates dissemination and disrupts protective mucociliary transport (MCT) function, an innate defense of the respiratory tract. Thus, drugs that augment MCT could improve the barrier function of the airway epithelium and reduce viral replication and, ultimately, COVID-19 outcomes. We tested five agents known to increase MCT through distinct mechanisms for activity against SARS-CoV-2 infection using a model of human respiratory epithelial cells terminally differentiated in an air/liquid interphase. Three of the five mucoactive compounds tested showed significant inhibitory activity against SARS-CoV-2 replication. An archetype mucoactive agent, ARINA-1, blocked viral replication and therefore epithelial cell injury; thus, it was further studied using biochemical, genetic, and biophysical methods to ascertain the mechanism of action via the improvement of MCT. ARINA-1 antiviral activity was dependent on enhancing the MCT cellular response, since terminal differentiation, intact ciliary expression, and motion were required for ARINA-1-mediated anti-SARS-CoV2 protection. Ultimately, we showed that the improvement of cilia movement was caused by ARINA-1-mediated regulation of the redox state of the intracellular environment, which benefited MCT. Our study indicates that intact MCT reduces SARS-CoV-2 infection, and its pharmacologic activation may be effective as an anti-COVID-19 treatment.

The synthetic aminoglycoside ELX-02 induces readthrough of G550X-CFTR producing superfunctional protein that can be further enhanced by CFTR modulators.

Ten percent of cystic fibrosis (CF) patients carry a premature termination codon (PTC); no mutation-specific therapies exist for these individuals. ELX-02, a synthetic aminoglycoside, suppresses translation termination at PTCs (i.e., readthrough) by promoting the insertion of an amino acid at the PTC and restoring expression of full-length CFTR protein. The identity of amino acids inserted at PTCs affects the processing and function of the resulting full-length CFTR protein. We examined readthrough of the rare G550X-CFTR nonsense mutation due to its unique properties. We found that forskolin-induced swelling in G550X patient-derived intestinal organoids (PDOs) was significantly higher than in G542X PDOs (both UGA PTCs) with ELX-02 treatment, indicating greater CFTR function from the G550X allele. Using mass spectrometry, we identified tryptophan as the sole amino acid inserted in the G550X position during ELX-02- or G418-mediated readthrough, which differs from the three amino acids (cysteine, arginine, and tryptophan) inserted in the G542X position after treatment with G418. Compared with wild-type CFTR, Fischer rat thyroid (FRT) cells expressing the G550W-CFTR variant protein exhibited significantly increased forskolin-activated Cl- conductance, and G550W-CFTR channels showed increased PKA sensitivity and open probability. After treatment with ELX-02 and CFTR correctors, CFTR function rescued from the G550X allele in FRTs reached 20-40% of the wild-type level. These results suggest that readthrough of G550X produces greater CFTR function because of gain-of-function properties of the CFTR readthrough product that stem from its location in the signature LSGGQ motif found in ATP-binding cassette (ABC) transporters. G550X may be a particularly sensitive target for translational readthrough therapy.NEW & NOTEWORTHY We found that forskolin-induced swelling in G550X-CFTR patient-derived intestinal organoids (PDOs) was significantly higher than in G542X-CFTR PDOs after treatment with ELX-02. Tryptophan (W) was the sole amino acid inserted in the G550X position after readthrough. Resulting G550W-CFTR protein exhibited supernormal CFTR activity, PKA sensitivity, and open probability. These results show that aminoglycoside-induced readthrough of G550X produces greater CFTR function because of the gain-of-function properties of the CFTR readthrough product.

Baseline PSMA-PET/CT as a predictor of PSA persistence following radical prostatectomy in high-risk nonmetastatic prostate cancer patients receiving neoadjuvant therapy.

