Targeting SHP2 phosphatase in breast cancer overcomes RTK-mediated resistance to PI3K inhibitors.

BACKGROUND: PI3K signaling is frequently activated in breast cancer and is targeted by PI3K inhibitors. However, resistance of tumor cells to PI3K inhibition, often mediated by activated receptor tyrosine kinases, is commonly observed and reduces the potency of PI3K inhibitors. Therefore, new treatment strategies to overcome resistance to PI3K inhibitors are urgently needed to boost their efficacy. The phosphatase SHP2, which plays a crucial role in mediating signal transduction between receptor tyrosine kinases and both the PI3K and MAPK pathways, is a potential target for combination treatment. METHODS: We tested combinations of PI3K and SHP2 inhibitors in several experimental breast cancer models that are resistant to PI3K inhibition. Using cell culturing, biochemical and genetic approaches, we evaluated tumor cell proliferation and signaling output in cells treated with PI3K and SHP2 inhibitors. RESULTS: Combination treatment with PI3K and SHP2 inhibitors counteracted both acquired and intrinsic breast cancer cell resistance to PI3K inhibition that is mediated by activated receptor tyrosine kinases. Dual PI3K and SHP2 inhibition blocked proliferation and led to sustained inactivation of PI3K and MAPK signaling, where resistant cells rapidly re-activated these pathways upon PI3K inhibitor monotreatment. In addition, we demonstrate that overexpression of SHP2 induced resistance to PI3K inhibition, and that SHP2 was frequently activated during the development of PI3K inhibitor resistance after prolonged treatment of sensitive cells. CONCLUSIONS: Our results highlight the importance of SHP2 as a player in resistance to PI3K inhibitors. Combination treatment with PI3K and SHP2 inhibitors could pave the way for significant improvements in therapies for breast cancer.

Patient-Derived Xenografts from Solid Tumors (PDX) for Models of Metastasis.

In cancer research, availability of clinically relevant tumor models is still essential for drug testing, proof of concept studies, and also molecular analyses. To achieve this, models are of advantage, which more closely reflect heterogeneity of tumors. In this regard, patient-derived xenograft (PDX) models more closely recapitulate the native tumor biology, tissue composition, and molecular characteristics. Since metastasis is still the major challenge of tumor therapy, models are pivotal, which resemble this particular property. In this context, PDX model-derived metastasis is of particular interest for testing antimetastatic therapies for their efficacy to better target this systemic disease. This protocol describes the establishment of PDX models from tumor specimen and their applicability for PDX-derived metastasis at metastatic sites such as liver and lung, which are also clinically relevant for the systemic spread of cancer. Analysis of metastasis and methods for quantification of metastatic spread are provided.

Patient-derived xenograft (PDX) models of colorectal carcinoma (CRC) as a platform for chemosensitivity and biomarker analysis in personalized medicine.

Patient-derived xenograft (PDX) tumor models represent a valuable platform for identifying new biomarkers and novel targets, to evaluate therapy response and resistance mechanisms. This study aimed at establishment, characterization and therapy testing of colorectal carcinoma-derived PDX. We generated 49 PDX and validated identity between patient tumor and corresponding PDX. Sensitivity of PDX toward conventional and targeted drugs revealed that 92% of PDX responded toward irinotecan, 45% toward 5-FU, 65% toward bevacizumab, and 61% toward cetuximab. Expression of epidermal growth factor receptor (EGFR) ligands correlated to the sensitivity toward cetuximab. Proto-oncogene B-RAF, EGFR, Kirsten rat sarcoma virus oncogene homolog gene copy number correlated positively with cetuximab and erlotinib sensitivity. The mutational analyses revealed an individual mutational profile of PDX and mainly identical profiles of PDX from primary tumor vs corresponding metastasis. Mutation in PIK3CA was a determinant of accelerated tumor doubling time. PDX with wildtype Kirsten rat sarcoma virus oncogene homolog, proto-oncogene B-RAF, and phosphatidylinositol-4,5-bisphosphate 3-kinaseM catalytic subunit alfa showed higher sensitivity toward cetuximab and erlotinib. To study the molecular mechanism of cetuximab resistance, cetuximab resistant PDX models were generated, and changes in HER2, HER3, betacellulin, transforming growth factor alfa were observed. Global proteome and phosphoproteome profiling showed a reduction in canonical EGFR-mediated signaling via PTPN11 (SHP2) and AKT1S1 (PRAS40) and an increase in anti-apoptotic signaling as a consequence of acquired cetuximab resistance. This demonstrates that PDX models provide a multitude of possibilities to identify and validate biomarkers, signaling pathways and resistance mechanisms for clinically relevant improvement in cancer therapy.

