A first-in-human study of [68Ga]Ga-CDI: a positron emitting radiopharmaceutical for imaging tumour cell death.

PURPOSE: This study assesses human biodistribution, radiation dosimetry, safety and tumour uptake of cell death indicator labelled with 68Ga ([68Ga]Ga-CDI), a novel radiopharmaceutical that can image multiple forms of cell death. METHODS: Five participants with at least one extracranial site of solid malignancy > 2 cm and no active cancer treatment in the 8 weeks prior to the study were enrolled. Participants were administered 205 ± 4.1 MBq (range, 200-211 MBq) of [68Ga]Ga-CDI and 8 serial PET scans acquired: the first commencing immediately and the last 3 h later. Participants were monitored for clinical, laboratory and electrocardiographic side effects and adverse events. Urine and blood radioactivity was measured. Spherical volumes of interest were drawn over tumour, blood pool and organs to determine biodistribution and calculate dosimetry. In one participant, tumour specimens were analysed for cell death using terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) staining. RESULTS: [68Ga]Ga-CDI is safe and well-tolerated with no side effects or adverse events. [68Ga]Ga-CDI is renally excreted, demonstrates low levels of physiologic uptake in the other organs and has excellent imaging characteristics. The mean effective dose was 2.17E - 02 ± 4.61E - 03 mSv/MBq. It images constitutive tumour cell death and correlates with tumour cell death on histology. CONCLUSION: [68Ga]Ga-CDI is a novel cell death imaging radiopharmaceutical that is safe, has low radiation dosimetry and excellent biodistribution and imaging characteristics. It has potential advantages over previously investigated radiopharmaceuticals for imaging of cell death and has progressed to a proof-of-concept trial. TRIAL REGISTRATION: ACTRN12621000641897 (28/5/2021, retrospectively registered).

Elevation of fatty acid desaturase 2 in esophageal adenocarcinoma increases polyunsaturated lipids and may exacerbate bile acid-induced DNA damage.

BACKGROUND: The risk of esophageal adenocarcinoma (EAC) is associated with gastro-esophageal reflux disease (GERD) and obesity. Lipid metabolism-targeted therapies decrease the risk of progressing from Barrett's esophagus (BE) to EAC, but the precise lipid metabolic changes and their roles in genotoxicity during EAC development are yet to be established. METHODS: Esophageal biopsies from the normal epithelium (NE), BE, and EAC, were analyzed using concurrent lipidomics and proteomics (n = 30) followed by orthogonal validation on independent samples using RNAseq transcriptomics (n = 22) and immunohistochemistry (IHC, n = 80). The EAC cell line FLO-1 was treated with FADS2 selective inhibitor SC26196, and/or bile acid cocktail, followed by immunofluorescence staining for γH2AX. RESULTS: Metabolism-focused Reactome analysis of the proteomics data revealed enrichment of fatty acid metabolism, ketone body metabolism, and biosynthesis of specialized pro-resolving mediators in EAC pathogenesis. Lipidomics revealed progressive alterations (NE-BE-EAC) in glycerophospholipid synthesis with decreasing triglycerides and increasing phosphatidylcholine and phosphatidylethanolamine, and sphingolipid synthesis with decreasing dihydroceramide and increasing ceramides. Furthermore, a progressive increase in lipids with C20 fatty acids and polyunsaturated lipids with ≥4 double bonds were also observed. Integration with transcriptome data identified candidate enzymes for IHC validation: Δ4-Desaturase, Sphingolipid 1 (DEGS1) which desaturates dihydroceramide to ceramide, and Δ5 and Δ6-Desaturases (fatty acid desaturases, FADS1 and FADS2), responsible for polyunsaturation. All three enzymes showed significant increases from BE through dysplasia to EAC, but transcript levels of DEGS1 were decreased suggesting post-translational regulation. Finally, the FADS2 selective inhibitor SC26196 significantly reduced polyunsaturated lipids with three and four double bonds and reduced bile acid-induced DNA double-strand breaks in FLO-1 cells in vitro. CONCLUSIONS: Integrated multiomics revealed sphingolipid and phospholipid metabolism rewiring during EAC development. FADS2 inhibition and reduction of the high polyunsaturated lipids effectively protected EAC cells from bile acid-induced DNA damage in vitro, potentially through reduced lipid peroxidation.

