Jenkins, G. Douglas

ABSTRACTBackgroundInsomnia and substance use disorders (SUD) are common comorbidities of bipolar disorder (BD). Genome‐wide association studies (GWAS) have uncovered shared genetic contributions to insomnia and BD as well as SUDs and BD. Electronic health record (EHR) derived phenotypes (phecodes) and questionnaire data were used to examine the relationship between insomnia genetic liability and SUDs in BD.Methods40,839 participants from the Mayo Clinic Bipolar Disorder Biobank (BD Biobank; n = 774) and Mayo Clinic Biobank (n = 485 BD cases, n = 39,580 controls) were included in the analyses of diagnosis (phecode) outcomes (insomnia, SUD, alcohol use disorder [AUD] and tobacco use disorder [TUD]). Analyses of specific SUD outcomes obtained through the BD Biobank questionnaire included 1789 cases and considered BD subtype. Logistic regression was used to test for associations between insomnia polygenic risk scores (PRS) and insomnia and SUD outcomes in BD cases and controls.ResultsInsomnia PRS was associated with having an insomnia diagnosis (phecode) in the EHR in controls (OR = 1.19, p = 9.64e‐33) but not in BD cases (OR = 1, p = 0.95). Associations between insomnia PRS and SUD diagnoses were significant in BD cases and controls, with the association being stronger in BD cases (interaction p = 0.024). In the BD Biobank data, the insomnia PRS was associated with increased odds of AUD (OR = 1.19, p = 4.26e‐04), TUD (OR = 1.21, p = 1.25e‐05) and cannabis use disorder (OR = 1.16, p = 4.19e‐03).ConclusionThe effect of genetic predisposition to insomnia on SUD risk may be stronger in BD cases than in controls, which could have clinical care implications for individuals with BD and comorbid SUD.

Abstract Background: Monoclonal B-cell lymphocytosis (MBL) is characterized by the presence of a circulating population of clonal B-cells where the total B-cell count is <5 × 109/L and criteria for the diagnosis of other lymphoproliferative disorders are not present. In most patients the immunophenotype is similar to that of chronic lymphocytic leukemia with co-expression of CD19 and CD5. The natural history of MBL is unclear. We used the Mayo Clinic hematopathology database to identify patients with MBL and abstracted clinical information on natural history and prognostic parameters from clinical and research records. Methods: After IRB review and approval, we reviewed the hematopathology records of 2292 patients seen at Mayo Clinic between August 1998 and May 2006 who had a clonal B-cell population on flow cytometry phenotypically consistent with CLL and fewer than <5 × 109/L B-cells. Clinical records were then reviewed to identify patients who fulfilled the criteria for MBL (no evidence of other lymphoproliferative disorders, no adenopathy on exam). Patient characteristics, prognostic parameters, and clinical outcomes were abstracted from the clinical and research records of these patients. Cox regression analysis was used to evaluate association between prognostic markers and time to treatment. Results: A total of 118 patients meeting the criteria for MBL were identified. Median age at the time of MBL diagnosis was 69.4 years (range 39.0 – 91.4). A slight male preponderance was observed (54% male). CD38 status was available for 106 (90%) patients of whom 20% were CD38 positive using the 30% threshold to classify CD38. Cytogenetic analysis by fluorescent in situ hybridization (FISH) was available in 58 (49%) patients. Using the hierarchical classification system of Dohner and colleagues, 26 (44.8%) were 13q-, 10 (17.2%) were trisomy 12, 2 (3.4%) were 11q-, 1 (1.7%) was 17p-, and 19 were (32.8%) no defect was detected. ZAP-70 status was available for 32 patients of whom 22% were ZAP-70+ positive. IgVH gene mutation status was only available for 15 (12.7%) patients of whom 40% were unmutated. Time from MBL diagnosis to treatment is shown in Figure 1. Survival free of treatment was 92.7% (95%CI: 86.8–99.1%) at 1 year, 90.9% (95%CI: 84.2–98.2%) at 2 years, and 87.5% (95% CI: 78.7–97.4%) at 5 years. While age (p=0.68) and gender (p=0.14) were not found to be associated with time to treatment (TTT), CD38 status showed a strong relationship with TTT (p<0.001, HR: 22.7, 95% CI: 3.4–239.6). Conclusion: The distribution of prognostic parameters (e.g. CD38, FISH) in patients with MBL appears similar to that of patients with early stage CLL. We observed a low rate of progression, with 5 year survival free of treatment greater than 87%. TTT among newly diagnosed MBL patients was associated with the CD38 status of clonal B-cells. Figure Figure

