Achenbach, Sara J.

Abstract Introduction Chronic lymphocytic leukemia (CLL) is a B-cell malignancy that is known to have a familial component to disease risk. Although 31 loci have been found to be associated with CLL risk, the functional variant(s) driving these associations is mostly unknown. Here we set out to identify rare, highly-penetrant, cosegregating, susceptibility variants within the known GWAS discovered loci using whole exome sequencing (WES) data in CLL families from the Mayo Clinic family study of B-cell malignancies. Methods We performed WES on germ line DNA of 93 CLL families with two or more members with CLL, using Agilent capture kits and Illumina HiSeq2000. Bioinformatics analyses leveraged the following software packages: Novoalign, Picard, The Genome Analysis Toolkit (GATK), and the Biological Reference Repository (bioR). Quality control filters were implemented; subjects with mis-specified relationships were removed, as were variants with <75% call rate, <8X coverage, and those identified as sequencing artifacts. Each GWAS locus was defined by +/- 1Mb of the top GWAS hit within the locus. Linkage disequilibrium (LD) was calculated among the single nucleotide variants (SNVs) located within each locus. Potentially functional SNVs were identified based on: a) uncommon in public databases (< 5%), b) cosegregating in at least two CLL families, c) being highly conserved and in coding regions, and d) functional prediction status of deleterious (SIFT Score), damaging (PolyPhen Score), and a moderate, or high variant impact (SNP Effect). Results In our 93 CLL families, we sequenced 443 individuals: 160 with CLL, 73 with monoclonal B cell lymphocytosis (MBL), and 210 relatives that were not diagnosed with CLL or MBL at the time of sequencing. Median age of CLL diagnosis was 59 years (range 34-87), and 56% were male. Among the MBL individuals and relatives, the median age at recruitment was 55 years (range 18-93), and 40% were male. A total of 317,666 SNVs passed our sequencing quality control filters of which 10,731 were within +/- 1 Mb of known GWAS hits from 31 loci. Of these SNVs, 91% were in coding regions, 18% were reported to have high or moderate impact, 6% were estimated to be damaging and 6% were predicted to be deleterious. From these SNVs, we identified 76 putatively functional variants distributed across 25 GWAS loci that were cosegregating in the individuals with CLL or MBL in multiple CLL pedigrees. These SNVs were all located in coding regions with high or moderate impact and were predicted to be damaging and deleterious. Of these 76 variants, 56 had a frequency of <0.005 in 1000 Genomes’ European population while the remaining 20 had a frequency of 1%. Conclusions Through WES, we identified a number of rare, penetrant and potentially predisposing SNVs located within 25 of the 31 CLL GWAS-discovered loci. These segregating variants provide a list for future validation and functional studies. Citation Format: Sara Beiggi, Daniel R. O'Brien, Sara J. Achenbach, Kari G. Chaffee, Timothy G. Call, Neil E. Kay, Tait D. Shanafelt, Julie Cunningham, James R. Cerhan, Celine M. Vachon, Susan L. Slager. Cosegregating variants in chronic lymphocytic leukemia (CLL) families that are located in loci discovered by genome wide association studies (GWAS). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2764. doi:10.1158/1538-7445.AM2015-2764

Abstract Background Previous literature regarding the association between occupational exposure to agricultural pesticides and non-Hodgkin lymphoma (NHL) risk has been inconsistent, particularly when looking at specific lymphoma subtypes. Most recently, a large European case-control study showed an elevated risk of chronic lymphocytic leukemia (CLL) among those ever exposed to inorganic and organic pesticides with the strongest association among those exposed to organophosphates. However, when restricting the analysis to subjects with high confidence of exposure, these associations were attenuated (Cocco P., 2013). We used case-control data from the Mayo Clinic to evaluate whether there is an increased risk of CLL among individuals with known occupational exposure to agricultural pesticides. Methods We evaluated pesticide exposure and CLL risk in a clinic-based study of newly diagnosed CLL cases and frequency-matched controls enrolled at the Mayo Clinic from 2002-2012. 200 CLL cases and 474 controls returned a detailed farming and pesticide exposure questionnaire that was modeled after the Agricultural Health Study, where exposure was based on pesticide-specific application. Unconditional logistic regression, adjusted for age and sex, was used to estimate odds ratios (ORs) and 95% confidence intervals (CI). We evaluated the effect of each pesticide individually, by chemical class of pesticides, and by an aggregate of multiple pesticide exposure on CLL risk. Results Among those participants with detailed farming data, the mean age of CLL diagnosis was 66 years and 76% were male; for controls, the mean age at enrollment was 65 years and 66% were male. Risk of CLL was mildly elevated, but not significant, amongst those with application of any pesticide (OR 1.39; 95% C.I. 0.92-2.10). When broken down by chemical class, risk of CLL was also mildly elevated, but not significant, for application of organophosphates and carbamates (OR 1.21; 95% CI 0.84-1.73; OR 1.21; 95% 0.85-1.73, respectively). Organochlorines, phenoxy compounds, pyrethriods and triazines showed no evidence of an elevated risk, nor did we see evidence of a trend that CLL risk increases with number of pesticides applied. Discussion Our preliminary results provide inconclusive support for the role of pesticide exposure as a possible risk factor for CLL. However our sample size was small. Additional data will be incorporated, including confidence of exposure measures, as well as other modifying factors, to further evaluate these results. Citation Format: Emily J. Hallberg, Sara J. Achenbach, Kari G. Rabe, Timothy G. Call, Cristine Allmer, Tait D. Shanafelt, Mark Liebow, Neil E. Kay, James R. Cerhan, Susan L. Slager. Occupational exposure to agricultural pesticides and CLL risk. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 274. doi:10.1158/1538-7445.AM2014-274

