Abstract

Ras proteins are critical nodes in cellular signaling that integrate inputs from activated cell surface receptors and other stimuli to modulate cell fate through a complex network of effector pathways. Oncogenic RAS mutations are found in ∼ 25% of human cancers and are highly prevalent in hematopoietic malignancies. Because of their structural and biochemical properties, oncogenic Ras proteins are exceedingly difficult targets for rational drug discovery, and no mechanism-based therapies exist for cancers with RAS mutations. This article reviews the properties of normal and oncogenic Ras proteins, the prevalence and likely pathogenic role of NRAS, KRAS, and NF1 mutations in hematopoietic malignancies, relevant animal models of these cancers, and implications for drug discovery. Because hematologic malignancies are experimentally tractable, they are especially valuable platforms for addressing the fundamental question of how to reverse the adverse biochemical output of oncogenic Ras in cancer.

Introduction

Aberrant signal transduction resulting in reduced dependence on growth factors and other extracellular stimuli for the survival and proliferation of malignant cells is an established “hallmark of cancer.”1 RAS genes encode a family of 21-kDa proteins that are central nodes in signaling networks that regulate cell fate in many tissue lineages. RAS genes are also the most common targets of dominant somatic mutations in human cancer.2,3  The high prevalence of mutations in NRAS, KRAS, and molecules, such as CKIT, PTPN11, CBL, and BCR-ABL1, that interact with Ras biochemically strongly implicate aberrant Ras signaling as an important therapeutic target in these cancers.4-7  In this review, we focus on the pathobiology of mutations in hematologic cancers involving the “core” Ras-GTPase activating protein (GAP) complex, which includes Ras itself, guanine nucleotide exchange factors, and GAPs. At this writing, mutations in NRAS, KRAS, and the NF1 tumor suppressor, which encodes a GAP called neurofibromin,8  are strongly associated with myeloid malignancies. Increasing evidence also implicates these genes as “drivers” in lymphoid cancers with “high-risk” clinical features. No mechanism-based treatments exist for the ∼ 25% of human cancers with KRAS or NRAS mutations or for the growing number of malignancies showing NF1 inactivation, and we discuss potential therapeutic strategies for addressing the adverse biochemical consequences of aberrant Ras signaling.

Structural and functional properties of the Ras GTPase switch

Ras proteins are signal switch molecules that regulate cell fates by cycling between active guanosine triphosphate (GTP)–bound and inactive guanosine diphosphate (GDP)–bound conformations.9  On ligand binding, molecules, such as Shc, Grb2, Gab2, and SHP-2, are recruited to growth factor receptors and these complexes activate Ras guanine nucleotide exchange factors (GEFs; Figure 1).10  GEFs catalyze dissociation of guanine nucleotides from Ras, which is followed by passive rebinding. Because the concentration of free GTP in cells vastly exceeds that of GDP, GEF-induced nucleotide exchange increases Ras-GTP levels.10  Ras can be activated by a number of different GEFs in mammalian cells, including SOS1 and SOS2, RasGRFS1 and RasGRFS2, and RasGRP1 to RasGRP4 (Figure 1).2 

Figure 1

The Ras switch. Ras proteins are switches that relay signals initiated when transmembrane receptors bind ligand. Activated receptors recruit GEFs by assembly of multiprotein complexes (eg, including SOS) or more indirectly, by evoking lipid modifications that recruit GEFs, such as RasGRPs, to cytosolic membrane surfaces. GEFs promote exchange of GDP for GTP on Ras. When bound to GTP, Ras adopts a conformation in which the “switch” regions are stabilized and can interact productively with various downstream effectors. Thus, Ras connects extracellular stimuli to intracellular networks that compute and execute cell fate decisions. The Ras signal is terminated by GTP hydrolysis, which is largley dependent on GAPs, such as neurofibromin (NF1) or p120 RasGAP.

Figure 1

The Ras switch. Ras proteins are switches that relay signals initiated when transmembrane receptors bind ligand. Activated receptors recruit GEFs by assembly of multiprotein complexes (eg, including SOS) or more indirectly, by evoking lipid modifications that recruit GEFs, such as RasGRPs, to cytosolic membrane surfaces. GEFs promote exchange of GDP for GTP on Ras. When bound to GTP, Ras adopts a conformation in which the “switch” regions are stabilized and can interact productively with various downstream effectors. Thus, Ras connects extracellular stimuli to intracellular networks that compute and execute cell fate decisions. The Ras signal is terminated by GTP hydrolysis, which is largley dependent on GAPs, such as neurofibromin (NF1) or p120 RasGAP.

GTP binding stabilizes the switch I and switch II domains of Ras, which then interacts productively with effectors.9  These effector molecules are activated biochemically through complex mechanisms that involve recruitment to discrete subcellular compartments, increasing intrinsic catalytic activity and/or inducing conformational changes that allow effectors to act as scaffolds for the assembly of signaling complexes.11,12  Three canonical Ras effectors have been the focus of intense study: PI3-kinase (PI3K), Raf, and Ral-GDS proteins (Figure 1). Of these, aberrant activation of the Raf/MEK/ERK pathway and the PI3K/Akt/mTOR cascade is most strongly implicated in malignant transformation and tumor maintenance. We refer interested readers to detailed reviews of individual effector pathways.2,13 

Signaling is terminated when Ras-GTP is hydrolyzed to Ras-GDP. This “off” reaction is catalyzed by intrinsic Ras GTPase activity, which is inefficient in the absence of GAPs. GAPs bind to the switch domains of Ras-GTP and insert an “arginine finger” into the phosphate binding loop of Ras that stabilizes a transition state between Ras-GTP and Ras-GDP.2,8  Because GAPs accelerate GTP hydrolysis thousands of fold, some view the Ras-GTPase as enzyme complex composed of Ras and a GAP. Neurofibromin, the protein encoded by NF1, and p120 GAP are the predominant Ras GAPs in mammalian cells, and both are expressed broadly.8,14  Given the central role of Ras signaling in cellular homeostasis, it is perhaps not surprising that germline mutations in NF1 and other genes that regulate Ras-GTP levels also cause developmental disorders.2,10 

Ras isoforms, posttranslational modifications, and intracellular trafficking

RAS genes encode 4 highly homologous proteins (H-Ras, N-Ras, K-Ras4a, and K-Ras4b) that are identical in the first 85 amino acids. This “G” domain includes the P-loop, which interacts with the γ-phosphate of GTP, and the switch regions (Figure 2).2,10  Ras proteins share 85% identity over the next 80 amino acids and only diverge substantially at the C-terminal “hypervariable region” (Figure 2).2  The hypervariable region of all Ras isoforms terminates with a CAAX motif, where the cysteine is prenylated by farnesyltransferase (FTase). After prenylation, the —AAX of the CAAX motif is removed by Ras-converting enzyme 1, and the carboxyl group of the newly exposed isoprenylcysteine is methylated by isoprenylcysteine-carboxyl-methyltransferase (ICMT). These lipid modifications provide weak membrane binding affinity that is stabilized by a second signal motif. For K-Ras4b, this is provided by a polybasic lysine domain, which interacts strongly with the phospholipid membrane. By contrast, H-, N-, and K-Ras4a are palmitoylated at cysteine(s) adjacent to the CAAX motif (Figure 2). These modifications are essential for correct trafficking and subcellular localization of each isoform.15  In particular, whereas H-, N-, and K-Ras4a are directed from the endoplasmic reticulum (ER) to the Golgi en route to the plasma membrane (PM), K-Ras4b localizes directly to the PM through an unknown mechanism. A significant portion of H-Ras, N-Ras, and K-Ras4a reside and signal in the Golgi and ER, and these proteins traffic between the Golgi, ER, and PM in a dynamic cycle that is regulated by depalmitoylation and repalmitoylation.15  Depalmitoylation is mediated by acyl protein thioesterase 1 and acyl protein thioesterase 2.15  By contrast, there are at least 20 palmitoyl transferases, and it is uncertain which of these act on individual Ras proteins.16  Perturbation of palmitate turnover leads to nonspecific distribution of H-Ras, N-Ras, and K-Ras4a to endomembranes and decreases signaling from the PM.17 

Figure 2

Ras processing and trafficking to subcellular compartments. H-Ras, N-Ras, K-Ras4a, and K-Ras4b proteins are identical in the first 85 amino acids, a region that includes the P loop (phosphate binding loop, amino acids 10-16), which binds the γ-phosphate of GTP, and the switch I (amino acids 30-38) and switch II (amino acids 60-76) regions, which regulate binding to Ras regulators and effectors. The next 78 amino acids show ∼ 85%-90% sequence homology. Amino acids that are shared by all isoforms are depicted in light gray, those that differ are depicted in dark gray. The final 24 (23 for K-Ras4b) amino acids, called the hypervariable region (HVR), specify posttranslational modifications and trafficking for each Ras isoform. All 4 isoforms have a C-terminal —CAAX motif, which is farnesylated (F) by FTase. The —AAX is removed by Ras-converting enzyme 1 (RCE1) and the cysteine methylated (M) by ICMT. K-Ras4b is then shuttled directly to the PM where it is stabilized by its polylysine domain (KKKKKK). The other 3 isoforms are shuttled to the Golgi apparatus, where they are palmitoylated (P) at one or more cysteines near the C-terminus before reaching the plasma membrane. On the membrane, H-Ras, N-Ras, and K-Ras4a can be depalmitoylated by acyl protein thioesterases 1 and 2 (APT1/APT2), directing them back to the Golgi. This palmitoylation-depalmitoylation, Golgi-plasma membrane cycle continues in a delicate balance until the proteins are degraded.

