The Adverse Event Landscape of Stem Cell Transplant: Evidence for aGVHD Driving Early Transplant Associated Toxicities.

Although unrelated-donor (URD) hematopoietic cell transplantation (HCT) is associated with many toxicities, a detailed analysis of adverse events, as defined by the Common Terminology Criteria for Adverse Events (CTCAE), has not previously been curated. This represents a major unmet need, especially as it relates to assessing the safety of novel agents. We analyzed a detailed AE database from the "ABA2" randomized, double-blind, placebo-controlled clinical trial of abatacept for acute graft-versus-host disease (aGVHD) prevention, for which the FDA mandated a detailed AE assessment through Day +180, and weekly neutrophil and platelet counts through Day +100. These were analyzed for their relationship to key transplant outcomes, with a major focus on the impact of aGVHD on the development/severity of AEs. A total of 2102 AEs and 1816 neutrophil/platelet counts were analyzed from 142 8/8-HLA-matched URD HCT recipients on ABA2 (placebo cohort, n = 69, abatacept cohort, n = 73). This analysis resulted in 2 major observations. (1) Among graft source, conditioning intensity, age, and Grade 2 to 4 aGVHD, only aGVHD impacted Grade 3 to 5 AE acquisition after the first month post-transplant. (2) The development of Grade 3 to 4 aGVHD was associated with thrombocytopenia. We have created a detailed resource for the transplant community by which to contextualize clinical toxicities after transplant. It has identified aGVHD as a major driver of post-HCT Grade 3 to 5 AEs, and underscored a link between aGVHD and thrombocytopenia. This establishes a critical safety framework upon which the impact of novel post-transplant aGVHD therapeutics should be evaluated. This trial was registered at www.clinicaltrials.gov (#NCT01743131).

Overlap chronic GVHD is associated with adverse survival outcomes compared to classic chronic GVHD.

Chronic graft-versus-host-disease (cGVHD) is divided into two subtypes: classic (absence of acute GVHD features) and overlap cGVHD ('ocGVHD'), in which both chronic and acute GVHD clinical features are present simultaneously. While worse outcomes with ocGVHD have been reported, there are few recent analyses. We performed a secondary analysis of data from the ABA2 trial (N = 185), in which detailed GVHD data were collected prospectively and systematically adjudicated. Analyses included cumulative incidence of classic versus ocGVHD, their specific organ manifestations, global disease severity scores, non-relapse mortality (NRM), disease-free survival (DFS) and overall survival (OS) in these two cGVHD subtypes. Of 92 patients who developed cGVHD, 35 were classified as ocGVHD. The 1-year cumulative incidence, organ involvement, and global severity of classic and ocGVHD were similar between ABA2 patients receiving CNI/MTX+placebo and CNI/MTX+abatacept; thus, cohorts were combined for ocGVHD evaluation. This analysis identified ocGVHD as having significantly higher severity at presentation and at maximum global severity compared to classic cGVHD. OS and DFS were significantly lower for ocGVHD versus classic cGVHD. OcGVHD is associated with increased cGVHD severity scores, and is associated with decreased OS and DFS compared to classic cGVHD, underscoring the high risks with this cGVHD subtype.

A novel phosphorylation site involved in dissociating RAF kinase from the scaffolding protein 14-3-3 and disrupting RAF dimerization.

