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Background: Gonadotoxic treatment-related infertility has a significant impact on quality of life in childhood cancer survivors. Genome-wide association analyses to delineate the risk of infertility in childhood cancer survivors have not been previously reported. Methods: Leveraging genotype data from a large survivor cohort, the Childhood Cancer Survivor Study (CCSS), we investigated the role of SNPs on future pregnancy or siring a pregnancy in survivors without pelvic, testicular, or brain radiation who had ever been married. We calculated sex-stratified hazard ratios, using Cox proportional hazards modeling, adjusting for birth cohort (before 1965 vs. 1965 or later) and doses of relevant chemotherapies; replication was attempted in the independent St. Jude Lifetime Cohort study (SJLIFE). Results: In the CCSS cohort, nine SNPs were found to be suggestive (P < 10–7) or statistically significantly (P < 5 × 10–8) associated with pregnancy, however, none of the SNPs were replicated in SJLIFE. Cohorts differed based on the overall pregnancy rate, frequency of sterilizing procedures, and birth cohort. Conclusions: We were not able to replicate our findings of SNPs associated with pregnancy in childhood cancer survivors. Impact: Future attempts at replication should be considered in cohorts treated in a comparable era. In addition, understanding the role of genetics in fertility in childhood cancer survivors may be better approached using more advanced sequencing techniques.

Breast cancer is the most common cancer in females, affecting one in every eight women and accounting for the majority of cancer-related deaths in women worldwide. Germline mutations in theBRCA1andBRCA2genes are significant risk factors for specific subtypes of breast cancer.BRCA1mutations are associated with basal-like breast cancers, whereasBRCA2mutations are associated with luminal-like disease. There are currently few chemoprevention strategies available forBRCA1/2mutation carriers, and irreversible prophylactic mastectomy is the primary option. Designing chemo-preventive strategies requires an in-depth understanding of the physiological processes underlying tumor initiation. Here, we employ spatial transcriptomics to investigate defects in mammary epithelial cell differentiation accompanied by distinct microenvironmental alterations in preneoplastic breast tissues fromBRCA1/2mutation carriers and normal breast tissues from non-carrier controls. We uncovered spatially defined receptor-ligand interactions in these tissues for the investigation of autocrine and paracrine signaling. We discovered that β1-integrin-mediated autocrine signaling inBRCA2-deficient mammary epithelial cells differs fromBRCA1-deficient mammary epithelial cells. In addition, we found that the epithelial-to-stromal paracrine signaling in the breast tissues ofBRCA1/2mutation carriers is greater than in control tissues. More integrin-ligand pairs were differentially correlated inBRCA1/2-mutant breast tissues than non-carrier breast tissues with more integrin receptor-expressing stromal cells. These results reveal alterations in the communication between mammary epithelial cells and the microenvironment inBRCA1andBRCA2mutation carriers, laying the foundation for designing innovative breast cancer chemo-prevention strategies for high-risk patients.

Epithelial ovarian cancer (EOC) is the leading cause of gynecologic cancer death. Despite initial responses to intervention, up to 80% of patient tumors recur and require additional treatment. Retrospective clinical analysis of patients with ovarian cancer indicates antibiotic use during chemotherapy treatment is associated with poor overall survival. Here, we assessed whether antibiotic (ABX) treatment would impact growth of EOC and sensitivity to cisplatin. Immunocompetent or immunocompromised mice were given untreated control or ABX-containing (metronidazole, ampicillin, vancomycin, and neomycin) water prior to intraperitoneal injection with EOC cells, and cisplatin therapy was administered biweekly until endpoint. Tumor-bearing ABX-treated mice exhibited accelerated tumor growth and resistance to cisplatin therapy compared with control treatment. ABX treatment led to reduced apoptosis, increased DNA damage repair, and enhanced angiogenesis in cisplatin-treated tumors, and tumors from ABX-treated mice contained a higher frequency of cisplatin-augmented cancer stem cells than control mice. Stool analysis indicated nonresistant gut microbial species were disrupted by ABX treatment. Cecal transplants of microbiota derived from control-treated mice was sufficient to ameliorate chemoresistance and prolong survival of ABX-treated mice, indicative of a gut-derived tumor suppressor. Metabolomics analyses identified circulating gut-derived metabolites that were altered by ABX treatment and restored by recolonization, providing candidate metabolites that mediate the cross-talk between the gut microbiome and ovarian cancer. Collectively, these findings indicate that an intact microbiome functions as a tumor suppressor in EOC, and perturbation of the gut microbiota with ABX treatment promotes tumor growth and suppresses cisplatin sensitivity.Significance:Restoration of the gut microbiome, which is disrupted following antibiotic treatment, may help overcome platinum resistance in patients with epithelial ovarian cancer.See related commentary by Hawkins and Nephew, p. 4511

Supplemental Figures and Tables supporting manuscript.

