TGF-ß-mediated silencing of genomic organizer SATB1 promotes Tfh cell differentiation and formation of intra-tumoral tertiary lymphoid structures.

The immune checkpoint receptor PD-1 on T follicular helper (Tfh) cells promotes Tfh:B cell interactions and appropriate positioning within tissues. Here, we examined the impact of regulation of PD-1 expression by the genomic organizer SATB1 on Tfh cell differentiation. Vaccination of CD4CreSatb1f/f mice enriched for antigen-specific Tfh cells, and TGF-ß-mediated repression of SATB1 enhanced Tfh differentiation of human T cells. Mechanistically, high Icos expression in Satb1-/- CD4+ T cells promoted Tfh cell differentiation by preventing T follicular regulatory cell skewing and resulted in increased isotype-switched B cell responses in vivo. Ovarian tumors in CD4CreSatb1f/f mice accumulated tumor antigen-specific, LIGHT+CXCL13+IL-21+ Tfh cells and tertiary lymphoid structures (TLS). TLS formation decreased tumor growth in a CD4+ T cell and CXCL13-dependent manner. The transfer of Tfh cells, but not naive CD4+ T cells, induced TLS at tumor beds and decreased tumor growth. Thus, TGF-ß-mediated silencing of Satb1 licenses Tfh cell differentiation, providing insight into the genesis of TLS within tumors.

The Unfolded Protein Response Mediator PERK Governs Myeloid Cell-Driven Immunosuppression in Tumors through Inhibition of STING Signaling.

The primary mechanisms supporting immunoregulatory polarization of myeloid cells upon infiltration into tumors remain largely unexplored. Elucidation of these signals could enable better strategies to restore protective anti-tumor immunity. Here, we investigated the role of the intrinsic activation of the PKR-like endoplasmic reticulum (ER) kinase (PERK) in the immunoinhibitory actions of tumor-associated myeloid-derived suppressor cells (tumor-MDSCs). PERK signaling increased in tumor-MDSCs, and its deletion transformed MDSCs into myeloid cells that activated CD8+ T cell-mediated immunity against cancer. Tumor-MDSCs lacking PERK exhibited disrupted NRF2-driven antioxidant capacity and impaired mitochondrial respiratory homeostasis. Moreover, reduced NRF2 signaling in PERK-deficient MDSCs elicited cytosolic mitochondrial DNA elevation and, consequently, STING-dependent expression of anti-tumor type I interferon. Reactivation of NRF2 signaling, conditional deletion of STING, or blockade of type I interferon receptor I restored the immunoinhibitory potential of PERK-ablated MDSCs. Our findings demonstrate the pivotal role of PERK in tumor-MDSC functionality and unveil strategies to reprogram immunosuppressive myelopoiesis in tumors to boost cancer immunotherapy.

Non-histone protein methylation as a regulator of cellular signalling and function.

Methylation of Lys and Arg residues on non-histone proteins has emerged as a prevalent post-translational modification and as an important regulator of cellular signal transduction mediated by the MAPK, WNT, BMP, Hippo and JAK-STAT signalling pathways. Crosstalk between methylation and other types of post-translational modifications, and between histone and non-histone protein methylation frequently occurs and affects cellular functions such as chromatin remodelling, gene transcription, protein synthesis, signal transduction and DNA repair. With recent advances in proteomic techniques, in particular mass spectrometry, the stage is now set to decode the methylproteome and define its functions in health and disease.

IgA transcytosis and antigen recognition govern ovarian cancer immunity.

Most ovarian cancers are infiltrated by prognostically relevant activated T cells1-3, yet exhibit low response rates to immune checkpoint inhibitors4. Memory B cell and plasma cell infiltrates have previously been associated with better outcomes in ovarian cancer5,6, but the nature and functional relevance of these responses are controversial. Here, using 3 independent cohorts that in total comprise 534 patients with high-grade serous ovarian cancer, we show that robust, protective humoral responses are dominated by the production of polyclonal IgA, which binds to polymeric IgA receptors that are universally expressed on ovarian cancer cells. Notably, tumour B-cell-derived IgA redirects myeloid cells against extracellular oncogenic drivers, which causes tumour cell death. In addition, IgA transcytosis through malignant epithelial cells elicits transcriptional changes that antagonize the RAS pathway and sensitize tumour cells to cytolytic killing by T cells, which also contributes to hindering malignant progression. Thus, tumour-antigen-specific and -antigen-independent IgA responses antagonize the growth of ovarian cancer by governing coordinated tumour cell, T cell and B cell responses. These findings provide a platform for identifying targets that are spontaneously recognized by intratumoural B-cell-derived antibodies, and suggest that immunotherapies that augment B cell responses may be more effective than approaches that focus on T cells, particularly for malignancies that are resistant to checkpoint inhibitors.

