Refractive outcomes using Barrett formulas and patient characteristics of cataract surgery patients with and without prior LASIK/PRK.

PURPOSE: The goal of this study is to describe characteristics of cataract surgery patients who previously underwent laser in situ keratomileusis/photorefractive keratectomy (LASIK/PRK) in comparison to non-LASIK/PRK cataract surgery patients including psychiatric comorbidities, as well as describe refractive prediction error after cataract surgery while accounting for axial length (AL) using the Barrett True-K and Barrett Universal II formulas. METHODS: This was a retrospective study of patients from the University of Colorado Cataract Outcomes Registry. The primary outcomes were refraction prediction error (RPE), mean absolute RPE, and median absolute RPE. Outcomes were stratified by five axial length groups. Univariate and multivariate models for RPE were stratified by the AL group. RESULTS: Two hundred eighty-one eyes with prior LASIK/PRK and 3101 eyes without are included in the study. Patients with prior LASIK/PRK were significantly younger: 67.0 vs 69.9 years, p < 0.0001. The LASIK/PRK group had significantly better mean pre-operative BCVA in comparison to the non-LASIK group, logMAR 0.204 vs logMAR 0.288, p = 0.003. The LASIK/PRK group had significantly lower rates of cardiovascular disease (18.5% vs 29.3%, p < 0.001), hypertension (49.1% vs 59.3%, p < 0.012), and type 2 diabetes (10.7% vs 26.0%, p < 0.001), and no significant difference in psychiatric disease. The absolute RPE was higher for the LASIK group for all ALs, but only significantly higher for eyes with AL less than 25 mm. CONCLUSION: Patient eyes with prior LASIK/PRK surgery undergoing cataract surgery were significantly younger, had significantly less comorbidities, and a significantly better pre-operative BCVA. Using the Barrett formulas, absolute prediction error for eyes with longer ALs was not significantly worse for LASIK/PRK eyes than those without and the difference was smaller for eyes with longer AL.

Association Between Quantitative and Qualitative Imaging Biomarkers and Geographic Atrophy Growth Rate.

PURPOSE: Investigate associations between geographic atrophy (GA) growth rate and multimodal imaging biomarkers and patient demographics in patients with advanced non-neovascular age-related macular degeneration (nnAMD). DESIGN: Prospective cohort study. METHODS: 121 eyes of 66 patients with advanced nnAMD with GA enrolled in the University of Colorado AMD Registry from August 2014 to June 2021, with follow-up through June 2023. Multimodal images were reviewed by two graders for imaging biomarkers at enrollment. GA growth rate and square-root transformed (SQRT) GA growth rate were measured between enrollment and final visit. Associations between the outcome SQRT GA growth rate and imaging biomarkers, baseline GA lesions characteristics, and patient demographics were evaluated. RESULTS: Average GA growth rate was 1.430mm2/year and SQRT GA growth rate was 0.268mm/year over a mean of 3.7years. SQRT GA growth rate was positively associated with patient age (p=0.010) and female sex (0.035), and negatively associated with body mass index (0.041). After adjustment for these demographic factors, SQRT GA growth rate was positively associated with presence of non-exudative subretinal fluid (p<0.001), non-exudative subretinal hyperreflective material (p=0.037), and incomplete retinal pigment epithelium and outer retina atrophy (p=0.022), and negatively associated with subfoveal choroidal thickness (p=0.031) and presence of retinal pseudocysts (p=0.030). Larger baseline GA size at enrollment was associated with faster GA growth rate (p=0.002) but not SQRT GA growth rate. CONCLUSIONS: Select patient demographic factors and basic clinically-relevant imaging biomarkers were associated with GA growth rate. These biomarkers may guide patient selection when considering treating GA patients with novel therapeutics.

Using Natural Language Processing to Identify Different Lens Pathology in Electronic Health Records.