Figure 2

Ras processing and trafficking to subcellular compartments. H-Ras, N-Ras, K-Ras4a, and K-Ras4b proteins are identical in the first 85 amino acids, a region that includes the P loop (phosphate binding loop, amino acids 10-16), which binds the γ-phosphate of GTP, and the switch I (amino acids 30-38) and switch II (amino acids 60-76) regions, which regulate binding to Ras regulators and effectors. The next 78 amino acids show ∼ 85%-90% sequence homology. Amino acids that are shared by all isoforms are depicted in light gray, those that differ are depicted in dark gray. The final 24 (23 for K-Ras4b) amino acids, called the hypervariable region (HVR), specify posttranslational modifications and trafficking for each Ras isoform. All 4 isoforms have a C-terminal —CAAX motif, which is farnesylated (F) by FTase. The —AAX is removed by Ras-converting enzyme 1 (RCE1) and the cysteine methylated (M) by ICMT. K-Ras4b is then shuttled directly to the PM where it is stabilized by its polylysine domain (KKKKKK). The other 3 isoforms are shuttled to the Golgi apparatus, where they are palmitoylated (P) at one or more cysteines near the C-terminus before reaching the plasma membrane. On the membrane, H-Ras, N-Ras, and K-Ras4a can be depalmitoylated by acyl protein thioesterases 1 and 2 (APT1/APT2), directing them back to the Golgi. This palmitoylation-depalmitoylation, Golgi-plasma membrane cycle continues in a delicate balance until the proteins are degraded.

Structural properties of oncogenic Ras proteins and therapeutic implications

Somatic point mutations of RAS genes at codons 12, 13, and 61 are the most common dominant oncogenic lesions in human cancer.2  Substitutions in these residues favor the GTP-bound conformation because of reduced intrinsic GTP hydrolysis and resistance to GAPs.2  The oncogenic Ras/GAP switch is an exceedingly difficult target for rational drug discovery. The most tractable molecular targets in cancer, such as BCR-ABL, the mutant epidermal growth factor receptor (EGFR), and B-RafV600E, are aberrantly activated kinases that are inhibited by selective small-molecule inhibitors. By contrast, oncogenic Ras proteins have reduced GTPase activity.2  Restoring enzymatic function, the fundamental biochemical problem posed by oncogenic Ras, is extremely challenging. Developing mechanism-based inhibitors of oncogenic Ras is further complicated by the picomolar affinity of Ras for GTP and by high intracellular GTP concentrations.18  Finally, the P loop of Ras is structurally constrained, with any amino acid substitution at glycines 12 or 13 introducing space-occupying side chains that distort the interaction with GAPs.9  It is therefore difficult to envision how small molecules might bind to Ras and restore GAP-mediated GTP catalysis. For these reasons, recent drug discovery efforts have largely focused on inhibiting kinase components of downstream effector pathways, such as PI3K, Akt, mammalian target of rapamycin (mTOR), and MEK.

NRAS, KRAS, and NF1 mutations in hematologic cancers

Different RAS genes are preferentially mutated in distinct tumor types, with KRAS mutations highly prevalent in common epithelial malignancies.2  As summarized in the next 5 sections and in Table 1, hematologic cancers are unusual in that NRAS and KRAS are both mutated at significant frequencies, with NRAS mutations predominating. NF1 is the only Ras GAP implicated in tumorigenesis to date. Although mutations in Ras GEFs have been linked to the development of drug resistance in mouse models of hematologic malignancies,19  other mutations in this family of molecules are rare in human cancers (COSMIC database).

Table 1

Incidence of NRAS, KRAS, and NF1 mutations in hematologic malignancies

NRASKRASNF1
JMML 17% (9-30) 8% (3-18) 7% (4-15) 
CMML 19% (2-43) 11% (6-15) ND 
AML 11% (10-44) 5% (1-13) ∼ 5% (0-7) 
MDS 5% (4-9) ∼ 1% ∼ 3% (0-9) 
ALL 10% (0-18) 4% (0-5) 6% (3-8)* 
MM 18% (17-30) 8% (6-9) ND 
NRASKRASNF1
JMML 17% (9-30) 8% (3-18) 7% (4-15) 
CMML 19% (2-43) 11% (6-15) ND 
AML 11% (10-44) 5% (1-13) ∼ 5% (0-7) 
MDS 5% (4-9) ∼ 1% ∼ 3% (0-9) 
ALL 10% (0-18) 4% (0-5) 6% (3-8)* 
MM 18% (17-30) 8% (6-9) ND 

These data are tabulated from published series that used the current definition for each disease, distinguished clearly between NRAS and KRAS mutations and, with 1 exception (*T-ALL only), did not restrict inclusion based on an underlying genetic alteration or specific subtype of disease. The supplemental Materials (available on the Blood Web site; see the Supplemental Materials link at the top of the online article) include tables and references listing the primary sources for the data shown here by disease category, along with the number of cases sequenced in each report.

ND indicates not determined.

JMML and CMML

These aggressive myeloid malignancies are classified as myelodysplastic syndrome/myeloproliferative neoplasia (MDS/MPN) “overlap” syndromes by the World Health Organization. A distinctive pattern of hypersensitive CFU-GM progenitor growth in methylcellulose cultures containing GM-CSF is a cellular hallmark of juvenile myelomonocytic leukemia (JMML) that is also observed in some cases of chronic myelomonocytic leukemia (CMML).20 

The association between neurofibromatosis type 1 (NF1) and JMML pointed the way toward our current understanding of the underlying molecular pathogenesis.20  Genetic analysis of JMML bone marrow specimens from children with NF1 revealed somatic loss of the normal NF1 allele, elevated levels of Ras-GTP, and activation of the Raf/MEK/ERK effector pathway.20  Sequencing revealed NF1 mutations in ∼ 15% of patients with JMML, suggesting that other genes involved in Ras signaling might also be mutated. Indeed, molecular analysis demonstrating somatic NRAS and KRAS in 25%-30% of JMML patients, and subsequent research identifying PTPN11 and CBL mutations firmly established JMML as a disease of hyperactive Ras.20  Furthermore, the observation that children with NF1 and certain other developmental disorders are predisposed to JMML provided strong evidence that hyperactive Ras/Raf/MEK/ERK signaling initiates this disease.2 

NRAS and CBL mutations are highly prevalent in CMML.4,21  Whereas RAS mutations are found in > 20% of patients with the myeloproliferative variant of CMML, they are rare in the more benign myelodysplastic variant.22,23  It is uncertain whether hyperactive Ras signaling can initiate leukemogenesis in CMML or whether mutations in genes that are involved in epigenetic programming are also required.24  However, there is some evidence to support the role of cooperation between hyperactive Ras signaling and transcription factor mutations in acute transformation of CMML.25 

AML

The 2 largest published series identified NRAS codon 12, 13, or 61 mutations in 10%-11% of adult acute myeloid leukemia (AML), and KRAS mutations in an additional 5%.26,27  A similar incidence of NRAS/KRAS mutations is observed in pediatric patients.28,29 NRAS/KRAS mutations do not have independent prognostic significance in patients treated with current antileukemia regimens, although patients with AML harboring a RAS mutation may respond better to high-dose cytarabine as post-remission therapy.30  Recently, high-throughput sequencing revealed novel NRAS and KRAS mutations in AML and CMML that introduce substitutions at amino acids 14, 60, 74, and 146 in 2%-5% of cases.31  This discovery suggests that 20%-25% of AML carries somatic NRAS/KRAS mutations. Loss of NF1 expression was reported in 7 of 95 patients with AML, and 7 additional cases had heterozygous NF1 loss.32 

Observations in JMML and CMML and studies of genetically engineered mice summarized in “Insights from mouse models” provide compelling evidence that NF1, NRAS, and KRAS mutations are early or initiating events in some myeloid malignancies, particularly MDS/MPN overlap disorders. The picture is more complex in AML, with existing data suggesting that these mutations may function as either early/initiating or as cooperating mutations that are acquired during disease progression. RAS and NF1 mutations are over-represented in the French-American-British (FAB) M4 and M5 monocytic subtypes, with an overall incidence of > 30%.26,27,32  Interestingly, JMML and CMML also evolve to M4/M5 AML,31,33  and leukemia cells from patients with JMML, CMML, and FAB M4/M5 AML exhibit a similar signature of phospho-protein activation characterized by aberrant STAT5 activation at low GM-CSF concentrations.34  Together with the absence of consistent transcription factor mutations in FAB M4/M5 AML, these data raise the possibility that M4/M5 AML, JMML, and CMML are related malignancies in which NRAS/KRAS/NF1 play an early and essential role in leukemic growth.34,35 

A popular model suggests that initiation of AML requires both a “class 1” mutation that aberrantly activates signal transduction and a “class 2” mutation in a transcription factor.36  Recent studies demonstrating a high frequency of heterozygous and homozygous mutations in genes that broadly regulate the eipgenome, such as TET2, EZH2, IDH1, IDH2, and DNMT3A, and other lines of evidence indicate that this simple paradigm only partially explains the complex pathogenesis of AML.37  However, it is useful for considering the potential role of NRAS/KRAS/NF1 mutations as cooperating events in leukemogenesis. The observation that activating mutations in FLT3 and CKIT, which encode receptor tyrosine kinases upstream of Ras, are common in AML and largely confined to cases without RAS mutations supports the broad relevance of “class 1” mutations in AML. Furthermore, several groups have shown that RAS and FLT3 mutations can be lost or gained during disease evolution in AML and MDS.38,39  Indeed, cases with an NRAS mutation at diagnosis may demonstrate a FLT3 mutation at relapse (and vice versa), and NRAS mutations are associated with acquired resistance to Flt3 kinase inhibitors in vitro.40  These studies are consistent with the idea that mutations that alter signaling molecules are secondary events that contribute to clonal evolution in many patients with AML.

