Beyond the phenotype: Aqueous humor sampling moves retinoblastoma toward personalized therapy

Retinoblastoma has long stood apart from other solid tumors in one critical way: it has never been safe to biopsy, because passing a needle into the tumor risks seeding extraocular spread. As a result, clinicians have staged and treated the disease almost entirely on what the eye looks like. Aqueous humor liquid biopsy upends that limitation—sampling tumor-derived cell-free DNA from the anterior chamber without touching the tumor—and gives physicians their first molecular window into a living, at-risk eye. At the Retina Society Annual Meeting, Jesse L. Berry, MD, presented prospective data on implementing the aqueous humor liquid biopsy platform in retinoblastoma and other intraocular lesions.

In the prospective cohort, the molecular test was 98% sensitive for active disease, and no benign lesion masquerading as retinoblastoma returned a positive result—findings with direct implications for the retina surgeon weighing whether an unexplained intraocular mass can be safely approached. Berry discussed how markers such as RB1 alterations, 6p gain, MYCN amplification, and BCOR mutation are beginning to inform recurrence risk and personalized treatment intensity, why the clinically validated LBSeq4Kids assay already accepts samples from centers worldwide, and how the same aqueous window may help resolve diagnostic dilemmas well beyond retinoblastoma, including RB mimickers and intraocular lymphoma.

The Q&A

Note: Transcript lightly edited for clarity and length.

Retinoblastoma has never been biopsiable. What does sampling the aqueous humor let you see that you couldn't before?

Jesse L. Berry, MD: Before aqueous humor liquid biopsy, we treated retinoblastoma based almost entirely on what we could see—the clinical phenotype—because putting a needle into the tumor itself is contraindicated due to the risk of extraocular spread.

Now, the aqueous gives us access to tumor-derived cell-free DNA without touching the tumor. So for the first time, we can see the molecular phenotype of the living eye: RB1 alterations, copy-number changes such as 6p gain, MYCN amplification, and potentially tumor fraction longitudinally (tumor fraction remains in the research space—not clinically validated at this time, but the others are).

For the first time, we can interrogate the tumor molecularly while the child still has his or her eye—and while we're actively trying to save it.

In this prospective work, what did the molecular result change about a patient's management?

Berry: I think one of the most important things this prospective study tells us is how much confidence we can put in the test when we're faced with an uncertain diagnosis. If you're taking care of a child and you're asking, “Could this be retinoblastoma?” the test was 98% sensitive for active disease. And importantly, not a single eye with a benign lesion masquerading as retinoblastoma (RB) had a positive test.

That has real clinical implications. If the molecular test is negative, we can feel much more comfortable that we're not dealing with retinoblastoma. And that's particularly important for our retina colleagues. Historically, operating on an eye with an unexplained intraocular mass carries this concern that you could inadvertently operate on a retinoblastoma. Having a molecular test that helps distinguish those lesions from RB can give us much greater confidence to move forward with appropriate retinal surgery.

And the opposite is also true. We had children with very atypical presentations—where cataract or hemorrhage obscured our ability to see the tumor clearly—and a positive molecular result helped us establish the diagnosis of RB earlier and move promptly to appropriate treatment.

When the biopsy flags a high-risk eye that still looks salvageable, how do you weigh the marker against the exam?

Berry: Right now, the molecular result is an additional piece of information—it doesn't replace the clinical exam. If an eye looks salvageable, a high-risk molecular marker alone isn't going to make us give up on that eye. But it does tell us that the risk of recurrence is higher, so we're going to watch that child very closely and we're not going to extend the interval between examinations for some time.

It also becomes particularly important when the clinical course isn't going the way we would like. If an eye is responding poorly and we also identify a high-risk molecular marker, such as MYCN or a BCOR mutation, that weighs into our decision-making because we know we're dealing with a higher-risk tumor.

Where I think this ultimately takes us is toward truly personalized treatment for retinoblastoma. I believe one day we'll know the best treatment for each molecular type of RB based on the markers we identify in the aqueous humor. It won't be the same one-size-fits-all treatment for every child and every eye. We're not there yet—but that's where I think we're headed.

How do you explain a molecular risk signal to a family when the eye still looks fine?

Berry: We generally have this molecular information within a few weeks of diagnosis, and I explain to families that we've identified certain markers that may suggest that the eye will be more difficult to treat—that it may not respond as well to initial therapy or may require more treatment to achieve a cure.