SATB1 Expression Governs Epigenetic Repression of PD-1 in Tumor-Reactive T Cells.

Despite the importance of programmed cell death-1 (PD-1) in inhibiting T cell effector activity, the mechanisms regulating its expression remain poorly defined. We found that the chromatin organizer special AT-rich sequence-binding protein-1 (Satb1) restrains PD-1 expression induced upon T cell activation by recruiting a nucleosome remodeling deacetylase (NuRD) complex to Pdcd1 regulatory regions. Satb1 deficienct T cells exhibited a 40-fold increase in PD-1 expression. Tumor-derived transforming growth factor ß (Tgf-ß) decreased Satb1 expression through binding of Smad proteins to the Satb1 promoter. Smad proteins also competed with the Satb1-NuRD complex for binding to Pdcd1 enhancers, releasing Pdcd1 expression from Satb1-mediated repression, Satb1-deficient tumor-reactive T cells lost effector activity more rapidly than wild-type lymphocytes at tumor beds expressing PD-1 ligand (CD274), and these differences were abrogated by sustained CD274 blockade. Our findings suggest that Satb1 functions to prevent premature T cell exhaustion by regulating Pdcd1 expression upon T cell activation. Dysregulation of this pathway in tumor-infiltrating T cells results in diminished anti-tumor immunity.

SnapShot: Lysine Methylation beyond Histones.

Lysine methylation is a prevalent post-translational modification (PTM) used by the cell to reversibly regulate protein function. Although it has been extensively studied in the context of histones and the associated chromatin, the remaining methyllysine proteome remains largely unexplored. This SnapShot provides an overview of the current state of lysine methylation research and its emergence as a dynamic PTM occurring on histone and non-histone proteins.

Discovery and Characterization of Pyridoxal 5'-Phosphate-Dependent Cycloleucine Synthases.

Pyridoxal 5'-phosphate (PLP)-dependent enzymes are the most versatile biocatalysts for synthesizing nonproteinogenic amino acids. α,α-Disubstituted quaternary amino acids, such as 1-aminocyclopentane-1-carboxylic acid (cycloleucine), are useful building blocks for pharmaceuticals. In this study, starting with the biosynthesis of fusarilin A, we discovered a family of PLP-dependent enzymes that can facilitate tandem carbon-carbon forming steps to catalyze an overall [3 + 2]-annulation. In the first step, the cycloleucine synthases use SAM as the latent electrophile and an in situ-generated enamine as the nucleophile for γ-substitution. Whereas previously characterized γ-replacement enzymes protonate the resulting α-carbon and release the acyclic amino acid, cycloleucine synthases can catalyze an additional, intramolecular aldol or Mannich reaction with the nucleophilic α-carbon to form the substituted cyclopentane. Overall, the net [3 + 2]-annulation reaction can lead to 2-hydroxy or 2-aminocycloleucine products. These studies further expand the biocatalytic scope of PLP-dependent enzymes.

IRE1α-XBP1 controls T cell function in ovarian cancer by regulating mitochondrial activity.

Tumours evade immune control by creating hostile microenvironments that perturb T cell metabolism and effector function1-4. However, it remains unclear how intra-tumoral T cells integrate and interpret metabolic stress signals. Here we report that ovarian cancer-an aggressive malignancy that is refractory to standard treatments and current immunotherapies5-8-induces endoplasmic reticulum stress and activates the IRE1α-XBP1 arm of the unfolded protein response9,10 in T cells to control their mitochondrial respiration and anti-tumour function. In T cells isolated from specimens collected from patients with ovarian cancer, upregulation of XBP1 was associated with decreased infiltration of T cells into tumours and with reduced IFNG mRNA expression. Malignant ascites fluid obtained from patients with ovarian cancer inhibited glucose uptake and caused N-linked protein glycosylation defects in T cells, which triggered IRE1α-XBP1 activation that suppressed mitochondrial activity and IFNγ production. Mechanistically, induction of XBP1 regulated the abundance of glutamine carriers and thus limited the influx of glutamine that is necessary to sustain mitochondrial respiration in T cells under glucose-deprived conditions. Restoring N-linked protein glycosylation, abrogating IRE1α-XBP1 activation or enforcing expression of glutamine transporters enhanced mitochondrial respiration in human T cells exposed to ovarian cancer ascites. XBP1-deficient T cells in the metastatic ovarian cancer milieu exhibited global transcriptional reprogramming and improved effector capacity. Accordingly, mice that bear ovarian cancer and lack XBP1 selectively in T cells demonstrate superior anti-tumour immunity, delayed malignant progression and increased overall survival. Controlling endoplasmic reticulum stress or targeting IRE1α-XBP1 signalling may help to restore the metabolic fitness and anti-tumour capacity of T cells in cancer hosts.