Rapidly accelerated fibrosarcoma (ARAF, BRAF, CRAF) kinase is central to the MAPK pathway (RAS-RAF-MEK-ERK). Inactive RAF kinase is believed to be monomeric, autoinhibited, and cytosolic, while activated RAF is recruited to the membrane via RAS-GTP, leading to the relief of autoinhibition, phosphorylation of key regulatory sites, and dimerization of RAF protomers. Although it is well known that active and inactive BRAF have differential phosphorylation sites that play a crucial role in regulating BRAF, key details are still missing. In this study, we report the characterization of a novel phosphorylation site, BRAFS732 (equivalent in CRAFS624), located in proximity to the C-terminus binding motif for the 14-3-3 scaffolding protein. At the C terminus, 14-3-3 binds to BRAFpS729 (CRAFpS621) and enhances RAF dimerization. We conducted mutational analysis of BRAFS732A/E and CRAFS624A/E and revealed that the phosphomimetic S→E mutant decreases 14-3-3 association and RAF dimerization. In normal cell signaling, dimerized RAF phosphorylates MEK1/2, which is observed in the phospho-deficient S→A mutant. Our results suggest that phosphorylation and dephosphorylation of this site fine-tune the association of 14-3-3 and RAF dimerization, ultimately impacting MEK phosphorylation. We further characterized the BRAF homodimer and BRAF:CRAF heterodimer and identified a correlation between phosphorylation of this site with drug sensitivity. Our work reveals a novel negative regulatory role for phosphorylation of BRAFS732 and CRAFS624 in decreasing 14-3-3 association, dimerization, and MEK phosphorylation. These findings provide insight into the regulation of the MAPK pathway and may have implications for cancers driven by mutations in the pathway.

Ambulatory Orbital Fracture Repair: An Analysis of ER Visits After Surgery From a Multistate Study.

OBJECTIVE: To determine the 30-day postoperative emergency room (ER) visit rate following ambulatory orbital fracture repair with same-day discharge, and the causes and risk factors associated with ER visit. STUDY DESIGN: Database study. SETTING: State Ambulatory Surgery and Services Database (SASD) and State Emergency Department Database (SEDD) for California, New York, and Florida for 2011. METHODS: We identified orbital fracture repair procedures among adults from the SASD, which was linked to the SEDD to identify the incidence and causes of ER visits within 30 days. Univariate and multivariable logistic regression models were used to determine the factors associated with ER visit. RESULTS: Among 762 patients, the 30-day postoperative ER visit rate was 4.5%. Most ER visits (58.9%) occurred during the first week after surgery. The most common reasons for ER visits were related to pain, swelling, headache, dizziness, and fatigue (29.4%), followed by ophthalmologic etiologies including visual disturbances and infection of the eye (14.7%). There was no case of retrobulbar hematoma. In the multivariate analysis, patients living in Florida were at a significantly higher risk for ER visit compared to those in California (odds ratio: 4.48 [1.43-14.10], p = .010). CONCLUSION: Ambulatory orbital fracture repair appears to be safe. Common reasons for ER visit included pain, swelling, and ophthalmic symptoms. An increased risk for ER visit was seen with certain geographic regions but not with medical comorbidities or concurrent facial fractures or procedures.

Higher abatacept exposure after transplant decreases acute GVHD risk without increasing adverse events.

In the ABA2 study, the T-cell costimulation blockade agent, abatacept, was safe and effective in preventing acute graft-versus-host disease (aGVHD) after unrelated-donor hematopoietic cell transplant (HCT), leading to US Food and Drug Administration approval. Here, we performed a determination of abatacept pharmacokinetics (PK), which enabled an examination of how abatacept exposure-response relationships affected clinical outcomes. We performed a population PK analysis of IV abatacept using nonlinear mixed-effect modeling and assessed the association between abatacept exposure and key transplant outcomes. We tested the association between the trough after dose 1 (Ctrough_1) and grade (GR) 2 or 4 aGVHD (GR2-4 aGVHD) through day +100. An optimal Ctrough_1 threshold was identified via recursive partitioning and classification tree analysis. This demonstrated that abatacept PK was characterized by a 2-compartment model with first-order elimination. The ABA2 dosing regimen was based on previous work targeting a steady-state abatacept trough of 10 µg/mL. However, a higher Ctrough_1 (≥39 µg/mL, attained in ∼60% of patients on ABA2) was associated with a favorable GR2-4 aGVHD risk (hazard ratio, 0.35; 95% confidence interval, 0.19-0.65; P < .001), with a Ctrough_1 <39 µg/mL associated with GR2-4 aGVHD risk indistinguishable from placebo (P = .37). Importantly, no significant association was found between Ctrough_1 and key safety indicators, including relapse, and cytomegalovirus or Epstein-Barr virus viremia. These data demonstrate that a higher abatacept Ctrough_1 (≥39 µg/mL) was associated with a favorable GR2-4 aGVHD risk, without any observed exposure-toxicity relationships. This trial was registered at www.clinicaltrials.gov as #NCT01743131.

