Analysis of patient‐specific
            <i>NF1</i>
            variants leads to functional insights for Ras signaling that can impact personalized medicine

Long, Ashlee; Liu, Hui; Liu, Jian; Daniel, Michael; Bedwell, David M.; Korf, Bruce R.; Kesterson, Robert A.; Wallis, Deeann

doi:10.1002/humu.24290

Cited by 12 publications

(13 citation statements)

References 37 publications

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“…1 B ) in NF1 knockout 293T cells. This is consistent with previous reports of lower mutant protein expression observed in a G848R mouse model of NF1 ( 13 , 14 ) and in cells transiently expressing mouse Nf1 cDNA (complementary DNA) with L847P and G848R ( 15 ). Despite lower levels of expression, these mutants are still able to form heterodimers with wild-type protein ( Fig.…”

Section: Resultssupporting

confidence: 93%

Destabilizing NF1 variants act in a dominant negative manner through neurofibromin dimerization

Young

Salazar

Han

et al. 2023

Proc. Natl. Acad. Sci. U.S.A.

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The majority of pathogenic mutations in the neurofibromatosis type I ( NF1 ) gene reduce total neurofibromin protein expression through premature truncation or microdeletion, but it is less well understood how loss-of-function missense variants drive NF1 disease. We have found that patient variants in codons 844 to 848, which correlate with a severe phenotype, cause protein instability and exert an additional dominant-negative action whereby wild-type neurofibromin also becomes destabilized through protein dimerization. We have used our neurofibromin cryogenic electron microscopy structure to predict and validate other patient variants that act through a similar mechanism. This provides a foundation for understanding genotype–phenotype correlations and has important implications for patient counseling, disease management, and therapeutics.

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Section: Resultssupporting

confidence: 93%

Destabilizing NF1 variants act in a dominant negative manner through neurofibromin dimerization

Young

Salazar

Han

et al. 2023

Proc. Natl. Acad. Sci. U.S.A.

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“…There is a disconnect between the level of full-length transcript generated upon PMO treatment, and the amount of neurofibromin protein produced in the cell. Based on recently published data, mNf1 cDNA containing the Y489C missense mutation was able to produce a stable protein product [8], and thus, we suspect that this disconnect is not the result of a protein stability issue, but rather some other mechanism.…”

Section: Discussionmentioning

confidence: 82%

“…Previously, we created a cDNA of this variant (Y489C) that was not subject to splicing and showed that the missense variant alone did not disrupt Ras signaling [7]. More recently, we have shown that the variant had no effect on neurofibromin's expression, indicating that if the aberrant splicing could be corrected, the missense variant may not be deleterious [7,8]. Hence, a PMO approach might be feasible as a treatment strategy.…”

Section: Introductionmentioning

confidence: 99%

Restoration of Normal NF1 Function with Antisense Morpholino Treatment of Recurrent Pathogenic Patient-Specific Variant c.1466A>G; p.Y489C

Awad

Moore

Liu

et al. 2021

JPM

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Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder with almost 3000 different disease-causing variants within the NF1 gene identified. Up to 44% of these variants cause splicing errors to occur within pre-mRNA. A recurrent variant in exon 13, c.1466A>G; p.Y489C (Y489C) results in the creation of an intragenic cryptic splice site, aberrant splicing, a 62 base pair deletion from the mRNA, and subsequent frameshift. We investigated the ability of phosphorodiamidate morpholino oligomers (PMOs) to mask this variant on the RNA level, thus restoring normal splicing. To model this variant, we have developed a human iPS cell line homozygous for the variant using CRISPR/Cas9. PMOs were designed to be 25 base pairs long, and to cover the mutation site so it could not be read by splicing machinery. Results from our in vitro testing showed restoration of normal splicing in the RNA and restoration of full length neurofibromin protein. In addition, we observe the restoration of neurofibromin functionality through GTP-Ras and pERK/ERK testing. The results from this study demonstrate the ability of a PMO to correct splicing errors in NF1 variants at the RNA level, which could open the door for splicing corrections for other variants in this and a variety of diseases.

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“…One potential dominant-negative mechanism represents dimerization of mutant and WT neurofibromin [53,54] , with resultant degradation of WT neurofibromin at the faster rate of the mutant [110] . In this scenario, an NF1-HCT approach that decreases neurofibromin degradation may be preferable to one that increases NF1 transcription.…”

Section: Implications For Nf1-hctmentioning

confidence: 99%

Rationale for haploinsufficiency correction therapy in neurofibromatosis type 1

Frøst¹,

Serra²,

Viskochil³

et al. 2022

J Transl Genet Genom

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Neurofibromatosis type 1 (NF1) is a genetic disorder with a wide range of manifestations and severity. Currently, the few available NF1 treatments target specific manifestations, with no available therapies targeted to correct the underlying driver of all NF1 manifestations. Evidence supports that haploinsufficiency in NF1 caused by a decreased amount of wild-type (WT) neurofibromin in all Nf1+/- cells directly causes or facilitates a range of NF1 manifestations. Consequently, NF1 haploinsufficiency correction therapy (NF1-HCT) represents a potentially effective approach to treat some NF1 manifestations. NF1-HCT would normalize the level of WT neurofibromin in all NF1-haploinsufficient cells, including those integral to the NF1 phenotype such as Schwann cells (SCs), melanocytes, neurons, bone cells, and cells of the tumor microenvironment. This would correct altered cellular signaling pathways and, in turn, restore normal function to cells with a retained WT allele. NF1-HCT will not restore WT neurofibromin in NF1-/- cells; however, by restoring function in the surrounding Nf1+/- microenvironment cells, NF1-HCT is predicted to have a beneficial effect on NF1-/- cells. NF1-HCT is expected to have a clinical effect in some NF1 manifestations, as follows: (i) prevention, or delay of onset, of potential manifestations; and (ii) reversal, or halting/slowing progression, of established manifestations. This review describes the rationale for NF1-HCT, including specific NF1 considerations (e.g., NF1 clinical phenotype, neurofibromin function/regulation, NF1 mutational spectrum, genotype-phenotype correlation, and the impact of haploinsufficiency in NF1), HCT in other haploinsufficient diseases, potential NF1-HCT drug treatment strategies, and the potential advantages/challenges of NF1-HCT.

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Analysis of patient‐specific NF1 variants leads to functional insights for Ras signaling that can impact personalized medicine

Cited by 12 publications

References 37 publications

Destabilizing NF1 variants act in a dominant negative manner through neurofibromin dimerization

Destabilizing NF1 variants act in a dominant negative manner through neurofibromin dimerization

Restoration of Normal NF1 Function with Antisense Morpholino Treatment of Recurrent Pathogenic Patient-Specific Variant c.1466A>G; p.Y489C

Rationale for haploinsufficiency correction therapy in neurofibromatosis type 1

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