Landscape and Clinicopathologic Features of RAS Pathway Mutations in Chronic Myelomonocytic Leukemia

Introduction: Somatic mutations in RAS pathway genes are present in ~30% of patients with chronic myelomonocytic leukemia (CMML) and are usually associated with myeloproliferative features. Whether mutations in distinct RAS pathway genes share similar genomic context and have similar implications remains unknown. Such an understanding might inform future studies evaluating genotype-specific therapies (RAS-codon specific and pan-RAS inhibitors) or cooperative leukemogenic mechanisms. This multicenter study examines the landscape of RAS pathway mutations (RAS^MT) in CMML.

Methods: Newly diagnosed CMML patients evaluated at MD Anderson Cancer Center (n=449, 54%) and Mayo Clinic (n=383, n=46%) were included after IRB approval. Clonal relationships were tested using Pearson goodness-of-fit. Clones with the highest variant allele frequency (VAF), or with VAF close to 40% were defined as dominant, and those present at VAF <20% in the presence of another dominant clone were defined as subclonal. RAS^MT genes included NRAS, KRAS, BRAF, CBL, CBLC, PTPN11 and NF1. Standard statistical measures were applied.

Results: Among 832 CMML patients (median age 71 years, 68% male) 387 (47%) had myeloproliferative (MP) and 445 (53%) had myelodysplastic (MD) CMML. A total of 350 (42%) patients had RAS^MT with higher prevalence among MP-CMML compared to MD-CMML (55% vs 31%, p<0.001). Among patients with RAS^MT, 262 (32%), 59 (7%), 20 (2%), 6 (<1%) and 3 (<1%) had 1, 2, 3, 4 or 5 distinct RAS^MT, respectively. Importantly, 17 patients harbored a RASmt as the only detectable somatic mutation, with KRAS^MT being the most frequent (n=10, 59%), including 3 (18%) patients with >1 RAS^MT. Among RAS^MT patients, a trend for higher hemoglobin (12.7 vs 10.8 g/dL, p=0.054) was observed in NF1^MT CMML and lower WBC (14.4 vs 27.9 x10⁹/L, p=0.067) values in CBL^MT CMML. In total, 479 somatic mutations in RAS pathway genes were identified the most frequent involving NRAS (n=169, 35%) followed by CBL (n=120, 25%), KRAS (n=102, 21%), PTPN11 (n=38, 8%), NF1 (n=34, 7%), BRAF (n=11, 2%) and CBLC (n=5, 1%). Most frequently involved codons for each RAS pathway gene were as follows: G594 (n=3) for BRAF, R420 (n=22) and C404 (n=15) for CBL, G12 (n=41; G12D: 13, G12C: 3) and A146 (n=16) for KRAS, G12 (n=116; G12D: 74, G12C: 5) for NRAS and A72 (n=8) for PTPN11.

CBL^MT CMML had higher ASXL1 (58% vs 44%, p=0.011), BRAF (8% vs 4%, p=0.017), IDH2 (10% vs 4%, p=0.024), KIT (5% vs 1%, p=0.038), SRSF2 (54% vs 40%, p=0.004), TET2 (59% vs 47%, p=0.019) and ZRSR2 (10% vs 4%, p=0.014) co-mutation frequency. Of note, DNMT3A mutations were mutually exclusive of CBL^MT CMML (0% vs 7%, p=0.007), and multihit TET2 was more common among TET2^MT/CBL^MT vs TET2^MT/CBL^WT CMML (82% vs 67%, p =0.039). CMML with NF1 mutations had higher BCORL1 (8% vs 0%, p=0.005), PTPN11 (15% vs 3%, p=0.014) and SETBP1 (23% vs 8%, p=0.022) mutational cooccurrences. NRAS^MT CMML was enriched for ASXL1 (53% vs 43%, p=0.042), ETNK1 (5% vs 1%, p=0.004), GATA2 (4% vs 1%, p=0.034) and STAG2 (4% vs 1 %, p=0.023) mutations but had lower JAK2 (1% vs 6%, p=0.036) and TP53 (1% vs 5%, p=0.022) mutational frequency. PTPN11 mutations were associated with higher NF1 (29% vs 7%, p=0.014), SETBP1 (27% vs 9%, p=0.01) and WT1 (6% vs <1%, p=0.014) mutational cooccurrences and lower SRSF2 (23% vs 42%, p=0.05) frequency.

We evaluated RAS mutant clonal dominance in 253 (72%) RAS^MT patients. NRAS (63% vs 38%), KRAS (63% vs 38%) and CBL (65% vs 35%) mutations were more frequently observed as dominant clones while PTPN11 (44% vs 56%), BRAF (25% vs 75%) and NF1 (35% vs 65%) mutations were more frequently subclonal events, particularly of TET2, ASXL1 or SRSF2 mutant clones.

Presence of a RAS pathway mutation was associated with shorter LFS (36 vs 53 months, HR 1.5, 95% CI 1.1-2.2, p=0.020), with presence of multiple RAS mutations shortening LFS further (53, 43, 27 months for 0, 1 and >1, p=0.017). PTPN11 mutations predicted for shorter OS (18 vs 39 months, HR 1.7, 95% CI 1.0-2.8, p=0.035) and LFS (14 vs 36 months, HR 1.9, 95% CI 1.2-2.9, p=0.008). Presence of >1 RAS pathway mutation was associated with shorter OS in ASXL1^MT/TET2^MT CMML compared to those with no RAS mutations (24 vs 53 months, HR 4.2, 95% CI 1.6-10.8, p=0.004).

Conclusions: RAS mutations exhibit distinct genomic features and clonal hierarchy in CMML and cooperate with other somatic mutations to drive clinical outcomes. This study forms the foundation for assessment of novel RAS pathway inhibitors in CMML.