IntroductionOsimertinib is an irreversible EGFR tyrosine kinase inhibitor approved for the first-line treatment of patients with metastatic NSCLC harboring EGFR exon 19 deletions or L858R mutations. Patients treated with osimertinib invariably develop acquired resistance by mechanisms involving additional EGFR mutations, MET amplification, and other pathways. There is no known involvement of the oncogenic MUC1-C protein in acquired osimertinib resistance.MethodsH1975/EGFR (L858R/T790M) and patient-derived NSCLC cells with acquired osimertinib resistance were investigated for MUC1-C dependence in studies of EGFR pathway activation, clonogenicity, and self-renewal capacity.ResultsWe reveal that MUC1-C is up-regulated in H1975 osimertinib drug-tolerant persister cells and is necessary for activation of the EGFR pathway. H1975 cells selected for stable osimertinib resistance (H1975-OR) and MGH700-2D cells isolated from a patient with acquired osimertinib resistance are found to be dependent on MUC1-C for induction of (1) phospho (p)-EGFR, p-ERK, and p-AKT, (2) EMT, and (3) the resistant phenotype. We report that MUC1-C is also required for p-EGFR, p-ERK, and p-AKT activation and self-renewal capacity in acquired osimertinib-resistant (1) MET-amplified MGH170-1D #2 cells and (2) MGH121 Res#2/EGFR (T790M/C797S) cells. Importantly, targeting MUC1-C in these diverse models reverses osimertinib resistance. In support of these results, high MUC1 mRNA and MUC1-C protein expression is associated with a poor prognosis for patients with EGFR-mutant NSCLCs.ConclusionsOur findings reveal that MUC1-C is a common effector of osimertinib resistance and is a potential target for the treatment of osimertinib-resistant NSCLCs.
Background/Aim: Women with breast cancer are at increased risk of subsequent primary malignancies, specifically lung cancer. The aim of this study was to report the frequency of lung cancer in patients with breast cancer, and patients9 characteristics and surgical outcomes. Patients and Methods: We investigated 1,066 consecutive female patients undergoing surgical resection for breast cancer and 666 undergoing surgical resection for lung cancer. Results: Lung cancer with breast cancer was observed in 14 patients (1.3% of breast cancer and 2.1% of lung cancer cases; mean age=65 years), and 3/14 (21.4%) patients were smokers. Sixteen lung cancer lesions in 14 patients were adenocarcinomas and one was squamous cell carcinoma. All 14 patients were alive at the time of this report; 4/14 (28.6%) patients had recurrent breast cancer and 1/14 (7.1%) had recurrent lung cancer. In synchronous cases, 5/6 (83.3%) patients received concomitant surgery for both breast cancer and lung cancer. Patients9 postoperative courses were uneventful. In metachronous cases, eight patients had lung cancer a mean of 33 months after breast cancer surgery. All eight patients received adjuvant therapies and 4/8 (50%) patients received adjuvant therapies for recurrent breast cancer, including chemotherapy, radiotherapy, hormonal therapy, and anti-HER2 therapy. All patients had early-stage lung adenocarcinoma and underwent surgical resection. Conclusion: Concomitant surgery for synchronous lung and breast cancer was feasible and safe. In metachronous cases, lung cancers tended to be detected within 3 years after surgery for breast cancer. Careful follow-up for postoperative breast cancer may contribute to the detection of early-stage lung cancer.
<p>MUC1-C regulates a global transcriptional program enriched for STAT/IRF signaling in HNSCC cells. <b>A,</b> RNA-seq was performed on CAL27/tet-MUC1shRNA and HSC3/tet-MUC1shRNA cells treated with vehicle or DOX for 7 days. Volcano plots depicting downregulated (blue) and upregulated (red) DEGs (FDR<0.05; FC>2). Highlighted are the top DEGs by significance and magnitude. <b>B,</b> Common downregulated and upregulated genes in CAL27 and HSC3 cells with MUC1-C silencing. <b>C,</b> Purified chromatin from CAL27/tet-MUC1shRNA cells treated with vehicle or DOX for 7 days was immunoblotted with antibodies against the indicated proteins. <b>D</b> and <b>E,</b> Candidate enrichment plots for the HALLMARK INTERFERON ALPHA RESPONSE (<b>D</b>), HALLMARK INTERFERON GAMMA RESPONSE (<b>E</b>) gene signatures. <b>F</b> and <b>G,</b> Box plots of selected ISG expression in CAL27 (<b>F</b>) and HSC3 (<b>G</b>) cells with MUC1-C silencing.</p>
<p>MUC1-C regulates NOTCH3 expression and HNSCC cell self-renewal capacity. <b>A,</b> CAL27/tet-MUC1shRNA cells treated with vehicle or DOX for 7 days were analyzed for NOTCH3 mRNA levels. The results (mean ± SD of four determinations) are expressed as relative levels compared with that obtained for vehicle-treated cells (assigned a value of 1; left). Lysates were immunoblotted with antibodies against the indicated proteins (right). <b>B,</b> GSEA of RNA-seq data from CAL27/tet-MUC1shRNA cells treated with vehicle or DOX for 7 days using the REACTOME SIGNALING BY NOTCH3 gene signature. <b>C,</b> The indicated CAL27 cells treated with vehicle or DOX for 7 days were analyzed for tumorsphere formation. Shown are representative images of the tumorspheres (bar represents 100 µm). SFE is expressed as the mean ± SD of three independent replicates relative to that obtained for vehicle-treated cells (assigned a value of 1). <b>D,</b> Lysates from CAL27 cells treated with vehicle or 5 µmol/L GO-203 for 3 days were immunoblotted with antibodies against the indicated proteins. <b>E,</b> CAL27 cells were treated with vehicle or 5 µmol/L GO-203 while they were analyzed for colony formation. Shown are representative photomicrographs of stained colonies (left). The