LLY-283

Two of the most effective inhibitors, LLY-283 and GSK591, were targeted against PRMT5 using distinct binding mechanisms and reduced the viability of a subset of DMG cells expressing wild-type TP53 and mutant ACVR1. RNA-sequencing and phenotypic analyses revealed that LLY-283 could reduce the viability, clonogenicity and invasion of DMG cells in vitro, representing three clinically important phenotypes, but failed to prolong survival in an orthotopic xenograft model. Together, these data show the challenges of DMG treatment and highlight PRMT5 inhibitors for consideration in future studies of combination treatments.

The combination of GSK-591 or LLY-283 with either of the FLT3 TKIs Quizartinib or Giltertinib resulted in synergistically enhanced inhibition of FLT3-ITD+ MOLM-13 and MV4-11 AML cells, compared to TKI or PRMT5 inhibitor alone. An epigenetic probe screen identified PRMT5 as a key regulator of FLT3-ITD AML cell viability. We show an important role for PRMT5 in maintenance of murine and human FLT3-ITD AML stem cells and in their persistence following FLT3 TKI treatment. Treatment with a PRMT5 inhibitor in combination with a FLT3 TKI could be a promising approach to enhance elimination of FLT3-ITD AML stem cells.

LLY-283 inhibits the expression of PRMT5 and Ki-67, thereby decreases the proliferation and metastasis of HNSCC and the ability to form transplanted tumors in nude mice, exerting anti-tumor effects.

Finally, we observed that treatment of LLY-283 alleviated neurological deficits and reduced infarct volume in the MCAO/R mice. Taken together, PRMT5 may be a potential therapeutic target for cerebral I/R injury.

The 2 most effective compounds inhibit PRMT5 (GSK591 and LLY-283) reducing viability >50% in adherent and spheroid screens. However a reduction in stemness, as seen in vitro¸ does not always reduce primary tumor burden. Therefore proteomic characterization of neural differentiation in the primary PDX samples is underway, and investigation into secondary tumor initiation would be warranted.Our data shows PRMT5 inhibitors are a promising new target for DIPG, specifically in ACVR1 mutant DIPG, and justifies further in vivo investigations into changes in tumor initiation capacity and exploration of rational combinations to improve survival outcome.

Using two orthogonal-acting inhibitors of PRMT5 (GSK591 or LLY-283), we show that pharmacological inhibition of PRMT5 suppresses the growth of a cohort of 46 patient-derived GBM stem cell cultures, with the proneural subtype showing greater sensitivity. Importantly, we demonstrate that LLY-283 is brain-penetrant and significantly prolongs the survival of mice with orthotopic patient-derived xenografts. Collectively, our findings provide a rationale for the clinical development of brain penetrant PRMT5 inhibitors as treatment for GBM.

We further show that the combination of PRMT5 inhibitors (GSK591, LLY283 5μM) with AC220 (500pM) resulted in increased inhibition of cell proliferation and increased induction of apoptosis compared to TKI alone, p<0.05 (Figure 1a-c). In conclusion, our studies using an unbiased approach have identified PRMT5 inhibition as a novel and selective approach to enhance targeting of FLT3-ITD AML cells in combination with FLT3 TKI. These results support our ongoing studies to evaluate the cellular and molecular mechanisms underlying these combinatorial effects, and to determine the translational therapeutic potential of this combination using primary patient samples and mouse models.