Background/ObjectiveRetinal involvement in autoimmune polyendocrine syndrome type 1 (APS1), a rare monogenic autoimmune disorder caused by mutations in the AIRE gene, is increasingly recognised but remains poorly defined. Prior reports suggest a variable phenotype, ranging from mild changes to severe vision loss, often presumed untreatable. We explored the range of retinal phenotypes associated with AIRE gene deficiency in a multicentre case series of patients with APS1.MethodsWe performed a retrospective case note review of patients with molecularly confirmed APS1 from tertiary ophthalmic centres. Clinical history, multimodal retinal imaging, electrophysiology, genetic data, and treatment regimens were analysed. Histopathology was available in one case postmortem.ResultsRecords were reviewed from five unrelated female patients. Median age was 14 years at onset of ocular involvement and 33 years at most recent follow up. Some findings from two cases have been previously reported. Three distinct pathogenic AIRE variants contributing to biallelic genotypes were observed. Retinal findings ranged from structurally and functionally normal to advanced degeneration. One patient demonstrated sharp zonal atrophy on histopathology. Inflammatory features predominated in two cases, both showing durable vision preservation with periocular or systemic immunomodulation. One patient demonstrated four years of disease stabilisation with rituximab. No consistent genotype-phenotype correlation emerged.ConclusionAIRE-associated retinopathy encompasses a diverse spectrum, from clinically silent to profound degeneration. Early, targeted immunomodulation might preserve vision in selected cases. These findings advocate for ophthalmic surveillance in APS1, and support further investigation into predictive biomarkers and possible tailored immunotherapy in this vision-threatening autoimmune disorder.

Objective To evaluate and compare epidemiology, clinical and histopathologic features, and outcomes of conjunctival papilloma in historical and modern cohorts. Design Retrospective comparative cohort study. Participants Patients with conjunctival papilloma at a tertiary referral center between 2015 and 2024. Methods A retrospective review of Oncology and Pathology records was conducted to identify all patients with conjunctival papilloma over a 10-year period (2015–2024). Data collected included demographics, HPV vaccination status, clinical tumor characteristics, treatment modalities, outcomes, and histopathologic findings of excised lesions. Patients were stratified by age (≤20 vs.> 20 years). Findings were compared with a historical cohort managed at the same institution (1970–2012). Results Fifty-six conjunctival lesions were identified in 40 patients (mean age, 50 years; range, 8–89). Those older than 20-years-old demonstrated thicker tumors (4.6 vs. 2.2 mm, p = 0.03) with a predilection for the fornix (36%vs 0%; p = 0.02), whereas younger patients more commonly had bulbar lesions (46%vs 13%; p = 0.03). There was no difference in configuration (sessile vs. pedunculated, p > 0.99) between the age groups. The most common primary treatments included excision with cryotherapy (38%) or excision with cryotherapy plus interferon alfa-2b (38%). A combination of excision, cryotherapy and adjuvant oral cimetidine and interferon alfa-2b was significantly more common in the children and adolescent cohort compared to adults (56%vs. 0%, p < 0.01). Three tumors (9%) recurred following treatment. None of the lesions underwent malignant transformation. Compared with the historical cohort, patients in this study were older (mean 50 vs 43 years, p = 0.04), with a higher proportion of diffuse lesions (14%vs 3%, p < 0.01). Historically, most papillomas were treated with excision and cryotherapy alone (62%vs. 38%, p < 0.01), whereas, in our modern cohort there was a significantly increased use of excision and cryotherapy with adjuvant interferon alfa-2b (38%vs. 18%, p < 0.001). Conclusion This study demonstrates evolving demographic, clinical characteristics, and treatment trends in conjunctival papilloma over the past decade, including a shift toward an older patient population. Despite these changes, treatment outcomes remain favorable.