Context A sex discordance in COVID exists, with males disproportionately affected. Although sex steroids may play a role in this discordance, no definitive genetic data exist to support androgen-mediated immune suppression neither for viral susceptibility nor for adrenally produced androgens. Objective The common adrenal-permissive missense-encoding variant HSD3B1(1245C) that enables androgen synthesis from adrenal precursors and that has been linked to suppression of inflammation in severe asthma was investigated in COVID susceptibility and outcomes reported in the UK Biobank. Methods The UK Biobank is a long-term study with detailed medical information and health outcomes for over 500 000 genotyped individuals. We obtained COVID test results, inpatient hospital records, and death records and tested for associations between COVID susceptibility or outcomes and HSD3B1(1245A/C) genotype. Primary analyses were performed on the UK Biobank Caucasian cohort. The outcomes were identification as a COVID case among all subjects, COVID positivity among COVID-tested subjects, and mortality among subjects identified as COVID cases. Results Adrenal-permissive HSD3B1(1245C) genotype was associated with identification as a COVID case (odds ratio (OR): 1.11 per C allele, 95% CI: 1.04–1.18, P = 0.0013) and COVID-test positivity (OR: 1.09, 95% CI: 1.02–1.17, P = 0.011) in older (≥70 years of age) women. In women identified as COVID cases, there was a positive linear relationship between age and 1245C allele frequency (P < 0.0001). No associations were found between genotype and mortality or between genotype and circulating sex hormone levels. Conclusion Our study suggests that a common androgen synthesis variant regulates immune susceptibility to COVID infection in women, with increasingly strong effects as women age.

Germline PTEN variants (PTEN hamartoma tumor syndrome [PHTS]) confer up to 85% lifetime risk of female breast cancer (BC). BCs arising in PHTS are clinically distinct from sporadic BCs, including younger age of onset, multifocality, and an increased risk of second primary BCs. Yet, there is no previous investigation into the underlying genomic landscape of this entity. We sought to address the hypothesis that BCs arising in PHTS have a distinct genomic landscape compared to sporadic counterparts. We performed and analyzed exome sequencing data from 44 women with germline PTEN variants who developed BCs. The control cohort comprised of 497 women with sporadic BCs from The Cancer Genome Atlas (TCGA) dataset. We demonstrate that PHTS-derived BCs have a distinct somatic mutational landscape compared to the sporadic counterparts, namely second somatic hits in PTEN, distinct mutational signatures, and increased genomic instability. The PHTS group had a significantly higher frequency of somatic PTEN variants compared to TCGA (22.7% versus 5.6%; odds ratio [OR] 4.93; 95% confidence interval [CI] 2.21 to 10.98; p

Background Acetylation and methylation of histones alter the chromatin structure and accessibility that affect transcriptional regulators binding to enhancers and promoters. The binding of transcriptional regulators enables the interaction between enhancers and promoters, thus affecting gene expression. However, our knowledge of these epigenetic alternations in patients with heart failure remains limited. Methods and results From the comprehensive analysis of major histone modifications, 3-dimensional chromatin interactions, and transcriptome in left ventricular (LV) tissues from dilated cardiomyopathy (DCM) patients and non-heart failure (NF) donors, differential active enhancer and promoter regions were identified between NF and DCM. Moreover, the genome-wide average promoter signal is significantly lower in DCM than in NF. Super-enhancer (SE) analysis revealed that fewer SEs were found in DCM LVs than in NF ones, and three unique SE-associated genes between NF and DCM were identified. Moreover, SEs are enriched within the genomic region associated with long-range chromatin interactions. The differential enhancer-promoter interactions were observed in the known heart failure gene loci and are correlated with the gene expression levels. Motif analysis identified known cardiac factors and possible novel players for DCM. Conclusions We have established the cistrome of four histone modifications and chromatin interactome for enhancers and promoters in NF and DCM tissues. Differential histone modifications and enhancer-promoter interactions were found in DCM, which were associated with gene expression levels of a subset of disease-associated genes in human heart failure.

