Anatomy of a crime: how IL7R and NRAS join forces to drive T-cell acute lymphoblastic leukemia

In this issue of Haematologica, Winer et al.¹ demonstrate that MYC is crucial to the oncogenic cooperation between IL7R and NRAS driving T-cell acute lymphoblastic leukemia (T-ALL) development.² They further show that the kinase PLK-1 may contribute to MYC protein stability and that MYC-modulating drugs can be of therapeutic value against T-ALL driven by activating mutations in both IL7R and NRAS. Why the relevance of diving deeper into the mechanisms underlying this collaboration? Roughly 10% of T-ALL patients display IL7R gain-of-function mutations, and a much larger fraction (some 50-80% of the cases) express IL7R and may benefit from IL7 produced in the leukemia milieu.^3-7 IL7R-mediated signaling (because of IL7, high IL7R levels or mutational activation of the receptor or downstream effectors) can promote T-ALL establishment and maintenance, and resistance to glucocorticoids.⁵ RAS activating mutations in general occur in around 2% of the cases, and NRAS alterations are infrequent. However, full appreciation of the importance of RAS signaling in T-ALL must consider other lesions, including inactivating mutations in the RAS-MEK-ERK pathway negative regulator NF1 or RasGRP1 overexpression. Interestingly, MEK-ERK pathway can be activated also by IL7/IL7R in T-ALL, contributing to IL7R-mediated resistance to glucocorticoids.⁸

To understand why IL7R or NRAS alone cannot drive T-ALL in a transplant mouse model, whereas their combination is clearly leukemogenic,² Winer et al. have now analyzed the transcriptome of immature mouse thymocytes transduced with mutant IL7R (a particular type 1a IL7R activating mutation, hereafter referred to as mutIL7R),^3,5 mutant NRAS (coding for NRAS G13D, which leads to NRAS activation; hereafter referred to as mutNRAS) or their combination. They found no evidence of Myc activation (as measured by the upregulation of Myc target genes) by mutIL7R alone, whereas mutNRAS activated Myc and, importantly, this effect was augmented by the combination of mutIL7R and mutNRAS. In agreement, Myc itself was up-regulated by mutNRAS and mutNRAS+mutIL7R, but not by mutIL7R alone. The fact that mutIL7R was unable to activate Myc is intriguing, as MYC is a downstream target of IL7R-mediated signaling in human thymocytes, and in zebrafish models of T-ALL.⁹ The reasons for these discrepancies are unclear. They may relate to the stage of disease development at which the analyses were conducted: D1 cells, being p53-null, are one step closer to transformation than healthy thymocytes, and the analyses in zebrafish and human T-ALL focused on fully transformed cells.

These considerations apart, the authors provide convincing evidence linking RAS signaling and MYC. They show that MYC overexpression phenocopied mutant NRAS in its ability to collaborate with mutIL7R to drive T-ALL. Both mutIL-7R+mutNRAS and mutIL7R+MYC led to CD4⁺CD8⁺ T-ALL, with relatively similar expression of TCR Vα and predominance of αβ over γδ T-cells. Whether this resemblance extends to the transcriptional profile was not addressed. Nonetheless, these findings align with previous studies showing that mutIL7R and MYC collaborate to drive T-ALL in zebrafish.⁹ The relevance of MYC in mutIL7R+mutNRAS-driven leukemias was further exposed by experiments showing that Myc deletion decreased leukemia burden in vivo and that Myc silencing decreased the fitness of D1 cells and primary thymocytes transduced with the combination of the two oncogenes.

