Citations

Background Trans-acting programmed death-ligand 1 (PD-L1) derives from malignant cells in three known forms. High levels of secreted splice variant PD-L1 (sPD-L1), ADAM10/ADAM17-shed sPD-L1, and PD-L1-positive extracellular vesicles (evPD-L1) each predict poor prognosis and limited response to PD-(L)1 checkpoint inhibitors in cancer. To our knowledge, no clinical intervention has reduced any of these circulating forms of extracellular PD-L1. Here, we explore therapeutic plasma exchange (TPE) as a treatment to reduce circulating extracellular PD-L1. Results In patients with melanoma, sPD-L1 levels above 0.277 ng/mL predicted inferior overall survival. In patients undergoing TPE for non-malignant indications, each TPE session removed a mean 70.8% sPD-L1 and 73.1% evPD-L1 detectable in plasma. TPE also reduced total and ADAM10-positive extracellular vesicles. Conclusion Here, we report the first known clinical intervention to remove either sPD-L1 or evPD-L1 from plasma in vivo. TPE reduces plasma sPD-L1 and evPD-L1 in vivo and may have a role in treatment with immunotherapy. TPE may also prove useful in patients with other extracellular vesicle-related conditions.

Abstract Background: Sex differences in non-small cell lung cancer (NSCLC) outcomes have been described. Immune-related adverse events (IRAEs) have emerged as a serious clinical problem in the use of immune checkpoint inhibitors (ICI). Risk factors for IRAEs and their association with response to therapy remain controversial. Sex differences in innate and adaptive immune responses have been observed but the association of these differences with IRAEs remains unclear. Therefore, we studied sex differences in IRAEs and their association with response to therapy. Methods: All patients with metastatic NSCLC treated with anti-PD1 and anti-PDL1 therapy at Mayo Clinic Rochester and Florida from 2015 to 2018 were reviewed. Patients receiving treatment at an outside facility with history autoimmune disorders or radiation-induced pneumonitis were excluded. Chi-square test was used to estimate differences in categorical data. Kaplan-Meier method was used for time-to-event analysis. Results: A total of 231 patients were identified; 120 (52%) were women and 111 (48%) were men. Baseline characteristics and ICI distribution were similar among groups. Women were more likely to experience IRAEs compared to men (48% vs. 31%, p<0.008). Among patients with IRAEs, women were more likely to be prescribed systemic steroids (63% vs. 41%, p<0.02). On the other hand, no significant differences were observed in the administration of intravenous steroids. Women were more likely to develop pneumonitis (23% vs. 12%, p<0.03) and arthralgias (17% vs. 3%, p<0.04). However, dermatologic toxicities (35% vs. 9%, p<0.002) were more commonly seen in men. In 17% of women the ICI was discontinued due to toxicity (men 7%). Besides sex, no other clinical characteristic was associated with increased IRAEs. Women with IRAEs were more likely to have a radiographic response compared with women without IRAEs (78% vs. 23%, p<0.0001), although this was not observed in men (37% vs. 26%, p>0.22). Better PFS was observed in women with IRAEs (10 months vs. 3.3 months, p<0.0006) compared to women without IRAEs. Conclusions: Women with metastatic NSCLC are more likely to experience IRAEs compared to men. We also observed sex differences in the frequency of certain IRAEs. In addition, an association between IRAEs and response to therapy was observed in women. Larger studies are needed to investigate the mechanisms underlying these associations. Citation Format: Narjust Duma, Abdel-Ghani Azzouqa, Siddhartha Yadav, Kahterine Hoversten, Clay Reed, Andrea Sitek, Elizabeth Enninga, Jonas Paludo, Lisa Kottschade, Aaron Mansfield, Rami Manochakian, Roxana Dronca, Alex Adjei. Sex differences in tolerability and response to immune checkpoint inhibitors in non-small cell lung cancer patients [abstract]. In: Proceedings of the Eleventh AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2018 Nov 2-5; New Orleans, LA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl):Abstract nr A120.

