Assessment of retinoblastoma RNA reflux after intravitreal injection of melphalan

WINTER, URSULA; NICOLAS, MICHAEL; SGROI, MARIANA; SAMPOR, CLAUDIA; TORBIDONI, ANA; FANDIÑO, ADRIANA; CHANTADA, GUILLERMO L; MUNIER, FRANCIS L; SCHAIQUEVICH, PAULA

BRITISH JOURNAL OF OPHTHALMOLOGY

B M J PUBLISHING GROUP

Año: 2018 vol. 102 p. 415 - 418

0007-1161

Background:Intravitreal injection of chemotherapy in retinoblastoma eyes with vitreousseeds may lead to a risk of extraocular tumor dissemination that has not been assessedso far.Aims:To develop a sensitive and clinically feasible technique to assess forpotential retinoblastoma-cell reflux after intravitreal injection of melphalan.Methods:Filter papers were cut in 6mm diameter circles and sterilized before use. Eyeswith retinoblastoma vitreous seeds (group D, International Classification)received weekly intravitreal melphalan injections (20mg or 30mg/dose)followed by cryotherapy as part of local treatment. Immediately after finishingthe injection and cryotherapy, filter papers were placed on the injection site andon the cryoprobe tip to assess for the expression of the cone-rod homeobox gene(CRX) by real-time qPCR as a surrogate of retinoblastoma RNA. The assay wasdeveloped and validated to determine sensitivity, linearity, recovery,repeatability, and reproducibility. Results:The assay for quantitation of CRX expression waslinear in the range of 1 to 1000 cells. The lowest limit of detection was oneretinoblastoma cell and allowed to recover 100% of the cell load in externalsupplementation. A total of 14 eyesreceived 22 cycles of intravitreal melphalan and were evaluated for potential extraoculartumor-cell dissemination using the developed technique. None of the cycles werepositive for CRX in samples from the scar or from the cryoprobe tip.Conclusions: Asensitive and simple method of tumor-cell assessment has been developed thatcan be used in the clinics to assess for potential extraocular disseminationafter intravitreal injections to assure its performance.