MDS/MPNs

Initial studies demonstrating somatic NRAS mutations in 20%-40% of MDS included patients with CMML, which undoubtedly inflated these numbers. Recent data indicate that NRAS is mutated in ∼ 5% of MDS where it is associated with adverse clinical features and a high risk of transformation to AML.41,42 KRAS mutations are rare in MDS.42  Although there are limited data on somatic NF1 inactivation in MDS, recurrent cryptic changes or deletions involving the NF1 locus were identified in 3 of 35 patients.43  Mutations in BRAF, PTPN11, CBL, FLT3, CKIT, and other genes encoding signaling molecules that interact with Ras are also uncommon in MDS.44  Taken together, although the existing data do not support an integral role of NRAS/KRAS/NF1 mutations in the development of MDS, they probably contribute to transformation to AML. By contrast, mutations that deregulate signaling networks are integral to the pathogenesis of MPN and NRAS/KRAS/NF1 mutations are therefore not implicated in progression to acute leukemia.45 

ALL

There are limited data regarding RAS mutations in lymphoid malignancies, particularly in adults. One small study suggested that ∼ 11% of adult T lineage acute lymphoblastic leukemia (T-ALL) patients harbor RAS mutations.46  Neri et al identified RAS mutations in 6 of 33 ALL cases (18%), all of which were in codon 12 or 13 of NRAS, although patient age was not reported.47  In the pediatric literature, estimates of RAS mutation frequency in both B- and T-lineage ALL approximate 15%, with NRAS mutations predominating.48,49 

Early T-cell precursor ALL is a recently identified subtype of pediatric ALL with a distinct gene expression profile and an immunophenotype notable for expression of stem cell and myeloid markers.50  Children with this form of leukemia have high rates of induction failure, as well as extremely high risk of relapse.50  Whole-genome sequencing recently uncovered mutations in genes involved in Ras signaling in 67% of early T-cell precursor ALL cases (19% NRAS, 3% KRAS, 14% FLT3, and the remainder in JAK1, JAK3, BRAF, and IGFR1).48  These data suggest that early T-cell precursor ALL may be initiated by NRAS/KRAS and other “driver” mutations that are commonly identified in myeloid malignancies in a multipotent hematopoietic stem/progenitor cell. Somatic NF1 mutations were also reported recently in some human T-ALLs.51 

MM

RAS is the single most commonly mutated gene family in multiple myeloma (MM).52  Whereas early publications estimated the frequency of NRAS/KRAS mutations in MM as high as 30%-40%,53-55  a comprehensive recent study identified codon 12, 13, or 61 RAS mutations in 23% of 529 patients.52  Of these, two-thirds were in the NRAS gene (mostly in codon 61), with the remainder in KRAS (largely in codons 12 and 13). RAS mutations predict shorter overall survival and progression-free survival in MM, but are rare in monoclonal gammopathy of unknown significance, a precursor to MM in some patients.52,53  Furthermore, as in AML, RAS gene mutations are occasionally not detected at diagnosis but found at relapse. Although not definitive, these data suggest that NRAS/KRAS mutations are more likely to be involved in MM progression rather than initiation.52,53,55 

Insights from mouse models

Ras oncogenes have been studied extensively in the mouse, both to elucidate the mechanisms underlying deregulated growth and to develop robust in vivo platforms for performing preclinical trials. Table 2 presents a partial list of informative models of hematologic malignancies. One strong theme that has emerged is that either inactivating Nf1 or expressing oncogenic Kras from its endogenous genetic locus in the hematopoietic compartment is sufficient to produce a myeloproliferative disorder that models human MDS/MPN. This highly penetrant phenotype progresses rapidly in Mx1-Cre; KrasG12D mice.35,56  Interestingly, similar models based on endogenous NrasG12D expression demonstrated a mild and variable myeloid phenotype,57-59  although mice that are homozygous for a conditional NrasG12D knock-in allele consistently develop aggressive MPN.59,60  Together, these data support the idea that differences in the structure and/or expression levels of different Ras isoforms influence the severity of myeloid growth dysregulation, which may have implications for therapeutic development.

Table 2

Mouse models of hematologic malignancies characterized by KRAS, NRAS, or NF1 mutations

AlleleModel typeGenotypePhenotype(s)References
KrasG12D Heterozygous conditional “knock in” Mx1-Cre; KrasG12D/+ Aggressive MPN 35, 56  
KrasG12D Adoptive transfer of conditional “knock in” marrow into irradiated recipients Mx1-Cre; KrasG12D/+ T-ALL 65, 108  
KrasG12D Conditional “knock in” + retroviral insertional mutagenesis Mx1-Cre; KrasG12D/+ T-ALL (∼ 80%); AML (∼ 20%) 66  
NrasG12D Retroviral transduction/transplantation WT cells expressing exogenous NrasG12D CMML, AML 69, 70, 71  
NrasG12D Heterozygous conditional “knock in” Mx1-Cre; NrasG12D/+ Indolent MPN, MDS, LPD, HS 57, 58, 75  
NrasG12D Homozygous conditional “knock in” Mx1-Cre; NrasG12D/G12D Aggressive MPN 59, 60  
NrasG12D Heterozygous conditional “knock in” + retroviral insertional mutagenesis Mx1-Cre; NrasG12D/+ AML 58  
Nf1 Heterozygous germline “knock out” Nf1+/− Late-onset MPN in ∼ 10% of mice 109  
Nf1 Adoptive transfer of homozygous “knock out” fetal liver cells Nf1−/− Subactute MPN 110  
Nf1 Heterozygous “knock out” + irradiation Nf1+/− MDS, MPN 72, 73  
Nf1flox Homozygous conditional “knock out” Mx1-Cre; Nf1flox/flox MPN 74  
Nf1flox Homozygous conditional “knock out” + retroviral insertional mutagenesis Mx1-Cre; Nf1flox/flox AML (∼ 70%); T-ALL (∼ 30%) 19  
AlleleModel typeGenotypePhenotype(s)References
KrasG12D Heterozygous conditional “knock in” Mx1-Cre; KrasG12D/+ Aggressive MPN 35, 56  
KrasG12D Adoptive transfer of conditional “knock in” marrow into irradiated recipients Mx1-Cre; KrasG12D/+ T-ALL 65, 108  
KrasG12D Conditional “knock in” + retroviral insertional mutagenesis Mx1-Cre; KrasG12D/+ T-ALL (∼ 80%); AML (∼ 20%) 66  
NrasG12D Retroviral transduction/transplantation WT cells expressing exogenous NrasG12D CMML, AML 69, 70, 71  
NrasG12D Heterozygous conditional “knock in” Mx1-Cre; NrasG12D/+ Indolent MPN, MDS, LPD, HS 57, 58, 75  
NrasG12D Homozygous conditional “knock in” Mx1-Cre; NrasG12D/G12D Aggressive MPN 59, 60  
NrasG12D Heterozygous conditional “knock in” + retroviral insertional mutagenesis Mx1-Cre; NrasG12D/+ AML 58  
Nf1 Heterozygous germline “knock out” Nf1+/− Late-onset MPN in ∼ 10% of mice 109  
Nf1 Adoptive transfer of homozygous “knock out” fetal liver cells Nf1−/− Subactute MPN 110  
Nf1 Heterozygous “knock out” + irradiation Nf1+/− MDS, MPN 72, 73  
Nf1flox Homozygous conditional “knock out” Mx1-Cre; Nf1flox/flox MPN 74  
Nf1flox Homozygous conditional “knock out” + retroviral insertional mutagenesis Mx1-Cre; Nf1flox/flox AML (∼ 70%); T-ALL (∼ 30%) 19  

Although the role of Ras pathway activation has been more thoroughly described in the development of myeloid disease, growing evidence implicates hyperactive Ras in the development of lymphoid cancers as well. Lymphoid transformation was observed in early studies of primary hematopoietic cells expressing mutant Ras,61  and spontaneous T-ALL is a common finding in transgenic mouse models bearing Ras oncogenes.62-64  In retrospect, such studies predicted the presence of Ras pathway mutations in human lymphoid leukemia (see “ALL”). T-ALL can be induced in KrasG12D mice by transplanting limiting numbers of hematopoietic stem cells into irradiated recipients or by retroviral insertional mutagenesis.65-67  Mutations leading to Notch1 activation are common in this setting, and strong synergy exists between these 2 human leukemia oncogenes.68 

It is probable that additional mutations contribute to leukemic outgrowth initiated by oncogenic Ras or Nf1 inactivation in other contexts. Bone marrow transduced with retroviral vectors encoding oncogenic Kras or Nras can cause AML in recipient mice after transplantation, and these cancers exhibit clonal retroviral integrations.69-71  Retroviral insertional mutagenesis in Nf1, Kras, and Nras mutant mice was deployed as a forward genetic strategy to identify mutations that might cooperate with hyperactive Ras in leukemogenesis.19,58,66  These studies implicated Evi1, Myb, Sox4, and Gfi1 in AML initiated by Nras or Nf1 inactivation, and Ikzf1 in Kras mutant mice with T-ALL. Consistent with reports of therapy-induced hematologic cancers in patients with NF1, heterozygous Nf1 inactivation cooperates with radiation to induce MDS and MPN in mice.72,73 

Hematopoietic cells that either express endogenous levels of oncogenic NrasG12D or KrasG12D or have inactivated Nf1 demonstrate a hypersensitive pattern of CFU-GM progenitor colony growth in response to GM-CSF that recapitulates what is observed in JMML and in some patients with CMML.20,35,58,74  Consistent with in vivo observations, this cellular phenotype is most pronounced in cells from Mx1-Cre; KrasG12D/+ and homozygous Mx1-Cre; NrasG12D/G12D mice, but relatively subtle in Mx1-Cre; NrasG12D/+ and Mx1-Cre; Nf1flox/flox cells. Whereas biochemical analysis of bone marrow cells from these mutant mice revealed elevated basal levels of phosphorylated ERK (pERK), the degree of activation was unexpectedly subtle and bone marrow cells markedly increased pERK levels in response to growth factor stimulation.35,59,60,74,75  Detailed biochemical profiling of Nf1 and Nras mutant AMLs has not been reported. Interestingly, however, although T-ALL cells from Kras mutant mice have constitutively high Ras-GTP levels, pERK, pAkt, and pS6 levels were highly variable.66  Together, biochemical studies of mouse leukemias support the idea that primary hematopoietic cells dynamically modulate signaling networks in response to increased Ras-GTP levels, which has implications for the use of targeted therapeutics.

