Diagnostic Accuracy of Circulating Tumor HPV DNA Testing in Patients With a Lateral Neck Mass.

Importance: The most frequent presenting symptom for patients with human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OPSCC) is a lateral neck mass. Circulating tumor tissue-modified viral (TTMV)-HPV DNA is a unique biomarker produced by the fragmentation of HPV DNA during the degradation of HPV-associated tumors, and its detection and quantitation are currently being used as an adjunct to imaging in monitoring for disease recurrence and may have utility for diagnosis. Objective: To measure the diagnostic characteristics of TTMV-HPV DNA compared with gold standard tissue biopsy for diagnosing HPV-OPSCC in patients presenting with an indeterminate lateral neck mass. Design, Setting, and Participants: This prospective diagnostic test study enrolled patients 18 years or older who presented with a lateral neck mass to a large urban tertiary health care system from December 2021 to June 2023. Participants underwent standard-of-care testing to obtain a tissue diagnosis and a single TTMV-HPV DNA measurement. Main Outcomes and Measures: The primary outcome of interest was sensitivity, while specificity, positive predictive value, and negative predictive value were secondary end points. A subset analysis was performed comparing test performance metrics between TTMV-HPV DNA testing and fine-needle aspiration. Results: A total of 138 patients were included, of whom 80 (58.0%) were men, with median age of 57.5 years (IQR, 43.3-67.0 years). Of 138 patients, 87 (63.0%) had neck masses in level 2 and 47 (34.1%) had HPV-OPSCC. TTMV-HPV DNA testing exhibited a sensitivity of 95.7% (95% CI, 85.5%-99.5% [45 of 47 patients]), specificity of 97.8% (95% CI, 92.3%-99.7% [89 of 91 patients]), positive predictive value of 95.7% (95% CI, 85.5%-99.5% [45 of 47 patients]), and negative predictive value of 97.8% (95% CI, 92.3%-99.7% [89 of 91 patients]). Conclusions and Relevance: In this diagnostic study of patients presenting with a lateral neck mass, circulating TTMV-HPV DNA demonstrated excellent diagnostic test characteristics for the detection of HPV-OPSCC. Such testing may have particular utility for patients in whom obtaining adequate tissue is problematic, as is often the case with cystic neck masses and unknown primary tumors.

A Scoping Review of Pain Management After Transoral Robotic Surgery.

OBJECTIVE: Pain following transoral robotic surgery (TORS) is a driver of adverse outcomes and can lead to readmission and treatment delays. A scoping review was conducted to characterize TORS-related pain and identify key management strategies utilized in the literature. DATA SOURCES: OVID Medline, CINAHL, Cochrane, Pubmed, and Embase databases were queried. REVIEW METHODS: Two team members independently screened titles and abstracts and completed full-text reviews. Studies examining TORS for OPSCC with quantitative pain data were included. The study followed the PRISMA guidelines. RESULTS: A total of 1467 studies were imported for screening and 25 studies were ultimately included. The average study sample size was 89 participants. 68% were conducted in a single-center academic setting. Pain was assessed on varying timelines up to 3 years using 13 different metrics. Pain peaks days-weeks postoperatively and returns to baseline thereafter. Postoperative pain is a significant cause of morbidity and limited data exist about optimal management. CONCLUSION: Prospective studies are needed to characterize and address TORS-related pain.

Selective Internal Radiotherapy Alters the Profiles of Systemic Extracellular Vesicles in Hepatocellular Carcinoma.