A 2-day-old boy presented with a large orbital mass, which showed radiological and pathological overlap between congenital cystic eye (CCE) and coloboma with cyst (CwC). Initially identified on prenatal imaging at 21 weeks, the patient underwent postnatal clinical and radiologic evaluation followed by histopathologic analysis of the enucleated eye. The coexistence of both anomalies within a single globe challenges the traditional view of CCE and CwC as distinct entities and supports a unifying theory of a developmental spectrum of early ocular malformations.

Purpose:Leiomyomas are benign smooth muscle tumors that commonly present in the uterus, soft tissue, skin, and gastrointestinal tract but in rare cases can also arise within the eye. Notably, intraocular leiomyomas often show slightly different histopathologic and immunohistochemical features, referred to as mesectodermal morphology, given their presumed neural crest origin. Genetic and cytogenetic alterations of intraocular leiomyomas, as well as their association with various clinical and histopathologic features, have not been previously studied. Methods:We identified eight patients diagnosed with intraocular leiomyoma and performed targeted next-generation sequencing, whole transcriptome RNA sequencing, and chromosomal copy number analysis on those with sufficient residual tissue. Results:Copy number analysis demonstrated multiple, recurrent whole-chromosome losses (including losses of 1, 2, 3, 10, 15q, 22q) suggestive of near haploidization of the genome. DNA sequencing did not reveal any pathogenic single-nucleotide variants, deep deletions, amplifications, or structural rearrangements. Whole-transcriptome RNA sequencing performed in a subset of cases did not show any pathogenic or likely pathogenic gene fusions. Tumors with mesenchymal versus mesectodermal morphology and those with or without histologically worrisome features did not show any differences in their copy number profile. Conclusions:Genetic and cytogenetic features of intraocular leiomyomas are different from the leiomyomas of uterine and deep soft tissues, as well as from uveal melanocytic tumors. Translational Relevance:Recurrent whole-chromosome losses in the absence of mutations or fusions are common among intraocular leiomyomas and may aid in the diagnosis.

PURPOSE:To report the clinical, pathologic, and genetic findings in a family with early-onset pterygia, corneal vascularization, and corneal myofibromatous lesions. METHODS:We performed clinical, pathologic, and genetic analysis of 12 members of a family originating in Puebla Mexico, who manifested with pterygia/pseudopterygia and corneal opacification transmitted in an autosomal-dominant inheritance pattern. Three unaffected family members also were evaluated. RESULTS:Clinical findings included isolated pterygia, isolated corneal subepithelial and anterior stromal opacities associated with varying degree of corneal vascularization, and a combination of pterygia and corneal opacities. Nine patients (17 eyes) underwent surgical procedures, including penetrating keratoplasty (9/17), superficial keratectomy (7/17), and pterygium excision (7/17). Five patients (eight eyes) had more than one surgery for recurrence of pterygia (2/8), recurrent corneal opacities obscuring the visual axis (3/8), penetrating keratoplasty failure (1/8), and indication not known (2/8). Documented recurrences occurred early, within 1 to 2 years of surgery. Histopathology of 21 specimens available for evaluation (nine penetrating keratoplasty corneas, seven superficial keratectomies, and five conjunctival-corneal tissues from pterygium/pseudopterygium excision) from seven patients showed varying degrees of myofibroblastic proliferation. Exome sequencing identified a heterozygous c.1610C>A (p.Ala537Asp) variant in the platelet-derived growth factor receptor beta ( PDGFRB ) gene in all 11 affected family members tested and in one of three unaffected family members. CONCLUSIONS:The combined clinical presentation, histopathologic features, and genetic findings suggest an autosomal dominantly inherited predisposition to exuberant corneal myofibroblastic proliferation, driven by the platelet-derived growth factor receptor activation.