The epigenetic regulator Mll1 is required for Wnt-driven intestinal tumorigenesis and cancer stemness.

Wnt/ß-catenin signaling is crucial for intestinal carcinogenesis and the maintenance of intestinal cancer stem cells. Here we identify the histone methyltransferase Mll1 as a regulator of Wnt-driven intestinal cancer. Mll1 is highly expressed in Lgr5+ stem cells and human colon carcinomas with increased nuclear ß-catenin. High levels of MLL1 are associated with poor survival of colon cancer patients. The genetic ablation of Mll1 in mice prevents Wnt/ß-catenin-driven adenoma formation from Lgr5+ intestinal stem cells. Ablation of Mll1 decreases the self-renewal of human colon cancer spheres and halts tumor growth of xenografts. Mll1 controls the expression of stem cell genes including the Wnt/ß-catenin target gene Lgr5. Upon the loss of Mll1, histone methylation at the stem cell promoters switches from activating H3K4 tri-methylation to repressive H3K27 tri-methylation, indicating that Mll1 sustains stem cell gene expression by antagonizing gene silencing through polycomb repressive complex 2 (PRC2)-mediated H3K27 tri-methylation. Transcriptome profiling of Wnt-mutated intestinal tumor-initiating cells reveals that Mll1 regulates Gata4/6 transcription factors, known to sustain cancer stemness and to control goblet cell differentiation. Our results demonstrate that Mll1 is an essential epigenetic regulator of Wnt/ß-catenin-induced intestinal tumorigenesis and cancer stemness.

CRISPR-Cas9-Mediated ELANE Mutation Correction in Hematopoietic Stem and Progenitor Cells to Treat Severe Congenital Neutropenia.

Severe congenital neutropenia (SCN) is a monogenic disorder. SCN patients are prone to recurrent life-threatening infections. The main causes of SCN are autosomal dominant mutations in the ELANE gene that lead to a block in neutrophil differentiation. In this study, we use CRISPR-Cas9 ribonucleoproteins and adeno-associated virus (AAV)6 as a donor template delivery system to repair the ELANEL172P mutation in SCN patient-derived hematopoietic stem and progenitor cells (HSPCs). We used a single guide RNA (sgRNA) specifically targeting the mutant allele, and an sgRNA targeting exon 4 of ELANE. Using the latter sgRNA, ∼34% of the known ELANE mutations can in principle be repaired. We achieved gene correction efficiencies of up to 40% (with sgELANE-ex4) and 56% (with sgELANE-L172P) in the SCN patient-derived HSPCs. Gene repair restored neutrophil differentiation in vitro and in vivo upon HSPC transplantation into humanized mice. Mature edited neutrophils expressed normal elastase levels and behaved normally in functional assays. Thus, we provide a proof of principle for using CRISPR-Cas9 to correct ELANE mutations in patient-derived HSPCs, which may translate into gene therapy for SCN.

Inhibiting WNT and NOTCH in renal cancer stem cells and the implications for human patients.

Current treatments for clear cell renal cell cancer (ccRCC) are insufficient because two-thirds of patients with metastases progress within two years. Here we report the identification and characterization of a cancer stem cell (CSC) population in ccRCC. CSCs are quantitatively correlated with tumor aggressiveness and metastasis. Transcriptional profiling and single cell sequencing reveal that these CSCs exhibit an activation of WNT and NOTCH signaling. A significant obstacle to the development of rational treatments has been the discrepancy between model systems and the in vivo situation of patients. To address this, we use CSCs to establish non-adherent sphere cultures, 3D tumor organoids, and xenografts. Treatment with WNT and NOTCH inhibitors blocks the proliferation and self-renewal of CSCs in sphere cultures and organoids, and impairs tumor growth in patient-derived xenografts in mice. These findings suggest that our approach is a promising route towards the development of personalized treatments for individual patients.