Therapeutic strategies for inhibiting Ras processing and activation

Inhibiting posttranslational processing of Ras engendered intense interest in the 1990s. Drug discovery efforts focused on the essential lipid modification at CAAX sequence that is catalyzed by FTase.76  Clinical trials of selective and potent FTase inhibitors were largely disappointing,76,77  probably because K-Ras and N-Ras undergo lipid modification via geranylgeranyl transferase when FTase is inhibited and therefore retain biologic activity.76  Indeed, a preclinical trial of the FTase inhibitor L744,832 in mice that developed MPN after adoptive transfer of Nf1-deficient hematopoietic cells revealed no inhibition of K- or N-Ras processing at the maximally tolerated dose and no improvement in leukocyte counts or splenomegaly.78  Off-target effects may explain the clinical responses to FTase inhibitors reported in some leukemias.77,79  Unfortunately, combining FTase and geranylgeranyl transferase inhibition proved too toxic to represent a viable therapeutic regimen.76 

Genetic studies in mice investigated the effects of interfering with C-terminal processing of prenylated Ras proteins by Ras-converting enzyme 1 and ICMT. Inactivating Rce1 in hematopoietic cells unexpectedly accelerated MPN in the Mx1-Cre, KrasG12D model of JMML/CMML.80  By contrast, conditional inactivation of Icmt attenuated myeloproliferation in this strain and also dramatically reduced growth-factor independent colony growth in vitro.81  Although the opposite effects of Rce1 and Icmt inactivation on KrasG12D-induced myeloid disease are perplexing, it should be recalled that these enzymes catalyze the posttranslational modification of many other cellular proteins that could modulate the phenotype. Small molecules have been identified that block ICMT activity; however, these compounds have not yet been tested in vivo.82 

The palmitoylation/depalmitoylation cycle is a potential therapeutic target for selectively inhibiting the growth of hematologic cancers with NRAS mutations (Figure 2). A recent study reported that mutating the palmitoylation site at cysteine 181 to serine (C181S) in N-RasG12D inhibited plasma membrane localization in fibroblasts and prevented the development of myeloid disease in a retroviral transduction/transplantation model of MPN.83  Xu et al found that the C181S mutation exhibited dominant negative activity on myeloid progenitor growth and ERK activation, probably by sequestering effectors.60  Palmostatin B is a novel APT inhibitor that causes entropy-driven diffusion of H-Ras and N-Ras throughout the cell.84  Exposing transduced hematopoietic cells from Nras and Kras mutant mice to palmostatin B mislocalized N-RasG12D away from the plasma membrane and reduced CFU-GM and AML blast colony growth. Importantly, this inhibition was selective for Nras mutant cells and mapped to the hypervariable region. Together, these studies identify APT inhibitors as a novel class of potential therapeutics in hematologic malignancies with NRAS mutations. Newer and more potent inhibitors in this chemical series were reported recently, although systemic toxicities have not been fully described.85 

Blocking growth-promoting signals that converge to activate GEFs is another potential therapeutic strategy for treating cancers characterized by NRAS/KRAS/NF1 mutations. In the case of oncogenic Ras, this raises the question of whether these proteins are truly “constitutively active” or remain dependent on guanine nucleotide exchange for transforming activity. The answer is not known and may depend, at least in part, on cell context and on the nature of the underlying RAS mutation. However, there is some evidence that the mutant Ras/GAP switch is at least partially dependent on growth factor signaling in hematopoietic cells. This dependence is clearest in Nf1-deficient cells where genetic ablation of either Gmcsf or the β-common chain of the GM-CSF receptor markedly attenuates the JMML-like MPN.86,87  Interestingly, primary hematopoietic cells from Mx1-Cre, KrasG12D mice show little basal activation of Raf/MEK/ERK or PI3K/Akt/S6 signaling, and these pathways respond robustly to growth factor stimulation.35,88  It is unclear exactly how upstream inputs promote growth of cells expressing mutant Ras proteins, but the rapid development of AML in Mx1-Cre, KrasG12D mice after Nf1 deletion implies that Ras-GTP levels may be limiting.89  Together, these observations suggest that interfering with growth factor-mediated Ras activation may be beneficial, at least in principle, in some cancers with RAS or NF1 mutations. Along these lines, Maurer et al18  recently identified chemical inhibitors to a novel binding pocket on the surface of Ras. DCA1, a compound from this screen, inhibits Ras activation by interfering with SOS-mediated guanine nucleotide exchange, and probably also has activity against other exchange factors.18  Although the long-term therapeutic potential of this approach is uncertain, treatment with drugs that block “upstream” proteins might be considered in hematologic cancers with NRAS/KRAS/NF1 mutations that respond to growth factor stimulation.

Targeting Ras effector pathways

Intensive effort has focused on developing small-molecule inhibitors of the Raf/MEK/ERK and PI3K/Akt/mTOR cascades. Effectors of oncogenic Ras are “indirect” targets that are activated by the mutant protein, introducing additional complexity. It is now appreciated that oncoprotein expression elicits compensatory responses with unpredictable consequences for downstream pathways. For example, whereas T-ALLs generated by insertional mutagenesis in Mx1-Cre, KrasG12D mice invariably show elevated levels of Ras-GTP, the activation of effectors, such as ERK, Akt, and S6, is highly variable in independent leukemias.66 

Although evidence is limited regarding which Ras effector pathways are essential for tumor initiation and which are required for maintenance in vivo, the spectrum of non-RAS mutations offers some insights. Whereas mutations that alter Ras-activated proteins are uncommon in JMML, CMML, and AML, “upstream” genes, such as FLT3, CKIT, CBL, and PTPN11, are frequently mutated in these cancers, suggesting that deregulation of multiple effector pathways is essential for myeloid transformation. By contrast, mutational analysis strongly implicates the PI3K pathway in T-ALL,90  and the recent discovery of activating BRAF mutations in ∼ 4% of MM91  suggests that aberrant Raf/MEK/ERK signaling is an important consequence of RAS mutations in this cancer.

Given the uncertainty regarding which Ras effector(s) are essential for the survival and proliferation of different hematopoietic cancers, evaluating targeted inhibitors in preclinical and clinical trials that include pharmacodynamic monitoring of biochemical endpoints can provide pathogenic as well as clinical insights. Sorafenib, a multikinase inhibitor with activity against Raf proteins, had promising single-agent efficacy in AML with FLT3 mutations in a phase 1 trial.92  However, this may be the result of non-Raf effects, as sorafenib also inhibits Flt3 and other kinases.93  More potent and selective Flt3 inhibitors have shown great promise in AML94  and will certainly supersede sorafenib for directly targeting this mutant protein.

CI-1040 and PD0325901 are allosteric MEK inhibitors that have been evaluated in preclinical models of JMML/CMML, AML, and T-ALL.19,66,95  Whereas CI-1040 was ineffective in Mx1-Cre, Nf1flox/flox mice with MPN,19  PD0325901 induced dramatic hematologic improvement and greatly prolonged survival in the Mx1-Cre, KrasG12D model of JMML/CMML.95  These differential effects may reflect the short duration of in vivo MEK inhibition in the bone marrow cells of mice treated with CI-1040 (∼ 4 hours) versus prolonged effects of PD0325901.19  The beneficial effects of PD0325901 in Mx1-Cre, KrasG12D mice were associated with persistence of Kras mutant cells.95  These observations imply that treatment modulates the biologic behavior of these cells and suggest that it will be important to assess molecular markers of the malignant clone in clinical trials of MEK inhibition in JMML and CMML.

MEK inhibitors were also tested in insertional mutagenesis models of AML initiated by Nf1 inactivation19  and of T-ALL characterized by oncogenic KrasG12D expression.66  Although CI-1040 had no beneficial therapeutic index in Mx1-Cre, Nf1flox/flox mice with MPN, it induced dramatic disease regression and markedly extended survival in recipients transplanted with primary Nf1-deficient AMLs.19  This suggests that cooperating mutations acquired during progression from MPN to AML rendered these aggressive cancers more dependent on Raf/MEK/ERK signaling. All of these mice that initially responded to MEK inhibition ultimately relapsed despite continued treatment. Genetic analysis revealed the emergence of preexisting drug-resistant clones, underscoring the importance of genetic heterogeneity as a cause of resistance to targeted inhibitors.