OBJECTIVE:Ciliary body medulloepithelioma (CBME), a pediatric intraocular tumor with potential for locally aggressive behavior and metastasis, may present with a diverse spectrum of clinical and histopathologic features leading to diagnostic and management challenges. Examination of unusual CBME cases highlights challenges and modern diagnostic techniques which facilitate accurate diagnosis and guide management. METHODS:A retrospective clinicopathologic analysis of 6 patients with unusual clinical or pathologic features of CBME was performed. RESULTS:The mean duration of delay in accurate diagnosis was 5.7 years (SD: 8.2, median: 3, range: 0-22). All patients developed cataract, 4 (67%) were diagnosed with glaucoma, and 4 (67%) underwent surgery prior to accurate diagnosis. At initial presentation, only one patient with a known history of genetically confirmed DICER1 syndrome underwent appropriate imaging leading to a timely identification of a ciliary body mass and no delay in diagnosis. Following identification of intraocular mass, 4 (67%) patients underwent enucleation. Two patients (33%) underwent exenteration for extraocular extension of CBME. Initial histopathologic differential diagnosis included CBME, melanoma, adenoma or adenocarcinoma of the pigmented ciliary body epithelium, retinoblastoma, sarcoma, and malignant teratoma. Immunohistochemistry and genetic testing assisted in the diagnosis of CBME. Two patients (33%) had a germline DICER1 variant; this was known prior to CBME diagnosis in one patient and discovered after CBME diagnosis in the second patient. CONCLUSION:This series highlights the unusual clinical and histopathologic features of CBME that contribute to delays in diagnosis. Modern aids including genetic testing, ancillary imaging studies, and immunohistochemistry facilitate a timely accurate diagnosis of CBME and guide management.

Concentric macular rings (CMR) and Henle fiber layer (HFL) corrugations, potential clinical biomarkers of foveal hypoplasia, have been observed in laser fundus photographs and optical coherence tomography (OCT) images, respectively. Some believe these findings represent true anatomical structural changes while others hypothesize that they are artifacts of interference; however, to our knowledge, no previous study has provided evidence to support either theory. This retrospective case series analyzed CMR and OCT corrugations in 3 patients (6 eyes) with foveal hypoplasia. Dark fringe diameters of CMR at different wavelengths (λ: red = 635, green = 532, blue = 488 nm) and OCT corrugations of the first five dark fringes were measured. Dark fringe diameter was compared to fringe order (n) and λ using linear regression and t-tests. CMR fringe measurements demonstrated linear relationships between radius2 (r2) and fringe order at all wavelengths (mean R2: 0.949, SD: 0.022, range: 0.911–0.985). OCT corrugation measurements also exhibited strong linearity with fringe order (R2 = 0.998, 0.946). Fringe r2 increased with wavelength but exceeded theoretical predictions (p < 0.05), especially among ratios including shorter λ (mean difference r2 red:green = 0.104, green:blue = 0.292, red:blue = 0.468). This study is the first to provide evidence supporting an etiologic mechanism of CMR and HFL corrugations, showing correlation with physical phenomena of interference. CMR and HFL corrugations correlate with fringe order (n) and λ in a manner consistent with Newton’s rings (r2 = nR’λ, r = radius, R’ = radius of curvature), suggesting these phenomena are imaging artifacts. Further research validating these results will be important. This study does not report the results of a health care intervention on human participants. This study does not report results of a clinical trial.

PURPOSE:New treatments for advanced retinoblastoma (RB) have offered alternatives to primary enucleation. We assessed the impact of these therapies on the indications for enucleation and the histopathological findings in enucleated eyes with RB. MATERIALS AND METHODS:Eyes of all patients who underwent enucleation for RB at a single institution between January 2005 and August 2021 were included. Data collected retrospectively included demographics, clinical and pathologic staging, pathologic findings, and management. Statistical analysis included Kendall's τb, Pearson χ 2, and Cramér's V. RESULTS:There were 254 eyes from 252 patients with information available for review. Annual enucleations decreased between 2005 and 2008 at a rate of 4.2 enucleations/year, increased from 2008 to 2013 at a rate of 2.7 enucleations/year and decreased from 2013 to 2019 at a rate of 1.5 enucleations/year, reflecting changes in RB therapies. When compared to earlier years, the eyes enucleated in recent years were more likely to be enucleated for patient symptoms (P < 0.001) and insufficient view (P = 0.019), were more likely to have prior treatment (P < 0.001), had lower tumor stage (P = 0.010) and grade (P = 0.006), contained no viable tumor (P < 0.001), and were phthisical (P = 0.003). Five of 252 patients (2%) developed metastases; one of these patients had no viable tumor in a previously treated enucleated eye. CONCLUSION:Therapeutic innovations shifted the management of RB from primary enucleation in favor of eye salvage. Enucleated eyes show less viable tumor and disease severity but more intraocular degeneration, emphasizing the importance of skilled pathologic interpretation.