PURPOSE: Nearly all published ophthalmology-related Big Data studies rely exclusively on International Classification of Diseases (ICD) billing codes to identify patients with particular ocular conditions. However, inaccurate or nonspecific codes may be used. We assessed whether natural language processing (NLP), as an alternative approach, could more accurately identify lens pathology. DESIGN: Database study comparing the accuracy of NLP versus ICD billing codes to properly identify lens pathology. METHODS: We developed an NLP algorithm capable of searching free-text lens exam data in the electronic health record (EHR) to identify the type(s) of cataract present, cataract density, presence of intraocular lenses, and other lens pathology. We applied our algorithm to 17.5 million lens exam records in the Sight Outcomes Research Collaborative (SOURCE) repository. We selected 4314 unique lens-exam entries and asked 11 clinicians to assess whether all pathology present in the entries had been correctly identified in the NLP algorithm output. The algorithm's sensitivity at accurately identifying lens pathology was compared with that of the ICD codes. RESULTS: The NLP algorithm correctly identified all lens pathology present in 4104 of the 4314 lens-exam entries (95.1%). For less common lens pathology, algorithm findings were corroborated by reviewing clinicians for 100% of mentions of pseudoexfoliation material and 99.7% for phimosis, subluxation, and synechia. Sensitivity at identifying lens pathology was better for NLP (0.98 [0.96-0.99] than for billing codes (0.49 [0.46-0.53]). CONCLUSIONS: Our NLP algorithm identifies and classifies lens abnormalities routinely documented by eye-care professionals with high accuracy. Such algorithms will help researchers to properly identify and classify ocular pathology, broadening the scope of feasible research using real-world data.

Are there sex-based disparities in cataract surgery?

AIM: To investigate sex-based differences in the occurrence of intra-operative and post-operative complications and associated visual outcomes following cataract surgery. METHODS: This was a retrospective study of patients who had phacoemulsification cataract surgery at the University of Colorado School of Medicine. Data collected included the patient's health history, ocular comorbidities, operative and post-operative complications, and the post-operative best corrected visual acuity (BCVA). The data were analyzed using univariate and multivariable logistic regression with generalized estimating equations to account for the correlation of some patients having two eyes included in the study. RESULTS: A total of 11 977 eyes from 7253 patients were included in the study. Ocular comorbidities differed by sex, with males having significantly higher percentages of traumatic cataracts (males 0.7% vs females 0.1%), prior ocular surgery (6.7% vs 5.5%), and mature cataracts (2.8% vs 1.9%). Conversely, females had significantly higher rates of pseudoexfoliation (2.0% vs 3.2%). In unadjusted analysis, males had higher rates of posterior capsular rupture (0.8% vs 0.4%) and vitreous loss (1.0% vs 0.6%), but this difference was not significant after adjustment for confounders. Males had a significantly increased risk of post-operative retinal detachment, but in multivariable analysis this was no longer significant. Males were significantly less likely to undergo post-operative neodymium-doped yttrium aluminum garnet (Nd:YAG) laser capsulotomy for posterior capsule opacification (OR=0.8, 95%CI=0.7-0.9, P=0.0005). The BCVA was slightly worse for males pre-operatively; but post-operatively, both sexes exhibited similar visual acuity of Snellen equivalent 20/25. CONCLUSION: The study finds that in a cohort of patients presenting for cataract surgery, sex differences exist in pre-operative comorbidities and surgical characteristics that contribute to higher rates of some complications for males. However, observed surgical complication rates exhibit almost no difference by sex after adjusting for pre-operative differences and post-operative BCVA is similar between sexes.

Rates of diabetic retinopathy among cluster analysis-identified type 2 diabetic mellitus subgroups.