Significance Although resistance to glucocorticoids is a major clinical problem, the underlying mechanisms are unknown. It is known that glucocorticoid use can suppress adrenal androgen production. In population studies, animal models, and cell culture experiments, androgens are associated with several benefits in asthma, but neither androgen use in glucocorticoid-resistant asthma nor the genetic determinants of androgen responsiveness have been studied in humans. A missense-encoding variant in HSD3B1 is known to regulate conversion from adrenal precursors to potent androgens and clinical outcomes in prostate cancer. This is the first genetic evidence to our knowledge that implicates an androgen synthesis variant in resistance to glucocorticoids for asthma or any other inflammatory disease. Furthermore, this study demonstrates an adverse consequence of adrenal androgen suppression with glucocorticoid therapy., Asthma resistance to glucocorticoid treatment is a major health problem with unclear etiology. Glucocorticoids inhibit adrenal androgen production. However, androgens have potential benefits in asthma. HSD3B1 encodes for 3β-hydroxysteroid dehydrogenase-1 (3β-HSD1), which catalyzes peripheral conversion from adrenal dehydroepiandrosterone (DHEA) to potent androgens and has a germline missense-encoding polymorphism. The adrenal restrictive HSD3B1(1245A) allele limits conversion, whereas the adrenal permissive HSD3B1(1245C) allele increases DHEA metabolism to potent androgens. In the Severe Asthma Research Program (SARP) III cohort, we determined the association between DHEA-sulfate and percentage predicted forced expiratory volume in 1 s (FEV1PP). HSD3B1(1245) genotypes were assessed, and association between adrenal restrictive and adrenal permissive alleles and FEV1PP in patients with (GC) and without (noGC) daily oral glucocorticoid treatment was determined (n = 318). Validation was performed in a second cohort (SARP I&II; n = 184). DHEA-sulfate is associated with FEV1PP and is suppressed with GC treatment. GC patients homozygous for the adrenal restrictive genotype have lower FEV1PP compared with noGC patients (54.3% vs. 75.1%; P < 0.001). In patients with the homozygous adrenal permissive genotype, there was no FEV1PP difference in GC vs. noGC patients (73.4% vs. 78.9%; P = 0.39). Results were independently confirmed: FEV1PP for homozygous adrenal restrictive genotype in GC vs. noGC is 49.8 vs. 63.4 (P < 0.001), and for homozygous adrenal permissive genotype, it is 66.7 vs. 67.7 (P = 0.92). The adrenal restrictive HSD3B1(1245) genotype is associated with GC resistance. This effect appears to be driven by GC suppression of 3β-HSD1 substrate. Our results suggest opportunities for prediction of GC resistance and pharmacologic intervention.

Epithelial ovarian cancer (EOC) is the leading cause of gynecologic cancer death. Despite initial responses to intervention, up to 80% of patient tumors recur and require additional treatment. Retrospective clinical analysis of patients with ovarian cancer indicates antibiotic use during chemotherapy treatment is associated with poor overall survival. Here, we assessed whether antibiotic (ABX) treatment would impact growth of EOC and sensitivity to cisplatin. Immunocompetent or immunocompromised mice were given untreated control or ABX-containing (metronidazole, ampicillin, vancomycin, and neomycin) water prior to intraperitoneal injection with EOC cells, and cisplatin therapy was administered biweekly until endpoint. Tumor-bearing ABX-treated mice exhibited accelerated tumor growth and resistance to cisplatin therapy compared with control treatment. ABX treatment led to reduced apoptosis, increased DNA damage repair, and enhanced angiogenesis in cisplatin-treated tumors, and tumors from ABX-treated mice contained a higher frequency of cisplatin-augmented cancer stem cells than control mice. Stool analysis indicated nonresistant gut microbial species were disrupted by ABX treatment. Cecal transplants of microbiota derived from control-treated mice was sufficient to ameliorate chemoresistance and prolong survival of ABX-treated mice, indicative of a gut-derived tumor suppressor. Metabolomics analyses identified circulating gut-derived metabolites that were altered by ABX treatment and restored by recolonization, providing candidate metabolites that mediate the cross-talk between the gut microbiome and ovarian cancer. Collectively, these findings indicate that an intact microbiome functions as a tumor suppressor in EOC, and perturbation of the gut microbiota with ABX treatment promotes tumor growth and suppresses cisplatin sensitivity. Significance: Restoration of the gut microbiome, which is disrupted following antibiotic treatment, may help overcome platinum resistance in patients with epithelial ovarian cancer. See related commentary by Hawkins and Nephew, p. 4511

Background Trimethylamine‐ N ‐oxide (TMAO) is a small molecule derived from the metabolism of dietary nutrients by gut microbes and contributes to cardiovascular disease. Plasma TMAO increases following consumption of red meat. This metabolic change is thought to be partly because of the expansion of gut microbes able to use nutrients abundant in red meat. Methods and Results We used data from a randomized crossover study to estimate the degree to which TMAO can be estimated from fecal microbial composition. Healthy participants received a series of 3 diets that differed in protein source (red meat, white meat, and non‐meat), and fecal, plasma, and urine samples were collected following 4 weeks of exposure to each diet. TMAO was quantitated in plasma and urine, while shotgun metagenomic sequencing was performed on fecal DNA. While the cai gene cluster was weakly correlated with plasma TMAO (rho=0.17, P =0.0007), elastic net models of TMAO were not improved by abundances of bacterial genes known to contribute to TMAO synthesis. A global analysis of all taxonomic groups, genes, and gene families found no meaningful predictors of TMAO. We postulated that abundances of known genes related to TMAO production do not predict bacterial metabolism, and we measured choline‐ and carnitine‐trimethylamine lyase activity during fecal culture. Trimethylamine lyase genes were only weakly correlated with the activity of the enzymes they encode. Conclusions Fecal microbiome composition does not predict systemic TMAO because, in this case, gene copy number does not predict bacterial metabolic activity. Registration URL: https://www.clinicaltrials.gov ; Unique identifier: NCT01427855.