Incidence of hepatocellular carcinoma (HCC) is increasing globally. Radioembolization (RE)/selective internal radiotherapy (SIRT) is a promising treatment for inoperable HCC. RE triggers an immune response, involving extracellular vesicles (EVs) which are crucial for cell communication and tumor development. This study explores EV immune profiles and origins in patients with inoperable HCC before and after SIRT/RE. Blood samples from 50 HCC-patients treated with SIRT/RE were collected before and after therapy to determine cytokines and isolate EVs using size exclusion chromatography. The dynamic range and EV quality required for detecting variations in surface markers were assessed. Thirty-seven EV surface markers were analyzed using flow cytometry and correlated with clinical parameters. Several immunological markers (CD4, CD2, CD40, CD45, CD49e, CD69, CD209-EVs) were present in the circulation of HCC patients. These markers positively correlated with therapy response and survival. Conversely, B cell CD20, endothelial cell CD146, platelet CD49e, and CD41b EV markers negatively correlated with 60-day survival. Elevated levels of IL-6 and IL-8 before therapy correlated negatively with patient survival, coinciding with a positive correlation with CD20-positive EVs. Plasma EVs from HCC patients exhibit immunological, cancer, and coagulation markers, including potential biomarkers (CD4, CD20, CD49e, CD146). These may enhance our understanding of cancer biology and facilitate SIRT therapy monitoring.

Selective Internal Radiotherapy Changes the Immune Profiles of Extracellular Vesicles and Their Immune Origin in Patients with Inoperable Cholangiocarcinoma.

The incidence of cholangiocellular carcinoma (CCA) is rising worldwide. As there are no specific early symptoms or specific markers of CCA, it is often diagnosed in later inoperable stages. Accumulating evidence underlines the importance of radiation therapy in the induction of antitumor immunity. The surface protein composition on extracellular vesicles (EVs) relates to originating cells and thus may play a role in vesicle function. We assessed immune profiles of EVs and their immune origin in patients with inoperable CCA prior and after selective internal radiotherapy (SIRT). A total of 47 CCA patients receiving SIRT and 12 healthy volunteers (HV) were included. Blood was withdrawn before therapy (pre T) and after T. EVs were purified from plasma by cluster of differentiation (CD)9-, CD63-, and CD81-immunobead isolation. To detect differently abundant surface markers, dynamic range and EVs input quality were assessed. A total of 37 EVs surface markers were measured by flow cytometry and correlated either with the administered activity dose (MBq) or with the interval until death (month). EVs phenotyping identified lymphocytes, B cells, NK cells, platelets, endothelial cells, leukocyte activation, B cell activation, T and B cell adhesion markers, stem/progenitor cells, and antigen-presenting cells (APC) as EVs-parenteral cells. CD4 and CD8 significantly declined, while other markers significantly increased in CCA patients pre T vs. HV. Platelets-deriving EVs significantly decreased, normalizing to levels of HV but still significantly increasing vs. HV post SIRT. B cells-deriving EVs significantly increased pre T vs. HV, positively correlating with administered activity dose. MHCII and CD40 EVs significantly increased pre SIRT and negatively correlated with administered activity dose, while EVs from antigen presenting cells and CD49e pre SIRT positively correlated with survival time after therapy. Increased levels of CD24 and CD44 in cancer pre T were significantly decreased post T. Among the heterogeneity of EVs that was demonstrated, in particular, B cells-deriving, MHCII, and CD40 positive or APC-deriving EVs need to be further studied for their diagnostic or prognostic relevance in clinical scenarios.

Prognosis of Distant Metastatic Sites in Anterior Skull Base Malignancies.