INTRODUCTION: Obesity is associated with lower pregnancy and live birth rates following in vitro fertilization (IVF). As the follicular fluid (FF) provides the microenvironment for the oocyte, immune dysregulation due to obesity related inflammation could disrupt the developing oocyte. The objective of this study was to compare differences in FF cytokines based on BMI in women undergoing IVF. METHODS: FF was collected from women undergoing IVF. Cytokines levels were measured from the FF using ELISA and compared based on BMI ≥35. Patient characteristics and cycle outcomes were compared between the groups. Wilcoxon rank sum and Fisher exact tests were used for continuous and categorical variables, respectively. Values are presented as medians. RESULTS: The analysis included 57 women with BMI <35 and 11 women with BMI ≥35. Women with BMI ≥35 had higher levels of IL-6 (129.23 vs 16.55 pg/mL, P=.019) and TNF-α (62.6 vs 9.54 pg/mL, P=.037) in the FF. Anti-müllerian hormone was lower with BMI ≥35 (2.0 vs 3.6 ng/mL, P=.049), but no other differences in patient characteristics were seen between the groups. There were no differences in stimulation duration (11 vs 11 days; P=.993), total gonadotropin (3075 vs 2475 units; P=.122), peak estradiol (2080 vs 1972 pg/mL, P=.521), mature oocytes (13 vs 12; P=.822), and oocyte maturity rate (82% vs 80%, P=.653) between the groups. CONCLUSION: Obese women undergoing IVF have higher levels of inflammatory cytokines IL-6 and TNF-α in the FF. These biochemical discrepancies may provide early insight into pathophysiologic differences in oocyte development in obese women undergoing IVF.

Abstract During human pregnancy, proinflammatory responses in the placenta can cause severe fetal complications, including growth restriction, preterm birth, and stillbirth. Villitis of unknown etiology (VUE), an inflammatory condition characterized by the infiltration of maternal CD8+ T cells into the placenta, is hypothesized to be secondary to either a tissue rejection response to the haploidentical fetus or from an undiagnosed infection. In this study, we characterized the global TCR β-chain profile in human T cells isolated from placentae diagnosed with VUE compared with control and infectious villitis–placentae by immunoSEQ. Immunosequencing demonstrated that VUE is driven predominantly by maternal T cell infiltration, which is significantly different from controls and infectious cases; however, these T cell clones show very little overlap between subjects. Mapping TCR clones to common viral epitopes (CMV, EBV, and influenza A) demonstrated that Ag specificity in VUE was equal to controls and significantly lower than CMV-specific clones in infectious villitis. Our data indicate VUE represents an allograft response, not an undetected infection. These observations support the development of screening methods to predict those at risk for VUE and the use of specific immunomodulatory therapies during gestation to improve outcomes in affected fetuses.

Congenital diaphragmatic hernia (CDH) occurs in approximately 1 in 2,500 live births; it is associated with high rates of neonatal mortality, especially in cases with severe pulmonary hypoplasia and pulmonary arterial hypertension. Treatments have focused on hernia repair and ameliorating lung hypoplasia, with the precise pathophysiological mechanisms of CDH remain unknown. We investigated whether or not diaphragm muscle (DIAm) weakness is apparent in CDH, evaluating DIAm force generation and ventilatory function in a Nitrofen rat model. 7 pregnant rats received 100 mg of Nitrofen in 2 mL of olive oil at 9 days gestational age and rat pups were collected at birth. Ventilatory function was assessed in isolated plethysmography chambers (Buxco). Following these functional assessments, neonatal pups were euthanized, and the DIAm with its rib attachments excised. The presence or absence of a CDH was confirmed and a DIAm strip was obtained either from control pups, or from the unaffected side in CDH pups. The isometric force generation of these DIAm strips and their fatigueability was assessed using field stimulation of muscle at various frequencies (5–100 Hz) and normalized to DIAm cross‐sectional area. Statistical significance was determined using a Student’s t‐test or Mann‐Whitney u test at the P <0.05 level. A marked reduction in both neonatal respiratory rate (−42%) and respiratory minute ventilation (−43%) was observed in CDH pups compared to controls ( P <0.01). We observed reduced DIAm twitch (−40%), maximum tetanic (−72%) and residual force following fatigue (−69%) in CDH pups compared to controls ( P <0.05). CDH neonatal rat pups exhibit DIAm weakness and ventilatory disturbances. Based on previous modelling of ventilatory function, it is possible that DIAm weakness contributes to the poor ventilation, as CDH DIAm forces following fatigue were less than control groups. This work has important considerations when assessing novel treatment outcomes in clinical CDH.