Influence of X-ray computed tomography (CT) exposure and reconstruction parameters on positron emission tomography (PET) quantitation.

BACKGROUND: The CT of PET CT provides diagnostic information, anatomic localisation and attenuation correction (AC). When only AC is required, very lose dose CT is desirable. CT iterative reconstruction (IR) improves image quality with lower exposures however there is little data on very low dose IR CT for AC of PET. This work assesses the impact of CT exposure and reconstruction algorithm on PET voxel values. METHOD: An anthropomorphic torso phantom was filled with physiologically typical [18]F concentrations in heart, liver and background compartments. A 17-mm-diameter right lung "tumour" filled with [18]F was included (surrounding lung contained no 18[F]). PET was acquired followed by 24 CT acquisitions with varying CT exposures (15-50 mAs, 80-120 kVp, pitch 0.671 or 0.828). Each CT was reconstructed twice using filtered back projection (FBP) or IR and these used for AC of PET. The reference PET reconstruction (RR) used CT acquired at 50 mAs, 120 kVp, pitch 0.828, IR, all others were test PET reconstructions (TR). Regions of interest (ROIs) were drawn in the liver, soft tissue and over "tumour" on each TR and compared with the RR. Voxel values in each TR were compared to the RR using a paired t test and by calculating which and what proportion of voxels in each TR differed by a quantitatively significant difference (QSD) from the RR. RESULTS: TRs reconstructed using lower dose CTs underestimated mean and maximum ROI activity relative to the RR; greater with IR than FBP. Once CT dose index (CTDI) increased to 1 mGy, differences were less than QSD. On voxel analysis, all TRs were significantly different to the RR (p < 0.0001). TRs reconstructed at the lowest CT exposure with IR had 6% of voxels that differed by greater than QSD. Differences were reduced with increasing CTDI and FBP reconstruction. Voxels which exceeded the QSD were spatially localised to regions of high activity, interfaces between different attenuation and areas of CT beam hardening. CONCLUSIONS: Very low dose CT exposures are feasible for accurate PET AC. Scanner- and reconstruction-specific validation should be employed prior very low dose CT AC for PET.

A prospective cohort study of immunogenicity of quadrivalent human papillomavirus vaccination among Alaska Native Children, Alaska, United States.

OBJECTIVE: In the United States, HPV vaccination is routinely recommended at age 11 or 12 years; the series can be started at age 9. We conducted a cohort study to assess long-term immunogenicity of quadrivalent HPV vaccine (4vHPV) in an American Indian/Alaska Native (AI/AN) Indigenous population. METHODS: During 2011-2014, we enrolled AI/AN girls and boys aged 9-14 years, who were vaccinated with a 3-dose series of 4vHPV. Serum specimens were collected at five time points: immediately prior to doses 2 and 3, and at one month, one year, and two years after series completion. Antibody testing was performed using a multiplex virus-like-particle-IgG ELISA for 4vHPV types (HPV 6/11/16/18). RESULTS: Among 477 children (405 girls/72 boys) completing the 3-dose series, median age at enrollment was 11.2 years. Of the 477, 72 (15%) were tested before dose 2 and 70 (15%) before dose 3. Following series completion, 435 (91%) were tested at one month, 382 (80%) at one year, and 351 (74%) at two years. All tested participants had detectable antibody to 4vHPV types at all time points measured. Geometric mean concentrations (GMCs) for 4vHPV types at one month and two years post-series completion were 269.9 and 32.7 AU/ml for HPV6, 349.3 and 42.9 AU/ml for HPV11, 1240.2 and 168.3 IU/ml HPV16, and 493.2 and 52.2 IU/ml for HPV18. Among children tested after each dose, GMCs after doses 1 and 2 were 3.9 and 32.2 AU/ml for HPV6, 5.3 and 45.6 AU/ml for HPV11, 20.8 and 187.9 IU/ml for HPV16; and 6.6 and 49.7 IU/ml for HPV18. No serious adverse events were reported. CONCLUSION: All AI/AN children developed antibodies to all 4vHPV types after vaccination. GMCs rose after each dose, then decreased to a plateau over the subsequent two years. This cohort will continue to be followed to determine duration of antibody response.

Chronic High-Fat Diet Induces Early Barrett's Esophagus in Mice through Lipidome Remodeling.