Objective This study aimed to provide information regarding the prognosis of patients presenting with metastatic anterior skull base malignancies based upon histology and site of distant metastasis (DM). Patients and Methods The National Cancer Database was queried for patients with anterior skull base malignant neoplasms with DM. Outcomes Prognosis was compared between site of DM and tumor histologies. A multivariable Cox proportional hazards model was used to identify prognostic factors for overall survival (OS). Results A total of 481 patients were identified. Lung was the most common site of DM (24.9%), followed by bone (22.2%), liver (5.6%), and brain (2.5%). Lung was the most common site for squamous cell carcinoma (SCCa) (28.3%), melanoma (37.7%), and adenoid cystic carcinoma (ACC; 31.4%). The median survival for patients presenting with metastatic disease regardless of tumor histology was 9.0 months (95% confidence interval [CI]: 8.2-10.3), and patients with metastasis to the liver had the best median survival at 15.5 months (95% CI: 10.5-25.6). The median survivals for the most common histologies, SCCa, melanoma, and ACC were 8.2 months (95% CI: 5.5-10.2), 10.5 months (95% CI: 8.7-14.1), and 15.0 months (95% CI: 11.1-61.1), respectively. Multivariable analysis demonstrated worse overall survival (OS) for older patients, higher Charlson-Deyo comorbidity scores, and tumors with higher grade and T stage. Compared with metastasis to bone, lung metastasis had better OS on multivariable analysis (hazard ratio [HR]: 0.70, 95% CI: 0.51-97). Adenoid cystic carcinoma had improved OS compared with SCCa (HR: 0.62, 95% CI: 0.39-99). Conclusion Tumor histology, metastatic sites, and several disease factors affected prognosis in anterior skull base malignancies with DM.

Applicant Perspectives on Virtual Otolaryngology Residency Interviews.

OBJECTIVE: Residency interviews serve as an opportunity for prospective applicants to evaluate programs and to determine their potential fit within them. The 2019 SARS-CoV2 pandemic mandated programs conduct interviews virtually for the first time. The purpose of this study was to assess applicant perspectives on the virtual interview. METHODS: A Qualtrics survey assessing applicant characteristics and attitudes toward the virtual interview was designed and disseminated to otorhinolaryngology applicants from 3 large academic institutions in the 2020 to 2021 application cycle. RESULTS: A total of 33% of survey applicants responded. Most applicants were satisfied with the virtual interview process. Applicants reported relatively poor quality of interactions with residents and an inability to assess the "feel" of a geographic area. Most applicants received at least 11 interviews with over a third of applicants receiving >16 interviews. Only 5% of applicants completed >20 interviews. Most applicants believed interviews should be capped between 15 and 20 interviews. Most applicants reported saving >$5000, with over a quarter of applicants saving >$8000, and roughly one-third of applicants saving at least 2 weeks of time with virtual versus in-person interviews. CONCLUSIONS: While virtual interviews have limitations, applicants are generally satisfied with the experience. Advantages include cost and time savings for both applicants and programs, as well as easy use of technology. Continuation of the virtual interview format could be considered in future application cycles; geographical limitations may be overcome with in-person second looks, and increased emphasis should be placed on resident interactions during and prior to interview day.

The MarrowMiner: A Novel Minimally Invasive and Effective Device for the Harvest of Bone Marrow.

Bone marrow (BM) is a rich source of hematopoietic stem cells (HSCs), mesenchymal stem cells (MSCs), and other important stem/progenitor cells. It is the traditional source of cells used in hematopoietic cell transplantation, which is a proven curative treatment for many blood and immune diseases. BM-derived cells have also been shown to have other diverse clinical uses and are increasingly being used in orthopedic medicine, regenerative medicine, and gene therapy applications. Traditional methods for harvesting BM are crude, tedious, time-consuming, and expensive, requiring multiple bone punctures under general anesthesia with serial small-volume aspirates often diluted with peripheral blood. The MarrowMiner (MM) is a novel device designed for rapid and minimally invasive BM harvest. Here we show the safety and efficacy of the MM in both preclinical and clinical settings. In a large-animal porcine model, the MM enabled effective BM collection with similar total nucleated cell collection and increased colony formation compared with standard methods. The MM was subsequently evaluated in a clinical study showing effective and complication-free anterior and posterior BM collection of 20 patients under only local anesthesia or light sedation. Increased total nucleated and mononucleated cell collection was achieved with the MM compared with standard methods in the same patients. Importantly, stem cell content was high with trends toward increased HSC, MSC, and endothelial progenitor cells with similar T cell content. Given the MM is a novel device approved by the US Food and Drug Administration, enabling safe, effective, and minimally invasive harvest of BM, we anticipate rapid adoption for various applications.