Notch signaling drives intestinal graft-versus-host disease in mice and nonhuman primates.

Notch signaling promotes T cell pathogenicity and graft-versus-host disease (GVHD) after allogeneic hematopoietic cell transplantation (allo-HCT) in mice, with a dominant role for the Delta-like Notch ligand DLL4. To assess whether Notch's effects are evolutionarily conserved and to identify the mechanisms of Notch signaling inhibition, we studied antibody-mediated DLL4 blockade in a nonhuman primate (NHP) model similar to human allo-HCT. Short-term DLL4 blockade improved posttransplant survival with durable protection from gastrointestinal GVHD in particular. Unlike prior immunosuppressive strategies tested in the NHP GVHD model, anti-DLL4 interfered with a T cell transcriptional program associated with intestinal infiltration. In cross-species investigations, Notch inhibition decreased surface abundance of the gut-homing integrin α4ß7 in conventional T cells while preserving α4ß7 in regulatory T cells, with findings suggesting increased ß1 competition for α4 binding in conventional T cells. Secondary lymphoid organ fibroblastic reticular cells emerged as the critical cellular source of Delta-like Notch ligands for Notch-mediated up-regulation of α4ß7 integrin in T cells after allo-HCT. Together, DLL4-Notch blockade decreased effector T cell infiltration into the gut, with increased regulatory to conventional T cell ratios early after allo-HCT. Our results identify a conserved, biologically unique, and targetable role of DLL4-Notch signaling in intestinal GVHD.

Unplanned 30-Day ER Visit Rate and Factors Associated With ER Visits After Ambulatory Sinus Surgery.

BACKGROUND: Emergency room (ER) visits after surgery can be inconvenient and costly to the patient and the healthcare system. Estimates of the 30-day ER visit rate following ambulatory sinus procedures and their risk factors are largely unknown in the literature. OBJECTIVE: To determine the 30-day postoperative ER visit rate following ambulatory sinus procedures and the causes and risk factors associated with ER visits. METHODS: This is a retrospective, cohort study using data from the State Ambulatory Surgery and Services Databases (SASD) and the State Emergency Department Databases (SEDD) for California, New York, and Florida in 2019. We identified adult (18 years old) patients with chronic rhinosinusitis who underwent ambulatory sinus procedures from the SASD. Cases were linked to the SEDD to identify ER visits occurring within 30 days after the procedure. Logistic regression models were used to identify patient- and procedure-related risk factors associated with the 30-day postoperative ER visit. RESULTS: Among the 23 239 patients, the 30-day postoperative ER visit rate was 3.9%. The most common reason for ER visit was bleeding (32.7%). A total of 56.9% of the ER visits occurred within the first week. In the multivariate analysis, factors associated with ER visits included Medicare (odds ratio [OR] 1.29 [1.09-1.52], P = .003), Medicaid (OR 2.06 [1.69-2.51], P < .001), self-pay/no insurance (OR 1.44 [1.03-2.00], P = .031), chronic kidney disease/end-stage renal disease (OR 1.63 [1.06-2.51], P = .027), chronic pain/opioid use (OR 2.70 [1.02-7.11], P = .045), and a disposition other than home (OR 12.61 [8.34-19.06], P < .001). CONCLUSION: The most common reason for ER visit after ambulatory sinus procedures was bleeding. An increased ER visit rate was associated with certain demographic factors and medical comorbidities but not with procedure characteristics. This information can help us identify the patient populations who are at higher risk for ER visits to improve their postoperative recovery.

Human papillomavirus detection in oral rinses and history of tonsillectomy in U.S. adults.