Esophageal adenocarcinoma (EAC) incidence has been rapidly increasing, potentially associated with the prevalence of the risk factors gastroesophageal reflux disease (GERD), obesity, high-fat diet (HFD), and the precursor condition Barrett's esophagus (BE). EAC development occurs over several years, with stepwise changes of the squamous esophageal epithelium, through cardiac metaplasia, to BE, and then EAC. To establish the roles of GERD and HFD in initiating BE, we developed a dietary intervention model in C57/BL6 mice using experimental HFD and GERD (0.2% deoxycholic acid, DCA, in drinking water), and then analyzed the gastroesophageal junction tissue lipidome and microbiome to reveal potential mechanisms. Chronic (9 months) HFD alone induced esophageal inflammation and metaplasia, the first steps in BE/EAC pathogenesis. While 0.2% deoxycholic acid (DCA) alone had no effect on esophageal morphology, it synergized with HFD to increase inflammation severity and metaplasia length, potentially via increased microbiome diversity. Furthermore, we identify a tissue lipid signature for inflammation and metaplasia, which is characterized by elevated very-long-chain ceramides and reduced lysophospholipids. In summary, we report a non-transgenic mouse model, and a tissue lipid signature for early BE. Validation of the lipid signature in human patient cohorts could pave the way for specific dietary strategies to reduce the risk of BE in high-risk individuals.

Antimicrobial resistance among Helicobacter pylori isolates in Alaska, 2000-2016.

OBJECTIVES: Alaska Natives experience a high burden of Helicobacter pylori infection and concomitant high rates of gastric cancer. Additionally, the prevalence of antimicrobial-resistant H. pylori has been shown to be high in Alaska. In this study, antimicrobial resistance over time among sentinel surveillance isolates was evaluated and risk factors for carrying antimicrobial-resistant H. pylori were assessed. METHODS: Through Alaska's H. pylori sentinel surveillance system, antral and fundal biopsies from Alaska Native patients undergoing esophagogastroduodenoscopy for clinical indications during 2000-2016 were collected and cultured. For positive cultures, minimum inhibitory concentrations (MICs) of metronidazole, amoxicillin, clarithromycin, tetracycline and levofloxacin were determined. RESULTS: A total of 800 H. pylori isolates obtained from 763 patients were tested. Resistance to metronidazole was most common (342/800; 42.8%), followed clarithromycin (238/800; 29.8%), both clarithromycin and metronidazole (128/800; 16.0%) and levofloxacin (113/800; 14.1%). Low proportions of isolates were resistant to amoxicillin and tetracycline. Levofloxacin resistance increased between 2000 and 2016 (P<0.001), but resistance to other antimicrobials did not change over time. Metronidazole and clarithromycin resistance were more common among women (P<0.001 for both), whilst levofloxacin resistance was more common among those with an urban residence (P=0.003). Metronidazole and levofloxacin resistance were more common among older patients (P<0.05). CONCLUSION: Between 2000 and 2016, a large percentage of H. pylori isolates received by the Alaska Sentinel Surveillance System demonstrated resistance to common antimicrobials. The surveillance system provides valuable information for clinicians to make informed treatment choices for patient with H. pylori.

In Vivo Measurement and Characterization of a Novel Formulation of [177Lu]-DOTA-Octreotate.

OBJECTIVES: Lutetium-177 can be made with high specific activity and with no other isotopes of lutetium present, referred to as "No Carrier Added" (NCA) 177Lu. We have radiolabelled DOTA-conjugated peptide DOTA-(Tyr3)-octreotate with NCA 177Lu ("NCA-LuTATE") and used it in nearly 40 therapeutic administrations for subjects with neuroendocrine tumours or meningiomas. In this paper, we report on our initial studies on aspects of the biodistribution and dosimetry of NCA-LuTATE from gamma camera 2D whole body (WB) and quantitative 3D SPECT (qSPECT) 177Lu imaging. METHODS: Thirteen patients received 39 NCA-LuTATE injections. Extensive WB planar and qSPECT imaging was acquired at approximately 0.5, 4, 24 and 96 h to permit estimates of clearance and radiation dose estimation using MIRD-based methodology (OLINDA-EXM). RESULTS: The average amount of NCA-Lutate administered per cycle was 7839±520 MBq. Bi-exponential modelling of whole body clearance showed half lives for the fast & slow components of t½=2.1±0.6 h and t½=58.1±6.6 h respectively. The average effective dose to kidneys was 3.1±1.0 Gy per cycle. In eight patients completing all treatment cycles the average total dose to kidneys was 11.7±3.6 Gy. CONCLUSIONS: We have shown that NCA-LuTATE has an acceptable radiation safety profile and is a suitable alternative to Carrier-Added 177Lu formulations. The fast component of the radiopharmaceutical clearance was closely correlated with baseline renal glomerular filtration rate, and this had an impact on radiation dose to the kidneys. In addition, it has less radioactive waste issues and requires less peptide per treatment.