BACKGROUND Genetics of estrogen synthesis and breast cancer risk has been elusive. The 1245A→C missense-encoding polymorphism in HSD3B1, which is common in White populations, is functionally adrenal permissive and increases synthesis of the aromatase substrate androstenedione. We hypothesized that homozygous inheritance of the adrenal-permissive HSD3B1(1245C) is associated with postmenopausal estrogen receptor–positive (ER-positive) breast cancer. METHODS A prospective study of postmenopausal ER-driven breast cancer was done for determination of HSD3B1 and circulating steroids. Validation was performed in 2 other cohorts. Adrenal-permissive genotype frequency was compared between postmenopausal ER-positive breast cancer, the general population, and postmenopausal ER-negative breast cancer. RESULTS Prospective and validation studies had 157 and 538 patients, respectively, for the primary analysis of genotype frequency by ER status in White female breast cancer patients who were postmenopausal at diagnosis. The adrenal-permissive genotype frequency in postmenopausal White women with estrogen-driven breast cancer in the prospective cohort was 17.5% (21/120) compared with 5.4% (2/37) for ER-negative breast cancer (P = 0.108) and 9.6% (429/4451) in the general population (P = 0.0077). Adrenal-permissive genotype frequency for estrogen-driven postmenopausal breast cancer was validated using Cambridge and The Cancer Genome Atlas data sets: 14.4% (56/389) compared with 6.0% (9/149) for ER-negative breast cancer (P = 0.007) and the general population (P = 0.005). Circulating androstenedione concentration was higher with the adrenal-permissive genotype (P = 0.03). CONCLUSION Adrenal-permissive genotype is associated with estrogen-driven postmenopausal breast cancer. These findings link genetic inheritance of endogenous estrogen exposure to estrogen-driven breast cancer. FUNDING National Cancer Institute, NIH (R01CA236780, R01CA172382, and P30-CA008748); and Prostate Cancer Foundation Challenge Award.

Glucocorticoids are a highly effective first-line treatment option for many inflammatory diseases, including asthma. Some patients develop a steroid-resistant condition, yet, the cellular and molecular mechanisms underlying steroid resistance remain largely unknown. In this study, we used a murine model of steroid-resistant airway inflammation and report that combining systemic dexamethasone and intranasal IL-27 is able to reverse the inflammation. Foxp3+ regulatory T cells (Tregs) were required during dexamethasone/IL-27 treatment of steroid-resistant allergic inflammation, and importantly, direct stimulation of Tregs via glucocorticoid or IL-27 receptors was essential. Mechanistically, IL-27 stimulation in Tregs enhanced expression of the agonistic glucocorticoid receptor-α isoform. Overexpression of inhibitory glucocorticoid receptor-β isoform in Tregs alone was sufficient to elicit steroid resistance in a steroid-sensitive allergic inflammation model. Taken together, our results demonstrate for the first time, to our knowledge, that Tregs are instrumental during steroid resistance and that manipulating steroid responsiveness in Tregs may represent a novel strategy to treat steroid refractory asthma.

Introduction: Despite the exceptional promise of anti-CD19 CAR T-cell therapy, 40-60% of relapsed or refractory large B-cell lymphoma (R/R LBCL) patients exhibit poor response and survival. Therefore, identification of pre-treatment markers of response to anti-CD19 CAR T-cell therapy is essential. Immune responses to infections and many autoimmune diseases exhibit gender bias, but it is not known if gender influences response to CAR T-cell therapy. We performed serum profiling to identify pre-treatment cytokines and chemokines associated with response and survival in male and female LBCL patients treated with axicabtagene ciloleucel (axi-cel). Methods: Serum samples were collected from peripheral blood at the time of leukapheresis from 42 R/R LBCL patients treated with axi-cel. Multiplexed bead arrays were used to quantify baseline levels of 51 serum cytokines, chemokines and cytotoxic proteins. Unpaired Wilcoxon test was used to analyze the association of baseline cytokine levels with 3-month response, and Kaplan-Meier survival analysis for association with survival outcomes. Statistical analyses were performed using R and p Results: The median age of patients was 61 y, and 62% of the patients were male. All patients had been previously treated with R-CHOP or R-EPOCH, and had a median of 3 prior lines of therapy, including 47.6% with autologous stem cell transplantation. Male non-responders (stable/progressive disease) had significantly higher baseline levels of IL-6, IL-8, IL-1RA, MIP-1α, GM-CSF and CRP compared to responders (complete/partial response). Higher levels of IL-6, IL-8, IL-27, and CRP were significantly associated with poor overall survival (OS), and IL-6, IL-8, MIP-1β, and CRP with poor progression-free survival (PFS) in male patients. Baseline metabolic tumor volume (MTV) and lactate dehydrogenase (LDH) levels were also significantly higher in male patients with poor OS and PFS. Finally, baseline IL-8 and CRP were significantly associated with baseline MTV and LDH levels in male patients. Conclusions: Our data show that pre-treatment proinflammatory proteins IL-8, IL-6, CRP are associated with poor response and survival in axi-cel-treated male but not female LBCL patients. The correlation of high levels of these factors with high pretreatment tumor burden in male non-responder R/R LBCL patients is suggestive of an immunosuppressive tumor microenvironment that is unlikely to support optimal expansion of anti-CD19 CAR T cells, resulting in poor response and survival outcomes. Citation Format: Manishkumar S. Patel, Akansha Jalota, Agrima Mian, Peter Bazeley, Sara A. Hunter, Brian T. Hill, Neetu Gupta. Gender bias in the association of pre-treatment cytokine signatures with response and survival in B cell lymphoma patients treated with anti-CD19 CAR T-cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1262.