PURPOSE: This study aims to elucidate any relationship between prior tonsillectomy and the presence of oropharyngeal HPV DNA found in screening mouth rinses. MATERIALS AND METHODS: A cross sectional study was conducted using the 2011-2014 National Health and Nutrition Examination Survey (NHANES). Participants between 40 and 69 were included in the study and medical, surgical, and sexual health history were recorded. Multivariable analyses were conducted to examine factors associated with HPV prevalence in oral rinse samples. RESULTS: A total of 4825 participants were recorded with 21.1 % having a history of tonsillectomy. In the no tonsillectomy group, 8.6 % of respondents had a positive oral rinse for HPV, while 7.2 % of those with a tonsillectomy had a positive rinse sample. There was no association between age and HPV prevalence (OR = 1.04, 95 % CI: [1.00-1.07]). When controlling for demographics, medical history, and sexual behaviors, tonsillectomy history was not shown to have an association with HPV (OR = 0.86, 95 % CI: [0.53-1.40]). However, men, Hispanics, smokers, and those with higher lifetime sexual partners had increased odds of having a positive HPV oral rinse sample which was statistically significant. CONCLUSION: Our data showed that a history of tonsillectomy was not significantly associated with the presence of HPV in an oral rinse. However, a significant relationship was seen between the presence of HPV in oral rinses and certain demographic factors such as male gender, Hispanic race, smoking history, and increased sexual partners.

Association Between Insurance Type and Outcomes of Reconstructive Head and Neck Cancer Surgery.

OBJECTIVES/HYPOTHESIS: Although the benefits of expanding health insurance coverage are clear, there are limited studies comparing the different types of insurance. This study aims to determine the association between insurance type and outcomes in patients with head and neck cancer undergoing reconstructive surgery in the United States. METHODS: Population-based cross-sectional study of the 2012-2014 National Inpatient Sample. We identified 1,314 patients with head and neck cancers undergoing tumor ablative surgery followed by pedicled or free flap reconstruction of oncologic defects. Insurance type was classified as private, Medicare, Medicaid, self-pay, or other. The primary outcome was extended length of stay (LOS), defined as greater than 14 days, which represented the 75th percentile of the study sample. Secondary outcomes included acute medical complications, surgical complications, morbidities, and costs. Analyses were adjusted for gender, geographic location, and various medical comorbidities. RESULTS: In univariate analysis, insurance type was associated with extended LOS (P = .001), medical complications (P = <.001), and mortalities (P = .020). After controlling for other covariates in the multivariate analysis, compared to private insurance, Medicare and Medicaid were both associated with significantly higher odds of extended LOS (adjusted odds ratio [OR] [95% confidence interval (CI)] = 1.73 [1.09-2.76] and 2.22 [1.38-3.58], respectively). Medicare was associated with significantly higher odds of medical complications, but Medicaid was not (adjusted OR [95% CI] = 1.53 [1.02-2.31] and 1.64 [0.97-2.78], respectively). CONCLUSIONS: Medicaid and Medicare were independently associated with extended LOS after reconstructive head and neck cancer surgery. Medicare was associated with higher rates of medical complications. Efforts to address LOS should target care planning and coordination. LEVEL OF EVIDENCE: NA Laryngoscope, 132:1946-1952, 2022.

Phase II Trial of Costimulation Blockade With Abatacept for Prevention of Acute GVHD.

PURPOSE: Severe (grade 3-4) acute graft-versus-host disease (AGVHD) is a major cause of death after unrelated-donor (URD) hematopoietic cell transplant (HCT), resulting in particularly high mortality after HLA-mismatched transplantation. There are no approved agents for AGVHD prevention, underscoring the critical unmet need for novel therapeutics. ABA2 was a phase II trial to rigorously assess safety, efficacy, and immunologic effects of adding T-cell costimulation blockade with abatacept to calcineurin inhibitor (CNI)/methotrexate (MTX)-based GVHD prophylaxis, to test whether abatacept could decrease AGVHD. METHODS: ABA2 enrolled adults and children with hematologic malignancies under two strata: a randomized, double-blind, placebo-controlled stratum (8/8-HLA-matched URD), comparing CNI/MTX plus abatacept with CNI/MTX plus placebo, and a single-arm stratum (7/8-HLA-mismatched URD) comparing CNI/MTX plus abatacept versus CNI/MTX CIBMTR controls. The primary end point was day +100 grade 3-4 AGVHD, with day +180 severe-AGVHD-free-survival (SGFS) a key secondary end point. Sample sizes were calculated using a higher type-1 error (0.2) as recommended for phase II trials, and were based on predicting that abatacept would reduce grade 3-4 AGVHD from 20% to 10% (8/8s) and 30% to 10% (7/8s). ABA2 enrolled 142 recipients (8/8s, median follow-up = 716 days) and 43 recipients (7/8s, median follow-up = 708 days). RESULTS: In 8/8s, grade 3-4 AGVHD was 6.8% (abatacept) versus 14.8% (placebo) (P = .13, hazard ratio = 0.45). SGFS was 93.2% (CNI/MTX plus abatacept) versus 82% (CNI/MTX plus placebo, P = .05). In the smaller 7/8 cohort, grade 3-4 AGVHD was 2.3% (CNI/MTX plus abatacept, intention-to-treat population), which compared favorably with a nonrandomized matched cohort of CNI/MTX (30.2%, P < .001), and the SGFS was better (97.7% v 58.7%, P < .001). Immunologic analysis revealed control of T-cell activation in abatacept-treated patients. CONCLUSION: Adding abatacept to URD HCT was safe, reduced AGVHD, and improved SGFS. These results suggest that abatacept may substantially improve AGVHD-related transplant outcomes, with a particularly beneficial impact on HLA-mismatched HCT.

