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Extensive clinical data from liver-mediated gene therapy trials have shown that dose-dependent immune responses against the vector capsid may impair or even preclude transgene expression if not managed successfully with prompt immune suppression. The goal of this preclinical study was to generate an adeno-associated viral (AAV) vector capable of expressing therapeutic levels of B-domain deleted factor VIII (FVIII) at the lowest possible vector dose to minimize the potential Risk of a capsid-mediated immune response in the clinical setting. Here, we describe the studies that identified the investigational agent SPK-8011, currently being evaluated in a phase 1/2 study (NCT03003533) in individuals with hemophilia A. In particular, the potency of our second-generation expression cassettes was evaluated in mice and in non-human primates using two different bioengineered capsids (AAV-Spark100 and AAV-Spark200). At 2 weeks after gene transfer, primates transduced with 2 × 1012 vg/kg AAV-Spark100-FVIII or AAV-Spark200-FVIII expressed FVIII antigen levels of 13% ± 2% and 22% ± 6% of normal, respectively. Collectively, these preclinical results validate the feasibility of lowering the AAV capsid dose for a gene-based therapeutic approach for hemophilia A to a dose level orders of magnitude lower than the first-generation vectors in the clinic., Graphical Abstract, The studies presented here represent the proof-of-concept work that generated the investigational agent SPK-8011, currently being evaluated in a phase 1/2 study (NCT03003533) for treatment of hemophilia A. Through a multi-pronged optimization approach, an adeno-associated viral vector capable of expressing therapeutic levels of hFVIII at the lowest possible vector dose was generated.

BACKGROUND: Small bowel neuroendocrine tumors (SB-NET) frequently metastasize to regional lymphatic or distant sites. While most prognostication of SB-NET focuses on lymph node involvement, findings from studies of NETs from other primary sites have suggested that preoperative serum chromogranin-A (CgA) levels may provide a more accurate metric. STUDY DESIGN: Using the National Cancer Database (2004–2016), we analyzed patients with locoregional SB-NET who underwent curative resection including an adequate lymphadenectomy (n = 1,274). A statistically optimized cut-point was used to dichotomize CgA cohort based on preoperative serum CgA levels. RESULTS: We determined that a CgA ≥139ng/mL identified patients with significantly shorter estimated mean overall survival (6.6 years vs. 7.6 years, log-rank p = 0.00001). These patients were also older (63 vs. 57 years, p < 0.001) and more likely to have poorly-differentiated tumors (2.1% vs. 0.7%, p = 0.04) or primary tumors >1cm (88.2% vs. 79.2%, p = 0.001). Clinical features associated with shorter overall survival included preoperative CgA ≥139ng/mL (HR = 2.19, 95% CI 1.22 – 3.92; p = 0.009), age at diagnosis (HR = 1.06, 95% CI 1.03 – 1.09; p < 0.001), Charlson-Deyo score ≥2 (HR = 3.93, 95% CI 1.71 – 9.01; p = 0.001), and poorly-differentiated tumors (HR = 11.22, 95% CI 4.16 – 30.24; p < 0.001). Neither lymph node metastasis nor T-stage were independently associated with shorter overall survival in patients with locoregional SB-NET. CONCLUSIONS: Elevated preoperative serum CgA is an adverse prognostic marker associated with shorter overall survival in patients with locoregional SB-NET.

Noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP) are follicular-patterned thyroid neoplasms defined by nuclear atypia and indolent behavior. They harbor RAS mutations, rather than BRAFV600E mutations as is observed in papillary thyroid carcinomas with extensive follicular growth. Reliably identifying NIFTPs aids in safe therapy de-escalation, but has proven to be challenging due to interobserver variability and morphologic heterogeneity. The genomic scoring system BRS (BRAF-RAS score) was developed to quantify the extent to which a tumor’s expression profile resembles a BRAFV600E or RAS-mutant neoplasm. We proposed that deep learning prediction of BRS could differentiate NIFTP from other follicular-patterned neoplasms. A deep learning model was trained on slides from a dataset of 115 thyroid neoplasms to predict tumor subtype (NIFTP, PTC-EFG, or classic PTC), and was used to generate predictions for 497 thyroid neoplasms within The Cancer Genome Atlas (TCGA). Within follicular-patterned neoplasms, tumors with positive BRS (RAS-like) were 8.5 times as likely to carry an NIFTP prediction than tumors with negative BRS (89.7% vs 10.5%, P