Abstract Background: Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL) is the most common adult leukemia in the United States. Prior studies in Olmsted County, Minnesota, showed a CLL age and sex adjusted incidence rate of 6.6 per 100,000 during 1985-89. Subsequently, the definition of CLL is now based on the 2008 International Workshop on CLL (IWCLL) Guidelines. Individuals having a CLL immunophenotype by flow cytometry are classified as CLL if the absolute B-cell count (ABC) is >5 x 109/L or classified as monoclonal B-cell lymphocytosis (MBL) if ABC is <5 x 109/L. To assess the impact of these changes on the incidence of CLL and clinically discovered MBL, we conducted an incidence study in Olmsted County for the years 2000-2010. Methods Using diagnostic indexes available through the Rochester Epidemiology Project and the Mayo CLL database, we identified all CLL and clinically identified MBL cases from 2000-2010. Incidence rates and demographics were compared between the older National Cancer Institute-Working Groups 1996 guidelines (NCI-WG 96) defined as absolute lymphocyte count (ALC)> 5 x 109/L and the IWCLL 2008 guidelines defined as ABC > 5 x 109/L. Incidence rates were age and sex adjusted using the 2010 US white population. Time to treatment (TTT) survival analyses was from time of diagnosis to date of first treatment or last follow-up and Cox regression analysis was used. Results Using the NCI-WG 96 classification, there were 8 MBL cases and 115 CLLs in whom 61% were Rai stage 0, 34% were Rai stage 1-2, and 5% were Rai stage 3-4. Using the IWCLL 2008 classification, there were 44 MBL cases and 79 CLLs in whom 43% Rai stage 0, 49% were Rai stage 1-2 and 8% Rai stage 3-4. Median ALC at diagnosis was 8.1 x 109/L for CLL and 5.9 x 109/L for MBL. Median ABC count at diagnosis was 7.1 x 109/L for CLL and 2.8 x 109/L for MBL. The age and sex adjusted incidence rate (per 100,000) for CLL for 2000-2010 was 10.0 using NCI-WG 96 and 6.8 using IWCLL 2008. Rates for MBL were 0.66 using NCI-WG 96 and 3.5 using IWCLL 2008. The median TTT for CLL was 9.2 years using NCI-WG 96 and 6.5 years by IWCLL 2008. Discussion This study demonstrates that reclassification of CLL using the IWCLL 2008 guidelines decreases CLL incidence relative to the NCI-WG 96 criteria. For the first time, this population based study also establishes an incidence rate for clinically identified MBL. Approximately 50% of previously labeled Rai 0 CLL are now reclassified as MBL. Re-classification increases the percent of Rai stage 1-2 and decreases the percent of Rai 0 cases at diagnosis. Reclassification also substantially shortens the median TTT for newly diagnosed CLL cases classified by the 2008 criteria relative to the NCI-WG 96 criteria. In summary, the IWCLL 2008 reclassification has significant implications for incidence studies, staging, and prognosis of CLL and MBL. Citation Format: Aaron D. Norman, Timothy G. Call, Curtis A. Hanson, Neil E. Kay, Clive S. Zent, Wei Ding, James R. Cerhan, Sara J. Achenbach, Kari G. Rabe, Celine M. Vachon, Emily J. Hallberg, Tait D. Shanafelt, Susan L. Slager. Incidence of chronic lymphocytic leukemia and monoclonal B-cell lymphocytosis in Olmsted county, 2000-2010: impact of the 2008 International Workshop on CLL Guidelines. [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 2287. doi:10.1158/1538-7445.AM2013-2287

