Preliminary evidence of CAR T-cell therapy activity in vitreoretinal lymphomas: An LOC network study

Vitreoretinal lymphoma (VRL) is a rare subtype of large B-cell lymphoma (LBCL) that may present as an isolated condition or with central nervous system (CNS) or systemic involvement. Like the brain, the eye is an immune-privileged site, posing unique therapeutic challenges according to the blood–eye barrier. Despite its typically indolent course, VRL is difficult to eradicate, and the long-term prognosis is poor because of the high risk of CNS involvement.¹ The median overall survival (OS) is 36–75 months for primary VRL and approximately 57 months for isolated vitreoretinal relapses of primary CNS lymphomas.² Therefore, new therapeutic strategies are urgently needed.

Anti-CD19 chimeric antigen receptor (CAR) T-cells represent a major advancement in managing systemic LBCL.^{3, 4} Studies show that CAR T-cells can cross the blood–brain barrier and demonstrate high efficacy in CNS lymphomas,^5-8 but their potential in VRL treatment is not fully understood, and their efficiency has never been described from an ophthalmologic point of view with the newest validated exams.

Since 2020, CAR T-cell therapy has been considered within the French oculocerebral lymphoma network (LOC network) for relapsed or refractory VRL. This study retrospectively analyzed patients with primary or secondary VRL treated with CAR T-cells.

We retrospectively identified all VRL patients treated with commercial anti-CD19 CAR T-cells at Pitié-Salpêtrière Hospital (Paris) via the LOC network database until November 2023. Eligible patients had active vitreoretinal involvement, either isolated or with CNS involvement. VRL could be primary or secondary to systemic or CNS lymphoma. Data were retrospectively collected from medical records between April and October 2024. The LOC database was approved by the Institutional Ethical Committee of the coordinating center and the French “Commission Nationale de l'Informatique et des Libertés” (CNIL). All patients provided written informed consent. Responses were evaluated using International Primary CNS Lymphoma Collaborative Group (IPCG) criteria,⁹ and all ophthalmologic examinations of the patient performed immediately before and after the CAR T-cells were systematically reviewed by a single ophthalmologist from the Pitié-Salpétrière Hospital, based on the photographs taken at each consultation (Supplementary Methods). Progression-free survival (PFS), ophthalmic-free survival (OFS), and OS were defined as the time from CAR T-cell infusion: “to CNS, ophthalmic or systemic disease progression or relapse/to ophthalmic progression or relapse/and to death respectively.” Survival rates were calculated using the Kaplan–Meier method. Statistical analyses were conducted using GraphPad Prism v10.0. Cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome (ICANS) were graded per 2019 American Society for Transplantation and Cellular Therapy (ASTCT) guidelines.¹⁰

Interleukin (IL)-10 and IL-6 assays were performed on the aqueous humor (AH) using a sensitive Cytometric Bead Array kit (BD Biosciences) on a FACSCanto II cytometer. The AH IL-10 level was classified as detectable (≥2.5 pg/mL) or undetectable (<2.5 pg/mL) based on the laboratory's standard threshold. The presence of CAR T-cells in the blood and AH was measured by flow cytometry. Each sample was incubated for 15 min with 1 µL of CD19 CAR Detection Reagent (Miltenyi Biotec), followed by staining with the following antibodies: anti-Biotin-PE (Miltenyi Biotec), CD45-KrO (clone J33), CD3-Alexa Fluor A750 (clone UCHT1), CD4-ECD (clone SFCI12T4D11), and CD8-Alexa Fluor 700 (clone B9.11). The final sample was analyzed on a DxFLEX flow cytometer (Beckman Coulter), and data were processed using CytExpert (Beckman Coulter) software version 2.0.2.18.

Between August 2020 and December 2023, seven patients with VRL were treated with CAR T-cells. Their main baseline characteristics and individual outcomes are summarized in Tables 1 and S1.