But I also emphasize that this is one piece of information. We're going to monitor the child very closely and watch clinically how the tumor is—or isn’t—responding to therapy. Then we use all of that information together as we make treatment decisions.

Ultimately, our goal is to work together with the family to save the eye, as long as it remains safe for the child to do so.

Four centers just published where they disagree on managing retinoblastoma in 2026. Where could liquid biopsy settle those debates?

Berry: I love this question because it highlights just how much variability there still is in the treatment of retinoblastoma. And I think, in large part, that's because we've never had molecular data on this disease. We couldn't biopsy these tumors, so historically we've based treatment on how the eye looks—and different centers have developed very different approaches.

We still see significant differences among leading centers in the U.S. and world today. For example, some centers treat essentially every eye with high-dose, three-drug intra-arterial chemotherapy. But some eyes almost certainly don't need that much therapy. We see eyes with a relatively flat genomic profile and low mutational burden that respond beautifully to far less aggressive treatment.

There's also ongoing debate about systemic chemotherapy, particularly for children with bilateral disease, where one systemic treatment can treat both eyes. These are exactly the kinds of questions where molecular information may eventually help us move beyond treating patients primarily according to what their eyes look like.

I believe that in the not-too-distant future, we'll use the molecular features of each child's tumor to help guide both the type and intensity of treatment. We're moving toward a much more precise and much more personalized approach to retinoblastoma therapy.

Retinoblastoma care lives in a few centers. What must happen for a molecular approach to reach the average patient?

Berry: The exciting thing is that this can already reach patients outside of a major retinoblastoma center. LBSeq4Kids is a clinically validated test performed through the Center for Personalized Medicine at Children's Hospital Los Angeles. And I think it's important to emphasize that this isn't a research assay being run in my laboratory—it’s a clinical test performed in a clinical laboratory.

Any physician, anywhere, can obtain an aqueous sample and send it to CHLA for clinical testing. We already receive aqueous samples from centers throughout the United States and Canada, and we've received samples internationally from places including Slovenia and Australia.

So the molecular testing doesn't have to live only where the expertise to develop the assay lives. That's really important for a rare disease like retinoblastoma. We can centralize sophisticated molecular testing while making it accessible to physicians—and therefore children—around the world.

You're extending this to other intraocular lesions. Which diagnostic dilemma stands to benefit most?

Berry: I think one of the biggest opportunities is exactly what retina surgeons already encounter: the unexplained intraocular mass or other lesion that could potentially be retinoblastoma. These RB mimickers are not infrequent, and before proceeding with intraocular surgery, retina surgeons almost always want an evaluation to exclude retinoblastoma—and they should.

That's a setting where aqueous humor testing with LBSeq4Kids can be particularly helpful. We've worked with surgeons around the world when the molecular test has been negative, giving them additional confidence that they're not dealing with RB and helping them move forward with appropriate management.

But the positive results can be equally important. Sometimes the test tells us that the lesion really is retinoblastoma, and in other cases the molecular findings can point toward another intraocular malignancy, such as lymphoma. So we're beginning to use the aqueous not simply as a liquid biopsy for retinoblastoma but as a molecular window into an otherwise diagnostically inaccessible intraocular lesion.

Jesse L. Berry, MD

is Professor Ophthalmology, Chief of Ophthalmology at Children’sHospital Los Angeles (CHLA), and Director of Ocular Oncology at the University ofSouthern California (USC) Roski Eye Institute. She received her undergraduate andmedical degree from Harvard University followed by ophthalmology and fellowshiptraining at CHLA+USC under the mentorship of A. Linn Murphree, MD. Dr. Berry is thefounder of Women in Ocular Oncology, President of the International Society of OcularOncology (ISOO, 2024-26) and President of the American Association for OphthalmicOncology and Pathology (AAOOP, 2026-2028). Her clinical expertise focuses onretinoblastoma and uveal melanoma. She is the director of the Amy Elizabeth CorwinAqueous Humor Liquid Biopsy Laboratory at CHLA and holds the A. Linn Murphree, MDChair in Retinoblastoma and the Berle and Lucy Adams Chair in Cancer Research. Shehas pioneered development of the aqueous humor as a liquid biopsy for retinoblastomaand other ocular tumors; this research is supported by the National Cancer Institute andDepartment of Defense