Author Correction: Discovery of stimulation-responsive immune enhancers with CRISPR activation.

In this Letter, analysis of steady-state regulatory T (Treg) cell percentages from Il2ra enhancer deletion (EDEL) and wild-type (WT) mice revealed no differences between them (Extended Data Fig. 9d). This analysis included two mice whose genotypes were incorrectly assigned. Even after correction of the genotypes, no significant differences in Treg cell percentages were seen when data across experimental cohorts were averaged (as was done in Extended Data Fig. 9d). However, if we normalize the corrected data to account for variation among experimental cohorts, a subtle decrease in EDEL Treg cell percentages is revealed and, using the corrected and normalized data, we have redrawn Extended Data Fig. 9d in Supplementary Fig. 1. The Supplementary Information to this Amendment contains the corrected and reanalysed Extended Data Fig. 9d. The sentence "This enhancer deletion (EDEL) strain also had no obvious T cell phenotypes at steady state (Extended Data Fig. 9)." should read: "This enhancer deletion (EDEL) strain had a small decrease in the percentage of Treg cells (Extended Data Fig. 9).". This error does not affect any of the main figures in the Letter or the data from mice with the human autoimmune-associated single nucleotide polymorphism (SNP) knocked in or with a 12-base-pair deletion at the site (12DEL). In addition, we stated in the Methods that we observed consistent immunophenotypes of EDEL mice across three founders, but in fact, we observed consistent phenotypes in mice from two founders. This does not change any of our conclusions and the original Letter has not been corrected.

Cellular stress responses and metabolic reprogramming in cancer progression and dormancy.

Recurrent disease after prolonged cancer dormancy is a major cause of cancer associated mortality, yet many of the mechanisms that are engaged to initiate dormancy as well as later recurrence remain incompletely understood. It is known that cancer cells initiate adaptation mechanisms to adapt tightly regulated cellular processes to non-optimal growth environments; Recent investigations have begun to elucidate the contribution of these mechanisms to malignant progression, with intriguing studies now defining cellular stress as a key contributor to the development and maintenance of cancer dormancy. This review will focus on our current understanding of stress responses facilitating malignant cell adaptation and metabolic reprogramming to establish cancer dormancy.

The CDK7 inhibitor CT7001 (Samuraciclib) targets proliferation pathways to inhibit advanced prostate cancer.

BACKGROUND: Current strategies to inhibit androgen receptor (AR) are circumvented in castration-resistant prostate cancer (CRPC). Cyclin-dependent kinase 7 (CDK7) promotes AR signalling, in addition to established roles in cell cycle and global transcription, providing a rationale for its therapeutic targeting in CRPC. METHODS: The antitumour activity of CT7001, an orally bioavailable CDK7 inhibitor, was investigated across CRPC models in vitro and in xenograft models in vivo. Cell-based assays and transcriptomic analyses of treated xenografts were employed to investigate the mechanisms driving CT7001 activity, alone and in combination with the antiandrogen enzalutamide. RESULTS: CT7001 selectively engages with CDK7 in prostate cancer cells, causing inhibition of proliferation and cell cycle arrest. Activation of p53, induction of apoptosis, and suppression of transcription mediated by full-length and constitutively active AR splice variants contribute to antitumour efficacy in vitro. Oral administration of CT7001 represses growth of CRPC xenografts and significantly augments growth inhibition achieved by enzalutamide. Transcriptome analyses of treated xenografts indicate cell cycle and AR inhibition as the mode of action of CT7001 in vivo. CONCLUSIONS: This study supports CDK7 inhibition as a strategy to target deregulated cell proliferation and demonstrates CT7001 is a promising CRPC therapeutic, alone or in combination with AR-targeting compounds.

Discovery of stimulation-responsive immune enhancers with CRISPR activation.