At the time of the CAR T-cell therapy decision, four patients had isolated VRL, whereas three also had brain parenchymal involvement. Their median age was 70 years (range, 50–76), and their median Eastern Cooperative Oncology Group performance status was 1 (range, 1–3). The median number of prior therapies before leukapheresis was 4 (range, 2–6), including autologous stem cell transplantation (ASCT) (thiotepa-based induction regimen) in four patients (two patients were ineligible for ASCT at the time of CAR T-cells). All but one patient received bridging therapy using drug regimens directed against ocular/brain disease such as methotrexate, temozolomide, ibrutinib, or cytotoxic chemotherapies (ifosfamide, etoposide, and carboplatin) (Table 1). There were no steroid-containing regimens.

At the time of CAR T-cell infusion, with regard to ophthalmic involvement, two patients had a complete response (CR), two had a partial response (PR), two had stable disease, and one had progressive disease. Additionally, three patients had CNS progressive disease.

Three patients received tisagenlecleucel and four received axicabtagene ciloleucel (axi-cel). At 1 month, an ophthalmologic response was observed in all five evaluable patients, including PR in two (40%) and CR in three (60%). Data were unavailable for two patients who developed severe neurotoxicity and could not undergo ophthalmologic assessment at this time (Table S1). During follow-up, the best response achieved was CR or CR unconfirmed for all seven patients. The median time from CAR T-cell infusion to first response was 1 month (range, 1–4). After a median follow-up of 23 months (range, 10–47), the 2-year OFS, PFS, and OS rates were 83%, 69% and 83%, respectively (Figure 1A,B). IL-10 dosage in the AH in six tested patients during the first 6 months post-CAR T-cell therapy was undetectable for all but one patient who relapsed a few months later (Figure S1). On the other hand, IL-6 levels exceeding 2.5 pg/mL were detected in the AH of all six patients tested during this period (range, 19–194 pg/mL), with no observed correlation to the occurrence of grade ≥ 3 ICANS. Two of seven patients experienced relapses: one patient had an isolated intraocular relapse 10 months after CAR T-cell infusion, and the other one experienced a relapse in the brain and cerebrospinal fluid 5 months after CAR T-cell infusion.

To better characterize the trafficking and persistence of CAR T-cells in the eye, flow cytometry was performed at different time points in three patients (one patient was assessed four times, and two were assessed twice). One patient demonstrated AH positivity for CAR T-cells at 11 days, 1 month, 4 months, and 10 months (last ophthalmologic follow-up) (Figure 1C). The other two patients tested negative for CAR T-cells at all time points. All three patients remained in CR at their last ophthalmologic follow-up (17, 16, and 10 months after CAR T-cell infusion, respectively). Peripheral blood CAR T-cell expansion during the first 30 days post-infusion is illustrated in Figure S2.

All patients developed cytokine release syndrome (Grade 1 in three patients, Grade 2 in four patients). ICANS of any grade occurred in six patients (Grade 3 in one, Grade 4 in two). The Grade 4 ICANS cases involved a 65-year-old patient with a history of cerebral involvement and whole-brain radiotherapy who received axi-cel for isolated ocular relapse, and a 71-year-old patient who received axi-cel with progressive bulky cerebral disease at the time of infusion. Four out of seven patients received steroids, and no patients developed ocular pseudoprogression.

This is the first series on CAR T-cell therapy in patients with VRL, and we acknowledge the small and heterogeneous series (VRL both primary and secondary, with or without CNS involvement), which may affect the generalizability of our findings. In particular, it is conceivable that the presence of occult systemic disease, especially in cases of previous systemic involvement, may have facilitated CAR T-cell expansion or that distinct molecular profiles may have influenced the duration of the therapeutic response.

To date, only a few case reports on CAR T-cell therapy for hematological malignancies with ocular involvement have been published.^11-13 Taher et al. reported a case of early ocular failure of CAR T-cells in a patient with diffuse large B-cell lymphoma (DLBCL) treated for intraocular and CNS localizations, where the CNS disease remained in CR.¹⁴ Additionally, two cases of CAR T-cell therapy for B-cell acute lymphoblastic leukemia with ophthalmic localization have been described: one demonstrated persistent CR after 1 year, whereas the other reported a flare reaction with sudden ocular deterioration following CAR T-cell therapy.^{12, 13} The flare reaction consisted of bilateral retinal detachment occurring on Day 7 after CAR T-cell infusion, in the absence of blasts in the AH, but with the presence of white blood cells, 35% of which expressing CD3.