Breast and endometrial cancer are both malignancies that can be classified into subtypes based on their hormone dependence, such as estrogen receptor (ER)-positive vs. ER-negative breast cancer and low-grade vs. high-grade endometrial cancer. The gene HSD3B1 encodes the steroid metabolism enzyme 3β-hydroxysteroid dehydrogenase 1, which catalyzes a critical step in conversion of adrenal precursor steroids to potent androgens and estrogens. Associations between the missense-encoding germline variant rs1047303 in HSD3B1 have been described to affect outcomes of castration-resistant prostate cancer and associated with ER positivity in postmenopausal breast cancer. The adrenal-permissive 1245C allele enables greater production of androgens and estrogens from adrenal precursors, whereas the adrenal-restrictive 1245A allele limits this reaction pathway. Using data from The Cancer Genome Atlas’s TCGA-UCEC and TCGA-BRCA projects, we interrogated whether the adrenal-restrictive rs1047303 genotype was associated with less hormone-driven genomic clusters of endometrial and breast cancer. We determined the genotype frequencies for different tumor subtypes, as classified by the genomic analysis of Sanchez-Vega, et al. (2018). The adrenal-restrictive HSD3B1(1245A) allele was associated with the copy number (CN) high cluster of endometrial cancer (OR 1.59, 95% CI 1.12-2.26, p = 0.0098), notably composed of serous and high-grade endometrioid endometrial cancers which are less estrogen-dependent than low-grade endometrioid tumors. Similarly, the adrenal-restrictive HSD3B1(1245A) was associated with the basal subtype of breast cancer (OR 1.54 [1.13-2.09], p = 0.0057), a subtype that is largely ER-negative and is known to share genomic features with serous-type endometrial cancers. Lastly, we interrogated overall survival of endometrial cancer patients in the UK Biobank and found that patients homozygous for the adrenal-restrictive HSD3B1(1245A) had worse overall survival (HR 1.39 [1.16-1.68], p = 0.00049) and cancer-specific survival (HR 1.39 [1.14-1.70], p = 0.0013), a finding consistent with the A allele being enriched in the more aggressive CN-high subtype of endometrial cancer. Taken together, these findings suggest a role for adrenal-restrictive vs. adrenal-permissive steroidogenesis, by way of rs1047303 genotype, in the development of and/or outcomes from at least three different commonly hormone-associated types of cancer: prostate, breast, and endometrial. Citation Format: Jeffrey M. McManus, Roberto Vargas, Peter Bazeley, Navin Sabharwal, Nima Sharifi. Association between adrenal-restrictive HSD3B1 inheritance and hormone independent subtypes of endometrial and breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 647.

Aims While most patients with myocardial infarction (MI) have underlying coronary atherosclerosis, not all patients with coronary artery disease (CAD) develop MI. We sought to address the hypothesis that some of the genetic factors which establish atherosclerosis may be distinct from those that predispose to vulnerable plaques and thrombus formation. Methods and results We carried out a genome-wide association study for MI in the UK Biobank (n∼472 000), followed by a meta-analysis with summary statistics from the CARDIoGRAMplusC4D Consortium (n∼167 000). Multiple independent replication analyses and functional approaches were used to prioritize loci and evaluate positional candidate genes. Eight novel regions were identified for MI at the genome wide significance level, of which effect sizes at six loci were more robust for MI than for CAD without the presence of MI. Confirmatory evidence for association of a locus on chromosome 1p21.3 harbouring choline-like transporter 3 (SLC44A3) with MI in the context of CAD, but not with coronary atherosclerosis itself, was obtained in Biobank Japan (n∼165 000) and 16 independent angiography-based cohorts (n∼27 000). Follow-up analyses did not reveal association of the SLC44A3 locus with CAD risk factors, biomarkers of coagulation, other thrombotic diseases, or plasma levels of a broad array of metabolites, including choline, trimethylamine N-oxide, and betaine. However, aortic expression of SLC44A3 was increased in carriers of the MI risk allele at chromosome 1p21.3, increased in ischaemic (vs. non-diseased) coronary arteries, up-regulated in human aortic endothelial cells treated with interleukin-1β (vs. vehicle), and associated with smooth muscle cell migration in vitro. Conclusions A large-scale analysis comprising ∼831 000 subjects revealed novel genetic determinants of MI and implicated SLC44A3 in the pathophysiology of vulnerable plaques.