Supplementary Figure 3. Mouse retina expression of GPC2 and GD2, retinal CAR T-cell infiltration, systemic toxicities, and antigen loss in the intraocular model (see related main Fig. 4).

Purpose: The BRCA-associated protein1 (BAP1) immunohistochemical (IHC) stain has emerged as a powerful and inexpensive prognostic tool in uveal melanoma (UM), correlating with UM genetics and outcome. The data on the reliability of BAP1 immunohistochemistry in previously irradiated UM is scant. We aim to assess BAP1 IHC in post-Iodine-125 plaque brachytherapy-treated UM-enucleated eyes. Methods: In a case-control study, the medical records of all patients who underwent enucleation for UM at a major Ocular Oncology Service from December 1st, 2007 to December 31st, 2014 were reviewed. All cases with either chromosome 3 (ch3) status or sufficient follow-up (>5 years or metastasis) were selected. Nuclear BAP1 (nBAP1) immunoreactivity was interpreted as intact (positive in >90% of nuclei), lost (positive in <5% of nuclei), or heterogeneous (positive in 5–90% of nuclei). Retina and intratumoral blood vessels served as internal positive controls. Results: A comparison of 34 postbrachytherapy UM secondary-enucleated eyes with 47 nonbrachytherapy primary enucleated controls revealed no significant difference with respect to nBAP1 IHC (lost in 41% vs 51%, P = 0.19), ch3 status (ch3 monosomy in 59% vs 60%, P = 0.48), and outcome (metastatic disease in 44% vs 47%, P = 0.8). Association of nBAP1 IHC with ch3 status and outcome [intact nBAP1/(ch3 disomy and/or no metastasis) and lost nBAP1 (ch3 monosomy and/or metastasis)] in post-brachytherapy UM was significantly lower when compared with non-brachytherapy tumors [21/30 (70%) vs 41/44 (93%), P = 0.004*]. Conclusion: Although nBAP1 IHC stain is a strong prognostic tool in UM, its association with ch3 status, and outcome in postbrachytherapy UM was significantly lower compared with nonbrachytherapy tumors due to pitfalls in the interpretation of nBAP1 immunoreactivity in irradiated UM. This test should be used judiciously in the prognostication of postbrachytherapy-enucleated UM.