MiR-1287-5p inhibits triple negative breast cancer growth by interaction with phosphoinositide 3-kinase CB, thereby sensitizing cells for PI3Kinase inhibitors.

BACKGROUND: Non-coding RNAs and especially microRNAs have been discovered to act as master regulators of cancer initiation and progression. The aim of our study was to discover and characterize the function of yet functionally uncharacterized microRNAs in human breast carcinogenesis. METHODS: In an unbiased approach, we utilized an established model system for breast cancer (BC) stem cell formation ("mammosphere assay") to identify whole miRNome alterations in breast carcinogenesis. Clinical samples of BC patients were used to evaluate the human relevance of the newly identified miRNA candidates. One promising candidate, miR-1287-5p, was further explored on its impact on several hallmarks of cancer. The molecular mode of action was characterized by whole transcriptome analysis, in silico prediction tools, miRNA-interaction assays, pheno-copy assays, and drug sensitivity assays. RESULTS: Among several other microRNAs, miR-1287-5p was significantly downregulated in mammospheres and human BC tissue compared to normal breast tissue (p < 0.0001). Low expression levels were significantly associated with poor prognosis in BC patients. MiR-1287-5p significantly decreased cellular growth, cells in S phase of cell cycle, anchorage-independent growth, and tumor formation in vivo. In addition, we identified PIK3CB as a direct molecular interactor of miR-1287-5p and a novel prognostic factor in BC. Finally, PI3Kinase pathway chemical inhibitors combined with miR-1287-5p mimic increased the pharmacological growth inhibitory potential in triple negative BC cells. CONCLUSION: Our data identified for the first time the involvement of miR-1287-5p in human BC and suggest a potential for therapeutic interventions in difficult to treat triple negative BC.

The Wnt-Driven Mll1 Epigenome Regulates Salivary Gland and Head and Neck Cancer.

We identified a regulatory system that acts downstream of Wnt/ß-catenin signaling in salivary gland and head and neck carcinomas. We show in a mouse tumor model of K14-Cre-induced Wnt/ß-catenin gain-of-function and Bmpr1a loss-of-function mutations that tumor-propagating cells exhibit increased Mll1 activity and genome-wide increased H3K4 tri-methylation at promoters. Null mutations of Mll1 in tumor mice and in xenotransplanted human head and neck tumors resulted in loss of self-renewal of tumor-propagating cells and in block of tumor formation but did not alter normal tissue homeostasis. CRISPR/Cas9 mutagenesis and pharmacological interference of Mll1 at sequences that inhibit essential protein-protein interactions or the SET enzyme active site also blocked the self-renewal of mouse and human tumor-propagating cells. Our work provides strong genetic evidence for a crucial role of Mll1 in solid tumors. Moreover, inhibitors targeting specific Mll1 interactions might offer additional directions for therapies to treat these aggressive tumors.

miR-196b-5p Regulates Colorectal Cancer Cell Migration and Metastases through Interaction with HOXB7 and GALNT5.

Purpose: miR-196b-5p has been previously implicated in malignant transformation; however, its role in colorectal cancer has not been fully explored. In this study, we examine the clinical and biological relevance of miR-196b-5p, and the molecular pathways regulated by miR-196b-5p in colorectal cancer.Experimental Design: miR-196b-5p expression was quantitated by qRT-PCR in 2 independent cohorts composed of 292 patients with colorectal cancer in total, to explore its biomarker potential. Transient and stable gain- and loss-of-function experiments were conducted in a panel of colorectal cancer cell lines and mice, to evaluate the impact of miR-196b-5p on proliferation, chemosensitivity, migration/invasion, and metastases formation in vitro and in vivo The molecular pathways influenced by miR-196b-5p were characterized using whole transcriptome profiling, in silico target prediction tools, luciferase interaction assays, and phenocopy/rescue gene knockdown experiments.Results: Low miR-196b-5p expression was significantly associated with metastases and poor outcomes in 2 independent colorectal cancer patient cohorts (P < 0.05, log-rank test). miR-196b-5p inhibition led to significantly increased colorectal cancer cell migration/invasion and metastases formation in mice, whereas ectopic overexpression showed the opposite phenotype. Molecular profiling and target confirmation identified an interaction between miR-196b-5p and HOXB7 and GALNT5, which in turn regulated colorectal cancer cell migration.Conclusions: The association of low levels of miR-196b-5p and poor prognosis in patients with colorectal cancer can be explained by its influence on cancer cell migration and metastases formation. miR-196b-5p has an impact on colorectal cancer progression pathways through direct interaction with genes involved in cancer cell migration. Clin Cancer Res; 23(17); 5255-66. ©2017 AACR.