Background Peptide receptor radionuclide therapy is a targeted therapy used to treat unresectable somatostatin receptor-positive neuroendocrine tumors. The objective of this study was to evaluate response rates among neuroendocrine tumors of different primaries and identify factors relevant to future treatment strategies. Methods We retrospectively reviewed patients who received peptide receptor radionuclide therapy for neuroendocrine tumors from 2018 to 2019 at our institution. Patients were assessed with computed tomography/magnetic resonance imaging and 68Ga-DOTATATE-positron emission tomography before and after 2 or 4 cycles of peptide receptor radionuclide therapy. Tumor response was evaluated by RECIST 1.1. Statistics included multinomial logistic regression models and Fisher exact test. Results Twenty-seven patients underwent 92 cycles of peptide receptor radionuclide therapy: pancreas (n = 11), small bowel (n = 7), and other (n = 9) neuroendocrine tumors. Overall, 30% (8 of 27) had partial response, 59% (16 of 27) stable disease, and 11% (3 of 27) progressed. Pancreatic neuroendocrine tumors responded differently from small bowel neuroendocrine tumors regardless of cycle number (P = .01). The majority of pancreatic neuroendocrine tumors (6 of 11) had partial response to peptide receptor radionuclide therapy, while all small bowel neuroendocrine tumors had stable disease. Pancreatic neuroendocrine tumors stable after 2 cycles were more likely to respond to additional cycles versus other neuroendocrine tumors (probability: 60% vs 11%). Conclusion Patients with unresectable advanced or metastatic pancreatic neuroendocrine tumors may benefit from a full course of peptide receptor radionuclide therapy, whereas other neuroendocrine tumors appear less likely to respond. Large prospective studies are needed to confirm these findings.

Background Indoleamine 2,3-dioxygenase 1 (IDO-1) is overexpressed in many human carcinomas and a successful target for therapy in mouse models. Prognosis of patients with advanced adrenocortical carcinoma (ACC) is poor due to the lack of effective treatments, and new therapies are therefore needed. Herein, we investigate whether IDO-1 is expressed in human ACC tissues. Methods 53 tissue samples from patients with ACC, adrenal adenoma (AA), adrenocortical tumors (ACTs), and normal adrenal were identified. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded slides for IDO-1. Samples were scored for cytoplasmic staining as per intensity and the percent of positive cells and for stromal staining by percent of positive cells. Tumor characteristics, PD-L1, PDL-2, and CD-8+ T-lymphocyte expression were also determined. Results Samples from 32 ACC, 3 ACT, 15 AA, and 3 normal adrenal were analyzed. IDO-1 was expressed in tumor tissue in 22 of 32 ACC samples, compared with 8 of 15 AA sample (P = 0.344). IDO-1 expression was significantly increased in stromal tissue of ACC samples (16 of 33), compared with AA samples (0 of 15) (P = 0.001). IDO-1 expression in ACC and AA samples was associated with PD-L2 expression (P = 0.034). IDO-1 expression in ACC stromal tissue was associated with CD8+ T-lymphocyte infiltration (P = 0.028). Conclusions IDO-1 is expressed in a majority of ACC samples. Its expression in tumor tissue is associated with PD-L2 expression, and expression in stroma is associated with CD8+ cell infiltration. IDO-1 inhibition, alone or in combination with PD-1 inhibition, could therefore be an interesting target in treatment of ACC.

Positive resection margins are associated with worse survival after surgery for adrenocortical carcinoma (ACC). We aimed to identify risk factors for positive margins post-resection.The NCDB was queried for ACC patients from 2006 to 2015. Patients with positive versus negative resection margins post-surgery were compared using Chi-square tests. Survival based on adjuvant treatment was assessed using Kaplan-Meier curves.1,973 patients with ACC were identified, 217 (11.0%) with positive margins. Multivariable analysis identified extra-adrenal extension (HR 4.92, p 0.001), lymph node metastases (HR 2.64, p = 0.001), and distant metastases (HR 1.53, p = 0.03) as risk factors for positive margins. No significant difference in margin status existed between patients who had an open versus minimally invasive procedure (p = 0.6). Positive margin patients receiving adjuvant radiation (p = 0.007) or combined chemo-radiation (p = 0.001) had the longest survival.No modifiable risk factors were identified, but patients with positive margins receiving adjuvant radiation or chemo-radiation had the longest survival.