PURPOSE: To determine whether phenotypic clustering of patients with diabetes mellitus (DM) is associated with more advanced diabetic retinopathy (DR). METHODS: Retrospective cohort study of 495 patients with no prior DR treatment seen at a tertiary care clinic 2014-2020. Four previously identified clusters from Ahlqvist's 2018 paper were reproduced utilizing baseline hemoglobin A1c, body mass index, and age at DM diagnosis. Age-adjusted Cox proportional hazard ratios were used to compare clusters with reference as the lowest risk cluster. RESULTS: All four type 2 DM clusters were replicated with our cohort. There was a significant difference in racial distribution among clusters (p = 0.018) with severe insulin-resistant diabetes (SIRD) having the higher percentage of Caucasians and lower percentage of Hispanics compared to other groups and a higher percentage of African Americans comprising the severe insulin-deficient diabetes (SIDD) cluster than other groups. Rates of proliferative diabetic retinopathy were higher in mild obesity-related diabetes (MOD) (28%), SIDD (24%), mild age-related diabetes (MARD) (20%), and lowest in SIRD (7.9%), overall p = 0.004. Rates of vitreous hemorrhage were higher in MOD (p = 0.032) and MARD (0.005) compared to SIRD. CONCLUSION: Baseline clinical measures may be useful in risk stratifying patients for progression to retinopathy requiring intervention.

Cataract Phacoemulsification in People with Dementia: Characterization and Outcomes.

PURPOSE: To characterize cataract surgery in people with dementia (PWD) using a cataract surgery outcomes database. METHODS: Demographics, medical and ocular history, surgical characteristics, and postoperative measures were analyzed for differences between PWD and non-PWD cohorts. Patient-level data were analyzed with Fisher's Exact Test, and eye-level data were analyzed with logistic regression using generalized estimating equations to account for correlation of eyes from the same individual. RESULTS: 507 eyes from 296 PWD were identified using appropriate ICD codes and cross-referenced to a cataract surgery outcomes database containing 12,949 eyes from 7,853 patients who underwent cataract phacoemulsification at a single center between January 2014 and October 2019. PWD were older (p < .001), had shorter duration cataract surgeries (p = .006), and were more likely to have mature cataract (p = .017). The rate of general anesthesia was higher in PWD (p = .005). There were no differences in complication rates between PWD and non-PWD cohorts. Both preoperative best corrected LogMAR distance visual acuity (CDVA) (p < .001) and postoperative CDVA (p < .001) were worse in PWD. CDVA significantly improved in both groups (p < .001); however, the average magnitude of improvement in CDVA was not significantly different between groups (p = .169). CONCLUSIONS: PWD present for cataract surgery at a later age and were more likely to have mature cataracts and general anesthesia, but did not have higher rates of complication, and showed significant improvement in CDVA following surgery. These findings should be encouraging to PWD undergoing counseling for cataract surgery, and for the potential for improved function in PWD.

Limited English Proficiency Is Associated With Diabetic Retinopathy in Patients Presenting for Cataract Surgery.

Purpose: To investigate the relationship between limited English proficiency (LEP) and diabetic retinopathy (DR) in patients presenting for cataract surgery. Methods: This is a retrospective observational study of patients who underwent cataract surgery between January 2014 and February 2020. Patients who self-identified as needing or preferring an interpreter were defined as having LEP. Differences in demographics, characteristics, and outcomes including history of type 2 diabetes (T2DM), DR, preoperative best corrected visual acuity (BCVA), macular edema, and anti-vascular endothelial growth factor injections were analyzed. Statistical comparisons were assessed using logistic regression with generalized estimating equations. Results: We included 13,590 eyes. Of these, 868 (6.4%) were from LEP patients. Patients with LEP were more likely to be Hispanic (P < 0.001), female sex (P = 0.008), or older age (P = 0.003) and have worse mean BCVA at presentation (P < 0.001). Patients with LEP had a significantly higher rate of T2DM (P < 0.001), macular edema (P = 0.033), and DR (18.1% vs. 5.8%, P < 0.001). Findings remained significant when controlling for age, sex, race/ethnicity, and type of health insurance. Patients with LEP and DR were more likely to have had later stages of DR (P = 0.023). Conclusions: Patients with LEP presenting for cataract surgery had a higher rate of DR and associated complications compared to patients with English proficiency. Further studies are needed to understand how language disparities influence health and what measures could be taken to improve healthcare in this vulnerable population. Translational Relevance: Our study highlights healthcare disparities within ophthalmology and emphasizes the importance of advocating for improved healthcare delivery for patients with LEP.