Cancer Stem Cells Regulate Cancer-Associated Fibroblasts via Activation of Hedgehog Signaling in Mammary Gland Tumors.

Many tumors display intracellular heterogeneity with subsets of cancer stem cells (CSC) that sustain tumor growth, recurrence, and therapy resistance. Cancer-associated fibroblasts (CAF) have been shown to support and regulate CSC function. Here, we investigate the interactions between CSCs and CAFs in mammary gland tumors driven by combined activation of Wnt/ß-catenin and Hgf/Met signaling in mouse mammary epithelial cells. In this setting, CSCs secrete the Hedgehog ligand SHH, which regulate CAFs via paracrine activation of Hedgehog signaling. CAFs subsequently secrete factors that promote expansion and self-renewal of CSCs. In vivo treatment of tumors with the Hedgehog inhibitor vismodegib reduce CAF and CSC expansion, resulting in an overall delay of tumor formation. Our results identify a novel intracellular signaling module that synergistically regulates CAFs and CSCs. Targeting CAFs with Hedgehog inhibitors may offer a novel therapeutic strategy against breast cancer. Cancer Res; 77(8); 2134-47. ©2017 AACR.

Genome-Wide miRNA Analysis Identifies miR-188-3p as a Novel Prognostic Marker and Molecular Factor Involved in Colorectal Carcinogenesis.

Purpose: Characterization of colorectal cancer transcriptome by high-throughput techniques has enabled the discovery of several differentially expressed genes involving previously unreported miRNA abnormalities. Here, we followed a systematic approach on a global scale to identify miRNAs as clinical outcome predictors and further validated them in the clinical and experimental setting.Experimental Design: Genome-wide miRNA sequencing data of 228 colorectal cancer patients from The Cancer Genome Atlas dataset were analyzed as a screening cohort to identify miRNAs significantly associated with survival according to stringent prespecified criteria. A panel of six miRNAs was further validated for their prognostic utility in a large independent validation cohort (n = 332). In situ hybridization and functional experiments in a panel of colorectal cancer cell lines and xenografts further clarified the role of clinical relevant miRNAs.Results: Six miRNAs (miR-92b-3p, miR-188-3p, miR-221-5p, miR-331-3p, miR-425-3p, and miR-497-5p) were identified as strong predictors of survival in the screening cohort. High miR-188-3p expression proves to be an independent prognostic factor [screening cohort: HR = 4.137; 95% confidence interval (CI), 1.568-10.917; P = 0.004; validation cohort: HR = 1.538; 95% CI, 1.107-2.137; P = 0.010, respectively]. Forced miR-188-3p expression increased migratory behavior of colorectal cancer cells in vitro and metastases formation in vivo (P < 0.05). The promigratory role of miR-188-3p is mediated by direct interaction with MLLT4, a novel identified player involved in colorectal cancer cell migration.Conclusions: miR-188-3p is a novel independent prognostic factor in colorectal cancer patients, which can be partly explained by its effect on MLLT4 expression and migration of cancer cells. Clin Cancer Res; 23(5); 1323-33. ©2016 AACR.

New Wnt/ß-catenin target genes promote experimental metastasis and migration of colorectal cancer cells through different signals.

