Mark G. Kris

Purpose: All targeted therapies approved for ROS1 fusion–positive non–small cell lung cancer (NSCLC) are type I inhibitors. These tyrosine kinase inhibitors (TKI) may induce resistance amenable to type II inhibition. Cabozantinib is a type II ROS1/MET TKI with preclinical activity against on-target ROS1 resistance mutations and MET alterations. Patients and Methods: This single-center, open-label, phase II study (NCT01639508) enrolled patients with pretreated ROS1 fusion–positive NSCLC. The primary endpoint was the objective response rate (ORR). Secondary endpoints were progression-free survival (PFS), overall survival (OS), and safety. The first phase of the Simon two-stage design is reported. Results: Seven ROS1 TKI pretreated patients were enrolled. Previous treatment included crizotinib, entrectinib, repotrectinib, and zidesamtinib. The ORR was 43% [95% confidence interval (CI), 10%–81.5%]. The best response was −92% partial response (PR) in a patient with ROS1 D2033N–mutant cancer. Two additional PRs were achieved, one (−49%) in an ROS1 G2032R/L2086F–mutant cancer. The median PFS was 4.5 months [95% CI, 4–not estimable (NE)] with a median duration of response and OS of 8 (95% CI, 3–13) and 11 months (95% CI, 6–NE), respectively. Resistance mechanisms to cabozantinib were identified in two patients: MET overexpression and MET D1228N. The most frequent treatment-related toxicities observed were fatigue, nausea, and hypertension. Grade 3 treatment-related adverse events occurred in 57%, mostly palmar–plantar erythrodysesthesia (PPE) and transaminitis. All patients required a dose reduction. Conclusions: Cabozantinib achieved responses in pretreated ROS1 fusion–positive NSCLCs, even in the presence of multidrug-resistant mutations, highlighting the potential utility of type II ROS1 TKI therapy.

Supplementary Figure 1: Characteristics of the IMPACT cohort. Violin plots of (A) age at sample acquisition and (B) age at patient’s first diagnosis separated by amplification status of MYCf genes. Bar plot showing (C) the stage of the cancer at the patient’s first diagnosis and (D) patient sex separated by MYCf amplification status. (E-G) Scatter plot showing the correlation between fraction of genome altered (FGA) and segment size for amplifications in (E) MYC, (F) MYCN, and (G) MYCL. On each plot is shown Pearson’s R and p-value. (H) Violin plots showing the clonality (cellular fraction divided by sample purity derived from FACETS) of MYCf amplifications.

Supplementary Figure 2: Expression of MYC and MYCL in various cancer types. (A) Boxplot of the expression of MYCL separated by cancer type. Cancer types were included if they contained at least five samples with MYCL amplifications. Each point represents one sample, and samples are separated by amplification status of MYCL. BH-adjusted p-values are shown. (B-C) Boxplots of ssGSEA for samples in (Figure 3A) for two Hallmark pathways: MYC Targets V1 (B) and MYC Targets V2 (C). Nominal p-values are shown when below 0.05. (D-H) Boxplots comparing expression of MYC across cancer types, separated by alteration status of five genes that were identified through cross-validation GLM-NET as correlating with MYC expression: CDKN2A, ARID4A, PIK3CA, TERT, and TP53, respectively. (I) Boxplots comparing expression of MYC in HR positive breast carcinoma, separated by amplification status of MYC and alterations in PIK3CA. For (A-I), expression is shown as natural log of RSEM, the size of each point represents the ploidy-corrected total copy number of MYC, and p-values are derived from Wilcoxon tests.

Supplementary Figure 3: Overall survival analysis for six cancer types. Multivariate analysis of overall survival for patients with MYC amplifications. Cancer types are as follows: (A) hormone receptor (HR) positive breast cancer, (B) colorectal cancer, (C) lung adenocarcinoma, (D), ovarian cancer, (E) prostate cancers, (F) and uterine cancer. Covariates include fraction of genome altered (FGA), patient sex, age at sample acquisition, age and stage at first diagnosis, and binary variables for focality of the amplification, the amplitude of the total copy number compared to the median, and alterations in cooccurring genes. Hazard Ratios (HR), 95% confidence intervals (CI) and nominal p-values are shown to the right. BH-adjusted p-values can be found in Supplementary Table 7.

Summarizes considerations relevant to the representativeness and generalizability of patient cohorts in studies of the cancer types evaluated.

Number of samples from TCGA cohort used in expression analysis separated by type of MYCf amplification. If cancer type was not used in a comparison, values appear as NA.

Values derived from the cross-validated GLM-NET modeling expression by ploidy-corrected copy number, cancer type, and genes with alterations co-occurring with MYC amplifications across cancer types in TCGA samples.

Values derived from the cross-validated GLM-NET modeling expression by ploidy-corrected copy number, and genes with alterations co-occurring with MYC amplifications in each of six cancer types in TCGA samples.

Comparison of focal and broad MYC amplification frequency across the most common cancer types in Figure 1A. For each cancer type, the ratio was calculated as the number of focal divided by the number of total MYC amplifications. The background, expected ratio was based on all cancer types in Figure 1A, excluding the cancer type in question. Binomial test was used for comparison with nominal and BH-corrected p-values shown.

Co-occurrence and mutual exclusivity of MYCf genes as depicted in Figure 2. Columns include gene or gene focality from Figure 2A and B, respectively, alteration being tested, the number of samples per alteration combination, odds ratio (OR), q-value, and cancer type.