The majority of genetic variants associated with common human diseases map to enhancers, non-coding elements that shape cell-type-specific transcriptional programs and responses to extracellular cues. Systematic mapping of functional enhancers and their biological contexts is required to understand the mechanisms by which variation in non-coding genetic sequences contributes to disease. Functional enhancers can be mapped by genomic sequence disruption, but this approach is limited to the subset of enhancers that are necessary in the particular cellular context being studied. We hypothesized that recruitment of a strong transcriptional activator to an enhancer would be sufficient to drive target gene expression, even if that enhancer was not currently active in the assayed cells. Here we describe a discovery platform that can identify stimulus-responsive enhancers for a target gene independent of stimulus exposure. We used tiled CRISPR activation (CRISPRa) to synthetically recruit a transcriptional activator to sites across large genomic regions (more than 100 kilobases) surrounding two key autoimmunity risk loci, CD69 and IL2RA. We identified several CRISPRa-responsive elements with chromatin features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours an autoimmunity risk variant. Using engineered mouse models, we found that sequence perturbation of the disease-associated Il2ra enhancer did not entirely block Il2ra expression, but rather delayed the timing of gene activation in response to specific extracellular signals. Enhancer deletion skewed polarization of naive T cells towards a pro-inflammatory T helper (TH17) cell state and away from a regulatory T cell state. This integrated approach identifies functional enhancers and reveals how non-coding variation associated with human immune dysfunction alters context-specific gene programs.

Unmet need for gender-affirming care as a social determinant of mental health inequities for transgender youth in Aotearoa/New Zealand.

BACKGROUND: Past studies have demonstrated better mental health and well-being among transgender youth who had accessed gender-affirming care. However, few existing studies have assessed unmet need for gender-affirming care as a social determinant of mental health inequities. METHODS: Data on unmet need for gender-affirming care, distress and suicidality were analysed from the 2018 Counting Ourselves nationwide community-based survey of transgender people in Aotearoa/New Zealand. Associations between unmet need for gender-affirming care and mental health indicators were tested for transgender youth within the sample (aged 14-26 years; n = 608; Mage = 20.5). RESULTS: Transgender youth reported unmet needs ranging from 42% for gender-affirming hormone to 100% for feminizing surgeries and voice surgeries. Overall unmet need for gender-affirming care was associated with worse mental health. Trans men with an unmet need for chest reconstruction (84%) scored an average of 7.13 points higher on the K10 Psychological Distress Scale relative to those whose need had been met. Participants reporting unmet need for hormones (42%) had twice the odds (adjusted odds ratios = 2.01; CI = 1.02-3.98) of having attempted suicide in the last 12 months. CONCLUSIONS: Dismantling barriers to accessing gender-affirming care could play a crucial role in reducing mental health inequities faced by transgender youth.

Higher Computed Tomography (CT) Scan Resolution Improves Accuracy of Patient-specific Mandibular Models When Compared to Cadaveric Gold Standard.

BACKGROUND: 3D-printed patient-specific anatomical models are becoming an increasingly popular tool for planning reconstructive surgeries to treat oral cancer. Currently there is a lack of information regarding model accuracy, and how the resolution of the computed tomography (CT) scan affects the accuracy of the final model. PURPOSE: The primary objective of this study was to determine the CT z-axis resolution necessary in creating a patient specific mandibular model with clinically acceptable accuracy for global bony reconstruction. This study also sought to evaluate the effect of the digital sculpting and 3D printing process on model accuracy. STUDY DESIGN: This was a cross-sectional study using cadaveric heads obtained from the Ohio State University Body Donation Program. INDEPENDENT VARIABLES: The first independent variable is CT scan slice thickness of either 0.675 , 1.25, 3.00, or 5.00 mm. The second independent variable is the three produced models for analysis (unsculpted, digitally sculpted, 3D printed). MAIN OUTCOME VARIABLE: The degree of accuracy of a model as defined by the root mean square (RMS) value, a measure of a model's discrepancy from its respective cadaveric anatomy. ANALYSES: All models were digitally compared to their cadaveric bony anatomy using a metrology surface scan of the dissected mandible. The RMS value of each comparison evaluates the level of discrepancy. One-way ANOVA tests (P < .05) were used to determine statistically significant differences between CT scan resolutions. Two-way ANOVA tests (P < .05) were used to determine statistically significant differences between groups. RESULTS: CT scans acquired for 8 formalin-fixed cadaver heads were processed and analyzed. The RMS for digitally sculpted models decreased as slice thickness decreased, confirming that higher resolution CT scans resulted in statistically more accurate model production when compared to the cadaveric gold standard. Furthermore, digitally sculpted models were significantly more accurate than unsculpted models (P < .05) at each slice thickness. CONCLUSIONS: Our study demonstrated that CT scans with slice thicknesses of 3.00 mm or smaller created statistically significantly more accurate models than models created from slice thicknesses of 5.00 mm. The digital sculpting process statistically significantly increased the accuracy of models and no loss of accuracy through the 3D printing process was observed.