Several nomenclature and grading systems have been proposed for conjunctival melanocytic intraepithelial lesions (C-MIL). The fourth "WHO Classification of Eye Tumors" (WHO-EYE04) proposed a C-MIL classification, capturing the progression of noninvasive neoplastic melanocytes from low- to high-grade lesions, onto melanoma in situ (MIS), and then to invasive melanoma. This proposal was revised to the WHO-EYE05 C-MIL system, which simplified the high-grade C-MIL, whereby MIS was subsumed into high-grade C-MIL. Our aim was to validate the WHO-EYE05 C-MIL system using digitized images of C-MIL, stained with hematoxylin and eosin and immunohistochemistry. However, C-MIL cases were retrieved from 3 supraregional ocular pathology centers. Adequate conjunctival biopsies were stained with hematoxylin and eosin, Melan-A, SOX10, and PReferentially expressed Antigen in Melanoma. Digitized slides were uploaded on the SmartZoom platform and independently scored by 4 ocular pathologists to obtain a consensus score, before circulating to 14 expert eye pathologists for independent scoring. In total, 105 cases from 97 patients were evaluated. The initial consensus diagnoses using the WHO-EYE04 C-MIL system were as follows: 28 benign conjunctival melanoses, 13 low-grade C-MIL, 37 high-grade C-MIL, and 27 conjunctival MIS. Using this system resulted in 93% of the pathologists showing only fair-to-moderate agreement (kappa statistic) with the consensus score. The WHO-EYE05 C-MIL system (with high-grade C-MIL and MIS combined) improved consistency between pathologists, with the greatest level of agreement being seen with benign melanosis (74.5%) and high-grade C-MIL (85.4%). Lowest agreements remained between pathologists for low-grade C-MIL (38.7%). Regarding WHO-EYE05 C-MIL scoring and clinical outcomes, local recurrences of noninvasive lesions developed in 8% and 34% of the low- and high-grade cases. Invasive melanoma only occurred in 47% of the cases that were assessed as high-grade C-MIL. This extensive international collaborative study is the first to undertake a comprehensive review of the WHO-EYE05 C-MIL scoring system, which showed good interobserver agreement and reproducibility.

PURPOSE:To report the detailed histopathology of 2 enucleated eyes from 2 patients who developed severe visual loss associated with retinal hemorrhages, vessel sheathing, and vascular nonperfusion after administration of an initial dose of intravitreal pegcetacoplan, and propose, with supportive histopathology, the pathogenesis of the clinical syndrome previously termed hemorrhagic occlusive retinal vasculitis. DESIGN:Case series. SUBJECTS:Two enucleated eyes from 2 patients treated with intravitreal pegcetacoplan. METHODS:Retrospective, multicenter, consecutive clinical-pathologic analysis. MAIN OUTCOME MEASURES:Histopathologic review and immunophenotypic characterization. RESULTS:Both patients presented with inflammation and significant vision loss 9 days after the initial injection of pegcetacoplan with no subsequent improvement and underwent enucleation for pain control. Histologic examination of the enucleated eyes (patient 1 at 4 months postinjection and patient 2 at 40 days) revealed extensive vascular thrombosis, retinal hemorrhages and necrosis, and a dense inflammatory infiltrate in the uvea and, variably, the optic nerve, episclera, and muscle tendons composed of predominantly of T cells, macrophages, and eosinophils. Notably, the inflammatory infiltrate was absent from the retina. In addition, 1 eye demonstrated multiple foci of glomerular-like vascular proliferations in the uveal tract and thrombosis with focal recanalization of vessels in the optic nerve. CONCLUSIONS:Drug-induced, immune-mediated, retinal vasculopathy and choroiditis (DIRVAC) is a rare complication after pegcetacoplan injection. Although some limitations arise in interpretation of histopathologic findings because of compensatory changes in the eyes over time (before enucleation), the authors propose that the combined clinical, histopathologic, and immunohistochemical findings suggest a mixed-type, delayed hypersensitivity reaction as the mechanism of initial injury. FINANCIAL DISCLOSURE(S):Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