Surgical management of pancreatic neuroendocrine tumors (PNETS) is steadily evolving and is influenced by multiple factors. Sporadic PNETs are generally managed more aggressively than those occurring in the background of hereditary syndromes, and functioning PNETs are almost always resected if they are not metastatic. Localized nonfunctioning PNETs less than 2 cm can often be observed. Surgical resection for localized PNET greater than 2 cm comprises parenchymal sparing pancreas resections, such as enucleations, or formal anatomic resection, such as distal pancreatectomy or pancreaticoduodenectomy. PNETs commonly metastasize to the liver, and several systemic and liver-directed options to treat hepatic metastases are available.

Many current guidelines recommend nonoperative management for pancreatic neuroendocrine tumors2 cm. The objective of this study was to evaluate the utilization and outcomes of resection for these pancreatic neuroendocrine tumors in the United States.Using the National Cancer Database (2004-2014), 3,243 cases of T1 (≤2.0 cm) pancreatic neuroendocrine tumors were identified. Additional patient and tumor characteristics were examined. Multivariate models were used to identify factors that predicted resection and to assess patient survival after resection.75% of pancreatic neuroendocrine tumors measuring 0 to 1.0 cm and 80% of pancreatic neuroendocrine tumors measuring1.0 and ≤2.0 cm were resected. Eighty-four pancreatic neuroendocrine tumors were functional, of which 82% were resected. Variables influencing resection included positive lymph nodes, tumor in body or tail of pancreas, well or moderately differentiated tumors, and resection at academic medical centers (odds ratio 1.5-4.9). When controlling for other variables, patients with pancreatic neuroendocrine tumors 1 to 2 cm who underwent resection had a prolonged 5-year survival rate (hazard ratio 0.51, confidence interval 0.34-0.75) when compared with those who did not undergo resection. This survival benefit of resection was not found for pancreatic neuroendocrine tumors 0 to 1 cm (hazard ratio = 0.63, confidence interval 0.36-1.11).Contrary to many current recommendations, most patients with pancreatic neuroendocrine tumors ≤2.0 cm undergo surgical resection in the United States. A survival benefit was found for resection of pancreatic neuroendocrine tumors 1 to 2 cm, suggesting that current recommendations should perhaps be revised.

Background 68Gallium-DOTATATE positron emission tomography–computed tomography (PET CT) has shown superior accuracy in detecting grade 1 and 2 neuroendocrine tumors over previous imaging modalities and was recently included in National Comprehensive Cancer Network guidelines. It remains unclear which patients benefit most from this imaging modality. We therefore reviewed our initial experience with 68Gallium-DOTATATE PET CT to evaluate its usefulness in diagnosing, staging, and surveilling neuroendocrine tumors. Methods Records of patients who underwent 68Gallium-DOTATATE PET CT from March to December 2017 were prospectively evaluated. The primary endpoint was whether 68Gallium-DOTATATE PET CT changes treatment in patients with neuroendocrine tumors. Descriptive statistics, Fisher exact tests, and nested logistic regressions were conducted. Results A total of 50 consecutive patients were included. Of these, 41 patients (82%) had a biopsy-proven neuroendocrine tumor at the time of imaging. The remaining 9 patients (18%) had symptoms or biochemistry suggestive of a neuroendocrine tumor with negative cross-sectional imaging. 68Gallium-DOTATATE PET CT changed management in 33 patients (66%). There were 24 patients with intermodality changes in management and 9 patients with intramodality changes in management. Patients with scans performed for staging had a higher likelihood of a change in management (P = .006). Conclusion Performing 68Gallium-DOTATATE PET CT should be considered for staging and surveillance of neuroendocrine tumors because it is frequently associated with changes in management.

We map glacial cirques, and analyse spatial variability in their altitude and aspect to derive a long-term, time-integrated, perspective on climate patterns during former periods of mountain glaciation (likely spanning multiple Quaternary glaciations) in Britain and Ireland. The data reveal that, although air temperatures were important, exposure to moisture-bearing air masses was the key factor in regulating sites of former mountain glacier formation, and indicate that during such periods, moisture supply was largely controlled by North Atlantic westerlies, with notable inland precipitation gradients (precipitation decreasing inland), similar to present day. In places, trends in cirque altitude may also reflect regional differences in the extent of cirque deepening, controlled by the dimensions and dynamics of the glaciers that came to occupy them. Specifically, comparatively deep cirques in coastal locations may reflect the former presence of dynamic (fed by moisture from the North Atlantic), but comparatively small, glaciers (largely confined to their cirques). By contrast, decreasing cirque depth further inland, may reflect the former presence of larger and/or less dynamic ice masses, occupying comparatively continental climatic conditions.