A Randomized Prospective Trial Comparing Repository Corticotropin Injection and Intravenous Methylprednisolone for Neuroprotection in Acute Optic Neuritis.

BACKGROUND: Repository corticotrophin injection (RCI, Acthar Gel) and intravenous methylprednisolone (IVMP) improve the rate but not the extent of visual recovery following acute optic neuritis. RCI has adrenal-stimulating and melanocortin receptor-stimulating properties that may endow it with unique anti-inflammatory properties relative to IVMP. METHODS: Individuals with acute optic neuritis of less than 2 weeks duration were prospectively enrolled and randomized 1:1 to receive either RCI or IVMP. Peripapillary retinal nerve fiber layer (pRNFL) and ganglion cell plus inner plexiform layer thickness (GC + IPL) were serially evaluated by OCT. In addition, patient-reported outcomes (PROs) for changes in fatigue, mood, visual function, depression, and quality of life (QOL) were measured, and high and low contrast visual acuity were recorded. RESULTS: Thirty-seven subjects were enrolled (19 RCI; 18 IVMP); the average time from symptom to treatment was 8.8 days. At 6 months, there was no difference in the primary outcome: loss of average pRNFL thickness in the affected eye (RCI vs IVMP: -13.1 vs -11.7 µm, P = 0.88) 6 months after randomization. Additional outcomes also showed no difference between treatment groups: 6-month attenuation of GC + IPL thickness (RCI vs IVMP: -13.8 vs -12.0 µm, P = 0.58) and frequency of pRNFL swelling at 1 month (RCI vs IVMP: 63% vs 72%, P = 0.73) and 3 months (RCI vs IVMP: 26% vs 31%, P = 0.99). Both treatments resulted in improvement in visual function and PROs. CONCLUSIONS: Treatment of acute optic neuritis with RCI or IVMP produced no clinically meaningful differences in optic nerve structure or visual function.

Comparison of cataract surgery outcomes in English proficient and limited English proficiency patients.

PURPOSE: To determine differences in cataract surgery outcomes between English proficient (EP) and limited English proficiency (LEP) patients. SETTING: Sue Anschutz-Rodgers Eye Center, Aurora, Colorado. DESIGN: Retrospective. METHODS: Patients who underwent phacoemulsification at the Sue Anschutz-Rogers Eye Center between January 2014 and February 2020 were included. Patients who self-identified as needing or preferring an interpreter in medical encounters were defined as LEP. Differences in surgical characteristics and outcomes including cataract maturity, surgical complexity, and surgical complications were analyzed. RESULTS: 868 eyes (6.4%) were identified from LEP patients. LEP patients were more likely to have mature cataracts (5.1% vs 2.3%, P < .0001). LEP patients' surgeries were more likely to be considered complex (27.8% vs 15.3%, P < .0001) and use higher cumulative dissipated energy (mean of 9.5 [SD = 9.5] vs 7.2 [SD = 7.1], P < .0001). Preoperative visual acuity was worse in LEP patients (logMAR 0.566 [SD = 0.64] vs 0.366 [SD = 0.51], P < .0001) but showed greater improvement after surgery (logMAR 0.366 [SD = 0.54] vs 0.254 [SD = 0.41], P < .0001). There were no significant differences in operative time, intraoperative or postoperative complications. More LEP patients were on steroids 4 weeks postoperatively when compared with EP patients (14.6% vs 10.1%, P < .0002). LEP patients were less likely to undergo subsequent YAG capsulotomy (7.3% vs 12.8%, P < .0001). CONCLUSIONS: Disparities in cataract outcomes between EP and LEP patients was demonstrated. Further research into ophthalmic health disparities for LEP patients is needed to understand the root causes and how they can be addressed.