Many autoimmune diseases, and in some instances, immune response to infections exhibit gender bias. However, it is not known if gender influences the response of lymphoma patients to anti-CD19 CAR T-cell therapy. To address this, we profiled 51 pre-treatment serum cytokines and chemokines to identify those that were differentially associated with response and survival in male vs. female relapsed/refractory large B-cell lymphoma (R/R LBCL) patients treated with Axicabtagene ciloleucel. Male non-responders (stable/progressive disease) had significantly higher baseline levels of IL-6, IL-8, IL-1RA, MIP-1α, GM-CSF and CRP compared to responders (complete/partial response). Higher levels of IL-6, IL-8, IL-27, and CRP were significantly associated with poor overall survival (OS), and IL-6, IL-8, MIP-1β, and CRP with poor progression-free survival (PFS) in male but not female patients. Baseline metabolic tumor volume (MTV) and lactate dehydrogenase (LDH) levels were also significantly higher in male patients with poor OS and PFS. Finally, baseline IL-8 and CRP were significantly correlated with baseline MTV and LDH levels in male but not female patients. Taken together, elevated baseline levels of IL-8, IL-6, CRP with high pretreatment tumor burden in male non-responder R/R LBCL patients point to an immunosuppressive tumor microenvironment that can potentially hinder anti-tumor activity prior to treatment and CAR T-cell expansion upon treatment in these patients, and result in poor outcomes.

Background: Epithelial Ovarian Cancer (EOC) is the second most common gynecologic malignancy in the United States, but the leading cause of gynecologic cancer death. Despite many achieving remission with first-line therapy, up to 80% of patients will recur and require additional treatment. Antibiotic therapy is frequently used during cancer treatments for both prophylaxis and treatment of infections, although this profoundly impacts the gut microbiome. Multiple studies suggest that an unperturbed gut microbiome may provide a protective microenvironment, and disruption may be permissive to tumor growth and chemotherapy resistance, including platinum agents. Experimental Design: We assessed whether antibiotic therapy would impact growth of EOC and sensitivity to cisplatin in murine models. Immune competent or compromised mice were given either metronidazole, ampicillin, vancomycin, and neomycin (ABX) containing or control water for two weeks before being intraperitoneally injected with murine ID8 or ID8-VEGF EOC cells. Tumors were monitored and cisplatin therapy was administered weekly until endpoint. Stool was collected throughout the study to asses for microbial population effects over time.Results: Both immune competent and immune compromised ID8 and ID8 VEGF tumor-bearing mice demonstrated a decreased response to cisplatin therapy in ABX treated groups with an increase in overall tumor burden. RNAseq analysis showed enrichment of multiple cell proliferation and stem cell pathways, and stem cell genes SOX2, WNT and PAX2. The self-renewal of ABX treated tumor cells was also increased.Conclusion: Collectively, these studies indicate an intact microbiome provides a tumor suppressive microenvironment and enhances sensitivity to cisplatin.

Objective Memory dysfunction is prevalent in many neurological disorders and can have a significant negative impact on quality of life. The genetic contributions to memory impairment in epilepsy, particularly temporal lobe epilepsy (TLE), remain poorly understood. Here, we compare the brain transcriptome between TLE patients with and without verbal memory impairments to identify genes and signaling networks important for episodic memory. Methods Brain tissues were resected from 23 adults who underwent dominant temporal lobectomy for treatment of pharmacoresistant epilepsy. To control for potential effects of APOE on memory, only those homozygous for the APOE e3 allele were included. A battery of memory tests was performed, and patients were stratified into two groups based on preoperative memory performance. The groups were well matched on demographic and disease-related variables. Total RNA-Seq and small RNA-Seq were performed on RNA extracted from the brain tissues. Pathway and integrative analyses were subsequently performed. Results We identified 1092 differentially expressed transcripts (DETs), with the majority (71%) being underexpressed in brain tissues from patients with impaired memory compared to those from patients with intact memory. Enrichment analysis revealed overrepresentation of genes in pathways pertaining to brain-related neurological dysfunction, including a subset associated with neurodegenerative diseases, memory, and cognition (APP, MAPT, PINK1). Despite including patients with identical APOE genotypes, we identify APOE as a differentially expressed gene associated with memory status. Small RNA-Seq identified four differentially expressed microRNAs (miRNAs) that were predicted to target a subset (22%) of all DETs. Integrative analysis showed that these miRNA-predicted DET targets impact brain-related pathways and biological processes also pertinent to memory and cognition. Significance TLE-associated memory status may be influenced by differences in gene expression profiles within the temporal lobe. Upstream processes influencing differential expression signatures, such as miRNAs, could serve as biomarkers and potential treatment targets for memory impairment in TLE.

Mitochondrial dysfunction has emerged to be associated with a broad spectrum of diseases, and there is an increasing demand for accurate detection of mitochondrial DNA (mtDNA) variants. Whole genome sequencing (WGS) has been the dominant sequencing approach to identify genetic variants in recent decades, but most studies focus on variants on the nuclear genome. Whole genome sequencing is also costly and time consuming. Sequencing specifically targeted for mtDNA is commonly used in the diagnostic settings and has lower costs. However, there is a lack of pairwise comparisons between these two sequencing approaches for calling mtDNA variants on a population basis. In this study, we compared WGS and mtDNA-targeted sequencing (targeted-seq) in analyzing mitochondrial DNA from 1499 participants recruited into the Severe Asthma Research Program (SARP). Our study reveals that targeted-sequencing and WGS have comparable capacity to determine genotypes and to call haplogroups and homoplasmies on mtDNA. However, there exists a large variability in calling heteroplasmies, especially for low-frequency heteroplasmies, which indicates that investigators should be cautious about heteroplasmies acquired from different sequencing methods. Further research is highly desired to improve variant detection methods for mitochondrial DNA.