Characterization of three ciliopathy pedigrees expands the phenotype associated with biallelic C2CD3 variants.

Whole exome sequencing (WES) is utilized in diagnostic odyssey cases to identify the underlying genetic cause associated with complex phenotypes. Recent publications suggest that WES reveals the genetic cause in ~25% of these cases and is most successful when applied to children with neurological disease. The residual 75% of cases remain genetically elusive until more information becomes available in the literature or functional studies are pursued. WES performed on three families with presumed ciliopathy diagnoses, including orofaciodigital (OFD) syndrome, fetal encephalocele, or Joubert-related disorder, identified compound heterozygous variants in C2CD3. Biallelic variants in C2CD3 have previously been associated with ciliopathies, including OFD syndrome type 14 (OFD14; MIM: 615948). As three of the six identified variants were predicted to affect splicing, exon-skipping analysis using either RNA sequencing or PCR-based methods were completed to determine the pathogenicity of these variants, and showed that each of the splicing variants led to a frameshifted protein product. Using these studies in combination with the 2015 ACMG guidelines, each of the six identified variants were classified as either pathogenic or likely pathogenic, and are therefore likely responsible for our patients' phenotypes. Each of the families had a distinct clinical phenotype and severity of disease, extending from lethal to viable. These findings highlight that there is a broad phenotypic spectrum associated with C2CD3-mediated disease and not all patients present with the typical features of OFD14.

Overcoming the Immunosuppressive Tumor Microenvironment of Hodgkin Lymphoma Using Chimeric Antigen Receptor T Cells.

Patients with otherwise treatment-resistant Hodgkin lymphoma could benefit from chimeric antigen receptor T-cell (CART) therapy. However, Hodgkin lymphoma lacks CD19 and contains a highly immunosuppressive tumor microenvironment (TME). We hypothesized that in Hodgkin lymphoma, CART should target both malignant cells and the TME. We demonstrated CD123 on both Hodgkin lymphoma cells and TME, including tumor-associated macrophages (TAM). In vitro, Hodgkin lymphoma cells convert macrophages toward immunosuppressive TAMs that inhibit T-cell proliferation. In contrast, anti-CD123 CART recognized and killed TAMs, thus overcoming immunosuppression. Finally, we showed in immunodeficient mouse models that CART123 eradicated Hodgkin lymphoma and established long-term immune memory. A novel platform that targets malignant cells and the microenvironment may be needed to successfully treat malignancies with an immunosuppressive milieu.Significance: Anti-CD123 chimeric antigen receptor T cells target both the malignant cells and TAMs in Hodgkin lymphoma, thereby eliminating an important immunosuppressive component of the tumor microenvironment. Cancer Discov; 7(10); 1154-67. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 1047.

Silent Tyrosinemia Type I Without Elevated Tyrosine or Succinylacetone Associated with Liver Cirrhosis and Hepatocellular Carcinoma.

Tyrosinemia type I (TYRSN1, TYR I) is caused by fumarylacetoacetate hydrolase (FAH) deficiency and affects approximately one in 100,000 individuals worldwide. Pathogenic variants in FAH cause TYRSN1, which induces cirrhosis and can progress to hepatocellular carcinoma (HCC). TYRSN1 is characterized by the production of a pathognomonic metabolite, succinylacetone (SUAC) and is included in the Recommended Uniform Screening Panel for newborns. Treatment intervention is effective if initiated within the first month of life. Here, we describe a family with three affected children who developed HCC secondary to idiopathic hepatosplenomegaly and cirrhosis during infancy. Whole exome sequencing revealed a novel homozygous missense variant in FAH (Chr15(GRCh38):g.80162305A>G; NM_000137.2:c.424A > G; NP_000128.1:p.R142G). This novel variant involves the catalytic pocket of the enzyme, but does not result in increased SUAC or tyrosine, making the diagnosis of TYRSN1 problematic. Testing this novel variant using a rapid, in vivo somatic mouse model showed that this variant could not rescue FAH deficiency. In this case of atypical TYRSN1, we show how reliance on SUAC as a primary diagnostic test can be misleading in some patients with this disease. Augmentation of current screening for TYRSN1 with targeted sequencing of FAH is warranted in cases suggestive of the disorder.

