Clinical and molecular characterization of AXL in colorectal cancer: CALGB (Alliance)/SWOG 80405 and real-world data.
Authors: Ashouri Karam , Yang Yan , Millstein Joshua , Xiu Joanne , Soni Shivani , Algaze Sandra , Mittal Pooja , Wong Alexandra , Lo Jae Ho , Torres-Gonzalez Lesly , Jayachandran Priya , Zhang Wu , Shields Anthony F. , Goldberg Richard M. , Lou Emil , Weinberg Benjamin Adam , Marshall John , Meyer Aaron , Battaglin Francesca , Lenz Heinz-Josef 
Published: June 2024

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Background: Dysregulation of AXL signaling is linked with chemotherapy and targeted therapy resistance. However, it remains unclear which patients may benefit from therapies targeting AXL, given its role in both tumor cell intrinsic resistance and immune microenvironmental reprogramming. Here, we present a clinical and molecular characterization of AXL in colorectal cancer (CRC). Methods: 24,257 CRC samples tested at Caris Life Sciences (Phoenix, AZ) with WTS (Illumina NovaSeq), NextGen DNA sequencing (NextSeq, 592 genes and NovaSEQ, WES), and PD-L1 expression (22C3 clone; TPS ≥ 1%) were analyzed. RNA deconvolution analysis with QuantiSEQ estimated cell infiltration in the tumor microenvironment (TME). In this cohort, AXL expression was categorized as high (H) or low (L) using median cutoff: overall survival (OS) was evaluated from treatment initiation using insurance claims data. Data from the phase III CALGB/SWOG 80405 trial on 433 metastatic CRC (mCRC) patients treated with bevacizumab (Bev, n = 226) or cetuximab (Cet, n = 207) in combination with first-line chemotherapy were also evaluated. RNA isolated from FFPE tumor samples were sequenced with HiSeq 2500 (Illumina). OS and progression-free survival (PFS) were compared using categorical gene expression tertiles (high (T3), medium (T2), and low (T1)) adjusting for age, demographics, ECOG, and tumor/treatment characteristics. Results: BRAF and RNF43 mutations were more prevalent in AXL-H, but APC, NRAS, KRAS, and PIK3CA correlated with AXL-L (all q< .001). AXL-H had increased B cells, M1/M2 macrophages, T-regs, and NK cells in the TME while increased neutrophils and dendritic cells were associated with AXL-L (all q< .001). AXL-H had increased PD-L1 positivity (5.1 vs 2.6%) and pathway activation of epithelial-mesenchymal transition (EMT), inflammatory/IFN-G response, and TNF-a signaling (all q< .005). AXL-H demonstrated worse OS in FOLFOX/FOLFIRI treated CRC (H: 35.7 vs L: 38.7 months [mo], P = .003; HR 1.08, 95% CI [1.03-1.14]). The effect persisted within KRAS wildtype (wt) CRC (45.4 vs 40.1 mo, P = .005; HR 1.12 [1.04-1.22]) but not KRASmutant (mt). In 80405, higher AXLexpression was associated with worse PFS and OS ( Plinear = 2E-04 and 2.2E-05, respectively). AXL-T3 showed significantly shorter PFS (T3: 9.2 vs T2: 11.7 vs T1: 12.9 mo, T3 vs T1 (reference) adjusted HR 1.53 [1.19-1.97]) and OS (24.2 vs 34.0 vs 34.7 mo, adjusted HR 1.73 [1.33-2.25]). No significant interaction between AXL expression and treatment was observed for PFS and OS (likelihood ratio test). Conclusions: Our results indicate increased AXL expression is associated with immune cell infiltration, EMT, and inflammatory signaling. AXL-H confers worse OS/PFS on first-line chemotherapy, with a more pronounced effect in KRASwt CRC. This data supports evaluation of AXL as a prognostic marker and potential therapeutic target in CRC. Trial Identifier: NCT00265850.