Background: Epithelial Ovarian Cancer (EOC) is the second most common gynecologic malignancy in the United States, but the leading cause of gynecologic cancer death. Despite many achieving remission with first-line therapy, up to 80% of patients will recur and require additional treatment. Antibiotic therapy is frequently used during cancer treatments for both prophylaxis and treatment of infections, although this profoundly impacts the gut microbiome. Multiple studies suggest that an unperturbed gut microbiome may provide a protective microenvironment, and disruption may be permissive to tumor growth and chemotherapy resistance, including platinum agents. Experimental Design: We assessed whether antibiotic therapy would impact growth of EOC and sensitivity to cisplatin in murine models. Immune competent or compromised mice were given control or antibiotic (ABX) containing water (metronidazole, ampicillin, vancomycin, and neomycin) for two weeks before being intraperitoneally injected with murine ID8 or ID8-VEGF EOC cells. Tumors were monitored and cisplatin therapy was administered weekly until endpoint. Stool was collected throughout the study to assess for microbial population effects over time. Cecal microbiota transplants (CMTs) of contents derived from ABX or control treated donor mice were performed on ABX treated recipient mice. Results: Both immune competent and immune compromised ID8 and ID8 VEGF tumor-bearing mice demonstrated a decreased response to cisplatin therapy in ABX treated groups with an increase in overall tumor burden. RNAseq analysis showed enrichment of multiple cell proliferation and stem cell pathways, and stem cell genes SOX2, WNT and PAX2. The self-renewal of cancer cells derived from tumors of ABX treated mice was increased compared to tumors from control mice, indicative of a microbiota derived tumor suppressor. Conclusion: Collectively, these studies indicate an intact microbiome provides a tumor suppressor and maintains chemosensitivity that is disrupted by ABX treatment.

IMPORTANCE: The role of Tumor Mutation Burden (TMB) as a prognostic and/or predictive biomarker for Immune Checkpoint Blockade (ICB) therapy in a real-world clinical setting is still unclear. OBJECTIVE: To assess whether TMB status provided by a clinically and commercially available tumor genomic profiling (TGP) assay is associated with overall survival of Non-Small Cell Lung Cancer (NSCLC) patients treated with ICB from a single institute. DESIGN, SETTING, AND PARTICIPANTS: Outcomes and genetic testing data were collected for 188 NSCLC patients treated within the Cleveland Clinic system between August 2012 and July 2017. MAIN OUTCOMES AND MEASURES: Overall survival (OS) from time receiving ICB therapy. RESULTS: Among 188 patients with NSCLC (median age, 62 years; 49.5% female), 86 (45.7%) received ICB therapy. Patients were grouped into three categories based on the status of TMB (in mutations/Mb): high (>= 20/Mb), intermediate (>=5 to 0.8], median OS difference: 9.7 months). CONCLUSIONS AND RELEVANCE: Among patients with NSCLC from a single institute with a longitudinal database of clinical data including TGP results, exploratory analyses do not show statistical significance for the prognostic utility of TMB. These findings indicate that there is a need for more prospective data on the use of TMB status as a guide for ICB therapy in a routine care setting.

IMPORTANCEThe role of Tumor Mutation Burden (TMB) as a prognostic and/or predictive biomarker for Immune Checkpoint Blockade (ICB) therapy in a real-world clinical setting is still unclear.OBJECTIVETo assess whether TMB status provided by a clinically and commercially available tumor genomic profiling (TGP) assay is associated with overall survival of Non-Small Cell Lung Cancer (NSCLC) patients treated with ICB from a single institute.DESIGN, SETTING, AND PARTICIPANTSOutcomes and genetic testing data were collected for 188 NSCLC patients treated within the Cleveland Clinic system between August 2012 and July 2017.MAIN OUTCOMES AND MEASURESOverall survival (OS) from time receiving ICB therapy.RESULTSAmong 188 patients with NSCLC (median age, 62 years; 49.5% female), 86 (45.7%) received ICB therapy. Patients were grouped into three categories based on the status of TMB (in mutations/Mb): high (>= 20/Mb), intermediate (>=5 to 0.8], median OS difference: 9.7 months).CONCLUSIONS AND RELEVANCEAmong patients with NSCLC from a single institute in a longitudinal database of clinical data including TGP results, exploratory analyses do not show statistical significance for the prognostic utility of TMB. These findings indicate that there is a need for more prospective data on the use of TMB status as a guide for ICB therapy in a routine care setting.