Clonal precursor of bone, cartilage, and hematopoietic niche stromal cells.

Organs are composites of tissue types with diverse developmental origins, and they rely on distinct stem and progenitor cells to meet physiological demands for cellular production and homeostasis. How diverse stem cell activity is coordinated within organs is not well understood. Here we describe a lineage-restricted, self-renewing common skeletal progenitor (bone, cartilage, stromal progenitor; BCSP) isolated from limb bones and bone marrow tissue of fetal, neonatal, and adult mice. The BCSP clonally produces chondrocytes (cartilage-forming) and osteogenic (bone-forming) cells and at least three subsets of stromal cells that exhibit differential expression of cell surface markers, including CD105 (or endoglin), Thy1 [or CD90 (cluster of differentiation 90)], and 6C3 [ENPEP glutamyl aminopeptidase (aminopeptidase A)]. These three stromal subsets exhibit differential capacities to support hematopoietic (blood-forming) stem and progenitor cells. Although the 6C3-expressing subset demonstrates functional stem cell niche activity by maintaining primitive hematopoietic stem cell (HSC) renewal in vitro, the other stromal populations promote HSC differentiation to more committed lines of hematopoiesis, such as the B-cell lineage. Gene expression analysis and microscopic studies further reveal a microenvironment in which CD105-, Thy1-, and 6C3-expressing marrow stroma collaborate to provide cytokine signaling to HSCs and more committed hematopoietic progenitors. As a result, within the context of bone as a blood-forming organ, the BCSP plays a critical role in supporting hematopoiesis through its generation of diverse osteogenic and hematopoietic-promoting stroma, including HSC supportive 6C3(+) niche cells.

In vivo functional and transcriptional profiling of bone marrow stem cells after transplantation into ischemic myocardium.

OBJECTIVE: Clinical trials of bone marrow-derived stem cell therapy for the heart have yielded variable results. The basic mechanism(s) that underlies their potential efficacy remains unknown. In the present study, we evaluated the survival kinetics, transcriptional response, and functional outcome of intramyocardial bone marrow mononuclear cell (BMMC) transplantation for cardiac repair in a murine myocardial infarction model. METHODS AND RESULTS: We used bioluminescence imaging and high-throughput transcriptional profiling to evaluate the in vivo survival kinetics and gene expression changes of transplanted BMMCs after their engraftment into ischemic myocardium. Our results demonstrate short-lived survival of cells following transplant, with less than 1% of cells surviving by 6 weeks posttransplantation. Moreover, transcriptomic analysis of BMMCs revealed nonspecific upregulation of various cell regulatory genes, with a marked downregulation of cell differentiation and maturation pathways. BMMC therapy caused limited improvement of heart function as assessed by echocardiography, invasive hemodynamics, and positron emission tomography. Histological evaluation of cell fate further confirmed findings of the in vivo cell tracking and transcriptomic analysis. CONCLUSIONS: Collectively, these data suggest that BMMC therapy, in its present iteration, may be less efficacious than once thought. Additional refinement of existing cell delivery protocols should be considered to induce better therapeutic efficacy.

Functional and transcriptional characterization of human embryonic stem cell-derived endothelial cells for treatment of myocardial infarction.