Abstract The FKBP5 gene encodes the FKBP51 immunophilin, a negative regulator of AKT. Variability in level of FKBP5 expression is a major factor contributing to variation in response to several classes of chemotherapeutic agents including gemcitabine, first line treatment for pancreatic cancer. Genetic variation of FKBP5 could influence its expression, function, and, ultimately, treatment response of pancreatic cancer. We set out to comprehensively study the role of genetic variation in FKBP5 identified by Next Generation DNA resequencing in response to gemcitabine treatment of pancreatic cancer by utilizing both tumor and germline DNA samples from 43 pancreatic cancer patients, including 19 paired tumor-normal samples. Genotype-phenotype association studies with overall survival (in tumor only) as well as with FKBP5 gene expression were performed using the same samples in which Next Generation DNA resequencing had been performed, followed by functional genomic studies of top selected single nucleotide polymorphisms (SNPs). In-depth resequencing identified 404 FKBP5 SNPs in both normal and tumor DNA patient samples. SNPs with the smallest p-values for association with FKBP5 survival or expression were subjected to functional genomic studies. Electromobility shift assay (EMSA) showed that the rs73748206 “T” allele altered DNA-protein binding patterns, consistent with a significantly increased reporter gene activity. The effect of rs73748206 was also confirmed by mRNA expression studies using lymphoblastoid cell lines (LCL) derived from the same patients from whom DNA used for resequencing had been obtained. This comprehensive FKBP5 resequencing study provides insight into the role of inheritance in variation in response to gemcitabine therapy of pancreatic cancer. Citation Format: Katarzyna A. Ellsworth, Bruce W. Eckloff, Liang Li, Irene Moon, Brooke L. Fridley, Greg D. Jenkins, Erin Carlson, Abra Brisbin, Ryan Abo, William Bamlet, Gloria Petersen, Eric D. Wieben, Liewei Wang. Contribution of FKBP5 genetic variation to gemcitabine treatment and survival in pancreatic cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2209. doi:10.1158/1538-7445.AM2013-2209

Abstract The advancement in genotyping technology and large SNP arrays has led to an increase in the number of genome-wide association studies (GWAS) for various complex traits. These GWAS have resulted in the implication of over 1500 SNPs for association with disease traits. However, the SNPs identified from these GWAS are not necessarily the functional variants. Therefore, the next phase in GWAS will involve the validation, augmentation and refining of these putative regions or loci. One possible “next phase” approach would be to catalog all variants found in the study subjects within the loci of interest using sequencing technology, followed by the association analysis. However, sequencing a locus in a large number of subjects is still relatively expensive. By sequencing only a portion of the samples for the follow-up of signals from GWAS, followed by imputation in the remaining samples, one can significantly reduced the cost to localize the punitive variant involved in the etiology of complex phenotypes. A potentially attractive alternative option would be to impute based only on the 1000 Genomes Project; however, this has the drawbacks of using a reference population that does not necessarily match with disease status and LD pattern. Therefore, we conducted a study to investigate the ability to carry-out genotype imputation using a reference panel consisting of sequencing data for a fraction of the study participants. The various approaches were implemented using data from the candidate gene sequencing study conducted at the Mayo Clinic and the 1000 Genomes Project. Our results show that imputation of untyped markers based on sequencing a portion of the study participants is a reasonable, cost-saving, approach for disease mapping and the refinement of putative regions detected with GWAS. We recommend a few guidelines in designing sequencing studies to take advantage of the recent advances in genotype imputation: (1) Select the largest and most diverse reference panel for sequencing, with respect to both haplotypes and phenotypes. One can also use the 1000 Genomes Project in addition to the sequencing of a portion of the study participants; (2) Genotype as many “anchor” markers as possible, in that, the number of markers genotyped on all subjects impacts accuracy. Therefore, additional genotyping of a few common SNP markers not already genotyped on all subjects using a cost effective platform may be needed if the GWAS SNP array does not provide adequate coverage in the locus to be sequenced. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4738.