Cost-effectiveness analysis of hepatocellular carcinoma screening by combinations of ultrasound and alpha-fetoprotein among Alaska Native people, 1983-2012.

BACKGROUND: The American Association for the Study of Liver Diseases (AASLD) recommends semi-annual hepatocellular carcinoma (HCC) screening using ultrasound (US) in persons with chronic hepatitis B (CHB) virus infection at high risk for HCC such as Asian males aged ≥40 years and Asian females aged ≥50 years. OBJECTIVE: To analyse the cost-effectiveness of 2 HCC screening methods in the Alaska Native (AN) health system: US-alone, or screening by alpha-fetoprotein (AFP) initially and switching to US for subsequent screenings if AFP >10 ng/mL (AFP→US). DESIGN: A spreadsheet-based model was developed for accounting the costs of 2 hypothetical HCC screening methods. We used epidemiologic data from a cohort of 839 AN persons with CHB who were offered HCC screening by AFP/US semi-annually during 1983-2012. We assumed that compared with AFP→US, US-alone identifies 33% more tumours at an early stage (defined as a single tumour ≤5 cm or ≤3 tumours ≤3 cm in diameter). Years of life gained (YLG) attributed to screening was estimated by comparing additional years of survival among persons with early- compared with late-stage tumours. Screening costs were calculated using Medicare reimbursement rates in 2012. Future screening costs and YLG were projected over a 30-year time horizon using a 3% discount rate. RESULTS: The total cost of screening for the cohort by AFP→US would have been approximately $357,000 ($36,000/early-stage tumour detected) compared to $814,000 ($59,000/early-stage tumour detected) by US-alone. The AFP→US method would have yielded an additional 27.8 YLG ($13,000/YLG) compared with 38.9 YLG ($21,000/YLG) for US-alone. Screening by US-alone would incur an additional $114,000 per extra early-tumour detected compared with AFP→US and $41,000 per extra YLG. CONCLUSIONS: Although US-alone HCC screening might have yielded more YLG than AFP→US, the reduced costs of the AFP→US method could expand access to HCC screening in resource constrained settings.

Helicobacter pylori infection and markers of gastric cancer risk in Alaska Native persons: a retrospective case-control study.

BACKGROUND: Alaska Native persons experience gastric cancer incidence and mortality rates that are three to four times higher than in the general United States population. OBJECTIVE: To evaluate pepsinogen I, pepsinogen I/II ratio, anti-Helicobacter pylori and cytotoxin-associated gene A (CagA) antibody levels, and blood group for their associations with gastric cancer development in Alaska Native people. METHODS: The present analysis was a retrospective case-control study that matched gastric cancers reported to the Alaska Native Tumor Registry from 1969 to 2008 to three controls on known demographic risk factors for H pylori infection, using sera from the Alaska Area Specimen Bank. Conditional logistic regression evaluated associations between serum markers and gastric cancer. RESULTS: A total of 122 gastric cancer cases were included, with sera predating cancer diagnosis (mean = 13 years) and 346 matched controls. One hundred twelve cases (91.8%) and 285 controls (82.4%) had evidence of previous or ongoing H pylori infection as measured by anti-H pylori antibody levels. Gastric cancer cases had a 2.63-fold increased odds of having positive anti-H pylori antibodies compared with their matched controls (P=0.01). In a multivariate model, noncardia gastric cancer (n=94) was associated with anti-H pylori antibodies (adjusted OR 3.92; P=0.004) and low pepsinogen I level (adjusted OR 6.04; P=0.04). No association between gastric cancer and blood group, anti-CagA antibodies or pepsinogen I/II ratio was found. CONCLUSION: Alaska Native people with gastric cancer had increased odds of previous H pylori infection. Low pepsinogen I level may function as a precancer marker for noncardia cancer.

HPV genotypes detected in cervical cancers from Alaska Native women, 1980-2007.