BACKGROUND: Differentiation of human embryonic stem cells into endothelial cells (hESC-ECs) has the potential to provide an unlimited source of cells for novel transplantation therapies of ischemic diseases by supporting angiogenesis and vasculogenesis. However, the endothelial differentiation efficiency of the conventional embryoid body (EB) method is low while the 2-dimensional method of co-culturing with mouse embryonic fibroblasts (MEFs) require animal product, both of which can limit the future clinical application of hESC-ECs. Moreover, to fully understand the beneficial effects of stem cell therapy, investigators must be able to track the functional biology and physiology of transplanted cells in living subjects over time. METHODOLOGY: In this study, we developed an extracellular matrix (ECM) culture system for increasing endothelial differentiation and free from contaminating animal cells. We investigated the transcriptional changes that occur during endothelial differentiation of hESCs using whole genome microarray, and compared to human umbilical vein endothelial cells (HUVECs). We also showed functional vascular formation by hESC-ECs in a mouse dorsal window model. Moreover, our study is the first so far to transplant hESC-ECs in a myocardial infarction model and monitor cell fate using molecular imaging methods. CONCLUSION: Taken together, we report a more efficient method for derivation of hESC-ECs that express appropriate patterns of endothelial genes, form functional vessels in vivo, and improve cardiac function. These studies suggest that hESC-ECs may provide a novel therapy for ischemic heart disease in the future.

Endochondral ossification is required for haematopoietic stem-cell niche formation.

Little is known about the formation of niches, local micro-environments required for stem-cell maintenance. Here we develop an in vivo assay for adult haematopoietic stem-cell (HSC) niche formation. With this assay, we identified a population of progenitor cells with surface markers CD45(-)Tie2(-)alpha(V)(+)CD105(+)Thy1.1(-) (CD105(+)Thy1(-)) that, when sorted from 15.5 days post-coitum fetal bones and transplanted under the adult mouse kidney capsule, could recruit host-derived blood vessels, produce donor-derived ectopic bones through a cartilage intermediate and generate a marrow cavity populated by host-derived long-term reconstituting HSC (LT-HSC). In contrast, CD45(-)Tie2(-)alpha(V)(+)CD105(+)Thy1(+) (CD105(+)Thy1(+)) fetal bone progenitors form bone that does not contain a marrow cavity. Suppressing expression of factors involved in endochondral ossification, such as osterix and vascular endothelial growth factor (VEGF), inhibited niche generation. CD105(+)Thy1(-) progenitor populations derived from regions of the fetal mandible or calvaria that do not undergo endochondral ossification formed only bone without marrow in our assay. Collectively, our data implicate endochondral ossification, bone formation that proceeds through a cartilage intermediate, as a requirement for adult HSC niche formation.

Efficient transplantation via antibody-based clearance of hematopoietic stem cell niches.

Upon intravenous transplantation, hematopoietic stem cells (HSCs) can home to specialized niches, yet most HSCs fail to engraft unless recipients are subjected to toxic preconditioning. We provide evidence that, aside from immune barriers, donor HSC engraftment is restricted by occupancy of appropriate niches by host HSCs. Administration of ACK2, an antibody that blocks c-kit function, led to the transient removal of >98% of endogenous HSCs in immunodeficient mice. Subsequent transplantation of these mice with donor HSCs led to chimerism levels of up to 90%. Extrapolation of these methods to humans may enable mild but effective conditioning regimens for transplantation.

Differentiation, survival, and function of embryonic stem cell derived endothelial cells for ischemic heart disease.