Predictors of Pseudophakic Retinal Tears at a Tertiary Care Academic Medical Center.

PURPOSE: To describe the incidence and identify risk factors for retinal tears (RTs) among patients who underwent cataract phacoemulsification surgery and to describe the timing and features of the RTs. DESIGN: Retrospective case-control study. SUBJECTS: Patients who underwent cataract phacoemulsification surgery between January 2014 and December 2019 at an academic eye center in Colorado. METHODS: Information was extracted from medical chart review into a cataract outcomes database. Measures of association and P values were determined from logistic regression models with generalized estimating equations to account for some patients having 2 eyes included in the database. MAIN OUTCOME MEASURES: The diagnosis of RTs within 1 year after cataract surgery. The incidences of RTs were analyzed by demographics, comorbid medical history, ocular characteristics, surgical characteristics, and intraoperative complications. RESULTS: In total, 13 007 cataract surgeries were included in the analysis, of which 79 (0.6%) eyes developed RTs within 1 year after cataract surgery. Men and patients aged <65 years had higher incidences of RTs (0.8% vs. 0.5% [men vs. women], P = 0.0175; 1.0% vs. 0.2% [age <65 years vs. age >75 years], P = 0.0001). The mean preoperative best-corrected visual acuity was better for the RT group (logarithm of the minimum angle of resolution 0.252 vs. 0.366, P = 0.0073). Patients with RTs had a higher rate of pre-operative high myopia (<=-6.0D) (1.4%) in comparison with patients with low myopia (<-1D and >-6D) (0.7%), emmetropia (<1.0D and >-1.0D) (0.4%), and hyperopia (>1.0D) (0.2%), P = 0.0006. Patients with intraoperative complications had a higher incidence of RTs (4.2%, P < 0.0001). The mean time from cataract surgery to RTs was 128 (standard deviation, 110) days. Most RTs were symptomatic (57.0%), located in the superior hemisphere (83.5%), and 40.5% were diagnosed concurrently with a retinal detachment. The multivariable model resulted in intraoperative complications, axial length, and preoperative visual acuity having the strongest association with RT. CONCLUSIONS: Patients with better preoperative visual acuity, eyes with longer axial length, and eyes with intraoperative complications were at an increased risk of RTs after cataract surgery. Careful counseling is important, and postoperative follow-up is advised for patients identified preoperatively as being at an elevated risk of RTs, given the large rate of asymptomatic RTs and high rate of progression to retinal detachment.

Gene Editing with Helper-Dependent Adenovirus Can Efficiently Introduce Multiple Changes Simultaneously over a Large Genomic Region.

Helper-dependent adenoviral vectors (HDAds) possess long homology arms that mediate high-efficiency gene editing. These long homology arms may permit simultaneous introduction of multiple modifications into a large genomic region or may permit a single HDAd to correct many different individual mutations spread widely across a gene. We investigated this important potential using an HDAd bearing 13 genetic markers in the region of homology to the target CFTR locus in human iPSCs and found that all markers can be simultaneously introduced into the target locus, with the two farthest markers being 22.2 kb apart. We found that genetic markers closer to the HDAd's selectable marker are more efficiency introduced into the target locus; a marker located 208 bp from the selectable marker was introduced with 100% efficiency. However, even markers 11 kb from the selectable marker were introduced at a relatively high frequency of 21.7%. Our study also revealed extensive heteroduplex DNA formation of up to 10 kb with no bias toward vector or chromosomal repair. However, mismatches escape repair at a frequency of up to 15%, leading to a genetically mixed colony and emphasizing the need for caution, especially if the donor and target sequences are not 100% homologous.

Homology Requirements for Efficient, Footprintless Gene Editing at the CFTR Locus in Human iPSCs with Helper-dependent Adenoviral Vectors.