BACKGROUND: Human papillomavirus (HPV) vaccine prevents cervical pre-cancers and cancers caused by HPV types 16 and 18. This study provides information on the HPV types detected in cervical cancers of Alaska Native (AN) women. METHODS: Cases of invasive cervical cancer diagnosed in AN women aged 18 and above between 1980 and 2007 were identified from the Alaska Native Tumor Registry. A representative formalin-fixed, paraffin-embedded archived pathology block was retrieved and serially sectioned to allow histologic confirmation of lesion (first and last sections) and PCR testing of intervening sections. Extracted DNA was tested for HPV using Linear Array HPV Genotyping Test (Roche Diagnostics) with additional INNO-LiPA HPV Genotyping Assay (Innogenetics) testing on negative or inadequate specimens. All specimens were tested for a minimum 37 HPV types. RESULTS: Of 62 cervical cancer specimens evaluated, 57 (91.9%) contained one or more HPV types. Thirty-eight (61.2%) cancers contained HPV types 16 or 18, and 18 (29%) contained an oncogenic type other than type 16 or 18. CONCLUSIONS: Overall, almost two-thirds (61.2%) of the archived cervical cancers had detectible HPV types 16 or 18, a finding similar to studies of US women. As expected, a proportion of cancers would not be prevented by the current vaccines. HPV vaccination and cervical cancer screening are important prevention strategies for AN women.

The effect of Helicobacter pylori infection on iron stores and iron deficiency in urban Alaska Native adults.

BACKGROUND: Helicobacter pylori (H. pylori) infection has been correlated with low serum ferritin and iron deficiency. As a secondary analysis of a study of H. pylori reinfection, we investigated the association of H. pylori infection and the effect of its eradication on serum ferritin and iron deficiency. METHODS: Alaska Native adults undergoing esophagogastroduodenoscopy had sera collected and a (13) C urea breath test (UBT) was performed. Those H. pylori positive were treated with an antibiotic regimen; those who tested negative 2 months after treatment were evaluated at 4, 6, 12, and 24 months by UBT and serum ferritin with an immunoradiometric assay. We excluded persons from further analysis if they were prescribed iron by their provider. RESULTS: We measured serum ferritin for 241 persons; 121/241 were H. pylori positive. The geometric mean ferritin (GMF) for persons with and without H. pylori infection was 37 µg/L and 50 µg/L, respectively (p = .04). At enrollment, 19/121 H. pylori-positive persons had iron deficiency compared with 8/120 H. pylori negative (p = .02). Among 66 persons tested at 24 months, the GMF was higher at 24 months (49.6 µg/L) versus enrollment (36.5 µg/L; p = .02). Six of 11 persons with iron deficiency at enrollment no longer had iron deficiency and had a higher GMF (p = .02) 24 months after treatment. CONCLUSIONS: H. pylori infection was correlated with lower serum ferritin and iron deficiency. After H. pylori eradication, serum ferritin increased and approximately half of persons resolved their iron deficiency. Testing for H. pylori infection and subsequent treatment of those positive could be considered in persons with unexplained iron deficiency.

Comparison of nasopharyngeal flocked swabs and nasopharyngeal wash collection methods for respiratory virus detection in hospitalized children using real-time polymerase chain reaction.

This paper describes the molecular detection of respiratory viruses from nasopharyngeal flocked swabs (flocked swabs) and nasopharyngeal washes (washes) in a clinical setting. Washes and flocked swabs collected from children<3 years old hospitalized with a lower respiratory tract infection were tested for parainfluenza virus 1-3, respiratory syncytial virus, influenza A and B and metapneumovirus (Group 1) and adenovirus, rhinovirus and coronavirus (Group 2) using real-time reverse transcriptase PCR (rRT-PCR). A consensuses standard was used to determine sensitivity and compare cycle thresholds (C(T)) of washes and flocked swabs for each virus and group of viruses. Sensitivities ranged from 79 to 89% and 69 to 94% for flocked swabs and washes, respectively, excluding AdV which had a sensitivity of 35% for flocked swabs. When the flocked swabs and washes of Group 1 viruses were collected on the day of admission, the sensitivity of both sample types was 100%. Wash specimens had a lower C(T) value and higher sensitivity than flocked swabs; however there was no statistical difference in the sensitivity of a flocked swab (89%) versus wash (93%) for the detection of Group 1 viruses, particularly when samples were collected on the same day. Flocked swabs may be a useful alternative to washes for detection of respiratory viruses in clinical settings.