BACKGROUND: Embryonic stem (ES) cells are distinguished by their capacity for self-renewal and pluripotency. Here we characterize the differentiation of ES cell-derived endothelial cells (ESC-ECs), use molecular imaging techniques to examine their survival in vivo, and determine the therapeutic efficacy of ESC-ECs for restoration of cardiac function after ischemic injury. METHODS AND RESULTS: Murine ES cells were transfected with a construct composed of a vascular endothelial cadherin promoter driving enhanced green fluorescence protein (pVE-cadherin-eGFP). Differentiation of ES cells to ECs was detected by FACS analysis using Flk-1 (early EC marker at day 4) and VE-cadherin (late EC marker at day 8). After isolation, these ESC-ECs express endothelial cell markers similar to adult mouse lung endothelial cells, form vascular-like channels, and incorporate DiI-labeled acetylated low-density lipoprotein (DiI-Ac-LDL). For in vivo imaging, ES cells were transduced with an ubiquitin promoter driving firefly luciferase and monomeric red fluorescence protein (pUb-Fluc-mRFP). A robust correlation exists between Fluc signals and cell numbers by ex vivo imaging analysis (R2=0.98) and by in vitro enzyme assay (R2=0.94). Afterward, 5x10(5) ESC-ECs or PBS (as control) was injected into the hearts of mice undergoing LAD ligation (n=15 per group). Bioluminescence imaging showed longitudinal survival of transplanted ESC-ECs for approximately 8 weeks. Echocardiogram demonstrated significant functional improvement in the ESC-EC group compared with control (P=0.04). Finally, postmortem analysis confirmed increased presence of small capillaries and venules in the infarcted zones by CD31 staining. CONCLUSIONS: This is the first study to track the fate and function of transplanted ESC-ECs in the heart. With further validation, these ESC-ECs could become a valuable source of cell therapy for induction of angiogenesis in the treatment of myocardial ischemia.

Porcine thymic grafts protect human thymocytes from HIV-1-induced destruction.

Human immunodeficiency virus type 1 (HIV-1) infection depletes thymocytes and destroys thymic structure. Functional, tolerant human T cells develop in vivo in immunodeficient mice receiving porcine thymus and human fetal liver fragments under the kidney capsule. In this model, we evaluated the potential of porcine thymus to protect human thymocytes from the effects of HIV-1. Compared with that observed in control mice with human thymic grafts, porcine thymus attenuated human thymocyte depletion by the CCR5-tropic isolate JR-CSF without preventing thymocyte infection. Porcine thymus protected human thymocytes from infection and depletion by a CXCR4-tropic HIV-1 isolate without reducing peripheral blood viral loads or T cell infection. Human thymocytes from human but not porcine grafts showed decreased Bcl-2 expression and increased apoptosis after NL4.3 infection. Thus, porcine thymus protects human thymocytes from the cytopathic effect of HIV-1, suggesting a possible approach to achieving immune restoration in patients with acquired immunodeficiency syndrome who have incomplete responses to antiretroviral therapy. The model allows analysis of the mechanisms of HIV-mediated thymic dysfunction.

Association of industry sponsorship to published outcomes in gastrointestinal clinical research.