10503 Background: Genetic factors that contribute to endogenous estrogen synthesis and postmenopausal breast cancer risk are unknown. We set out to test the hypothesis that homozygous inheritance of the common 1245A→C missense-encoding polymorphism in HSD3B1, which is common (8-10%) in White populations, functionally adrenal permissive and increases synthesis of the aromatase substrate, androstenedione, is associated with postmenopausal estrogen receptor-positive breast cancer. Methods: A prospective single institution study of postmenopausal estrogen receptor-driven breast cancer for determination of HSD3B1 genotype, circulating steroid concentrations, and adrenal-permissive genotype frequency compared with the genotype frequency in the general population and in estrogen receptor-negative breast cancer. Validation was performed in 2 breast cancer genomic studies with estrogen receptor documentation. The primary outcome is the adrenal-permissive genotype frequency in postmenopausal estrogen receptor-positive breast cancer and the general population. Genotype comparisons were also done with postmenopausal estrogen receptor-negative breast cancer and the association with circulating adrenal androgen concentrations determined. Results: The prospective and validation studies had 199 and 1628 women, respectively. The adrenal-permissive genotype frequency in postmenopausal White women with estrogen-driven breast cancer in the prospective cohort was 17.5% (21/120) compared with 9.6% (429/4451) in the general population [p = 0.0077]. The adrenal-permissive genotype frequency for estrogen-driven postmenopausal breast cancer was validated using the Cambridge and TCGA genomic datasets together: 14.4% (56/389) compared with 6.0% (9/149) for estrogen receptor-negative breast cancer (p = 0.007) and the general population (p = 0.005). Circulating androstenedione concentration was significantly higher for women with the adrenal-permissive genotype compared with the other genotypes (p = 0.03). Conclusions: The adrenal-permissive genotype is associated with estrogen-driven postmenopausal breast cancer. These findings link genetic inheritance of endogenous estrogen exposure to estrogen-driven breast cancer and have broad implications for risk stratification, prevention, potential biomarkers for hormonal therapy response and possibly other clinical outcomes related to estrogen physiology in postmenopausal women.

1507 Background: CHIP predisposes to a higher risk of developing hematological malignancies and cardiac events. Multiple germline mutations have been recognized as contributing to CHIP, most notably ASXL1, DNMT3A, and TET2. The frequency of CHIP mutations in solid tumor malignancies (STM) is unknown. We report the frequency and incidence of CHIP mutations in adult patients (pts) with STM. Methods: Data from 880 pts with STM who underwent next generation sequencing (NGS) at Foundation One from 2013-2017 was collected. This excluded two pts with known primary hematological malignancies who were removed. A list of CHIP mutations using NCCN guidelines as well as numerous original research articles was created, and tabulation of pathogenic or likely pathogenic mutations ( ASXL1, DNMT3A, TET2, JAK2, SF3B1, TP53, GNAS, N/KRAS) was performed. To date, only the lung, breast, and colorectal cancer pts have been annotated (N = 372) since these cancers have an overall higher incidence and prevalence in society. Results: Annotation of lung cancer pts (155/880), breast (118/880), and colorectal cancer pts (99/880) is collected and represents about 40% of all pts. At least one CHIP mutation was present in 44.5% (69/155) lung cancer pts, 32.2% (38/118) in breast cancer pts, and 7.1% (7/99) in colorectal cancer pts. Most common mutations found were TP53 and KRAS at 29.6% (110/372) and 28.0% (104/372), respectively. Mutations in genes not known to be somatic drivers for solid tumor malignancies, particularly SF3B1, DNMT3A, and JAK2, were found at very low frequencies 0.8% (3/372), 0.5% (2/372), and 0.3% (1/372), respectively. Notably, ASXL1 and TET2 mutations were not encountered in any pts. Conclusions: In tumor NGS testing, multiple CHIP mutations were noted to be present within the cohort of lung, breast, and colorectal cancers. There is a need to further understand clinical consequences of CHIP mutations incidentally found in pts with STM given known clinical implications of CHIP. We will report on clinical data (comorbidities), response/non-response to therapy, and identify specific molecular patterns of mutations to further understand the role of CHIP in pts with STM.

Magnetic resonance imaging (MRI) was used to evaluate within-subject variability in global mean cortical thickness over test-retest intervals of minutes-weeks in five healthy adults. Within-subject measures of global mean thickness were consistent over these intervals. Test-retest assessments of absolute thickness differences and percent thickness differences indicated variations of, respectively, < or =0.05-0.06 mm and < or =+/-1.9-2.3%. There have been few evaluations of normal within-subject variations in cortical thickness. The present results suggest that within-subject variability in global mean cortical thickness can be low over test-retest intervals of minutes-weeks, and that longitudinal scans can establish useful baseline estimates of variability from which to assess changes due to injury, disease, or other experiences.

Due to the small number of replicates in typical gene microarray experiments, the performance of statistical inference is often unsatisfactory. In this article, we present a scoring scheme, based on quantiles, that allows researchers to combine data from different platforms. We have applied the discrete-continuous normal distribution (DISCO) using the quantile scores on two publicly available data sets. Differentially expressed genes identified by DISCO are comparable to those identified by significance analysis of microarray (SAM) or Wilcoxon rank test. An algorithm based on DISCO and quantile scores is developed to combine results from Affymetrix and Illumina. Our results indicate that combining microarray data from different platforms is possible and straightforward.AVAILABILITY: R code implementing our methods is available from the authors.