Diagnostic accuracy of tests for Helicobacter pylori in an Alaska Native population.

AIM: To evaluate the accuracy of two non-invasive tests in a population of Alaska Native persons. High rates of Helicobacter pylori (H. pylori) infection, H. pylori treatment failure, and gastric cancer in this population necessitate documentation of infection status at multiple time points over a patient's life. METHODS: In 280 patients undergoing endoscopy, H. pylori was diagnosed by culture, histology, rapid urease test, (13)C urea breath test (UBT), and immunoglobulin G antibodies to H. pylori in serum. The performances of (13)C-UBT and antibody test were compared to a gold standard defined by a positive H. pylori test by culture or, in case of a negative culture result, by positive histology and a positive rapid urease test. RESULTS: The sensitivity and specificity of the (13)C-UBT were 93% and 88%, respectively, relative to the gold standard. The antibody test had an equivalent sensitivity of 93% with a reduced specificity of 68%. The false positive results for the antibody test were associated with previous treatment for an H. pylori infection [relative risk (RR) = 2.8]. High levels of antibodies to H. pylori were associated with chronic gastritis and male gender, while high scores in the (13)C-UBT test were associated with older age and with the H. pylori bacteria load on histological examination (RR = 4.4). CONCLUSION: The (13)C-UBT outperformed the antibody test for H. pylori and could be used when a non-invasive test is clinically necessary to document treatment outcome or when monitoring for reinfection.

Alaska sentinel surveillance study of Helicobacter pylori isolates from Alaska Native persons from 2000 to 2008.

Helicobacter pylori infection is more common in Alaska Native persons than in the general U.S. population, with seroprevalence to H. pylori approaching 75%. Previous studies in Alaska have demonstrated elevated proportions of antimicrobial resistance among H. pylori isolates. We analyzed H. pylori data from the Centers for Disease Control and Prevention's sentinel surveillance in Alaska from January 2000 to December 2008 to determine the proportion of culture-positive biopsy specimens with antimicrobial resistance from Alaska Native persons undergoing endoscopy. The aim of the present study was to monitor antimicrobial resistance of H. pylori isolates over time and by region in Alaska Native persons. Susceptibility testing of H. pylori isolates to metronidazole, clarithromycin, amoxicillin, and tetracycline was performed using agar dilution. Susceptibility testing for levofloxacin was performed by Etest. Overall, 45% (532/1,181) of persons undergoing upper endoscopy were culture positive for H. pylori. Metronidazole resistance was demonstrated in isolates from 222/531 (42%) persons, clarithromycin resistance in 159/531 (30%) persons, amoxicillin resistance in 10/531 (2%) persons, and levofloxacin resistance in 30/155 (19%) persons; no tetracycline resistance was documented. The prevalence of metronidazole, clarithromycin, and levofloxacin resistance varied by region. Female patients were more likely than male patients to demonstrate metronidazole (P < 0.05) and clarithromycin (P < 0.05) resistance. No substantial change in the proportion of persons with resistant isolates was observed over time. Resistance to metronidazole, clarithromycin, and levofloxacin is more common among H. pylori isolates from Alaska Native persons than those from elsewhere in the United States.

Characterization of the human gastric fluid proteome reveals distinct pH-dependent protein profiles: implications for biomarker studies.

Gastric fluid is a source of gastric cancer biomarkers. However, very little is known about the normal gastric fluid proteome and its biological variations. In this study, we performed a comprehensive analysis of the human gastric fluid proteome using samples obtained from individuals with benign gastric conditions. Gastric fluid proteins were prefractionated using ultracentrifuge filters (3 kDa cutoff) and analyzed by two-dimensional gel electrophoresis (2-DE) and multidimensional LC-MS/MS. Our 2-DE analysis of 170 gastric fluid samples revealed distinct protein profiles for acidic and neutral samples, highlighting pH effects on protein composition. By 2D LC-MS/MS analysis of pooled samples, we identified 284 and 347 proteins in acidic and neutral samples respectively (FDR ≤1%), of which 265 proteins (72.4%) overlapped. However, unlike neutral samples, most proteins in acidic samples were identified from peptides in the filtrate (i.e., <3 kDa). Consistent with this finding, immunoblot analysis of six potential gastric cancer biomarkers rarely detected full-length proteins in acidic samples. These findings have important implications for biomarker studies because a majority of gastric cancer patients have neutral gastric fluid compared to noncancer controls. Consequently, sample stratification, choice of proteomic approaches, and validation strategy can profoundly affect the interpretation of biomarker findings. These observations should help to refine gastric fluid biomarker studies.