A missense point mutation in nerve growth factor (NGFR100W) is associated with hereditary sensory autonomic neuropathy V (HSAN V), originally discovered in a Swedish family. These patients develop severe loss of perception to deep pain but with apparently normal cognitive functions. To better understand the disease mechanism, we have generated the first NGFR100Wknockin mouse model of HSAN V. Mice homozygous for the NGFR100Wmutation (NGFfln/fln) showed significant structural deficits in intra-epidermal nerve fibers (IENFs) at birth. These mice had a total loss of pain perception at ∼2 months of age and they often failed to survive to full adulthood. Heterozygous mice (NGF+/fln) developed a progressive degeneration of small sensory fibers both behaviorally and functionally: they showed a progressive loss of IENFs starting at the age of 9 months accompanied with progressive loss of perception to painful stimuli such as noxious temperature. Quantitative analysis of lumbar 4/5 dorsal root ganglia (DRG) revealed a significant reduction in small size neurons positive for calcitonin gene-related peptide, while analysis of sciatic nerve fibers revealed the mutant NGF+/flnmice had no reduction in myelinated nerve fibers. Significantly, the amount of NGF secreted from fibroblasts were reduced in heterozygous and homozygous mice compared to their wild-type littermates. Interestingly, NGF+/flnshowed no apparent structural alteration in the brain: neither the anterior cingulate cortex nor the medial septum including NGF-dependent basal forebrain cholinergic neurons. Accordingly, these animals did not develop appreciable deficits in tests for central nervous system function. Our study provides novel insights into the selective impact of NGFR100Wmutation on the development and function of the peripheral sensory system.

Malignant-functioning pancreatic neuroendocrine tumors (mFpNETs) are rare. Research analyzing the presentation, biological behavior, and patient outcomes of these tumors is limited.We used the Surveillance, Epidemiology, and End Results database to identify patients with malignant insulinomas, gastrinomas, glucagonomas, vasoactive intestinal peptide secreting tumors (VIPomas), somastatinomas, and mixed islet cell tumors (MICTs). The primary endpoint of this study was to identify factors affecting survival.We identified 401 patients with mFpNETs. Between histologic subtypes, there were significant differences in sex and age, and in tumor size, grade, location, and stage. Median survival time for insulinomas was 12.7 years; gastrinomas, 10.2 years; glucagonomas, 7.7 years; VIPomas, 7.9 years; and MICTs, 3.4 years. Multivariable analysis showed that histology (insulinoma, gastrinoma, and VIPoma; P = .009), absence of distant metastases (P = .002), age50 years (P = .001), surgical intervention (P = .001), and stage I/II disease (P = .011) were independently associated with prolonged survival. Subgroup analysis demonstrated that removal of the primary tumor in stage IV mFpNETs was associated with significantly prolonged survival (P = .01).mFpNETs are rare tumors that commonly present at an advanced stage despite hormonal secretion. Primary tumor resection is associated with longer survival in stages I-III as well as stage IV tumors.

Nonfunctioning pancreatic neuroendocrine tumors (NFpNET) present with distant metastases in up to 50% of patients. It is unknown whether removal of the primary tumor in patients with NFpNET and metastases is beneficial.We used the Surveillance, Epidemiology, and End Results database to identify patients with NFpNET and distant metastases. The primary outcome measure in this study was overall survival.We identified 882 patients with metastatic NFpNET who had survival data; 303 (34%) patients had operative removal of their primary tumor of which 243 (80%) were grade I or II. Median survival of patients undergoing resection of the primary site was 65 (95% confidence interval 60-86) versus 10 (8-12) months for those without resection (P.0001). Patients diagnosed after 2003 (n = 625, 71%) were more likely to undergo an operation than those diagnosed earlier (P = .001). Multivariable analysis showed that a lesser tumor grade (P.0001), younger age (P.0001), diagnosis during or after 2003 (P = .0003), tumor site in the body/tail (P = .009), and operative resection of the primary tumor site (P.0001) were associated with prolonged survival of patients with NFpNET and distant metastases.This study suggests that resection of the site of the primary NFpNET is associated with greater survival in patients with distant metastases and could therefore be considered as a additional treatment option in this patient population.