“First-generation” MEK inhibitors demonstrated efficacy against solid tumors with BRAF mutations but had minimal activity in RAS-mutant tumors.96  Based on these disappointing data and unexpected ocular toxicity, clinical development of CI-1040 and PD0325901 was suspended. Newer MEK inhibitors appear to have less toxicity and greater efficacy against solid tumors harboring RAS mutations. However, it is uncertain whether they are superior to standard therapies, and a recent phase 2 study of the MEK inhibitor AZD6244 in advanced AML showed only modest and transient antitumor activity.97  PI3K signaling is activated in AML blasts, and PI3K inhibitors demonstrated antiproliferative activity in vitro. Although there are a number of PI3K inhibitors in clinical trials for solid tumors, none has yet been tested in humans with hematologic malignancies.

One potential explanation for lack of efficacy of single-agent PI3K/Akt or Raf/MEK/ERK inhibition in RAS-mutant cancers tumors may be the ability of either pathway to overcome inhibition of the other through the release of negative feedback loops or other mechanisms. This provides a rationale for simultaneously inhibiting both major effector pathways in RAS-mutant malignancies. Studies demonstrating synergistic antitumor effects of combining MEK and PI3K/mTOR inhibition in genetically engineered murine solid tumor models supports this general strategy. Finally, other Ras effectors in addition to PI3K/Akt and Raf/MEK/ERK may promote tumor formation and/or maintenance in human malignancies and could also represent potential avenues for acquiring drug resistance.

Identifying synthetic lethal interactions with oncogenic Ras

Performing RNA interference (RNAi) screens to identify “synthetic lethal” interactions between oncogene mutations and other cellular proteins is an emerging strategy for identifying novel therapeutic targets in cancer. The underlying rationale is that 2 mutations compatible with survival on their own may be lethal when combined. The goal of these experiments is to identify genes that, when “knocked down,” selectively kill cells with RAS mutations with the hope that some of these will be tractable targets for drug discovery. Studies performed to date have uncovered novel candidate targets whose suppression results in selective apoptosis of oncogenic KRAS-mutant cell lines, including PLK1, STK33, TBK1, and WT1.98-100  It will be several years before we know whether patients with KRAS-mutant malignancies will benefit from drugs that inhibit any of these targets; however, a number of issues have already emerged. First, there has been little overlap in the candidate genes identified by different groups. Second, genetic dependencies in cancer cell lines may not accurately reflect reality in primary malignances, which, as clinicians know, are exceptionally resilient. Third, reducing the expression of a protein is not equivalent to pharmacologically inhibiting a specific activity of that molecule. For example, many kinases are multidomain proteins that regulate cell growth through enzymatic and nonenzymatic mechanisms. RNAi screens cannot distinguish which of these functions is essential for the growth of transformed cells. Finally, recent experience with RNAi screens emphasizes that it is essential to thoroughly validate “hits” by attempting to rescue the lethal phenotype by re-expressing candidate cDNA molecules that are not inhibited by the knockdown construct used in the initial screen. With these caveats in mind, in vitro and in vivo RNA interference screening is a promising approach for discovering new therapeutic targets in an unbiased manner. A recent study that applied this general strategy to KRAS mutant nonsmall cell lung cancer uncovered a strong synthetic lethal interaction with the GATA2 transcription factor.101  This work was particularly impressive as the authors identified drugable GATA-regulated signaling modules, which they validated by performing in vivo preclinical therapeutic trials in mice. Given the importance of GATA2 in hematopoietic lineage specification, these data have obvious implications for leukemia.

Summary and future directions

Developing therapeutic strategies to reverse the abnormal biochemical output of oncogenic Ras proteins without unacceptable toxicity to normal cellular signaling networks is one of the fundamental problems in cancer. NRAS and KRAS mutations contribute to a spectrum of hematologic malignances, and genetic analysis of human specimens as well as experiments in mouse models support the idea that RAS mutations can initiate leukemic growth or serve as cooperating mutations. The latter observation raises the question of whether targeting oncogenic Ras is worthwhile in cancers, such as AML, where it probably functions as a secondary mutation in most patients. Although the answer to this question is unknown, a reasonable working hypothesis is that any somatic genetic lesion that undergoes positive clonal selection in a substantial percentage of cancers contributes to malignant growth. The recent experience with Flt3 inhibitors in AML supports this idea. Like NRAS, FLT3 mutations have been classified as “class 1” mutations that cooperate with aberrant transcription to induce full leukemic transformation. Early-phase clinical trials of potent and selective Flt3 inhibitors, such as AC220, showed that monotherapy induced remission in some patients with refractory AML.102  Importantly, “gatekeeper” mutations in the Flt3 kinase occur with relapse,103  providing compelling genetic evidence that there is strong selective pressure for AML cells to restore aberrant Flt3 signaling in vivo. Effective biochemical inhibition of oncogenic Ras signaling in human acute leukemias with RAS mutations would probably have similar efficacy.

Treating advanced cancers with kinase inhibitors almost uniformly leads to genotype-specific response followed by regrowth of resistant cells.104,105  Based on this precedent, combining inhibitors of oncogenic Ras signaling with other anticancer agents will probably be essential for sustained efficacy. This general strategy was recently implemented in patients with “high risk” B-lineage ALL and BCR-ABL1 rearrangements with promising results.106  A recent study in lung cancer cells lines with EGFR mutations suggests that synthetic lethal screens might be particularly useful for identifying proteins that contribute to resistance to inhibitors of Ras effectors.107  In contrast to advanced cancers, such as AML and MM, it is possible that “early-stage” hematologic disorders, such as JMML, will respond to therapies that exclusively target oncogenic Ras signaling networks.

Targeting aberrant signaling downstream of oncogenic Ras remains a daunting challenge. Our current understanding of the Ras effector pathways that are essential for cancer initiation and maintenance is incomplete, and it is likely that these dependencies vary in specific cell types. A related question is how “addicted” different cancers are to oncogenic Ras signaling. At one end of the spectrum, mutant RAS might be required during early stages of leukemogenesis but dispensible thereafter. Alternatively, advanced hematologic cancers, such as AML, might become ever more dependent on oncogenic Ras as they evolve. If the latter proves true, it will be important fully elucidate the feedback mechanisms that cancer cells deploy to restrain aberrant signaling and to characterize how these adaptations contribute to oncogene dependence. In addition to their inherent value as potential anticancer drugs, potent and selective inhibitors of Ras-regulated kinases are invaluable tools for addressing these questions through in vitro and in vivo studies.

Hematopoietic malignancies provide exceptional platforms for addressing mechanistic and translational questions because of established systems for culturing primary cells, robust mouse models, and the ability to serially sample diseased tissues from patients enrolled on clinical trials. Thus, rigorously interrogating how targeted anticancer agents modulate oncogenic Ras signaling and how this correlates with therapeutic response in hematologic malignancies has broad implications for reducing the major contribution of oncogenic RAS mutations to the worldwide burden of cancer.

Acknowledgments

The authors thrank the members of the laboratories of B.S.B. and K.M.S. for helpful discussions. Limitations on the number of references prevented us from citing some primary sources.

The work in our laboratories on RAS and NF1 mutations in hematologic cancers was supported by the Children's Tumor Foundation (Neurofibromatosis Preclinical Consortium award), the National Institutes of Health (grant R37 CA72614), the Leukemia & Lymphoma Society of America (Specialized Center of Research award), a St Baldrick's Foundation Scholar award (B.S.B.), and a Hyundai Hope on Wheels Fellowship (A.F.W.). K.M.S. is an American Cancer Society Research Professor.

National Institutes of Health

Authorship

Contribution: A.F.W., B.S.B., and K.M.S. wrote the manuscript.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Kevin M. Shannon, Helen Diller Family Cancer Research Building, University of California, San Francisco, 1450 3rd St, Room 240, San Francisco, CA 94158-9001; e-mail: shannonk@peds.ucsf.edu.

References

1
Hanahan
 
D
Weinberg
 
RA
The hallmarks of cancer.
Cell
2000
, vol. 
100
 
1
(pg. 
57
-
70
)
2
Schubbert
 
S
Shannon
 
K
Bollag
 
G
Hyperactive Ras in developmental disorders and cancer.
Nat Rev Cancer
2007
, vol. 
7
 
4
(pg. 
295
-
308
)
3
Malumbres
 
M
Barbacid
 
M
RAS oncogenes: the first 30 years.
Nat Rev Cancer
2003
, vol. 
3
 
6
(pg. 
459
-
465
)
4
Dunbar
 
AJ
Gondek
 
LP
O'Keefe
 
CL
, et al. 
250K single nucleotide polymorphism array karyotyping identifies acquired uniparental disomy and homozygous mutations, including novel missense substitutions of c-Cbl, in myeloid malignancies.
Cancer Res
2008
, vol. 
68
 
24
(pg. 
10349
-
10357
)
5
Nakao
 
M
Yokota
 
S
Iwai
 
T
, et al. 
Internal tandem duplication of the flt3 gene found in acute myeloid leukemia.
Leukemia
1996
, vol. 
10
 
12
(pg. 
1911
-
1918
)
6
Nakata
 
Y
Kimura
 
A
Katoh
 
O
, et al. 
C-kit point mutation of extracellular domain in patients with myeloproliferative disorders.
Br J Haematol
1995
, vol. 
91
 
3
(pg. 
661
-
663
)
7
Tartaglia
 
M
Niemeyer
 
CM
Fragale
 
A
, et al. 
Somatic mutations in PTPN11 in juvenile myelomonocytic leukemia, myelodysplastic syndromes and acute myeloid leukemia.
Nat Genet
2003
, vol. 
34
 