center dot PURPOSE: To compare the clinical outcomes of children with unilateral retinoblastoma (Rb) and high-risk histopathology features (HRHF) following upfront enucleation with/without adjuvant chemotherapy, and investigate cases locally considered non-HRHF but converted to a standardized HRHF definition. center dot DESIGN: Retrospective multinational clinical cohort study. center dot METHODS: Children with Rb who presented to 21 centers from 12 countries between 2011-2020, and underwent primary enucleation were recruited. Centers retrieved clinical data and were asked to report detailed histopathology findings, as well as indicate cases defined locally as high-risk. For analysis, only unilateral cases with standardized HRHF, defined as retrolaminar optic nerve invasion, massive choroidal invasion, scleral invasion, anterior-segment involvement, and/or combined nonmassive choroidal and prelaminar/laminar optic nerve invasion, were included. Main outcome measures included orbital tumor recurrence, systemic metastasis, survival and number, and outcome of cases converted to standardized HRHF. center dot RESULTS: A total of 600 children presenting to 14 centers in 9 countries were included. Of these, 505 (84.2%) were considered locally as HRHF and received adjuvant chemotherapy. After a median follow-up period of 39.2 +/- 1.6 months (range: 0.8-60.0 months), 36 (6.0%) had orbital tumor recurrence, 49 (8.2%) metastasis, and 72 (12.0%) children died. Children not receiving adjuvant chemotherapy were at significantly increased risk of orbital tumor recurrence, metastasis, and death ( P <= .002). Of the study children, 63/600 (10.5%) were considered locally non-HRHF, but converted to standardized HRHF and included in the analysis. Of these, 6/63 (9.5%) had orbital tumor recurrence, 5/63 (7.9%) metastasis, and 6/63 (9.5%) children died. Isolated minor choroidal invasion with prelaminar/laminar optic nerve invasion was reported in 114 (19.0%) children, but considered locally as HRHF only in 68/114 (59.6%). Of these, 6/114 (5.3%) children developed metastasis and subsequently died, yielding a number needed to treat of 15. center dot CONCLUSION: Based on this multinational cohort of children with Rb, we recommend the use of adjuvant chemotherapy following upfront enucleation and diagnosis of HRHF. Variation exists worldwide among centers when defining HRHF, resulting in adverse patient outcomes, warranting standardization. (Am J Ophthalmol 2024;268: 399-408. (c) 2024 Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Context.— Ophthalmic pathology is a discipline that relies heavily on a knowledge of clinical ophthalmology. The diagnosis of ocular and periocular lesions can be challenging because some lesions and diseases are unique to this region, whereas others may demonstrate site-specific differences from nonocular counterparts. Because of these challenges, ocular and periocular biopsies are frequently referred to specialized ophthalmic pathology centers for second-opinion diagnoses. Objective.— To analyze the referral patterns, diagnostic challenges, and diagnostic discrepancies for second-opinion referrals at a dedicated ophthalmic pathology laboratory with an emphasis on lesions of special interest in ophthalmic pathology. Data Sources.— Data sources included the pathology records of all slides and blocks received in consultation at the referral eye pathology center between December 1, 2015, and December 1, 2022, the personal experience of senior authors, and published peer-reviewed literature. Conclusions.— Corneal, intraocular, and conjunctival biopsies are the most common types of cases received in consultation without the referring pathologist's diagnosis, likely reflecting diagnostic challenges. Degenerative intraocular processes occasionally raise concern for a neoplasm. Conjunctival melanocytic lesions are the most common conjunctival biopsies referred for second-opinion diagnosis and require careful tissue sampling and clinical-pathologic correlation. Careful clinical-pathologic correlation, a high level of suspicion, and adequate sampling also are required for the accurate diagnosis of periocular sebaceous carcinoma. The diagnostic discrepancies involving uveal, retinal, conjunctival, eyelid, and temporal artery biopsies are most likely to adversely influence patient management and possible outcome. Such specimens may benefit from referral to specialized ophthalmic pathology laboratories.

Supplementary Figure 2. GPC2 CAR expression, activation, and polyfunctionality (see related main Fig. 3).

