abstract # 1104 repotrectinib increases effectiveness of
TRANSCRIPT
Contact – Jim Mazzola: [email protected]
INTRODUCTION
• KRAS is the most frequently mutated oncogene in human cancers1 including ~30% of non-small cell lung cancer (NSCLC), ~50% of colorectal cancer (CRC) and ~90% of pancreatic cancer2
• Targeting downstream KRAS signaling by inhibiting MEK with trametinib showed limited activity in patients with mutant-KRAS NSCLC as a single agent or in combination with chemotherapy3,4
• Compensatory upregulation of the PI3K/AKT survival signaling or MEK inhibitor-induced JAK/STAT3 pathway activation are reported evasion mechanisms to MEK-targeted therapies5-7
• Repotrectinib is a compact macrocyclic inhibitor that potently inhibits SRC/FAK/JAK2 in biochemical, cellular, and in vivo assays8
• We previously screened 60 mutant-KRAS NSCLC, CRC, and pancreatic cell lines and found that repotrectinib in combination with trametinib resulted in a >10-fold increase in trametinib antiproliferative efficacy in 9 of 15 KRAS G12D mutant cancer models. We further evaluated repotrectinib combinations with other MAPK pathway inhibitors such as VS-6766 (MEK/RAFi)8
• Repotrectinib is currently being evaluated in a registrational phase 2 TRIDENT-1 study9 for patients with ROS1+ NSCLC and NTRK+ solid tumors and has been granted Breakthrough Therapy designation and 3 Fast Track designations by the FDA10
SYNERGY ANALYSIS FOR REPOTRECTINIB COMBINATIONSWITH MEK INHIBITORS IN VITRO
Figure 1. Repotrectinib Enhances Both Trametinib and VS-6766 Efficacy in A427 KRAS G12D NSCLC
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• Repotrectinib enhances inhibition of tumor cell viability by trametinib and VS-6766 in A427 KRAS G12D cancer cells grown in 2D and 3D spheroid cultures (Figures 1A and 1B)
• In 3D spheroid culture, repotrectinib synergizes with trametinib to inhibit tumor cell viability (Figure 1C). In contrast, VS-6766 has an additive effect (Figure 1D) (Bliss score 0-10 = additive; Bliss score >10 = synergistic)11
REPOTRECTINIB COMBINATIONS IN PATIENT-DERIVED KRASMUTANT EX VIVO MODELS
Figure 2. Repotrectinib Enhances Efficacy of MAPK Pathway Inhibitors in Patient-Derived KRAS G12D/V Lung and KRAS G12D/V/R Pancreatic Spheroid Models
LU6419ex KRAS G12VLU5178B KRAS G12DA
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• Repotrectinib enhances inhibition of tumor cell viability by trametinib and VS-6766 in patient-derived KRAS mutant lung and pancreatic cancer models (Figures 2A and 2C, respectively)
• Cell viability IC50 values of lung and pancreatic cancer spheroids treated with MAPK pathway inhibitors as single-agent treatment or in combination with repotrectinib are shown (Figures 2B and 2D)
• Repotrectinib combination with trametinib yielded superior potency compared to other MAPK pathway inhibitor combinations
SIGNALING MODULATION BY REPOTRECTINIB/TRAMETINIB COMBINATION
Figure 3. Repotrectinib/Trametinib Combination Suppresses Downstream Oncogenic Signaling in the A427 KRAS G12D NSCLC Cell Line
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• Repotrectinib potently inhibits SRC, FAK, and STAT3 phosphorylation (Figure 3A)
• Repotrectinib/trametinib combination suppresses both ERK and AKT phosphorylation, resulting in greater inhibition of S6 phosphorylation compared to either single-agent treatment (Figure 3B)
SIGNALING MODULATION BY REPOTRECTINIB/TRAMETINIB COMBINATION
Figure 4. Repotrectinib/Trametinib Combination Increases Cell Cycle Arrest and Induction of Apoptosis in A427 KRAS G12D NSCLC
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• Repotrectinib/trametinib combination yields greater inhibition of RB1 phosphorylation, along with upregulation of p27, p21 expression, and inhibition of cyclin E2 (CCNE2) expression (Figure 4A)
• Repotrectinib/trametinib combination yields greater induction of apoptosis, as evidenced by increased PARP and caspase 3 cleavage (Figure 4B)
• Flow cytometry analysis of A427 cells treated with a repotrectinib/trametinib combination for 48 h showed increased levels of annexin V+ early apoptotic cells relative to single-agent treatments (Figure 4C)
TRAMETINIB ACQUIRED-RESISTANCE MODELS
Figure 5. Repotrectinib Can Resensitize Trametinib-Resistant Cells