BACKGROUND: The precise staging and proper management of high-risk prostate cancer (PCa) continues to be a challenge. We aimed to demonstrate the prognostic value of baseline prostate-specific membrane antigen-ligand positron emission tomography/computed tomography (PSMA-PET/CT) in high-risk, nonmetastatic PCa patients who received neoadjuvant hormonal or chemohormonal treatment followed by radical prostatectomy (RP). METHODS: We performed retrospective analyses of 70 patients with high-risk, nonmetastatic PCa confirmed by biopsy between 2018 and 2021. All patients underwent neoadjuvant therapy followed by RP and pelvic lymph node dissection (PLND); PSMA-PET/CT was performed before initiation of neoadjuvant therapy. Acquired image information and clinical characteristics/outcomes were examined for possible associations. RESULTS: Among 70 high-risk PCa patients, median age was 69 years old and median prostate specific antigen (PSA) at presentation was 58.5 ng/mL. Thirty (42.9%) patients had uptake of the PSMA tracer only in the primary PCa lesions and 40 (57.1%) patients had PSMA-positive lesions in regional or distant sites. Sixteen (32%) localized PCa patients defined by pre-PET magnetic resonance imaging were found to have locally advanced PCa based on PSMA-PET/CT. Fifteen (30%) localized PCa patients and 7 (35%) locally advanced PCa patients were upstaged to metastatic PCa. The sensitivity and specificity of PSMA-PET/CT for the detection of lymph node involvement were 90.9% and 69.5%, respectively, with a positive prediction value of 35.7% and negative prediction value of 97.6%. The diagnostic accuracy was 72.9%. Univariate analysis showed upstaging, tumor stage, and metastasis location based on PSMA-PET/CT are predictors to PSA persistence after surgery, while multivariate logistic regression analysis showed only the tumor stage based on PSMA-PET/CT remained an independent predictor of the outcome. CONCLUSIONS: This study further highlights the accuracy and necessity of PSMA-PET/CT in newly diagnosed, high-risk, nonmetastatic PCa patients.

Test-retest repeatability of organ uptake on PSMA-targeted 18 F-DCFPyL PET/CT in patients with prostate cancer.

OBJECTIVES: We evaluated 18 F-DCFPyL test-retest repeatability of uptake in normal organs. METHODS: Twenty-two prostate cancer (PC) patients underwent two 18 F-DCFPyL PET scans within 7 days within a prospective clinical trial (NCT03793543). In both PET scans, uptake in normal organs (kidneys, spleen, liver, and salivary and lacrimal glands) was quantified. Repeatability was determined by using within-subject coefficient of variation (wCOV), with lower values indicating improved repeatability. RESULTS: For SUVmean , repeatability was high for kidneys, spleen, liver, and parotid glands (wCOV, range: 9.0%-14.3%) and lower for lacrimal (23.9%) and submandibular glands (12.4%). For SUVmax , however, the lacrimal (14.4%) and submandibular glands (6.9%) achieved higher repeatability, while for large organs (kidneys, liver, spleen, and parotid glands), repeatability was low (range: 14.1%-45.2%). CONCLUSION: We found acceptable repeatability of uptake on 18 F-DCFPyL PET for normal organs, in particular for SUVmean in the liver or parotid glands. This may have implications for both PSMA-targeted imaging and treatment, as patient selection for radioligand therapy and standardized frameworks for scan interpretation (PROMISE, E-PSMA) rely on uptake in those reference organs.

Lack of repeatability of radiomic features derived from PET scans: Results from a 18 F-DCFPyL test-retest cohort.

OBJECTIVES: PET-based radiomic metrics are increasingly utilized as predictive image biomarkers. However, the repeatability of radiomic features on PET has not been assessed in a test-retest setting. The prostate-specific membrane antigen-targeted compound 18 F-DCFPyL is a high-affinity, high-contrast PET agent that we utilized in a test-retest cohort of men with metastatic prostate cancer (PC). METHODS: Data of 21 patients enrolled in a prospective clinical trial with histologically proven PC underwent two 18 F-DCFPyL PET scans within 7 days, using identical acquisition and reconstruction parameters. Sites of disease were segmented and a set of 29 different radiomic parameters were assessed on both scans. We determined repeatability of quantification by using Pearson's correlations, within-subject coefficient of variation (wCOV), and Bland-Altman analysis. RESULTS: In total, 230 lesions (177 bone, 38 lymph nodes, 15 others) were assessed on both scans. For all investigated radiomic features, a broad range of inter-scan correlation was found (r, 0.07-0.95), with acceptable reproducibility for entropy and homogeneity (wCOV, 16.0% and 12.7%, respectively). On Bland-Altman analysis, no systematic increase or decrease between the scans was observed for either parameter (±1.96 SD: 1.07/-1.30, 0.23/-0.18, respectively). The remaining 27 tested radiomic metrics, however, achieved unacceptable high wCOV (≥21.7%). CONCLUSION: Many common radiomic features derived from a test-retest PET study had poor repeatability. Only Entropy and homogeneity achieved good repeatability, supporting the notion that those image biomarkers may be incorporated in future clinical trials. Those radiomic features based on high frequency aspects of images appear to lack the repeatability on PET to justify further study.