Spatiotemporal single-cell profiling reveals that invasive and tissue-resident memory donor CD8+ T cells drive gastrointestinal acute graft-versus-host disease.

Organ infiltration by donor T cells is critical to the development of acute graft-versus-host disease (aGVHD) in recipients after allogeneic hematopoietic stem cell transplant (allo-HCT). However, deconvoluting the transcriptional programs of newly recruited donor T cells from those of tissue-resident T cells in aGVHD target organs remains a challenge. Here, we combined the serial intravascular staining technique with single-cell RNA sequencing to dissect the tightly connected processes by which donor T cells initially infiltrate tissues and then establish a pathogenic tissue residency program in a rhesus macaque allo-HCT model that develops aGVHD. Our results enabled creation of a spatiotemporal map of the transcriptional programs controlling donor CD8+ T cell infiltration into the primary aGVHD target organ, the gastrointestinal (GI) tract. We identified the large and small intestines as the only two sites demonstrating allo-specific, rather than lymphodepletion-driven, T cell infiltration. GI-infiltrating donor CD8+ T cells demonstrated a highly activated, cytotoxic phenotype while simultaneously developing a canonical tissue-resident memory T cell (TRM) transcriptional signature driven by interleukin-15 (IL-15)/IL-21 signaling. We found expression of a cluster of genes directly associated with tissue invasiveness, including those encoding adhesion molecules (ITGB2), specific chemokines (CCL3 and CCL4L1) and chemokine receptors (CD74), as well as multiple cytoskeletal proteins. This tissue invasion transcriptional signature was validated by its ability to discriminate the CD8+ T cell transcriptome of patients with GI aGVHD from those of GVHD-free patients. These results provide insights into the mechanisms controlling tissue occupancy of target organs by pathogenic donor CD8+ TRM cells during aGVHD in primate transplant recipients.

Prevalence of oral human papillomavirus infections by race in the United States: An association with sexual behavior.

OBJECTIVES: To investigate the differences in oral HPV infection and sexual behaviors by race in the US. MATERIALS AND METHODS: We analyzed data from the 2011-2014 US National Health and Nutrition Examination Survey during which participants aged 18-69 years completed oral rinse exam for HPV detection (n = 8,229). Logistic regression was used to examine the associations of race with various types of oral HPV infection and sexual behaviors. RESULTS: The prevalence of overall oral HPV infection and HPV type16 infection was 7.5% [95% CI: 6.6-8.4] and 1.1% [95% CI: 0.7-1.3], respectively. Blacks were more likely to have any oral HPV infection [OR: 1.22, 95% CI: 1.01-1.47] and Asian Americans were less likely to have any oral HPV infection [OR: 0.33, 95% CI: 0.24-0.49] than Whites. In a multivariate model, Whites were less likely to have any oral HPV infections than Blacks while having higher order of impact by the number of lifetime sex partners. Overall, Asian Americans were less likely to have type16 infection [OR: 0.21, 95% CI: 0.06-0.67] than Whites; however, that difference disappears when adjusting for sexual behaviors. CONCLUSIONS: In this nationally representative sample of US adults, the prevalence of overall oral HPV infections was higher among Blacks and lower among Asians in comparison to Whites. Further analysis with sexual behavior data suggested that the racial differences in prevalence are likely due to different sexual behaviors.