Helper-dependent adenoviral vectors mediate high efficiency gene editing in induced pluripotent stem cells without needing a designer nuclease thereby avoiding off-target cleavage. Because of their large cloning capacity of 37 kb, helper-dependent adenoviral vectors with long homology arms are used for gene editing. However, this makes vector construction and recombinant analysis difficult. Conversely, insufficient homology may compromise targeting efficiency. Thus, we investigated the effect of homology length on helper-dependent adenoviral vector targeting efficiency at the cystic fibrosis transmembrane conductance regulator locus in induced pluripotent stem cells and found a positive correlation. With 23.8 and 21.4 kb of homology, the frequencies of targeted recombinants were 50-64.6% after positive selection for vector integration, and 97.4-100% after negative selection against random integrations. With 14.8 kb, the frequencies were 26.9-57.1% after positive selection and 87.5-100% after negative selection. With 9.6 kb, the frequencies were 21.4 and 75% after positive and negative selection, respectively. With only 5.6 kb, the frequencies were 5.6-16.7% after positive selection and 50% after negative selection, but these were more than high enough for efficient identification and isolation of targeted clones. Furthermore, we demonstrate helper-dependent adenoviral vector-mediated footprintless correction of cystic fibrosis transmembrane conductance regulator mutations through piggyBac excision of the selectable marker. However, low frequencies (≤ 1 × 10-3) necessitated negative selection for piggyBac-excision product isolation.

Helper virus-mediated downregulation of transgene expression permits production of recalcitrant helper-dependent adenoviral vector.

Helper-dependent adenoviral vectors (HDAd) that express certain transgene products are impossible to produce because the transgene product is toxic to the producer cells, especially when made in large amounts during vector production. Downregulating transgene expression from the HDAd during vector production is a way to solve this problem. In this report, we show that this can be accomplished by inserting the target sequence for the adenoviral VA RNAI into the 3' untranslated region of the expression cassette in the HDAd. Thus during vector production, when the producer cells are coinfected with both the helper virus (HV) and the HDAd, the VA RNAI produced by the HV will target the transgene mRNA from the HDAd via the endogenous cellular RNAi pathway. Once the HDAd is produced and purified, transduction of the target cells results in unimpeded transgene expression because of the absence of HV. This simple and universal strategy permits for the robust production of otherwise recalcitrant HDAds.

Transgene expression up to 7 years in nonhuman primates following hepatic transduction with helper-dependent adenoviral vectors.

Helper-dependent adenoviral vectors (HDAd) have been shown to mediate a considerably longer duration of transgene expression than first-generation adenoviral vectors. We have previously shown that transgene expression from HDAd-transduced hepatocytes can persist at high levels for up to 2.6 years in nonhuman primates following a single-vector administration. Because duration of transgene expression and long-term toxicity are critical for risk:benefit assessment, we have continued to monitor these animals. We report here that transgene expression has persisted for the entire observation period of up to 7 years for all animals without long-term adverse effects. However, in all cases, transgene expression level slowly declined over time to less than 10% of peak values by the end of the observation period but remained 2.3-111-fold above baseline values. These results will provide important information for a more informed risk:benefit assessment before clinical application of HDAd.

SR-A and SREC-I are Kupffer and endothelial cell receptors for helper-dependent adenoviral vectors.

Helper-dependent adenoviral (HDAd) vectors can mediate long-term, high-level transgene expression from transduced hepatocytes with no chronic toxicity. However, a toxic acute response with potentially lethal consequences has hindered their clinical applications. Liver sinusoidal endothelial cells (LSECs) and Kupffer cells are major barriers to efficient hepatocyte transduction. Understanding the mechanisms of adenoviral vector uptake by non-parenchymal cells may allow the development of strategies aimed at overcoming these important barriers and to achieve preferential hepatocyte gene transfer with reduced toxicity. Scavenger receptors on Kupffer cells bind adenoviral particles and remove them from the circulation, thus preventing hepatocyte transduction. In the present study, we show that HDAd particles interact in vitro and in vivo with scavenger receptor-A (SR-A) and with scavenger receptor expressed on endothelial cells-I (SREC-I) and we exploited this knowledge to increase the efficiency of hepatocyte transduction by HDAd vectors in vivo through blocking of SR-A and SREC-I with specific fragments antigen-binding (Fabs).