Characterization of Helicobacter pylori cagA and vacA genotypes among Alaskans and their correlation with clinical disease.

Helicobacter pylori infection is common in Alaska. The development of severe H. pylori disease is partially determined by the virulence of the infecting strain. Here we present vacA and cagA genotype data for H. pylori strains isolated from Alaskans and their correlation with clinical disease. We enrolled patients scheduled for esophagogastroduodenoscopy and positive for H. pylori infection. Gastric biopsy specimens from the stomach antrum and fundus were cultured. We performed PCR analysis of the H. pylori vacA gene and for the presence of the cagA gene and cagA empty site. We genotyped 515 H. pylori samples from 220 Native and 66 non-Native Alaskans. We detected the cagA gene in 242/286 (85%) persons; of 222 strains that could be subtyped, 95% (212) were non-Asian cagA and 3% (6) were East Asian cagA. After removing mixed infections (n = 17), 83% of H. pylori strains had either the vacA s1m1 (120/269) or s2m2 (103/269) genotype. Sixty-six percent (68/103) of H. pylori strains with the vacA s2m2 genotype also contained the cagA gene. Infection with an H. pylori strain having the cagA gene or vacA s1m1 genotype (compared with s1m2 and s2m2) was associated with a decreased risk of esophagitis (P = 0.003 and 0.0003, respectively). Infection with an H. pylori strain having the vacA s1m1 genotype (compared with s1m2 and s2m2) was associated with an increased risk of peptic ulcer disease (PUD) (P = 0.003). The majority of H. pylori strains in this study carried the non-Asian cagA gene and either the vacA s1m1 or s2m2 genotype. A majority of H. pylori strains with the vacA s2m2 genotype also contained the cagA gene. There was an association of H. pylori genotype with esophagitis and PUD.

Alaska Native adolescent views on cervical cancer, the human papillomavirus (HPV), genital warts and the quadrivalent HPV vaccine.

OBJECTIVES: To understand the knowledge levels, attitudes and perceptions of Alaska Native adolescent girls about cervical cancer, HPV, genital warts and the HPV vaccine. STUDY DESIGN: A qualitative study. METHODS: Seventy-nine in-depth interviews were conducted with adolescent females aged 11 through 18 years in 4 communities in Alaska. The convenience sample was recruited through word of mouth, posters and flyers distributed in community schools, medical clinics and stores. RESULTS: Many of those surveyed didn't know the purpose of a vaccine and were not familiar with basic knowledge about HPV, genital warts and cervical cancer. After learning about cervical cancer and HPV, most teens felt that someone their age had an average likelihood of contracting the diseases and that having the disease would be quite bad. Most teens said they were interested in vaccination. When asked if they would get a vaccine, older teens most commonly cited concerns about side effects or doubts about vaccine efficacy, while younger teens were afraid the shot would hurt. Most teens stated that they preferred to learn about health topics such as these through television programming, followed by the Internet, brochures and posters. CONCLUSIONS: The findings provide valuable information on how to inform adolescents about the vaccine and alleviate their concerns. The design of an educational campaign should vary depending on the age of the adolescents. For younger teens, distribution of information should be at school using a brochure or curriculum, while for older teens a web page may be more appropriate.

Factors associated with the progression of fibrosis on liver biopsy in Alaska Native and American Indian persons with chronic hepatitis C.