2
(pg. 
148
-
150
)
8
Cichowski
 
K
Jacks
 
T
NF1 tumor suppressor gene function: narrowing the GAP.
Cell
2001
, vol. 
104
 
4
(pg. 
593
-
604
)
9
Vetter
 
IR
Wittinghofer
 
A
The guanine nucleotide-binding switch in three dimensions.
Science
2001
, vol. 
294
 
5545
(pg. 
1299
-
1304
)
10
Braun
 
BS
Shannon
 
K
Targeting Ras in myeloid leukemias.
Clin Cancer Res
2008
, vol. 
14
 
8
(pg. 
2249
-
2252
)
11
Sawyers
 
CL
Denny
 
CT
Chronic myelomonocytic leukemia: Tel-a-kinase what Ets all about.
Cell
1994
, vol. 
77
 
2
(pg. 
171
-
173
)
12
Repasky
 
GA
Chenette
 
EJ
Der
 
CJ
Renewing the conspiracy theory debate: does Raf function alone to mediate Ras oncogenesis?
Trends Cell Biol
2004
, vol. 
14
 
11
(pg. 
639
-
647
)
13
Polak
 
R
Buitenhuis
 
M
The PI3K/PKB signaling module as key regulator of hematopoiesis: implications for therapeutic strategies in leukemia.
Blood
2012
, vol. 
119
 
4
(pg. 
911
-
923
)
14
Bollag
 
G
McCormick
 
F
Differential regulation of rasGAP and neurofibromatosis gene product activities.
Nature
1991
, vol. 
351
 
6327
(pg. 
576
-
579
)
15
Mor
 
A
Philips
 
MR
Compartmentalized Ras/MAPK signaling.
Annu Rev Immunol
2006
, vol. 
24
 (pg. 
771
-
800
)
16
Greaves
 
J
Chamberlain
 
LH
DHHC palmitoyl transferases: substrate interactions and (patho)physiology.
Trends Biochem Sci
2011
, vol. 
36
 
5
(pg. 
245
-
253
)
17
Rocks
 
O
Peyker
 
A
Kahms
 
M
, et al. 
An acylation cycle regulates localization and activity of palmitoylated Ras isoforms.
Science
2005
, vol. 
307
 
5716
(pg. 
1746
-
1752
)
18
Maurer
 
T
Garrenton
 
LS
Oh
 
A
, et al. 
Small-molecule ligands bind to a distinct pocket in Ras and inhibit SOS-mediated nucleotide exchange activity.
Proc Natl Acad Sci U S A
2012
, vol. 
109
 
14
(pg. 
5299
-
5304
)
19
Lauchle
 
JO
Kim
 
D
Le
 
DT
, et al. 
Response and resistance to MEK inhibition in leukaemias initiated by hyperactive Ras.
Nature
2009
, vol. 
461
 
7262
(pg. 
411
-
414
)
20
Loh
 
ML
Recent advances in the pathogenesis and treatment of juvenile myelomonocytic leukaemia.
Br J Haematol
2011
, vol. 
152
 
6
(pg. 
677
-
687
)
21
Onida
 
F
Kantarjian
 
HM
Smith
 
TL
, et al. 
Prognostic factors and scoring systems in chronic myelomonocytic leukemia: a retrospective analysis of 213 patients.
Blood
2002
, vol. 
99
 
3
(pg. 
840
-
849
)
22
Ricci
 
C
Fermo
 
E
Corti
 
S
, et al. 
RAS mutations contribute to evolution of chronic myelomonocytic leukemia to the proliferative variant.
Clin Cancer Res
2010
, vol. 
16
 
8
(pg. 
2246
-
2256
)
23
Onida
 
F
Beran
 
M
Chronic myelomonocytic leukemia: myeloproliferative variant.
Curr Hematol Rep
2004
, vol. 
3
 
3
(pg. 
218
-
226
)
24
Jankowska
 
AM
Makishima
 
H
Tiu
 
RV
, et al. 
Mutational spectrum analysis of chronic myelomonocytic leukemia includes genes associated with epigenetic regulation: UTX, EZH2, and DNMT3A.
Blood
2011
, vol. 
118
 
14
(pg. 
3932
-
3941
)
25
Kuo
 
M-C
Liang
 
D-C
Huang
 
C-F
, et al. 
RUNX1 mutations are frequent in chronic myelomonocytic leukemia and mutations at the C-terminal region might predict acute myeloid leukemia transformation.
Leukemia
2009
, vol. 
23
 
8
(pg. 
1426
-
1431
)
26
Bacher
 
U
Haferlach
 
T
Schoch
 
C
Kern
 
W
Schnittger
 
S
Implications of NRAS mutations in AML: a study of 2502 patients.
Blood
2006
, vol. 
107
 
10
(pg. 
3847
-
3853
)
27
Bowen
 
DT
Frew
 
ME
Hills
 
R
, et al. 
RAS mutation in acute myeloid leukemia is associated with distinct cytogenetic subgroups but does not influence outcome in patients younger than 60 years.
Blood
2005
, vol. 
106
 
6
(pg. 
2113
-
2119
)
28
Vogelstein
 
B
Civin
 
CI
Preisinger
 
AC
, et al. 
RAS gene mutations in childhood acute myeloid leukemia: a Pediatric Oncology Group study.
Genes Chromosomes Cancer
1990
, vol. 
2
 
2
(pg. 
159
-
162
)
29
Berman
 
JN
Gerbing
 
RB
Alonzo
 
TA
, et al. 
Prevalence and clinical implications of NRAS mutations in childhood AML: a report from the Children's Oncology Group.
Leukemia
2011
, vol. 
25
 
6
(pg. 
1039
-
1042
)
30
Neubauer
 
A
Maharry
 
K
Mrózek
 
K
, et al. 
Patients with acute myeloid leukemia and RAS mutations benefit most from postremission high-dose cytarabine: a Cancer and Leukemia Group B study.
J Clin Oncol
2008
, vol. 
26
 
28
(pg. 
4603
-
4609
)
31
Tyner
 
JW
Erickson
 
H
Deininger
 
MWN
, et al. 
High-throughput sequencing screen reveals novel, transforming RAS mutations in myeloid leukemia patients.
Blood
2009
, vol. 
113
 
8
(pg. 
1749
-
1755
)
32
Parkin
 
B
Ouillette
 
P
Wang
 
Y
, et al. 
NF1 inactivation in adult acute myelogenous leukemia.
Clin Cancer Res
2010
, vol. 
16
 
16
(pg. 
4135
-
4147
)
33
Kaneko
 
Y
Maseki
 
N
Sakurai
 
M
, et al. 
Chromosome pattern in juvenile chronic myelogenous leukemia, myelodysplastic syndrome, and acute leukemia associated with neurofibromatosis.
Leukemia
1989
, vol. 
3
 
1
(pg. 
36
-
41
)
34
Kotecha
 
N
Flores
 
NJ
Irish
 
JM
, et al. 
Single-cell profiling identifies aberrant STAT5 activation in myeloid malignancies with specific clinical and biologic correlates.
Cancer Cell
2008
, vol. 
14
 
4
(pg. 
335
-
343
)
35
Braun
 
BS
Tuveson
 
DA
Kong
 
N
, et al. 
Somatic activation of oncogenic Kras in hematopoietic cells initiates a rapidly fatal myeloproliferative disorder.
Proc Natl Acad Sci U S A
2004
, vol. 
101
 
2
(pg. 
597
-
602
)
36
Kelly
 
LM
Gilliland
 
DG
Genetics of myeloid leukemias.
Annu Rev Genomics Hum Genet
2002
, vol. 
3
 (pg. 
179
-
198
)
37
Vainchenker
 
W
Delhommeau
 
F
Constantinescu
 
SN
Bernard
 
OA
New mutations and pathogenesis of myeloproliferative neoplasms.
Blood
2011
, vol. 
118
 
7
(pg. 
1723
-
1735
)
38
Nakano
 
Y
Kiyoi
 
H
Miyawaki
 
S
, et al. 
Molecular evolution of acute myeloid leukaemia in relapse: unstable N-ras and FLT3 genes compared with p53 gene.
Br J Haematol
1999
, vol. 
104
 
4
(pg. 
659
-
664
)
39
Nakagawa
 
T
Saitoh
 
S
Imoto
 
S
, et al. 
Multiple point mutation of N-ras and K-ras oncogenes in myelodysplastic syndrome and acute myelogenous leukemia.
Oncology
1992
, vol. 
49
 
2
(pg. 
114
-
122
)
40
Piloto
 
O
Wright
 
M
Brown
 
P
, et al. 
Prolonged exposure to FLT3 inhibitors leads to resistance via activation of parallel signaling pathways.
Blood
2007
, vol. 
109
 
4
(pg. 
1643
-
1652
)
41
Bacher
 
U
Haferlach
 
T
Kern
 
W
Haferlach
 
C
Schnittger
 
S
A comparative study of molecular mutations in 381 patients with myelodysplastic syndrome and in 4130 patients with acute myeloid leukemia.
Haematologica
2007
, vol. 
92
 
6
(pg. 
744
-
752
)
42
Bejar
 
R
Stevenson
 
K
Abdel-Wahab
 
O
, et al. 
Clinical effect of point mutations in myelodysplastic syndromes.
N Engl J Med
2011
, vol. 
364
 
26
(pg. 
2496
-
2506
)
43
Kolquist
 
KA
Schultz
 
RA
Furrow
 
A
, et al. 
Microarray-based comparative genomic hybridization of cancer targets reveals novel, recurrent genetic aberrations in the myelodysplastic syndromes.
Cancer Genet
2011
, vol. 
204
 