Balloon catheter delivery of helper-dependent adenoviral vector results in sustained, therapeutic hFIX expression in rhesus macaques.

Hemophilia B is an excellent candidate for gene therapy because low levels of factor IX (FIX) (≥1%) result in clinically significant improvement of the bleeding diathesis. Helper-dependent adenoviral (HDAd) vectors can mediate long-term transgene expression without chronic toxicity. To determine the potential for HDAd-mediated liver-directed hemophilia B gene therapy, we administered an HDAd expressing hFIX into rhesus macaques through a novel and minimally invasive balloon occlusion catheter-based method that permits preferential, high-efficiency hepatocyte transduction with low, subtoxic vector doses. Animals given 1 × 10(12) and 1 × 10(11) virus particle (vp)/kg achieved therapeutic hFIX levels for the entire observation period (up to 1,029 days). At 3 × 10(10) and 1 × 10(10) vp/kg, only subtherapeutic hFIX levels were achieved which were not sustained long-term. Balloon occlusion administration of HDAd was well tolerated with negligible toxicity. Five of six animals developed inhibitors to hFIX. These results provide important information in assessing the clinical utility of HDAd for hemophilia B gene therapy.

Generation of a Kupffer cell-evading adenovirus for systemic and liver-directed gene transfer.

As much as 90% of an intravenously (i.v.) injected dose of adenovirus serotype 5 (Ad5) is absorbed and destroyed by liver Kupffer cells. Viruses that escape these cells can then transduce hepatocytes after binding factor X (FX). Given that interactions with FX and Kupffer cells are thought to occur on the Ad5 hexon protein, we replaced its exposed hypervariable regions (HVR) with those from Ad6. When tested in vivo in BALB/c mice and in hamsters, the Ad5/6 chimera mediated >10 times higher transduction in the liver. This effect was not due to changes in FX binding. Rather, Ad5/6 appeared to escape Kupffer cell uptake as evidenced by producing no Kupffer cell death in vivo, not requiring predosing in vivo, and being phagocytosed less efficiently by macrophages in vitro compared to Ad5. When tested as a helper-dependent adenovirus (Ad) vector, Ad5/6 mediated higher luciferase and factor IX transgene expression than either helper-dependent adenoviral 5 (HD-Ad5) or HD-Ad6 vectors. These data suggest that the Ad5/6 hexon-chimera evades Kupffer cells and may have utility for systemic and liver-directed therapies.

Modifications of adenovirus hexon allow for either hepatocyte detargeting or targeting with potential evasion from Kupffer cells.

In vivo gene transfer with adenovirus vectors would significantly benefit from a tight control of the adenovirus-inherent liver tropism. For efficient hepatocyte transduction, adenovirus vectors need to evade from Kupffer cell scavenging while delivery to peripheral tissues or tumors could be improved if both scavenging by Kupffer cells and uptake by hepatocytes were blocked. Here, we provide evidence that a single point mutation in the hexon capsomere designed to enable defined chemical capsid modifications may permit both detargeting from and targeting to hepatocytes with evasion from Kupffer cell scavenging. Vector particles modified with small polyethylene glycol (PEG) moieties specifically on hexon exhibited decreased transduction of hepatocytes by shielding from blood coagulation factor binding. Vector particles modified with transferrin or, surprisingly, 5,000 Da PEG or dextran increased hepatocyte transduction up to 18-fold independent of the presence of Kupffer cells. We further show that our strategy can be used to target high-capacity adenovirus vectors to hepatocytes emphasizing the potential for therapeutic liver-directed gene transfer. Our approach may lead to a detailed understanding of the interactions between adenovirus vectors and Kupffer cells, one of the most important barriers for adenovirus-mediated gene delivery.