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• HCT116 KRAS G13D CRC and PSN1 KRAS G12R pancreatic cancer cells were rendered resistant to trametinib via long-term step-up exposure to trametinib
• Trametinib-resistant cells (HCT116 and PSN1) are >13- and >20-fold less sensitive to trametinib, respectively, compared to parental cells, and exhibit cross-resistance to selumetinib (Figures 5A and 5C)
• Trametinib-resistant cells can be resensitized to trametinib by repotrectinib (1 µM) (Figures 5B and 5D)
SIGNALING MODULATION IN ACQUIRED-RESISTANCE MODELS
Figure 6. Repotrectinib Resensitizes Trametinib-Resistant Cells by Repressing Compensatory AKT and S6 Activation
• Repotrectinib suppresses trametinib- induced hyperphosphorylation of AKT and STAT3 in both parental and trametinib-resistant cells (Figures 6A and 6B)
• Repotrectinib synergizes with trametinib, yielding greater inhibition of RB1 phosphorylation and upregulation of p27 expression in trametinib-resistant cells (Figures 6A and 6B)
IN VIVO EVALUATION OF REPOTRECTINIB/TRAMETINIB COMBINATION
Figure 7. Repotrectinib/Trametinib Combination Exhibits Greater Tumor Growth Inhibition in KRAS G12D Syngeneic and PDX Lung Cancer Models
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• Repotrectinb/trametinib combination demonstrates enhanced tumor growth inhibition compared to single-agent treatment in a murine syngeneic lung cancer model harboring KRAS G12D/p53 null (MuPrime mLU6045) (**** P<0.0001) (Figure 7) and in a human LU0876 KRAS G12D lung PDX model (P<0.05; data not shown)
CONCLUSIONS
• Repotrectinib combinations with MEK inhibitors (trametinib, VS-6766) are more effective than single-agent treatment in patient-derived KRAS mutant lung (KRAS G12D/V) and pancreatic (KRAS G12D/V/R) spheroid models
• Simultaneous inhibition of SRC/FAK/JAK2 by repotrectinib in mutant-KRAS tumor cell lines suppresses trametinib-induced AKT and STAT3 feedback activation
• Repotrectinib can restore trametinib efficacy in trametinib-resistant cancer cells by suppressing AKT and JAK2/STAT3 signaling, resulting in greater inhibition of both S6 and cell cycle progression
• Repotrectinib enhances trametinib efficacy in mutant KRAS G12D in vivo lung tumor models
• Overall, these preclinical data suggest that combination with repotrectinib can enhance trametinib potency and may improve durability of response by reversing or preventing acquired resistance
• A phase 1/2 combination study of repotrectinib and trametinib in KRAS-mutated solid tumors is currently planned
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References1. Haigis KM, et al. Trends Cancer. 2017;3(10):686-97.2. Prior IA, et al. Cancer Res. 2020;80(14):2969-74.3. Blumenschein GR, et al. Ann Oncol. 2015;26(5):894-901.4. Gadgeel SM, et al. J Clin Oncol. 2019;37(15):9021. 5. Lee HJ, et al. Cancer Cell. 2014;26(2):207-21. 6. Tsubaki M, et al. Cancers (Basel). 2019;11(12):1866.7. Kun E, et al. Cancer Treat Rev. 2021;92:102137.
8. Murray B, et al. Cancer Res. 2020;80(16):Abstract 1957.9. Clinicaltrials.gov. NCT03093116. Accessed February 17, 2021.10. Turning Point Therapeutics granted FDA breakthrough therapy designation for
repotrectinib treatment in patients with ROS1-positive metastatic non-small cell lung cancer who have not been treated with a ROS1 tyrosine kinase inhibitor [press release]. San Diego, CA: Turning Point Therapeutics, Inc.; December 8, 2020. Accessed March 8, 2021. https://ir.tptherapeutics.com/node/7601/pdf
11. Ianevski A, et al. Bioinformatics. 2017;33(15):2413-15.
Acknowledgements & Disclosures This study was funded by Turning Point Therapeutics. All authors are employees of Turning Point Therapeutics.
Editorial support was provided by BluPrint Oncology Concepts, LLC.
ProliferationSurvival Cytokines
SRCFAK RTK
Inhibited by Repotrectinib
KRAS
STAT3
JAK2
PI3K Raf Ral GEF
AKT MEK NF-κB
Repotrectinib Increases Effectiveness of MEK Inhibitors in KRASKRAS Mutant Cancer ModelsNathan V. Lee, Wei Deng, Dayong Zhai, Laura Rodon, Ana Parra, Jessica Cowell, Afsheen Banisadr, Xin Zhang, Brion W. Murray
Turning Point Therapeutics, 10628 Science Center Drive, Suite 200, San Diego, CA 92121
Abstract # 1104
Presented at American Association for Cancer Research (AACR), April 10–15, 2021