11
(pg. 
603
-
628
)
44
Bejar
 
R
Levine
 
R
Ebert
 
BL
Unraveling the molecular pathophysiology of myelodysplastic syndromes.
J Clin Oncol
2011
, vol. 
29
 
5
(pg. 
504
-
515
)
45
Van Etten
 
RA
Shannon
 
KM
Focus on myeloproliferative diseases and myelodysplastic syndromes.
Cancer Cell
2004
, vol. 
6
 
6
(pg. 
547
-
552
)
46
Flex
 
E
Petrangeli
 
V
Stella
 
L
, et al. 
Somatically acquired JAK1 mutations in adult acute lymphoblastic leukemia.
J Exp Med
2008
, vol. 
205
 
4
(pg. 
751
-
758
)
47
Neri
 
A
Knowles
 
DM
Greco
 
A
McCormick
 
F
Dalla-Favera
 
R
Analysis of RAS oncogene mutations in human lymphoid malignancies.
Proc Natl Acad Sci U S A
1988
, vol. 
85
 
23
(pg. 
9268
-
9272
)
48
Zhang
 
J
Ding
 
L
Holmfeldt
 
L
, et al. 
The genetic basis of early T-cell precursor acute lymphoblastic leukaemia.
Nature
2012
, vol. 
481
 
7380
(pg. 
157
-
163
)
49
Perentesis
 
JP
Bhatia
 
S
Boyle
 
E
, et al. 
RAS oncogene mutations and outcome of therapy for childhood acute lymphoblastic leukemia.
Leukemia
2004
, vol. 
18
 
4
(pg. 
685
-
692
)
50
Coustan-Smith
 
E
Mullighan
 
CG
Onciu
 
M
, et al. 
Early T-cell precursor leukaemia: a subtype of very high-risk acute lymphoblastic leukaemia.
Lancet Oncol
2009
, vol. 
10
 
2
(pg. 
147
-
156
)
51
Balgobind
 
BV
Van Vlierberghe
 
P
van den Ouweland
 
AMW
, et al. 
Leukemia-associated NF1 inactivation in patients with pediatric T-ALL and AML lacking evidence for neurofibromatosis.
Blood
2008
, vol. 
111
 
8
(pg. 
4322
-
4328
)
52
Chng
 
WJ
Gonzalez-Paz
 
N
Price-Troska
 
T
, et al. 
Clinical and biological significance of RAS mutations in multiple myeloma.
Leukemia
2008
, vol. 
22
 
12
(pg. 
2280
-
2284
)
53
Liu
 
P
Leong
 
T
Quam
 
L
, et al. 
Activating mutations of N- and K-ras in multiple myeloma show different clinical associations: analysis of the Eastern Cooperative Oncology Group Phase III Trial.
Blood
1996
, vol. 
88
 
7
(pg. 
2699
-
2706
)
54
Corradini
 
P
Ladetto
 
M
Voena
 
C
, et al. 
Mutational activation of N- and K-ras oncogenes in plasma cell dyscrasias.
Blood
1993
, vol. 
81
 
10
(pg. 
2708
-
2713
)
55
Rasmussen
 
T
Kuehl
 
M
Lodahl
 
M
Johnsen
 
HE
Dahl
 
IMS
Possible roles for activating RAS mutations in the MGUS to MM transition and in the intramedullary to extramedullary transition in some plasma cell tumors.
Blood
2005
, vol. 
105
 
1
(pg. 
317
-
323
)
56
Chan
 
IT
Kutok
 
JL
Williams
 
IR
, et al. 
Conditional expression of oncogenic K-ras from its endogenous promoter induces a myeloproliferative disease.
J Clin Invest
2004
, vol. 
113
 
4
(pg. 
528
-
538
)
57
Haigis
 
KM
Kendall
 
KR
Wang
 
Y
, et al. 
Differential effects of oncogenic K-Ras and N-Ras on proliferation, differentiation and tumor progression in the colon.
Nat Genet
2008
, vol. 
40
 
5
(pg. 
600
-
608
)
58
Li
 
Q
Haigis
 
KM
McDaniel
 
A
, et al. 
Hematopoiesis and leukemogenesis in mice expressing oncogenic NrasG12D from the endogenous locus.
Blood
2011
, vol. 
117
 
6
(pg. 
2022
-
2032
)
59
Wang
 
J
Liu
 
Y
Li
 
Z
, et al. 
Endogenous oncogenic Nras mutation initiates hematopoietic malignancies in a dose- and cell type-dependent manner.
Blood
2011
, vol. 
118
 
2
(pg. 
368
-
379
)
60
Xu
 
J
Hedberg
 
C
Dekker
 
FJ
, et al. 
Inhibiting the palmitoylation/depalmitoylation cycle selectively reduces the growth of hematopoietic cells expressing oncogenic Nras.
Blood
2012
, vol. 
119
 
4
(pg. 
1032
-
1035
)
61
Holmes
 
KL
Pierce
 
JH
Davidson
 
WF
Morse
 
HC
Murine hematopoietic cells with pre-B or pre-B/myeloid characteristics are generated by in vitro transformation with retroviruses containing fes, ras, abl, and src oncogenes.
J Exp Med
1986
, vol. 
164
 
2
(pg. 
443
-
457
)
62
Dunbar
 
CE
Crosier
 
PS
Nienhuis
 
AW
Introduction of an activated RAS oncogene into murine bone marrow lymphoid progenitors via retroviral gene transfer results in thymic lymphomas.
Oncogene Res
1991
, vol. 
6
 
1
(pg. 
39
-
51
)
63
Hawley
 
RG
Fong
 
AZ
Ngan
 
BY
Hawley
 
TS
Hematopoietic transforming potential of activated ras in chimeric mice.
Oncogene
1995
, vol. 
11
 
6
(pg. 
1113
-
1123
)
64
Johnson
 
L
Mercer
 
K
Greenbaum
 
D
, et al. 
Somatic activation of the K-ras oncogene causes early onset lung cancer in mice.
Nature
2001
, vol. 
410
 
6832
(pg. 
1111
-
1116
)
65
Sabnis
 
AJ
Cheung
 
LS
Dail
 
M
, et al. 
Oncogenic Kras initiates leukemia in hematopoietic stem cells.
PLoS Biol
2009
, vol. 
7
 
3
pg. 
e59
 
66
Dail
 
M
Li
 
Q
McDaniel
 
A
, et al. 
Mutant Ikzf1, KrasG12D, and Notch1 cooperate in T lineage leukemogenesis and modulate responses to targeted agents.
Proc Natl Acad Sci U S A
2010
, vol. 
107
 
11
(pg. 
5106
-
5111
)
67
Zhang
 
J
Wang
 
J
Liu
 
Y
, et al. 
Oncogenic Kras-induced leukemogeneis: hematopoietic stem cells as the initial target and lineage-specific progenitors as the potential targets for final leukemic transformation.
Blood
2009
, vol. 
113
 
6
(pg. 
1304
-
1314
)
68
Chiang
 
MY
Xu
 
L
Shestova
 
O
, et al. 
Leukemia-associated NOTCH1 alleles are weak tumor initiators but accelerate K-ras-initiated leukemia.
J Clin Invest
2008
, vol. 
118
 
9
(pg. 
3181
-
3194
)
69
MacKenzie
 
KL
Dolnikov
 
A
Millington
 
M
Shounan
 
Y
Symonds
 
G
Mutant N-ras induces myeloproliferative disorders and apoptosis in bone marrow repopulated mice.
Blood
1999
, vol. 
93
 
6
(pg. 
2043
-
2056
)
70
Parikh
 
C
Subrahmanyam
 
R
Ren
 
R
Oncogenic NRAS rapidly and efficiently induces CMML- and AML-like diseases in mice.
Blood
2006
, vol. 
108
 
7
(pg. 
2349
-
2357
)
71
Parikh
 
C
Subrahmanyam
 
R
Ren
 
R
Oncogenic NRAS, KRAS, and HRAS exhibit different leukemogenic potentials in mice.
Cancer Res
2007
, vol. 
67
 
15
(pg. 
7139
-
7146
)
72
Chao
 
RC
Pyzel
 
U
Fridlyand
 
J
, et al. 
Therapy-induced malignant neoplasms in Nf1 mutant mice.
Cancer Cell
2005
, vol. 
8
 
4
(pg. 
337
-
348
)
73
Nakamura
 
JL
Phong
 
C
Pinarbasi
 
E
, et al. 
Dose-dependent effects of focal fractionated irradiation on secondary malignant neoplasms in Nf1 mutant mice.
Cancer Res
2011
, vol. 
71
 
1
(pg. 
106
-
115
)
74
Le
 
DT
Kong
 
N
Zhu
 
Y
, et al. 
Somatic inactivation of Nf1 in hematopoietic cells results in a progressive myeloproliferative disorder.
Blood
2004
, vol. 
103
 
11
(pg. 
4243
-
4250
)
75
Wang
 
J
Liu
 
Y
Li
 
Z
, et al. 
Endogenous oncogenic Nras mutation promotes aberrant GM-CSF signaling in granulocytic/monocytic precursors in a murine model of chronic myelomonocytic leukemia.
Blood
2010
, vol. 
116
 
26
(pg. 
5991
-
6002
)
76
Downward
 
J
Targeting RAS signalling pathways in cancer therapy.
Nat Rev Cancer
2003
, vol. 
3
 