Hostile home environment predicting early adolescent sexual harassment perpetration and potential school-related moderators.

Using family systems theory, this longitudinal study of middle school youth examined the effects of abuse, family conflict, and sibling aggression on sexual harassment perpetration (N = 1563; Mage 11.2, 51% boys; 39% Hispanic, 29% Black, and 19% White). Boys reported more sexual harassment than girls; perpetration increased for both. The association between a hostile home environment and sexual harassment perpetration was moderated by school experiences. School belonging buffered effects of hostile home environment on baseline sexual harassment perpetration for boys who experienced abuse and White adolescents with high sibling aggression. Academic grades moderated change in perpetration over time, but effects differed by sex and race. It is important to understand how early violence exposures relate to sexual violence perpetration during early adolescence.

Exposed Intramedullary Fixation Produces Similar Outcomes to Buried Fixation for Acute Pediatric Monteggia Fractures.

BACKGROUND: Treatment of acute pediatric Monteggia fractures requires ulnar length stability to maintain reduction of the radiocapitellar joint. When operative care is indicated, intramedullary ulna fixation can be buried or left temporarily exposed through the skin while under a cast. The authors hypothesized that treatment with exposed fixation yields equivalent results to buried fixation for Monteggia fractures while avoiding secondary surgery for hardware removal. METHODS: A retrospective review of children with acute Monteggia fractures at our Level 1 pediatric trauma center was performed. Patient charts and radiographs were evaluated for age, fracture type, fracture location, Bado classification, type of treatment, complications, cast duration, time to fracture union, time to hardware removal, and range of motion. RESULTS: Out of 59 acute Monteggia fractures surgically treated (average age 6 y, range 2 to 14), 15 (25%) patients were fixed with buried intramedullary fixation and 44 (75%) with exposed intramedullary fixation under a cast. There were no significant differences between buried and exposed intramedullary fixation in cast time after surgery (39 vs. 37 d; P =0.55), time to fracture union (37 vs. 35 d; P =0.67), pronation/supination (137 vs. 134 degrees; P =0.68) or flexion/extension (115 vs. 114 degrees; P =0.81) range of motion. The exposed fixation had a return to OR of 4.5% (2 out of 44), and the buried fixation returned to the OR for removal on all patients. CONCLUSION: Exposed intramedullary fixation yielded equivalent clinical outcomes to buried devices in the treatment of acute pediatric Monteggia fractures while eliminating the need for a second surgery to remove hardware, reducing the associated risks and costs of surgery and anesthesia, but had a higher complication rate. Open Monteggia fractures or patterns with a known risk of delayed union may benefit from buried instead of exposed intramedullary fixation for earlier mobilization. LEVEL OF EVIDENCE: III.

Uptake, experiences and barriers to cervical screening for trans and non-binary people in Aotearoa New Zealand.

Little is known about experiences and barriers for trans and non-binary (TGNB) people eligible for cervical screening in Aotearoa New Zealand. AIMS: To identify uptake, barriers and reasons for delaying cervical cancer screening among TGNB people in Aotearoa. MATERIALS AND METHODS: The 2018 Counting Ourselves data on TGNB people assigned female at birth aged 20-69 years who had ever had sex, were analysed to report on experiences of those who were eligible for cervical screening (n = 318). Participants answered questions about whether they had taken part in cervical screening and reasons behind any delays in receiving the test. RESULTS: Trans men were more likely than non-binary participants to report that they did not require cervical screening or were unsure if they needed it. For those who had delayed cervical screening, 30% did so due to feeling worried about how they would be treated as a trans or non-binary person and 35% due to another reason. Other reasons for delay related to general and gender-related discomfort, previous traumatic experiences, anxiety or fear of the test and pain. Material barriers to access included cost and lack of information. CONCLUSIONS: The current cervical screening program in Aotearoa does not consider the needs of TGNB people, leading to delayed and reduced uptake of cervical screening. Health providers require education on the reasons TGNB people delay or avoid cervical screening in order to provide appropriate information and affirmative healthcare environments. The human papillomavirus self-swab may address some of the existing barriers.