BACKGROUND: Various factors influence the development and rate of fibrosis progression in chronic hepatitis C virus (HCV) infection. OBJECTIVES: To examine factors associated with fibrosis in a longterm outcomes study of Alaska Native/American Indian persons who underwent liver biopsy, and to examine the rate of fibrosis progression in persons with subsequent biopsies. METHODS: A cross-sectional analysis of the demographic, inflammatory and viral characteristics of persons undergoing liver biopsy compared individuals with early (Ishak fibrosis score of lower than 3) with those with advanced (Ishak score of 3 or greater) fibrosis. Persons who underwent two or more biopsies were analyzed for factors associated with fibrosis progression. RESULTS: Of 253 HCV RNA-positive persons who underwent at least one liver biopsy, 76 (30%) had advanced fibrosis. On multivariate analysis, a Knodell histological activity index score of 10 to 14 and an alpha-fetoprotein level of 8 ng/mL or higher were found to be independent predictors of advanced liver fibrosis (P<0.0001 for each). When surrogate markers of liver inflammation (alanine aminotransferase, aspartate aminotransferase/alanine aminotransferase ratio and alpha-fetoprotein) were removed from the model, type 2 diabetes mellitus (P=0.001), steatosis (P=0.03) and duration of HCV infection by 10-year intervals (P=0.02) were associated with advanced fibrosis. Among 52 persons who underwent two or more biopsies a mean of 6.2 years apart, the mean Ishak fibrosis score increased between biopsies (P=0.002), with progression associated with older age at initial biopsy and HCV risk factors. CONCLUSIONS: The presence of type 2 diabetes mellitus, steatosis and duration of HCV infection were independent predictors of advanced fibrosis in the present cohort, with significant fibrosis progression demonstrated in persons who underwent serial biopsies.

Viral respiratory infections in hospitalized and community control children in Alaska.

Respiratory syncytial virus (RSV) in Alaska Native children from the Yukon Kuskokwim (YK) Delta is associated with a hospitalization rate five times higher than that reported for the general US child population. The role of other viral respiratory pathogens has not been studied in this population. YK Delta children <3 years of age hospitalized with respiratory infections and same aged community control children were prospectively enrolled between October 2005 and September 2007. Polymerase chain reaction detection of viruses was performed on nasopharyngeal samples. Characteristics of hospitalized and asymptomatic control children were analyzed. From October 2005 to September 2007, 440 hospitalized and 425 control children were analyzed. Respiratory viruses were detected in 90% (395) of hospitalized children: 194 (44%) rhinovirus, 131 (30%) adenovirus, 102 (23%) RSV, 77 (18%) para influenza viruses (PIV), 66 (15%) human metapneumovirus (hMPV), 23 (5%) influenza, and 25 (6%) coronavirus. Fifty-two percent (221) of control children had a virus detected, most commonly rhinovirus (33%), and adenovirus (16%). RSV, PIV, hMPV, and influenza were significantly more common in hospitalized cases than control children, but rhinovirus, adenovirus, and coronavirus were not. RSV and hMPV were associated with higher severity of illness. In this study, RSV remains the most important virus associated with respiratory hospitalization, although hMPV and PIV were also common. RSV and hMPV were associated with more severe illness. Rhinovirus and adenovirus were detected in two-thirds of hospitalized children, but their frequent detection in control children made their role in respiratory hospitalization uncertain.

Adverse outcomes in Alaska natives who recovered from or have chronic hepatitis C infection.

BACKGROUND & AIMS: The factors associated with adverse outcome from hepatitis C virus (HCV) infection are incompletely understood. To determine the incidence and risk factors associated with the development of end-stage liver disease (ESLD) and liver-related death (LRD), we conducted a retrospective/prospective population-based study in a cohort of Alaska Native persons chronically infected with HCV from 1994 to 2005. METHODS: We followed 960 persons prospectively for an average of 7.2 years and retrospectively for 12.1 years with data from medical records and serum samples. We compared data from subjects that were chronically infected with those who recovered from HCV infection, stratified by alcohol use. Survival models were used to examine factors associated with ESLD and LRD in chronically infected patients. RESULTS: During prospective follow-up, 80 (8.8%) and 47 (5.2%) patients developed ESLD and LRD, respectively. In examining incidence per 100 person-years, no difference was found among heavy alcohol users in the incidence of LRD (2.28 versus 3.50; P = .34) or ESLD (3.21 versus 5.69; P = .13) in persons with chronic HCV compared with those recovered from HCV infection. In subjects that consumed <50 g alcohol/d, the incidences of LRD were 0.77 and 0.09 (P = .01) and of ESLD were 1.58 versus 0.36 (P = .002), respectively, in subjects with chronic infection versus those that recovered. Multivariate analysis showed that older age, heavy alcohol use, and HCV genotype 3 were associated with ESLD. CONCLUSIONS: A history of heavy alcohol use is associated with the highest incidence of LRD and ESLD, regardless of whether patients are chronically infected or recover from HCV infection.