BACKGROUND & AIMS: Recent years have seen an increase in industry sponsorship of clinical trials throughout medicine. We conducted a study to evaluate the association of industry sponsorship to published outcomes in gastrointestinal (GI) clinical research. Our aims were (1) to evaluate the trends in the source of funding for GI clinical research during the period from 1992 to 2002-2003, (2) to determine whether the source of study funding predicted the likelihood that a study would publish results that favor the drug or device being tested, and (3) to determine whether differences exist in the methodologic quality of the investigational study methods used in studies funded by private industry versus other sources. METHODS: We selected all clinical studies evaluating a drug or device from 4 prominent GI journals (Gastroenterology, The American Journal of Gastroenterology, Hepatology, and Gastrointestinal Endoscopy). All studies were abstracted by using a standardized data abstraction form. We evaluated the trends in funding source for studies published during the years 1992, 2002, and 2003. All selected studies were scored for methodologic quality by using a previously validated scoring system. The percentage of studies that reported outcomes that favored the device or drug being tested against the standard therapy or placebo was determined for each funding source. A funnel plot was constructed to assess for the presence of negative publication bias. RESULTS: A total of 6326 studies were reviewed. For the 1992 trend data, 1860 studies were reviewed, and 135 were selected for inclusion in the study. Ninety-five studies were studies involving the investigation of drugs, and 40 studies involved the testing of a device. For the 2002-2003 data 4466 studies were reviewed, and 315 were selected for inclusion in the study. Two hundred twenty-two studies were clinical trials involving the investigation of drugs, and 93 were clinical trials involving devices. In comparing 1992 to 2002, the percentage of studies funded by industry sources more than doubled from 10% to greater than 28% of the total studies assessed. There was an associated decline in the proportion of studies with funding from non-industry sources during this period (62% to less than 48%). The percentage of studies that did not disclose a funding source fell modestly from 28% to 24%. We found that 86% of studies funded by private industry reported a result favorable to the study drug or device, and 83% of studies funded by academic sources reported a result favorable to the study drug or device (P=.572). On average, studies funded by private industry had a higher methodology score than studies funded by traditional academic sources (75 of 100 vs 65 of 100; P=.005). Analysis of the funnel plot did not reveal evidence of bias against the publication of small studies with insignificant results. CONCLUSIONS: The proportion of research funded by industry has more than doubled during the last decade and currently comprises almost half of the funding for GI clinical research. Industry-sponsored studies are, on average, of superior methodologic quality to studies funded by other sources. Industry-sponsored studies in leading GI journals were no more likely than other studies to publish results that favored the study sponsor, although an extremely high percentage of all studies in these journals reported positive results. There has been only a modest decline in studies not acknowledging a funding source.

Detection and characterization of cellular immune responses using peptide-MHC microarrays.

The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease. We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells. CD4(+) or CD8(+) lymphocytes can be captured in accordance with their ligand specificity using an array of peptide-MHC complexes printed on a film-coated glass surface. We have characterized the specificity and sensitivity of a peptide-MHC array using labeled lymphocytes from T cell receptor transgenic mice. In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse. This approach should be useful for epitope discovery, as well as for characterization and analysis of multiple epitope-specific T cell populations during immune responses associated with viral and bacterial infection, cancer, autoimmunity, and vaccination.

Identification of 58 novel mutations in Niemann-Pick disease type C: correlation with biochemical phenotype and importance of PTC1-like domains in NPC1.

The two known complementation groups of Niemann-Pick Type C disease, NPC1 and NPC2, result from non-allelic protein defects. Both the NPC1 and NPC2 (HE1) gene products are intimately involved in cholesterol and glycolipid trafficking and/or transport. We describe mutation analysis on samples from 143 unrelated affected NPC patients using conformation sensitive gel electrophoresis and DNA sequencing as the primary mutation screening methods for NPC1 and NPC2, respectively. These methods are robust, sensitive, and do not require any specialized laboratory equipment. Analyses identified two NPC1 mutations for 115 (80.4%) patients, one NPC1 mutation for 10 (7.0%) patients, two NPC2 mutations for five (3.5%) patients, one NPC2 mutation for one (0.7%) patient, and no mutations for 12 (8.4%) patients. Thus, mutations were identified on 251 of 286 (88%) disease alleles, including 121 different mutations (114 in NPC1 and seven in NPC2), 58 of which are previously unreported. The most common NPC1 mutation, I1061T, was detected on 18% of NPC alleles. Other NPC1 mutations were mostly private, missense mutations located throughout the gene with clustering in the cysteine-rich luminal domain. Correlation with biochemical data suggests classification of several mutations as severe and others as moderate or variable. The region between amino acids 1038 and 1253, which shares 35% identity with Patched 1, appears to be a hot spot for mutations. Additionally, a high percentage of mutations were located at amino acids identical to the NPC1 homolog, NPC1L1. Biochemical complementation analysis of cases negative for mutations revealed a high percentage of equivocal results where the complementation group appeared to be non-NPC1 and non-NPC2. This raises the possibilities of an additional NPC complementation group(s) or non-specificity of the biochemical testing for NPC. These caveats must be considered when offering mutation testing as a clinical service.