Butyrophilins: Dynamic Regulators of Protective T Cell Immunity in Cancer.

The efficacy of current immunotherapies remains limited in many solid epithelial malignancies. Recent investigations into the biology of butyrophilin (BTN) and butyrophilin-like (BTNL) molecules, however, suggest these molecules are potent immunosuppressors of antigen-specific protective T cell activity in tumor beds. BTN and BTNL molecules also associate with each other dynamically on cellular surfaces in specific contexts, which modulates their biology. At least in the case of BTN3A1, this dynamism drives the immunosuppression of αß T cells or the activation of Vγ9Vδ2 T cells. Clearly, there is much to learn regarding the biology of BTN and BTNL molecules in the context of cancer, where they may represent intriguing immunotherapeutic targets that could potentially synergize with the current class of immune modulators in cancer. Here, we discuss our current understanding of BTN and BTNL biology, with a particular focus on BTN3A1, and potential therapeutic implications for cancer.

Favorable outcomes of posterior cruciate-stabilized inserts over ultracongruent inserts in total knee arthroplasty.

OBJECTIVE: Posterior cruciate-stabilized (PS) and ultracongruent (UC) inserts are used during total knee arthroplasty (TKA), but superiority in gaining postoperative knee flexion and functionality remains contested. Therefore, this study compared postoperative outcomes between PS and UC inserts. METHODS: A retrospective review evaluated unilateral or bilateral TKAs with PS or UC inserts from August 2011 to March 2020. Nonparametric statistics were performed to evaluate differences in patient demographics, pre- and postknee flexion and Knee Society Knee (KSS-K) and Function Score (KSS-F). Univariate and multivariable regressions were performed to evaluate the influence on postoperative knee flexion ≥ 120°, presented as odds ratios (OR) and 95% confidence intervals (CI). RESULTS: Patient demographics were not significantly different between the 577 PS and 399 UC knees evaluated. Postoperatively, a larger proportion of UC knees demonstrated knee flexion < 120° (36.0% vs. 18.6%, p < 0.001) and lower KSS-K (91.0 ± 8.7 vs. 91.6 ± 10.3, p < 0.001) and KSS-F (76.8 ± 21.6 vs. 79.9 ± 21.6, p = 0.007) than the PS group. The PS group had greater improvement in flexion angle (4.9° ± 14.9° vs. 1.0° ± 15.6°, p < 0.001) and KSS-F (27.3 ± 23.3 vs. 23.1 ± 25.3, p = 0.007) as compared to that of UC patients. Patients with preoperative flexion < 120° (OR 2.787, CI 2.066-3.761; p < 0.001), higher body mass index (OR 1.033, CI 1.006-1.061; p = 0.017) and UC insert (OR 2.461, CI 1.832-3.307; p < 0.001) were less likely to achieve flexion ≥ 120°. CONCLUSION: Favorable clinical and functional outcomes were noted in the PS group as compared to UC inserts in TKA. The greater improvement in overall knee flexion may suggest the PS insert may be especially appropriate for patients with lower preoperative range of motion. LEVEL OF EVIDENCE: III, retrospective comparative study.

Impact of returning unsolicited genomic results to nongenetic health care providers in the eMERGE III Network.

PURPOSE: As genomic sequencing becomes more common, medically actionable secondary findings will increasingly be returned to health care providers (HCPs), who will be faced with managing the resulting patient care. These findings are generally unsolicited, ie, unrelated to the sequencing indication and/or ordered by another clinician. METHODS: To understand the impact of receiving unsolicited results, we interviewed HCPs who received genomic results for patients enrolled in the Electronic Medical Records and Genomics (eMERGE) Phase III Network, which returned results on >100 actionable genes to eMERGE participants and HCPs. RESULTS: In total, 16 HCPs across 3 eMERGE sites were interviewed about their experience of receiving a positive (likely pathogenic or pathogenic), negative, or variant of uncertain significance result for a patient enrolled in eMERGE Phase III and about managing their patient on the basis of the result. Although unsolicited, HCPs felt responsible for managing the patient's resulting medical care. HCPs indicated that clinical utility depended on the actionability of results, and whereas comfort levels varied, confidence was improved by the availability of subspecialist consults. HCPs were concerned about patient anxiety, insurability, and missing an actionable result in the electronic health record. CONCLUSION: Our findings help inform best practices for return of unsolicited genomic screening findings in the future.