ABSTRACT Children and adolescents who sustain a distal forearm fracture (DFF) owing to mild, but not moderate, trauma have reduced bone strength and cortical thinning at the distal radius and tibia. Whether these skeletal deficits track into adulthood is unknown. Therefore, we studied 75 women and 75 men (age range, 20 to 40 years) with a childhood (age <18 years) DFF and 150 sex-matched controls with no history of fracture using high-resolution peripheral quantitative computed tomography (HRpQCT) to examine bone strength (ie, failure load) by micro–finite element (µFE) analysis, as well as cortical and trabecular bone parameters at the distal radius and tibia. Level of trauma (mild versus moderate) was assigned using a validated classification scheme, blind to imaging results. When compared to sex-matched, nonfracture controls, women and men with a mild trauma childhood DFF (eg, fall from standing height) had significant reductions in failure load (p < 0.05) of the distal radius, whereas women and men with a moderate trauma childhood DFF (eg, fall while riding a bicycle) had values similar to controls. Consistent findings were observed at the distal tibia. Furthermore, women and men with a mild trauma childhood DFF had significant deficits in distal radius cortical area (p < 0.05), and significantly lower dual-energy X-ray absorptiometry (DXA)-derived bone density at the radius, hip, and total body regions compared to controls (all p < 0.05). By contrast, women and men with a moderate trauma childhood DFF had bone density, structure, and strength that did not differ significantly from controls. These findings in young adults are consistent with our observations in children/adolescents with DFF, and they suggest that a mild trauma childhood DFF may presage suboptimal peak bone density, structure, and strength in young adulthood. Children and adolescents who suffer mild trauma DFFs may need to be targeted for lifestyle interventions to help achieve improved skeletal health. © 2014 American Society for Bone and Mineral Research.

BACKGROUNDThe 1996 National Cancer Institute Working Group (NCI‐WG 96) guidelines classified disease in individuals who had a B‐cell clone with chronic lymphocytic leukemia (CLL) immunophenotype as CLL if their absolute lymphocyte count was ≥5 × 109/L. The 2008 International Workshop on CLL guidelines (IWCLL 2008) classified disease as CLL if the absolute B‐cell count was ≥5 × 109/L or as monoclonal B‐cell lymphocytosis (MBL) if the absolute B‐cell count was <5 × 109/L. The objective of the current study of Olmsted County, Minnesota, was to assess the effects of these changes on incidence rates and presentation from 2000 to 2010.METHODSUsing diagnostic indices available through the Rochester Epidemiology Project and the Mayo CLL database, the authors identified all patients with newly diagnosed CLL and high‐count MBL from 2000 to 2010. Age‐specific and sex‐specific incidence rates were determined.RESULTSAccording to NCI‐WG 96 criteria, there were 115 patients with CLL and 8 patients with MBL during the period studied. Using the IWCLL 2008 classification, there were 79 patients with CLL and 40 patients with MBL. Rai stage distribution (low risk, intermediate risk, and high risk) using NCI‐WG 96 criteria was 60.9%, 33.9%, and 5.2%, respectively, compared with 43%, 49.4%, and 7.6%, respectively, using IWCLL 2008 criteria. The age‐adjusted and sex‐adjusted incidence rates (per 100,000) for CLL and MBL were 10.0 and 0.66, respectively, using NCI‐WG 96 criteria versus 6.8 and 3.5, respectively, using IWCLL 2008 criteria. The median time to treatment according to NCI‐WG 96 criteria was 9.2 years versus 6.5 years with IWCLL 2008 criteria.CONCLUSIONSUse of the IWCLL 2008 guidelines reduced the incidence of CLL, altered the distribution of initial Rai stage at diagnosis, and shortened the median time to treatment. Cancer 2014;120:2000–2005. © 2014 American Cancer Society.

ABSTRACT To assess recent trends in fracture incidence from all causes at all skeletal sites, we used the comprehensive (inpatient and outpatient) data resources of the Rochester Epidemiology Project to estimate rates for Olmsted County, MN, USA, residents in 2009 to 2011 compared with similar data from 1989 to 1991. During the 3-year study period, 2009 to 2011, 3549 residents ≥50 years of age experienced 5244 separate fractures. The age- and sex-adjusted (to the 2010 US white population) incidence of any fracture was 2704 per 100,000 person-years (95% confidence interval [CI] 2614 to 2793) and that for all fractures was 4017 per 100,000 (95% CI 3908 to 4127). Fracture incidence increased with age in both sexes, but age-adjusted rates were 49% greater among the women. Overall, comparably adjusted fracture incidence rates increased by 11% (from 3627 to 4017 per 100,000 person-years; p = 0.008) between 1989 to 1991 and 2009 to 2011. This was mainly attributable to a substantial increase in vertebral fractures (+47% for both sexes combined), which was partially offset by a decline in hip fractures (−25%) among the women. There was also a 26% reduction in distal forearm fractures among the women; an increase in distal forearm fractures among men aged 50 years and over was not statistically significant. The dramatic increase in vertebral fractures, seen in both sexes and especially after age 75 years, was attributable in part to incidentally diagnosed vertebral fractures. However, the fall in hip fracture incidence, observed in most age groups, continues the steady decline observed among women in this community since 1950. More generally, these data indicate that the dramatic increases in the incidence of fractures at many skeletal sites that were observed decades ago have now stabilized. © 2014 American Society for Bone and Mineral Research.