Purpose The diagnosis of monoclonal B-cell lymphocytosis (MBL) is used to characterize patients with a circulating population of clonal B cells, a total B-cell count of less than 5 × 109/L, and no other features of a B-cell lymphoproliferative disorder including lymphadenopathy/organomegaly. The natural history of clinically identified MBL is unclear. The goal of this study was to explore the outcome of patients with MBL relative to that of individuals with Rai stage 0 chronic lymphocytic leukemia (CLL). Patients and Methods We used hematopathology records to identify a cohort of 631 patients with newly diagnosed MBL or Rai stage 0 CLL. Within this cohort, 302 patients had MBL (B-cell counts of 0.02 to 4.99 × 109/L); 94 patients had Rai stage 0 CLL with an absolute lymphocyte count (ALC) ≤ 10 × 109/L; and 219 patients had Rai stage 0 CLL with an ALC more than 10 × 109/L. Data on clinical outcome were abstracted from medical records. Results The percentage of MBL patients free of treatment at 1, 2, and 5 years was 99%, 98%, and 93%, respectively. B-cell count as a continuous variable (hazard ratio [HR] = 2.9, P = .04) and CD38 status (HR = 10.8, P = .006) predicted time to treatment (TTT) among MBL patients. The likelihood of treatment for MBL patients was lower (HR = 0.32, P = .04) than that of both Rai stage 0 CLL patients with an ALC less than 10 × 109/L (n = 94) and Rai stage 0 CLL patients with an ALC more than 10 × 109/L (n = 219; P = .0003). Conclusion Individuals with MBL identified in clinical practice have a low risk for progression at 5 years. Because B-cell count seems to relate to TTT as a continuous variable, additional studies are needed to determine what B-cell count should be used to distinguish between MBL and CLL.

Abstract The diagnosis of chronic lymphocytic leukemia (CLL) in asymptomatic patients has historically been based on documenting a characteristic lymphocyte clone and the presence of lymphocytosis. There are minimal data regarding which lymphocyte parameter (absolute lymphocyte count [ALC] or B-cell count) and what threshold should be used for diagnosis. We analyzed the relationship of ALC and B-cell count with clinical outcome in 459 patients with a clonal population of CLL phenotype to determine (1) whether the CLL diagnosis should be based on ALC or B-cell count, (2) what lymphocyte threshold should be used for diagnosis, and (3) whether any lymphocyte count has independent prognostic value after accounting for biologic/molecular prognostic markers. B-cell count and ALC had similar value for predicting treatment-free survival (TFS) and overall survival as continuous variables, but as binary factors, a B-cell threshold of 11 × 109/L best predicted survival. B-cell count remained an independent predictor of TFS after controlling for ZAP-70, IGHV, CD38, or fluorescence in situ hybridization (FISH) results (all P < .001). These analyses support basing the diagnosis of CLL on B-cell count and retaining the size of the B-cell count in the diagnostic criteria. Using clinically relevant criteria to distinguish between monoclonal B-cell lymphocytosis (MBL) and CLL could minimize patient distress caused by labeling asymptomatic people at low risk for adverse clinical consequences as having CLL.

Previous results following subfascial endoscopic perforator vein surgery were reported to be worse in post-thrombotic syndrome than in limbs with primary valvular incompetence. This report comprises a larger patient cohort with longer follow-up. The goal of this study was to determine if subfascial endoscopic perforator vein surgery is justified in patients with postthrombotic venous insufficiency. The clinical data of 91 consecutive patients who underwent subfascial endoscopic perforator vein surgery with or without superficial reflux ablation over a 7-year period from May 1993 to June 2000 were retrospectively analyzed. Fifty-four females and 37 males (median age, 53 years; range, 20-77) underwent 103 subfascial endoscopic perforator vein surgery procedures. Forty-two limbs were classified as C6 (active ulcer), 34 as C5 (healed ulcer), and 24 as C4 (lipodermatosclerosis). Thirty procedures were performed in post-thrombotic limbs. Concomitant superficial reflux ablation was performed in 74 limbs (72%); saphenous vein stripping had been previously performed in 29 (28%). Deep venous incompetence was present in 89% of limbs; 13% had venous outflow obstruction on plethysmography. Cumulative ulcer healing in post-thrombotic limbs was not significantly different from limbs with primary valvular incompetence; 30-, 60-, and 90-day healing rates were 44%, 72%, and 72% vs 39%, 70%, and 87%, respectively (p = 0.35). On univariate analysis, the presence of ulcer greater than 2 cm in diameter was associated with delayed ulcer healing (p = 0.02). Cumulative ulcer recurrence in all limbs was 4%, 20%, and 27% at 1, 3, and 5 years, respectively. Ulcer recurrence in post-thrombotic limbs was higher than in limbs with primary valvular incompetence at 1, 3, and 5 years; 16%, 47%, and 56% vs 0%, 8%, and 15%, respectively (p = 0.001). Recurrent ulcers were small, superficial, and easier to heal. Clinical improvement was significant even in post-thrombotic limbs; median clinical score decreased from 9.5 to 3 (p=0.001), and median outcome score was +2 (mean 1.9; range, -I to 3). Median clinical score in patients with primary valvular incompetence improved from 6 to 1.5 (p = 0.0001). Subfascial endoscopic perforator vein surgery with superficial reflux ablation promoted ulcer healing, improved clinical outcome, and resulted in a low long-term ulcer recurrence rate in limbs with primary valvular incompetence. Despite good clinical outcome in post-thrombotic limbs, ulcer recurrence was high. These results imply that the role of subfascial endoscopic perforator vein surgery with superficial reflux ablation in patients with post-thrombotic limbs continues to be controversial.