Association of Race/Ethnicity, Stage, and Survival in Oral Cavity Squamous Cell Carcinoma: A SEER Study.

OBJECTIVE: Survival differences in oral cancer between black and white patients have been reported, but the contributing factors, especially the role of stage, are incompletely understood. Furthermore, the outcomes for Hispanic and Asian patients have been scarcely examined. STUDY DESIGN: Retrospective, population-based national study. SETTING: Surveillance, Epidemiology, and End Results 18 Custom database (January 1, 2010, to December 31, 2014). SUBJECTS AND METHODS: In total, 7630 patients with primary squamous cell carcinoma in the oral cavity were classified as non-Hispanic white (white), non-Hispanic black (black), Hispanic, or Asian. Cox regression was used to obtain unadjusted and adjusted hazard ratios (HRs) of 5-year mortality for race/ethnicity with sequential adjustments for stage and other covariates. Logistic regression was used to examine the relationship between race/ethnicity and stage with adjusted odds ratios (aORs). RESULTS: The cohort consisted of 75.0% whites, 7.6% blacks, 9.1% Hispanics, and 8.3% Asians. Compared to whites, the unadjusted HR for all-cause mortality for blacks was 1.68 (P < .001), which attenuated to 1.15 (P = .039) after adjusting for stage and became insignificant after including insurance. The unadjusted HRs for all-cause mortality were not significant for Hispanics and Asians vs whites. Compared to whites, blacks and Hispanics were more likely to present at later stages (aORs of 2.63 and 1.42, P < .001, respectively). CONCLUSION: The greater mortality for blacks vs whites was largely attributable to the higher prevalence of later stages at presentation and being uninsured among blacks. There was no statistically significant difference in mortality for Hispanics vs whites or Asians vs whites.

Sinus irrigation penetration after balloon sinuplasty vs functional endoscopic sinus surgery in a cadaveric model.

BACKGROUND: Nasal irrigation is a cornerstone of treatment for chronic rhinosinusitis. The purpose of this study was to quantify irrigation penetration to the sinuses following balloon sinuplasty and functional endoscopic sinus surgery (FESS). METHODS: Balloon sinuplasty followed by FESS was performed on 4 cadaver heads. Using a high-volume, high-flow bottle, each head was irrigated with fluorescein-dyed water prior to and following each procedure, and recorded by rigid endoscopy through trephinations. Three blinded, fellowship-trained rhinologists reviewed videos and scored the extent of staining (using an accepted scale of 0 to 3) for each site. RESULTS: The mean score prior to any procedure was maxillary sinus 1.67, frontal sinus 1.29, and nasal cavity 1.71. After balloon sinuplasty the mean was maxillary 2.25, frontal 2.04, and nasal cavity 2.17. After FESS the mean was maxillary 2.75, frontal 2.08, and nasal cavity 2.63. There was a statistically significant increase for both maxillary (p = 0.005) and frontal sinuses (p = 0.006) following balloon sinuplasty. There was a statistically significant increase following FESS compared to balloon for the maxillary sinus (p = 0.003), but not the frontal sinus (p = 0.96). Interrater reliability was good, with Cronbach's alpha of 0.85. CONCLUSION: Irrigation improved in all sinuses following balloon sinuplasty and FESS. There was further improvement to the maxillary sinus after FESS; however, there was no difference in irrigation to the frontal sinuses following FESS compared to balloon sinuplasty. Extended frontal sinus approaches such as the Modified Lothrop procedure should be considered if more extensive access for irrigation is required.

Open Versus Endoscopic Approach for Sinonasal Melanoma: A Systematic Review and Meta-analysis.

