MDR1 Drug Efflux Pump Promotes Intrinsic and Acquired Resistance to PROTACs in Cancer Cells

ABSTRACTPROTACs (Proteolysis-Targeting Chimeras) represent a promising new class of drugs that selectively degrade proteins of interest from cells. PROTACs targeting oncogenes are avidly being explored for cancer therapies, with several currently in clinical trials. Drug resistance represents a significant challenge in cancer therapies, and the mechanism by which cancer cells acquire resistance to PROTACs remains poorly understood. Using proteomics, we discovered acquired and intrinsic resistance to PROTACs in cancer cells can be mediated by upregulation of the drug efflux pump MDR1. PROTAC-resistant cells could be re-sensitized to PROTACs through co-administering MDR1 inhibitors. Notably, co-treatment of MDR1-overexpressing colorectal cancer cells with MEK1/2 or KRASG12C degraders and the dual ErbB receptor/MDR1 inhibitor lapatinib exhibited potent drug synergy due to simultaneous blockade of MDR1 and ErbB receptor activity. Together, our findings suggest that concurrent blockade of MDR1 will likely be required in combination with PROTACs to achieve durable protein degradation and therapeutic response in cancer.

ErbB Receptor Stimulation Is Required for Mouse Colon Adenoma Organoids to Form Crypts

The majority of colon adenomas harbor genetic mutations in the APC gene. APC mutation leads to changes in Wnt signalling and cell-cell adhesion: as a consequence, intestinal crypt budding increases and the excess crypts accumulate to form adenomas that progress to colon cancer. When cultured with Wnt, R-spondin, EGF, Noggin, myofibroblast conditioned medium and Matrigel, crypts from normal mouse colon mucosa form crypt-producing organoids and can be passaged continuously. Under the same culture and passage conditions, crypts isolated from colon adenomas derived from Apcmin/+ mice typically grow as spheroidal cysts and do not produce crypts. The adenoma organoid growth requires EGF, but not Wnt, R-spondin or Noggin. However, when mouse colon adenoma spheroids are grown for more than 10 days in the presence of EGF, crypt formation occurs. EGF, EREG, β-cellulin, Neuregulin-1 or AREG are sufficient for initiating crypt formation, however, neuregulin-1 is more potent than the other EGF-family members. EGFR and ErbB2 inhibitors both prevent crypt formation in adenoma cultures. Either EGFR:ErbB2 or ErbB3:ErbB2 signalling is sufficient to initiate adenoma crypt budding and elongation. ErbB2 inhibitors may provide a therapeutic avenue for controlling and ablating colon adenomas.

A Computational Framework for Prediction and Analysis of Cancer Signaling Dynamics from RNA Sequencing Data—Application to the ErbB Receptor Signaling Pathway

A current challenge in systems biology is to predict dynamic properties of cell behaviors from public information such as gene expression data. The temporal dynamics of signaling molecules is critical for mammalian cell commitment. We hypothesized that gene expression levels are tightly linked with and quantitatively control the dynamics of signaling networks regardless of the cell type. Based on this idea, we developed a computational method to predict the signaling dynamics from RNA sequencing (RNA-seq) gene expression data. We first constructed an ordinary differential equation model of ErbB receptor → c-Fos induction using a newly developed modeling platform BioMASS. The model was trained with kinetic parameters against multiple breast cancer cell lines using autologous RNA-seq data obtained from the Cancer Cell Line Encyclopedia (CCLE) as the initial values of the model components. After parameter optimization, the model proceeded to prediction in another untrained breast cancer cell line. As a result, the model learned the parameters from other cells and was able to accurately predict the dynamics of the untrained cells using only the gene expression data. Our study suggests that gene expression levels of components within the ErbB network, rather than rate constants, can explain the cell-specific signaling dynamics, therefore playing an important role in regulating cell fate.

ErbB dysregulation impairs cognition via myelination-dependent and - independent oligodendropathy

White matter abnormalities are an emerging pathological feature of schizophrenia. However, their attributions to the disease remain largely elusive. ErbB receptors and their ligands, some of which are essential for peripheral myelination, confer susceptibility to schizophrenia. By synergistically manipulating ErbB receptor activities in a oligodendrocyte-stage-specific manner in mice after early development, we demonstrate the distinct effects of ErbB signaling on oligodendrocytes at various differentiation states. ErbB overactivation, in mature oligodendrocytes, induces necroptosis causing demyelination, whereas in oligodendrocyte precursor cells, induces apoptosis causing hypomyelination. In contrast, ErbB inhibition increases oligodendrocyte precursor cell proliferation but induces hypomyelination by suppressing the myelinating capabilities of newly-formed oligodendrocytes. Remarkably, ErbB inhibition in mature oligodendrocytes diminishes axonal conduction under energy stress and impairs working memory capacity independently of myelin pathology. This study reveals the etiological implications of oligodendrocyte vulnerability induced by ErbB dysregulation, and elucidates the pathogenetic mechanisms for variable structural and functional white matter abnormalities.

Enrichment of alterations in targetable molecular pathways in KRAS wild-type (WT) pancreatic cancer (PC).

4629 Background: Genomic profiling has identified KRAS mutations in 88-90% of PC. KRAS WT tumors represent a molecularly heterogeneous group that may harbor targetable alterations (TA). We studied KRAS WT PC using NextGen sequencing (NGS) and whole transcriptome sequencing (WTS) to characterize the molecular landscape of this unique group and to assess the prevalence of TA. Methods: A total of 1164 PC tumors were tested at Caris Life Sciences by NGS (592 genes), WTS (NovaSeq), IHC and fragment analysis. Comparison of KRAS WT vs. mutant (MT) was done by Fisher-Exact or Chi2 and was corrected for multiple tests. Results: The KRAS WT cohort included 144 tumors (12.4%). No differences were seen in gender (female: 46% in both WT & MT) and age (median: 66 & 67) compared to KRAS MT. In KRAS WT tumors, targetable fusions tested by WTS and pathogenic mutations by NGS were seen in 22% (32 of 144) and 52% (75 of 144) respectively; potentially targetable amplifications (amp) were seen in 5 additional tumors. No TA were seen in 22% of WT tumors. Key alterations are in Table. Alterations inducing MAPK activation, including BRAF, RAF1, NF1 and GNAS changes were seen in 38 (26%) tumors. Frequent alterations were seen in FGFR genes (11 tumors), MET (4, including 1 exon 14 skip), and ERBB receptor and ligands (10). Fusions in ALK, ROS1, RET and NOTCH1 were seen (8), largely exclusive of other drivers. Wnt, PI3K, chromatin remodeling (CR) and DDR changes were common and sometimes seen concurrent with other alterations. Compared to KRAS MT, no difference of PD-L1 or TMB-H was seen. BRAF, APC, PBRM1, CTNNB1 mutations, MDM2 amp, gene fusions and MSI-H/dMMR were all more frequent in KRAS WT tumors (corrected p < 0.05). Conclusions: KRAS WT PC is enriched with TA (e.g., BRAF, ALK, ROS1, NRG1, MSI-H). The use of WTS in combination with NGS identifies activated molecular pathways in the majority of KRAS WT tumors. Based on our findings, comprehensive profiling of PC at the DNA and RNA level is recommended to provide patients with therapeutic opportunities beyond standard treatments. [Table: see text]