AbstractObjectiveWe aimed to test for an association between the amount of circulating fetal cell‐free DNA and trisomy, and whether NIPS failure due to low fetal fraction indicates trisomy risk.MethodMaternal BMI, maternal age, fetal sex, gestational age, fetal cfDNA fraction, and NIPS results was collected on 2374 pregnancies. Additional clinical information was available for 1180 research consented patients. We investigated associations between fetal fraction and available variables and determined the success rate of repeat NIPS testing.ResultsFetal trisomy was marginally associated with decreased fetal fraction (P = .067). However, the proportions of trisomy events were not significantly increased in women who had failed NIPS due to low fetal fraction (<4%) (OR = 1.37 [0.3‐7.4]; P = .714). 66% of repeated NIPS after a second blood draw were successful.ConclusionFailure to meet the clinical cutoff of 4% fetal fraction established for NIPS accuracy did not suggest increased risk for trisomy in our cohort. Because repeat testing was successful in the majority of cases and most failures were explained by high BMI and low gestational age, a redraw may be an appropriate next step before invasive screening due to concerns for trisomic pregnancies.

Abstract The importance of fetal placental macrophages (Hofbauer cell [HCs]) is underscored by their appearance 18 d postconception and maintenance through term; however, how human HCs evolve during healthy pregnancy and how microenvironment and ontogeny impact phenotype and function remain unknown. In this study, we comprehensively classify human HCs ex vivo, interrogate phenotypic plasticity, and characterize antiviral immune responses through gestation. Activated HCs were abundant in early pregnancy and decreased by term; molecular signatures emphasize inflammatory phenotypes early in gestation. Frequency of HCs with regulatory phenotypes remained high through term. Furthermore, term HCs exhibited blunted responses to stimulation, indicating reduced plasticity. IFN-λ1 is a key placental IFN that appeared less protective than IFN-α, suggesting a potential weakness in antiviral immunity. Ligand-specific responses were temporally regulated: we noted an absence of inflammatory mediators and reduced antiviral gene transcription following RIG-I activation at term despite all HCs producing inflammatory mediators following IFN-γ plus LPS stimulation. Collectively, we demonstrate sequential, evolving immunity as part of the natural history of HCs through gestation.

Background: A female survival advantage in cutaneous melanoma has been long recognized. However, whether this extends across all age groups, with risk stratification using the latest prognostic staging system or in the current era of efficacious systemic therapies is unknown. Therefore, we evaluated whether sex-based differences in melanoma survival persisted within a recent population-based patient cohort with consideration of these factors. Materials and Methods: We identified stage II−IV cutaneous melanoma patients from 2010 to 2014 Surveillance, Epidemiology, and End Results cancer registries data. We recalculated stage per American Joint Committee on Cancer 8th edition guidelines. Cancer-specific survival (CSS) was estimated by using the Kaplan–Meier method and multivariable Cox proportional hazards regression. Results: Of 16,807 patients (39.8% female), 8,990 were stage II, 4,826 stage III, and 2,991 stage IV at diagnosis. Unadjusted 3-/5-year CSS estimates for females versus males were 64.2% versus 59.7%, and 53.5% versus 49.9%, respectively, p ≤ 0.0001. Five-year CSS varied within each stage and across age strata of <45, 45 − 59, and ≥60 years. Within each stage, females <45 had better CSS than all other sex/age groups ( p < 0.0001). In multivariable analysis of stage II/III patients, female sex, younger age, and lower mitotic index retained favorable CSS prognostic significance ( p < 0.001). Conclusions: Sex-based differences in melanoma survival persist in a contemporary patient cohort staged with the latest prognostic system. These data may guide decision marking regarding adjuvant therapy, highlight the importance of including sex as a pre-specified clinical trial variable, and suggest that investigation of underlying biologic mechanisms may drive discovery of biomarkers and therapeutic targets to improve patient care.