BACKGROUND: Open resection (OR) of sinonasal mucosal melanoma (SNM) traditionally has been the gold standard for treatment. However, endoscopic resection (ER) has recently become a surgical alternative. The aim of this study was to compare survival outcomes between OR and ER of SNM. METHODS: A literature search encompassing PubMed, Embase, Cochrane Library, Web of Science, ClinicalTrials.gov, and Google Scholar was performed. Two reviewers independently screened for original studies comparing survival outcomes between OR and ER for SNM. Data were systematically collected on study design, patient demographics, outcomes, and level of evidence. Quality assessment was performed using the Newcastle-Ottawa scale (NOS). Meta-analysis of overall survival and disease-free survival was performed using random-effects models. RESULTS: The initial search yielded 2078 abstracts, of which 9 cohort studies were included for a total of 510 patients from 6 different countries. The average quality of all included studies using the NOS was 7.7 stars. Six out of 7 studies reported no differences in the stages of SNM between patients receiving ER versus OR. Overall survival was longer in the ER group versus OR group (hazard ratio [HR]: 0.68, 95% confidence interval [CI]: 0.49-0.95). There was no significant difference in disease-free survival between groups (HR: 0.59, 95% CI: 0.28-1.25). CONCLUSION: Based on the available literature, an endoscopic approach for SNM resection has survival outcomes that are similar or greater compared to an open approach.

Characterization of gene deletion mutants of Cyprinid herpesvirus 3 (koi herpesvirus) lacking the immunogenic envelope glycoproteins pORF25, pORF65, pORF148 and pORF149.

Cyprinid herpesvirus 3 (CyHV-3) or koi herpesvirus is a global pathogen causing mass mortality in koi and common carp, against which improved vaccines are urgently needed. In this study we investigated the role of four nonessential, but immunogenic envelope glycoproteins encoded by members of the ORF25 gene family (ORF25, ORF65, ORF148 and ORF149) during CyHV-3 replication. Single deletion of ORF65 did not affect in vitro replication, and deletion of ORF148 even slightly enhanced virus growth on common carp brain (CCB) cells. Deletions of ORF25 or ORF149 led to reduced plaque sizes and virus titers, which was due to delayed entry into host cells. An ORF148/ORF149 double deletion mutant exhibited wild-type like growth indicating opposing functions of the two proteins. Electron microscopy of CCB cells infected with either mutant did not indicate any effects on virion formation and maturation in nucleus or cytoplasm, nor on release of enveloped particles. The ORF148, ORF149 and double deletion mutants were also tested in animal experiments using juvenile carp, and proved to be insufficiently attenuated for use as live virus vaccines. However, surviving fish were protected against challenge with wild-type CyHV-3, demonstrating that these antibody inducing proteins are dispensable for an efficient immune response in vivo.

Evidence for persistence of the SHIV reservoir early after MHC haploidentical hematopoietic stem cell transplantation.

Allogeneic transplantation (allo-HCT) has led to the cure of HIV in one individual, raising the question of whether transplantation can eradicate the HIV reservoir. To test this, we here present a model of allo-HCT in SHIV-infected, cART-suppressed nonhuman primates. We infect rhesus macaques with SHIV-1157ipd3N4, suppress them with cART, then transplant them using MHC-haploidentical allogeneic donors during continuous cART. Transplant results in ~100% myeloid donor chimerism, and up to 100% T-cell chimerism. Between 9 and 47 days post-transplant, terminal analysis shows that while cell-associated SHIV DNA levels are reduced in the blood and in lymphoid organs post-transplant, the SHIV reservoir persists in multiple organs, including the brain. Sorting of donor-vs.-recipient cells reveals that this reservoir resides in recipient cells. Moreover, tetramer analysis indicates a lack of virus-specific donor immunity post-transplant during continuous cART. These results suggest that early post-transplant, allo-HCT is insufficient for recipient reservoir eradication despite high-level donor chimerism and GVHD.

CD28 blockade controls T cell activation to prevent graft-versus-host disease in primates.