Ophthalmomyiasis is the result of fly larvae feeding on the tissues of the eye. Commonly associated with poor hygiene and open wounds, this condition is rare and often stigmatized. Treatment can be straightforward, and full recovery is common. Identifying the species responsible for ophthalmomyiasis is important for the medical, forensic, and entomological communities. Here, we present a case of ophthalmomyiasis where 30–40 blow fly (Diptera: Calliphoridae) larvae were removed from the eye of a human male. A representative subsample of five larvae was used for taxonomic identification via two approaches (a) DNA analysis, via sequencing of the complete mitochondrial genome (mtGenome) and comparison of the mtGenome and mitochondrial COI barcode region to GenBank, and (b) morphology, examination of the posterior spiracles using microscopy, and comparison to published larval descriptions of blow flies. Two species of blow flies were identified from the DNA analysis: Lucilia coeruleiviridis and Phormia regina. Morphological examination could only confirm L. coeruleiviridis as being present. To our knowledge, finding two blow fly species causing ophthalmomyiasis in a single individual has not been previously reported in the scientific literature. Neither P. regina nor L. coeruleiviridis prefers living tissue for larva development, but since they fill similar ecological niches, perhaps this was a show of competition rather than a normal feeding habit. Knowing these blow fly species can resort to this behavior, and that it can affect human populations, is valuable to the education of patients and providers.

Supplementary Figure 4. Mouse body weights and GPC2 expression in the brains of study endpoint mice included in the CNS efficacy study (see related main Fig. 5).

Purpose To report an atypical presentation of an epibulbar simple cartilaginous choristoma with a unique pigmented multicystic component. Case Description A 69-year-old African American female presented for evaluation of a right nasal epibulbar lesion that had progressed over the prior year. Slit-lamp evaluation revealed an immobile, mildly pigmented multicystic lesion measuring 6.0 × 4.5 mm that involved the nasal bulbar conjunctiva and the plica semilunaris. The lesion appeared benign, without feeder vessels or features of epithelial dysplasia. Given its recent growth and the patient's cosmetic concerns, the lesion was excised with ocular surface reconstruction. Histopathological evaluation disclosed a well-circumscribed nodule of well-differentiated cartilage in the substantia propria, consistent with a simple cartilaginous choristoma. The overlying conjunctival stroma contained multiple cysts lined by focally pigment epithelium. The patient recovered well from surgery, with satisfactory cosmetic results. Conclusions Our case of epibulbar simple cartilaginous choristoma includes a prominent superficial component of pigmented epithelial cysts, which has not been previously reported in the literature. This augments our knowledge on the spectrum of presentations of cartilaginous choristomas and underscores the importance of histopathological evaluation for definitive diagnosis.

Purpose: Retinoblastoma is the most common intraocular malignancy in children. Although new chemotherapeutic approaches have improved ocular salvage rates, novel therapies are required for patients with refractory intraocular and metastatic disease. Chimeric antigen receptor (CAR) T cells targeting glypican-2 (GPC2) are a potential new therapeutic strategy. Experimental Design: GPC2 expression and its regulation by the E2F1 transcription factor were studied in retinoblastoma patient samples and cellular models. In vitro, we performed functional studies comparing GPC2 CAR T cells with different costimulatory domains (4-1BB and CD28). In vivo, the efficacy of local and systemic administration of GPC2 CAR T cells was evaluated in intraocular and leptomeningeal human retinoblastoma xenograft models. Results: Retinoblastoma tumors, but not healthy retinal tissues, expressed cell surface GPC2, and this tumor-specific expression was driven by E2F1. GPC2-directed CARs with 4-1BB costimulation (GPC2.BBz) were superior to CARs with CD28 stimulatory domains (GPC2.28z), efficiently inducing retinoblastoma cell cytotoxicity and enhancing T-cell proliferation and polyfunctionality. In vivo, GPC2.BBz CARs had enhanced persistence, which led to significant tumor regression compared with either control CD19 or GPC2.28z CARs. In intraocular models, GPC2.BBz CAR T cells efficiently trafficked to tumor-bearing eyes after intravitreal or systemic infusions, significantly prolonging ocular survival. In central nervous system (CNS) retinoblastoma models, intraventricular or systemically administered GPC2.BBz CAR T cells were activated in retinoblastoma-involved CNS tissues, resulting in robust tumor regression with substantially extended overall mouse survival. Conclusions: GPC2-directed CAR T cells are effective against intraocular and CNS metastatic retinoblastomas.