ABSTRACT We evaluated two automated systems, MagNA Pure (Roche Molecular Biochemicals, Indianapolis, Ind.) and BioRobot 9604 (Qiagen, Inc., Chatsworth, Calif.) as effective replacements for the manual IsoQuick method (Orca Research, Inc., Bothell, Wash.) for extraction of herpes simplex virus (HSV) DNA from dermal and genital tract specimens prior to analysis by LightCycler PCR. Of 198 specimens (152 genital, 46 dermal), 92 (46.2%) were positive for HSV DNA by LightCycler PCR after automated extraction of specimens with either the MagNA Pure or BioRobot 9604 instrument. The manual IsoQuick method yielded HSV DNA (total n = 95) from three additional specimens that were negative by the automated method ( P = 0.25, sign test). Although the mean numbers of LightCycler PCR cycles required to reach positivity differed statistically significantly among all three of the methods of extraction, the estimated means differed by no more than 1.5 cycles ( P < 0.05). Seventy (76%) of the 92 specimens that were LightCycler PCR positive by all three extraction methods were also positive by shell vial cell culture assay. HSV DNA was detected by a lower LightCycler PCR cycle number (26.1 cycles) in specimens culture positive for the virus than in culture-negative samples (33.3 cycles) ( P < 0.0001). The manual IsoQuick and automated MagNA Pure and BioRobot 9604 methods provide standardized, reproducible extraction of HSV DNA for LightCycler PCR. The decision to implement a manual versus an automated procedure depends on factors such as costs related to the number of specimens processed rather than on the minimal differences in the technical efficiency of extraction of nucleic acids among these methods.

ABSTRACT A comparison of quantitative results expressed in hepatitis C virus (HCV) international units per milliliter, obtained from the VERSANT HCV RNA 3.0 (bDNA-3.0) assay, the QUANTIPLEX HCV RNA 2.0 (bDNA-2.0) assay, and the COBAS AMPLICOR HCV MONITOR version 2.0 (HCM-2.0) test was performed. A total of 168 patient specimens submitted to the Mayo Clinic Molecular Microbiology Laboratory for HCV quantification or HCV genotyping were studied. Of the specimens tested, 97, 88, and 79% yielded quantitative results within the dynamic range of the bDNA-3.0, bDNA-2.0, and HCM-2.0 assays, respectively. Overall, there was substantial agreement between the results generated by all three assays. A total of 15 out of 29 (52%) of the specimens determined to contain viral loads of <31,746 IU/ml by the bDNA-3.0 assay were categorized as containing viral loads within the range of 31,746 to 500,000 IU/ml by the bDNA-2.0 assay. Although substantial agreement was noted between the results generated by the bDNA-2.0 and bDNA-3.0 assays, a bias toward higher viral titer by the bDNA-2.0 assay was noted ( P = 0.001). Likewise, although substantial agreement was noted between the results generated by the HCM-2.0 and bDNA-3.0 assays, a bias toward higher viral titer by the bDNA-3.0 assay was noted ( P ≤ 0.001). The discrepancy between the HCM-2.0 and bDNA-3.0 results was more pronounced when viral loads were >500,000 IU/ml and resulted in statistically significant differences ( P ≤ 0.001) in determining whether viral loads were above or below 800,000 IU/ml of HCV RNA, the proposed threshold value for tailoring the duration of combination therapy. The expression of quantitative values in HCV international units per milliliter was a strength of both the bDNA-3.0 and HCM-2.0 assays.