results (mean ± SD of three determinations) are expressed as relative colony formation compared with that for vehicle-treated cells (assigned a value of 1; right). <b>F,</b> CAL27 cells were treated with vehicle or 5 µmol/L GO-203 while they were analyzed for tumorsphere formation. Shown are representative images of the tumorspheres (bar represents 100 µm). SFE is expressed as the mean ± SD of three independent replicates relative to that obtained for untreated cells (assigned a value of 1). <b>G</b> and <b>H,</b> Six-week-old nude mice were injected subcutaneously in the flank with 1 × 10<sup>7</sup> CAL27 cells. Mice pair-matched into groups of 5 mice each when tumors reached 100–150 mm<sup>3</sup> were treated with vehicle control or GO-203 for the indicated days. Tumor volumes are expressed as the mean ± SEM for 5 mice (<b>G</b>). Lysates from control and GO-203–treated tumors were immunoblotted with antibodies against the indicated proteins (<b>H</b>).</p>
<p>Expression of MUC1 in HNSCC tumors and effects of silencing MUC1-C on HNSCC cell clonogenicity. <b>A,</b> Analysis of primary (P) and metastatic (M) HNSCC tissues for MUC1 expression using the GSE136037 dataset. <b>B,</b> CAL27/CshRNA and MUC1shRNA cells were analyzed for MUC1-C mRNA levels (left). The results (mean ± SD of four determinations) are expressed as relative levels compared with that obtained for CshRNA cells (assigned a value of 1; left). Lysates were immunoblotted with antibodies against the indicated proteins (right). <b>C,</b> The indicated CAL27 cells were analyzed for colony formation. Shown are representative photomicrographs of stained colonies (left). The results (mean ± SD of three determinations) are expressed as relative colony formation compared with that for CshRNA cells (assigned a value of 1; right). <b>D,</b> Amino acid sequence of the MUC1-C cytoplasmic domain (CD) highlighting direct interactions with JAK1 and STAT1. <b>E</b> and <b>F,</b> CAL27 cells expressing the indicated vectors were treated with vehicle or DOX for 7 days. Lysates were immunoblotted with antibodies against the indicated proteins (<b>E</b>). Cells were analyzed for colony formation (<b>F</b>). Shown are representative photomicrographs of stained colonies (top). The results (mean ± SD of three determinations) are expressed as relative colony formation compared with that for control cells (assigned a value of 1; bottom).</p>
Synthetic peptides from the cross-region of the laminin A chain were prepared and tested for their biological activities, especially for neurite outgrowth. A synthetic 8-mer peptide (designated LMA-5) in the cross-region of the laminin A chain was found to promote neurite outgrowth in PC12 cells and cerebellar microexplant cultures. Furthermore, this peptide mediated cell attachment and heparin binding. In addition, an antibody against peptide LMA-5 inhibited laminin- and LMA-5-mediated cell attachment. This antibody also inhibited more than half of the LMA-5-promoted neurite outgrowth in cerebellar microexplant cultures. These data suggest that peptide LMA-5 is one of the active sites in laminin that regulate cell behaviour including neurite outgrowth, cell attachment and heparin binding.
Background Immune checkpoint inhibitors (ICIs) have had a profound impact on the treatment of many tumors; however, their effectiveness against triple-negative breast cancers (TNBCs) has been limited. One factor limiting responsiveness of TNBCs to ICIs is a lack of functional tumor-infiltrating lymphocytes (TILs) in ‘non-inflamed’ or ‘cold’ tumor immune microenvironments (TIMEs), although by unknown mechanisms. Targeting MUC1-C in a mouse transgenic TNBC tumor model increases cytotoxic tumor-infiltrating CD8+ T cells (CTLs), supporting a role for MUC1-C in immune evasion. The basis for these findings and whether they extend to human TNBCs are not known. Methods Human TNBC cells silenced for MUC1-C using short hairpin RNAs (shRNAs) were analyzed for the effects of MUC1-C on global transcriptional profiles. Differential expression and rank order analysis was used for gene set enrichment analysis (GSEA). Gene expression was confirmed by quantitative reverse-transcription PCR and immunoblotting. The The Cancer Genome Atlas Breast Invasive Carcinoma (TCGA-BRCA) and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) datasets were analyzed for effects of MUC1 on GSEA, cell-type enrichment, and tumor immune dysfunction and exclusion. Single-cell scRNA-seq datasets of TNBC samples were analyzed for normalized expression associations between MUC1 and selected genes within tumor cells. Results Our results demonstrate that MUC1-C is a master regulator of the TNBC transcriptome and that MUC1-C-induced gene expression is driven by STAT1 and IRF1. We found that MUC1-C activates the inflammatory interferon (IFN)-γ-driven JAK1→STAT1→IRF1 pathway and induces the IDO1 and COX2/PTGS2 effectors, which play key roles in immunosuppression. Involvement of MUC1-C in activating the immunosuppressive IFN-γ pathway was extended by analysis of human bulk and scRNA-seq datasets. We further demonstrate that MUC1 associates with the depletion and dysfunction of CD8+ T cells in the TNBC TIME. Conclusions These findings demonstrate that MUC1-C integrates activation of the immunosuppressive IFN-γ pathway with depletion of TILs in the TNBC TIME and provide support for MUC1-C as a potential target for improving TNBC treatment alone and in combination with ICIs. Of translational significance, MUC1-C is a druggable target with chimeric antigen receptor (CAR) T cells, antibody-drug conjugates (ADCs) and a functional inhibitor that are under clinical development.