1
(pg. 
11
-
22
)
77
Karp
 
JE
Lancet
 
JE
Kaufmann
 
SH
, et al. 
Clinical and biologic activity of the farnesyltransferase inhibitor R115777 in adults with refractory and relapsed acute leukemias: a phase 1 clinical-laboratory correlative trial.
Blood
2001
, vol. 
97
 
11
(pg. 
3361
-
3369
)
78
Mahgoub
 
N
Taylor
 
BR
Gratiot
 
M
, et al. 
In vitro and in vivo effects of a farnesyltransferase inhibitor on Nf1-deficient hematopoietic cells.
Blood
1999
, vol. 
94
 
7
(pg. 
2469
-
2476
)
79
Castleberry
 
RP
Loh
 
ML
Jayaprakash
 
N
, et al. 
Phase II window study of the farnesyltransferase inhibitor R115777 (Zarnestra) in untreated juvenile myelomonocytic leukemia (JMML): a Children's Oncology Group Study.
Blood
2005
, vol. 
106
 
Suppl 1
(pg. 
727a
-
728a
)
80
Wahlstrom
 
AM
Cutts
 
BA
Karlsson
 
C
, et al. 
Rce1 deficiency accelerates the development of K-RAS-induced myeloproliferative disease.
Blood
2007
, vol. 
109
 
2
(pg. 
763
-
768
)
81
Wahlstrom
 
AM
Cutts
 
BA
Liu
 
M
, et al. 
Inactivating Icmt ameliorates K-RAS-induced myeloproliferative disease.
Blood
2008
, vol. 
112
 
4
(pg. 
1357
-
1365
)
82
Wang
 
M
Hossain
 
MS
Tan
 
W
, et al. 
Inhibition of isoprenylcysteine carboxylmethyltransferase induces autophagic-dependent apoptosis and impairs tumor growth.
Oncogene
2010
, vol. 
29
 
35
(pg. 
4959
-
4970
)
83
Cuiffo
 
B
Ren
 
R
Palmitoylation of oncogenic NRAS is essential for leukemogenesis.
Blood
2010
, vol. 
115
 
17
(pg. 
3598
-
3605
)
84
Dekker
 
FJ
Rocks
 
O
Vartak
 
N
, et al. 
Small-molecule inhibition of APT1 affects Ras localization and signaling.
Nat Chem Biol
2010
, vol. 
6
 
6
(pg. 
449
-
456
)
85
Hedberg
 
C
Dekker
 
FJ
Rusch
 
M
, et al. 
Development of highly potent inhibitors of the Ras-targeting human Acyl protein thioesterases based on substrate similarity design [published online ahead of print September 9, 2011].
Angew Chem Int Ed Engl
 
86
Birnbaum
 
RA
O'Marcaigh
 
A
Wardak
 
Z
, et al. 
Nf1 and Gmcsf interact in myeloid leukemogenesis.
Mol Cell
2000
, vol. 
5
 
1
(pg. 
189
-
195
)
87
Kim
 
A
Morgan
 
K
Hasz
 
DE
, et al. 
Beta common receptor inactivation attenuates myeloproliferative disease in Nf1 mutant mice.
Blood
2007
, vol. 
109
 
4
(pg. 
1687
-
1691
)
88
Van Meter
 
MEM
Díaz-Flores
 
E
Archard
 
JA
, et al. 
K-RasG12D expression induces hyperproliferation and aberrant signaling in primary hematopoietic stem/progenitor cells.
Blood
2007
, vol. 
109
 
9
(pg. 
3945
-
3952
)
89
Cutts
 
BA
Sjogren
 
A-KM
Andersson
 
KME
, et al. 
Nf1 deficiency cooperates with oncogenic K-RAS to induce acute myeloid leukemia in mice.
Blood
2009
, vol. 
114
 
17
(pg. 
3629
-
3632
)
90
Gutierrez
 
A
Sanda
 
T
Grebliunaite
 
R
, et al. 
High frequency of PTEN, PI3K, and AKT abnormalities in T-cell acute lymphoblastic leukemia.
Blood
2009
, vol. 
114
 
3
(pg. 
647
-
650
)
91
Chapman
 
MA
Lawrence
 
MS
Keats
 
JJ
, et al. 
Initial genome sequencing and analysis of multiple myeloma.
Nature
2011
, vol. 
471
 
7339
(pg. 
467
-
472
)
92
Borthakur
 
G
Kantarjian
 
H
Ravandi
 
F
, et al. 
Phase I study of sorafenib in patients with refractory or relapsed acute leukemias.
Haematologica
2011
, vol. 
96
 
1
(pg. 
62
-
68
)
93
Wilhelm
 
SM
Carter
 
C
Tang
 
L
, et al. 
BAY 43–9006 exhibits broad spectrum oral antitumor activity and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases involved in tumor progression and angiogenesis.
Cancer Res
2004
, vol. 
64
 
19
(pg. 
7099
-
7109
)
94
Fathi
 
AT
Chabner
 
BA
FLT3 inhibition as therapy in acute myeloid leukemia: a record of trials and tribulations.
Oncologist
2011
, vol. 
16
 
8
(pg. 
1162
-
1174
)
95
Lyubynska
 
N
Gorman
 
MF
Lauchle
 
JO
, et al. 
A MEK inhibitor abrogates myeloproliferative disease in Kras mutant mice.
Sci Transl Med
2011
, vol. 
3
 
76
pg. 
76ra27
 
96
Solit
 
DB
Garraway
 
LA
Pratilas
 
CA
, et al. 
BRAF mutation predicts sensitivity to MEK inhibition.
Nature
2006
, vol. 
439
 
7074
(pg. 
358
-
362
)
97
Odenike
 
O
Curran
 
E
Iyengar
 
N
, et al. 
Phase II study of the oral MEK inhibitor AZD6244 in advanced acute myeloid leukemia (AML) [abstract].
Blood (ASH Annual Meeting Abstracts)
2009
, vol. 
114
 
98
Scholl
 
C
Fröhling
 
S
Dunn
 
IF
, et al. 
Synthetic lethal interaction between oncogenic KRAS dependency and STK33 suppression in human cancer cells.
Cell
2009
, vol. 
137
 
5
(pg. 
821
-
834
)
99
Barbie
 
DA
Tamayo
 
P
Boehm
 
JS
, et al. 
Systematic RNA interference reveals that oncogenic KRAS-driven cancers require TBK1.
Nature
2009
, vol. 
462
 
7269
(pg. 
108
-
112
)
100
Luo
 
J
Emanuele
 
MJ
Li
 
D
, et al. 
A genome-wide RNAi screen identifies multiple synthetic lethal interactions with the Ras oncogene.
Cell
2009
, vol. 
137
 
5
(pg. 
835
-
848
)
101
Kumar
 
MS
Hancock
 
DC
Molina-Arcas
 
M
, et al. 
The GATA2 transcriptional network is requisite for RAS oncogene-driven non-small cell lung cancer.
Cell
2012
, vol. 
149
 
3
(pg. 
642
-
655
)
102
Cortes
 
J
Perl
 
A
Smith
 
C
, et al. 
A phase II open-label, AC220 monotherapy efficacy (ACE) study in patients with acute myeloid leukemia (AML) with Flt3-ITD activating mutations: interim results.
Haematologica
2011
, vol. 
96
 
Suppl 2
pg. 
426
 
103
Smith
 
CC
Wang
 
Q
Chin
 
C-S
, et al. 
Validation of ITD mutations in FLT3 as a therapeutic target in human acute myeloid leukaemia.
Nature
2012
, vol. 
485
 
7397
(pg. 
260
-
263
)
104
Moore
 
AS
Faisal
 
A
de Castro
 
DG
, et al. 
Selective FLT3 inhibition of FLT3-ITD(+) acute myeloid leukaemia resulting in secondary D835Y mutation: a model for emerging clinical resistance patterns.
Leukemia
2012
, vol. 
26
 
7
(pg. 
1462
-
1470
)
105
Man
 
CH
Fung
 
TK
Ho
 
C
, et al. 
Sorafenib treatment of FLT3-ITD+ acute myeloid leukemia: favorable initial outcome and mechanisms of subsequent non-responsiveness associated with a D835 mutation.
Blood
2012
, vol. 
119
 
22
(pg. 
5133
-
5143
)
106
Ravandi
 
F
O'Brien
 
S
Thomas
 
D
, et al. 
First report of phase 2 study of dasatinib with hyper-CVAD for the frontline treatment of patients with Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia.
Blood
2010
, vol. 
116
 
12
(pg. 
2070
-
2077
)
107
Bivona
 
TG
Hieronymus
 
H
Parker
 
J
, et al. 
FAS and NF-kappaB signalling modulate dependence of lung cancers on mutant EGFR.
Nature
2011
, vol. 
471
 
7339
(pg. 
523
-
526
)
108
Kindler
 
T
Cornejo
 
MG
Scholl
 
C
, et al. 
K-RasG12D-induced T-cell lymphoblastic lymphoma/leukemias harbor Notch1 mutations and are sensitive to gamma-secretase inhibitors.
Blood
2008
, vol. 
112
 
8
(pg. 
3373
-
3382
)
109
Jacks
 
T
Shih
 
TS
Schmitt
 
EM
, et al. 
Tumour predisposition in mice heterozygous for a targeted mutation in Nf1.
Nat Genet
1994
, vol. 
7
 
3
(pg. 
353
-
361
)
110
Largaespada
 
DA
Brannan
 
CI
Jenkins
 
NA
Copeland
 
NG
Nf1 deficiency causes Ras-mediated granulocyte/macrophage colony stimulating factor hypersensitivity and chronic myeloid leukaemia.
Nat Genet
1996
, vol. 
12
 
2
(pg. 
137
-
143
)