ABSTRACT Distal forearm fractures are among the most common fractures during childhood, but it remains unclear whether they predict an increased fracture risk later in life. We studied a population-based cohort of 1776 children ≤18 years of age, from Olmsted County, MN, USA, who had a distal forearm fracture in 1935–1992. Incident fractures occurring at age ≥35 years were identified through review of complete medical records using the linkage system of the Rochester Epidemiology Project. Observed nonpathologic fractures resulting from no more than moderate trauma (fragility fractures) were compared with expected numbers estimated from fracture site–specific incidence rates, based on age, sex, and calendar year, for Olmsted County (standardized incidence ratios [SIR]). In 1086 boys (mean ± SD age; 11 ± 4 years) and 690 girls (10 ± 4 years) followed for 27,292 person-years after the age of 35 years, subsequent fragility fractures were observed in 144 (13%) men and 74 (11%) women. There was an increased risk for future fragility fractures in boys who had a distal forearm fracture (SIR, 1.9; 95% CI, 1.6–2.3) but not girls (SIR, 1.0; 95% CI, 0.8–1.2). Fragility fractures at both major osteoporotic (hip, spine, wrist, and shoulder) sites (SIR, 2.6; 95% CI, 2.1–3.3) and remaining sites (SIR, 1.7; 95% CI, 1.3–2.0) were increased in men, irrespective of age at distal forearm fracture as boys. A distal forearm fracture in boys, but not girls, is associated with an increased risk for fragility fractures as older adults. It is necessary to determine whether the increased fractures observed in men is due to persistent deficits of bone strength, continued high fracture risk activity, or both. Until then, men should be asked about a childhood distal forearm fracture and, if so, warrant further screening and counseling on measures to optimize bone health and prevent fractures.

ObjectivePulmonary disease represents an important extraarticular manifestation of rheumatoid arthritis (RA). While the association of RA and interstitial lung disease is widely acknowledged, obstructive lung disease (OLD) in RA is less well understood. We therefore aimed to assess the incidence, risk factors, and mortality of OLD in patients with RA.MethodsWe examined a population‐based incident cohort of patients with RA and a comparison cohort of individuals without RA. OLD was defined using a strict composite criterion. Cox proportional hazards models were used to compare OLD incidence between the RA and comparator cohorts to investigate risk factors and to explore the impact of OLD on patient survival.ResultsA total of 594 patients with RA and 596 subjects without RA were followed for a mean of 16.3 and 19.4 years, respectively. The lifetime risk of developing OLD was 9.6% for RA patients and 6.2% for subjects without RA (hazard ratio [HR] 1.54, 95% confidence interval [95% CI] 1.01–2.34). The risk of developing OLD was higher among male patients, among current or former smokers, and for individuals with more severe RA. Survival of RA patients diagnosed with OLD was worse compared to those without OLD (HR 2.09, 95% CI 1.47–2.97).ConclusionPatients with RA are at higher risk of developing OLD, which is significantly associated with premature mortality. Effective diagnostic and therapeutic strategies to detect and manage OLD in patients with RA may help to improve survival in these patients.

Abstract Background: Our genome-wide association study (GWAS) of chronic lymphocytic leukemia (CLL) identified 4 highly correlated intronic variants within the IRF8 gene that were associated with CLL. These results were further supported by a recent meta-analysis of our GWAS with two other GWAS of CLL, supporting the IRF8 gene as a strong candidate for CLL risk. Methods: To refine the genetic association of CLL risk, we conducted Sanger sequencing of IRF8 in 94 CLL cases and 96 controls. We then conducted fine mapping by genotyping 39 variants (of which 10 were identified from sequencing) in 745 CLL cases and 1,521 controls. We also assessed these associations with risk of other non-Hodgkin lymphoma (NHL) subtypes. Results: The strongest association with CLL risk was observed with a common single-nucleotide polymorphism (SNP) located within the 3′ untranslated region (UTR) of IRF8 (rs1044873, log additive OR = 0.7, P = 1.81 × 10−6). This SNP was not associated with the other NHL subtypes (all P &gt; 0.05). Conclusions: We provide evidence that rs1044873 in the IRF8 gene accounts for the initial GWAS signal for CLL risk. This association appears to be unique to CLL with little support for association with other common NHL subtypes. Future work is needed to assess functional role of IRF8 in CLL etiology. Impact: These data provide support that a functional variant within the 3′UTR of IRF8 may be driving the GWAS signal seen on 16q24.1 for CLL risk. Cancer Epidemiol Biomarkers Prev; 22(3); 461–6. ©2013 AACR.