4 Background: Membrane expression of PD-L1 on tumor cells is associated with poor survival in patients with advanced malignancies including metastatic melanoma (MM). In addition, tumor-associated PD-L1 has been proposed as a predictor of response to anti-PD-1 therapy, although responses are observed in some cases of PD-L1 negative MM. We hypothesize that tumor-derived PD-L1 has additional systemic effects through the release of biologically active soluble forms of PD-L1 (sPD-L1) into the circulation, which further impedes the anti-tumor immune response and contributes to poor clinical outcomes in MM. Methods: We developed a sPD-L1 ELISA and biochemically confirmed the identity of the detected protein. We measured the levels of sPD-L1 in 276 patients with MM enrolled on 3 clinical trials who had stored pre-treatment plasma samples and 37 healthy volunteers (HV) undergoing blood donation at Mayo Clinic. We also measured sPD-L1 in baseline samples from 38 MM patients treated with anti-PD1 (pembrolizumab) 2 mg/kg every 3 weeks. ROC analysis was used to compute sPD-L1 concentration cut-off value and Wald test was used to asses the difference of overall survival (OS) in patients with low versus high sPD-L1 concentration. Results: We found that sPD-L1 levels were significantly elevated in MM patients compared with HV (p=0.0011). Mean sPD-L1 level in MM was 1.73 ng/mL (range: 0.13-18.29) compared with 0.77 ng/mL (range: 0.11–6.02) for HV. Patients with higher levels (>0.293 ng/mL) had a median OS of 11.3 months compared to 14.8 months for patients with sPD-L1 level ≤ 0.293 ng/mL (p=0.040). Similarly to tumor-related PD-L1 findings, patients who had clinical benefit (OR/PR/SD) after 4 cycles of anti-PD1 had higher sPD-L1 levels at baseline (2.1 vs. 1.1 ng/mL). Conclusions: To our knowledge, our study is the first to report a correlation of sPD-L1 with MM patient outcome, although a larger study is needed to establish the utility of sPD-L1 as an independent prognostic and possibly predictive biomarker.

The Center for Disease Control and Prevention ranks diabetes mellitus (DM) as the seventh leading cause of death in the USA. The most prevalent forms of DM include Type 2 DM, Type 1 DM, and gestational diabetes mellitus (GDM). While the acute problem of diabetic hyperglycemia can be clinically managed through dietary control and lifestyle changes or pharmacological intervention with oral medications or insulin, long-term complications of the disease are associated with significant morbidity and mortality. These long-term complications involve nearly all organ systems of the body and share common pathologies associated with endothelial cell abnormalities. To better understand the molecular mechanisms underlying DM as related to future long-term complications following hyperglycemia, we have undertaken a study to determine the frequency that GDM did or did not occur in the second pregnancy of women who experienced GDM in their first pregnancy between 2013 and 2018 at Mayo Clinic, Rochester, MN. Within the five-year period of the study, the results indicate that 7,330 women received obstetrical care for pregnancy during the study period. Of these, 150 developed GDM in their first pregnancy and of these, 42 (28%) had a second pregnancy. Of these 42 women, 20 again developed GDM and 22 did not develop GDM in their second pregnancy within the study period. Following the occurrence of GDM in the first pregnancy, the study (1) established the number of women with and without GDM in the second pregnancy and (2) confirmed the feasibility to study diabetic metabolic memory using maternal placental tissue from GDM women. These studies represent Phase I of a larger research project whose goal is to analyze epigenetic mechanisms underlying true diabetic metabolic memory using endothelial cells isolated from the maternal placenta of women with and without GDM as described in this article.