Winer et al. further combined RNA-sequencing and mass-spectrometry to show that Bcl-2 transcript and protein were up-regulated.¹ Contrary to Myc, Bcl-2 up-regulation was due to mutIL7R and not mutNRAS, and the combination of the two oncogenes did not potentiate mutIL7R effects. That IL7-IL7R-mediated signaling up-regulates Bcl-2 is well known, although (in contrast to the illustration in Figure 8 of the paper) this is unlikely to be mediated by STAT5 in T-ALL.^8,10

Contrary to Bcl-2, Myc was up-regulated exclusively at the protein level, and the authors propose that NRAS collabo rated with IL7R essentially by stabilizing Myc. How activated NRAS, and its combination with mutant IL-7R, promoted Myc stabilization may involve two mechanisms. RAS-MEK-ERK signaling is known to phosphorylate Myc at Serine 62 (S62), and this phosphorylation contributes to Myc stability. Indeed, Myc S62 was up-regulated by mutIL7R and even more so by mutNRAS, although not further increased by the combination. On the other hand, AKT-dependent Myc Threonine 58 (T58) phosphorylation was up-regulated by each oncogene alone and by their combination. This should mark Myc for degradation, which obviously was not the case. So, there should be a mechanism counterbalancing the effects of Myc T58-phosphorylation. Winer et al. noticed that PLK-1 (which contributes to MYC stabilization by preventing its proteasomal degradation) was up-regulated by NRAS and its combination with IL7R. Thus, they used volasertib, a PLK-1 inhibitor, to test the impact on Myc expression. Volasertib not only down-regulated Myc protein but also, surprisingly, Myc transcript levels, suggesting that PLK-1 may up-regulate Myc via different mechanisms. In the absence of PLK-1 genetic manipulation, and given that pharmacological inhibitors often have off-target effects, it may be that volasertib impacted Myc expression, particularly at the transcript level, by PLK-1-independent mechanisms. Nonetheless, the authors provide good evidence that PLK-1 and MYC are likely involved in a positive-feedback loop that partakes in leukemia development promoted by the combination of mutIL7R and mutNRAS.

Figure 1.Winer et al. demonstrate that gain-of-function (i.e., activating) mutations in IL7R and NRAS cooperate to drive T-cell acute lymphoblastic leukemia (T-ALL) in mice mainly due to the ability to activate MYC, in particular via NRAS-dependent up-regulation of MYC protein levels. NRAS likely promotes MYC protein stabilization by at least two mechanisms: directly, by phosphorylation of MYC, and indirectly, via transcriptional activation of PLK-1, whose increased expression prevents MYC proteasomal degradation. Bcl-2 is also up-regulated (essentially due to IL7R-mediated signaling), although its exact role in the cooperative oncogenic effects of IL7R and NRAS is unclear. The use of a chemical inhibitor of PLK-1 (volasertib) or of a drug that down-regulates MYC (and IL7R) transcription (JQ-1) diminishes leukemia burden in vivo. Pre-clinical studies evaluating the value of these drugs against human T-ALL with IL7R and RAS signaling pathway mutations are warranted. Final version by somersault18:24.

Evidently, the question that remained was whether these findings have translational potential. Using their transplant mouse model of mutIL7R+mutNRAS T-ALL, Winer et al. show that volasertib diminished Myc levels and leukemia burden in vivo. There was no benefit in combining volasertib with the Bcl-2 inhibitor venetoclax. This is surprising, given the importance of Bcl-2 for IL-7R-mediated viability in T-ALL, and how MYC (which is down-regulated by volasertib in vivo) and BCL2 synergize to promote cancer development. The authors also tested JQ-1, a BET bromodomain BRD4 inhibitor, which down-regulates MYC and IL7R transcription. JQ-1 demonstrated similar in vivo effects to volasertib, yet another demonstration of the importance of MYC in these leukemias.

Overall, the studies by Winer et al. not only allow a better understanding of how oncogenic IL7R cooperates with RAS signaling in driving T-ALL (Figure 1), but also pave the way for preclinical studies testing the value of PLK-1 and/or MYC inhibitors in human T-ALL patient samples, and patient-derived xenograft models, with IL7R and RAS pathway mutations.

Footnotes

• Received January 9, 2024
• Accepted January 30, 2024

Correspondence

J.T. Barata joao_barata@medicina.ulisboa.pt

Disclosures

No conflicts of interest to disclose.

Funding

The work in João T. Barata’s lab was supported by grants from the following funding agencies: European Research Council (ERC-PoC-101069429), la Caixa Foundation (HR21-00761) and Fundação para a Ciência e a Tecnologia (PTDC/ MEC-ONC/4606/2021).

Acknowledgments

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