Controlling graft-versus-host disease (GVHD) remains a major unmet need in stem cell transplantation, and new, targeted therapies are being actively developed. CD28-CD80/86 costimulation blockade represents a promising strategy, but targeting CD80/CD86 with CTLA4-Ig may be associated with undesired blockade of coinhibitory pathways. In contrast, targeted blockade of CD28 exclusively inhibits T cell costimulation and may more potently prevent GVHD. Here, we investigated FR104, an antagonistic CD28-specific pegylated-Fab', in the nonhuman primate (NHP) GVHD model and completed a multiparameter interrogation comparing it with CTLA4-Ig, with and without sirolimus, including clinical, histopathologic, flow cytometric, and transcriptomic analyses. We document that FR104 monoprophylaxis and combined prophylaxis with FR104/sirolimus led to enhanced control of effector T cell proliferation and activation compared with the use of CTLA4-Ig or CTLA4-Ig/sirolimus. Importantly, FR104/sirolimus did not lead to a beneficial impact on Treg reconstitution or homeostasis, consistent with control of conventional T cell activation and IL-2 production needed to support Tregs. While FR104/sirolimus had a salutary effect on GVHD-free survival, overall survival was not improved, due to death in the absence of GVHD in several FR104/sirolimus recipients in the setting of sepsis and a paralyzed INF-γ response. These results therefore suggest that effectively deploying CD28 in the clinic will require close scrutiny of both the benefits and risks of extensively abrogating conventional T cell activation after transplant.

On-site detection of bovine leukemia virus by a field-deployable automatic nucleic extraction plus insulated isothermal polymerase chain reaction system.

Bovine leukemia virus (BLV) is a contagious, oncogenic deltaretrovirus of cattle with a worldwide distribution. In the US, over 40% of dairy cows are infected with the virus, and evidence of its economic impact is growing. This study evaluated the performance of a field-deployable automatic nucleic acid-extraction/insulated isothermal PCR (iiPCR) system for on-site BLV-proviral DNA detection in dairy cows compared with a conventional laboratory real-time PCR (rt-PCR). Assay performance was verified in parallel tests of 36 archived blood samples with 100% agreement (κ = 1.0; n = 36) between the iiPCR and conventional rt-PCR systems, and the limit of detection of the iiPCR assay was estimated to be 4 copies (genome equivalent) per reaction. The field-deployable iiPCR system was subsequently used on-farm to test freshly collected blood samples, and showed 100% agreement (κ = 1.0; n = 32) with the laboratory rt-PCR system. Fresh blood samples were collected on a second farm and tested on both systems, also with 100% agreement (κ = 1.0; n = 34). The field-deployable iiPCR/POCKIT™ combo system performs as well as a conventional laboratory-based rt-PCR system for detection of BLV proviral DNA in whole blood and may be a useful tool for on-farm evaluation of BLV-infection status in dairy cattle.

Chimeric Antigen Receptor T Cell-Mediated Neurotoxicity in Nonhuman Primates.

Chimeric antigen receptor (CAR) T-cell immunotherapy has revolutionized the treatment of refractory leukemias and lymphomas, but is associated with significant toxicities, namely cytokine release syndrome (CRS) and neurotoxicity. A major barrier to developing therapeutics to prevent CAR T cell-mediated neurotoxicity is the lack of clinically relevant models. Accordingly, we developed a rhesus macaque (RM) model of neurotoxicity via adoptive transfer of autologous CD20-specific CAR T cells. Following cyclophosphamide lymphodepletion, CD20 CAR T cells expand to 272 to 4,450 cells/µL after 7 to 8 days and elicit CRS and neurotoxicity. Toxicities are associated with elevated serum IL6, IL8, IL1RA, MIG, and I-TAC levels, and disproportionately high cerebrospinal fluid (CSF) IL6, IL2, GM-CSF, and VEGF levels. During neurotoxicity, both CD20 CAR and non-CAR T cells accumulate in the CSF and in the brain parenchyma. This RM model demonstrates that CAR T cell-mediated neurotoxicity is associated with proinflammatory CSF cytokines and a pan-T cell encephalitis.Significance: We provide the first immunologically relevant, nonhuman primate model of B cell-directed CAR T-cell therapy-mediated CRS and neurotoxicity. We demonstrate CAR and non-CAR T-cell infiltration in the CSF and in the brain during neurotoxicity resulting in pan-encephalitis, accompanied by increased levels of proinflammatory cytokines in the CSF. Cancer Discov; 8(6); 750-63. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 663.