Although our patients were highly pretreated and refractory, the efficacy of CAR T-cells appeared very promising, with outcomes comparable to those achieved with ASCT within our network (2-year PFS and OS rates of 68% and 87%, respectively).¹⁵ These results also compared favorably to those obtained in the first-line treatment of primary vitreoretinal lymphoma.¹ The impact of bridging therapy and the remission status at the time of CAR T-cell infusion may represent confounding factors, but the persistence of response after CAR T-cell infusion in the three highly refractory patients with PR before CAR T-cells (23, 16, and 17 months, respectively) strongly supports the major contribution of CAR T-cells in maintaining disease control. Notably, one patient in CR at 6 months exhibited high IL-10 in the AH (1246 pg/mL) without any clinical signs of relapse but subsequently relapsed at 10 months. The IPCG criteria for ophthalmic response do not account for IL-10 quantification; however, the relapse might have already been present in an infra-clinical state as early as 6 months.

AH CAR T-cell trafficking was demonstrated for the first time in this series in one patient evaluated at multiple time points, confirming their ability to migrate and persist in the AH despite low blood levels of the target antigen. We can hypothesize that CAR T-cells were detected in the AH of only one of three tested patients due to several factors, including the exceptional localizations in the AH of VRL, technical challenges associated with analyzing small-volume samples, sensitivity of flow cytometry, and potentially suboptimal sampling timing relative to the kinetics of CAR T-cell expansion. Further investigations are needed to better characterize CAR T-cell trafficking in these scenarios.

Finally, we noted unusual Grade 3–4 ICANS in th1ree of seven patients, while the rate and severity of neurotoxicity reported in CNS lymphomas are comparable to those in systemic lymphomas.^{5, 7, 16} This unexpectedly high rate of severe ICANS in our cohort may be attributed to multiple factors, including the small sample size, a high number of prior treatment lines, a history of ASCT and/or encephalic radiotherapy (both known to be associated with cognitive impairment), the presence of CNS disease (reflecting tumor burden), and the type of CAR T-cell construct used. Notably, among the three patients who developed Grade 3-4 ICANS in our cohort, all had progressive CNS disease; two had previously received ASCT with or without encephalic radiotherapy and were treated with axi-cel. Conversely, we did note any ocular immune-related adverse events during follow-up.

Larger studies are needed to evaluate the true risk of severe neurotoxicity following CAR T-cell therapy in VRLs. Nevertheless, the decision to pursue CAR T-cell therapy should carefully consider this risk.

In conclusion, this study is the first to report a significant response rate after CAR T-cell therapy in highly pretreated and refractory VRLs, with a promising 2-year PFS rate of 69%. Longer follow-up is essential to confirm these findings and define the role of CAR T-cell therapy within the broader therapeutic strategy for VRL.

Cécile Pivert: Investigation; formal analysis; writing—original draft; software. Adélaïde Toutée: Investigation; data curation. Christophe Parizot: Formal analysis; investigation; data curation. Denis Malaise: Investigation. Alexandre Matet: Investigation. Valérie Touitou: Investigation. Magali Le Garff-Tavernier: Formal analysis; data curation. Damien Roos-Weil: Investigation. Sarah Touhami: Investigation. Nabih Azar: Investigation. Ines Boussen: Investigation. Véronique Morel: Investigation. Madalina Uzunov: Investigation. Clotilde Bravetti: Formal analysis; data curation. Carole Metz: Investigation. Eve Todesco: Formal analysis; data curation. Delphine Sterlin: Formal analysis. Carole Soussain: Investigation. Kahina Ouis Tarhi: Investigation. Anne Besançon: Investigation. Khê Hoang-Xuan: Investigation. Sylvain Choquet: Conceptualization; supervision; writing—review and editing. Caroline Houillier: Conceptualization; supervision; writing—review and editing; formal analysis; validation; methodology; software. Marine Baron: Conceptualization; validation; supervision; writing—review and editing; methodology; software.

The authors declare no conflicts of interest.

No funding.