OBJECTIVES: We have previously identified a 115-gene signature that characterises the metastatic potential of human primary colon cancers. The signature included the canonical Wnt target gene BAMBI, which promoted experimental metastasis in mice. Here, we identified three new direct Wnt target genes from the signature, and studied their functions in epithelial-mesenchymal transition (EMT), cell migration and experimental metastasis. DESIGN: We examined experimental liver metastases following injection of selected tumour cells into spleens of NOD/SCID mice. Molecular and cellular techniques were used to identify direct transcription target genes of Wnt/ß-catenin signals. Microarray analyses and experiments that interfered with cell migration through inhibitors were performed to characterise downstream signalling systems. RESULTS: Three new genes from the colorectal cancer (CRC) metastasis signature, BOP1, CKS2 and NFIL3, were identified as direct transcription targets of ß-catenin/TCF4. Overexpression and knocking down of these genes in CRC cells promoted and inhibited, respectively, experimental metastasis in mice, EMT and cell motility in culture. Cell migration was repressed by interfering with distinct signalling systems through inhibitors of PI3K, JNK, p38 mitogen-activated protein kinase and/or mTOR. Gene expression profiling identified a series of migration-promoting genes, which were induced by BOP1, CKS2 and NFIL3, and could be repressed by inhibitors that are specific to these pathways. CONCLUSIONS: We identified new direct Wnt/ß-catenin target genes, BOP1, CKS2 and NFIL3, which induced EMT, cell migration and experimental metastasis of CRC cells. These genes crosstalk with different downstream signalling systems, and activate migration-promoting genes. These pathways and downstream genes may serve as therapeutic targets in the treatment of CRC metastasis.

A Small-Molecule Antagonist of the ß-Catenin/TCF4 Interaction Blocks the Self-Renewal of Cancer Stem Cells and Suppresses Tumorigenesis.

Wnt/ß-catenin signaling is a highly conserved pathway essential for embryogenesis and tissue homeostasis. However, deregulation of this pathway can initiate and promote human malignancies, especially of the colon and head and neck. Therefore, Wnt/ß-catenin signaling represents an attractive target for cancer therapy. We performed high-throughput screening using AlphaScreen and ELISA techniques to identify small molecules that disrupt the critical interaction between ß-catenin and the transcription factor TCF4 required for signal transduction. We found that compound LF3, a 4-thioureido-benzenesulfonamide derivative, robustly inhibited this interaction. Biochemical assays revealed clues that the core structure of LF3 was essential for inhibition. LF3 inhibited Wnt/ß-catenin signals in cells with exogenous reporters and in colon cancer cells with endogenously high Wnt activity. LF3 also suppressed features of cancer cells related to Wnt signaling, including high cell motility, cell-cycle progression, and the overexpression of Wnt target genes. However, LF3 did not cause cell death or interfere with cadherin-mediated cell-cell adhesion. Remarkably, the self-renewal capacity of cancer stem cells was blocked by LF3 in concentration-dependent manners, as examined by sphere formation of colon and head and neck cancer stem cells under nonadherent conditions. Finally, LF3 reduced tumor growth and induced differentiation in a mouse xenograft model of colon cancer. Collectively, our results strongly suggest that LF3 is a specific inhibitor of canonical Wnt signaling with anticancer activity that warrants further development for preclinical and clinical studies as a novel cancer therapy.

Shp2 signaling suppresses senescence in PyMT-induced mammary gland cancer in mice.

In this study, we have used techniques from cell biology, biochemistry, and genetics to investigate the role of the tyrosine phosphatase Shp2 in tumor cells of MMTV-PyMT mouse mammary glands. Genetic ablation or pharmacological inhibition of Shp2 induces senescence, as determined by the activation of senescence-associated ß-gal (SA-ß-gal), cyclin-dependent kinase inhibitor 1B (p27), p53, and histone 3 trimethylated lysine 9 (H3K9me3). Senescence induction leads to the inhibition of self-renewal of tumor cells and blockage of tumor formation and growth. A signaling cascade was identified that acts downstream of Shp2 to counter senescence: Src, focal adhesion kinase, and Map kinase inhibit senescence by activating the expression of S-phase kinase-associated protein 2 (Skp2), Aurora kinase A (Aurka), and the Notch ligand Delta-like 1 (Dll1), which block p27 and p53. Remarkably, the expression of Shp2 and of selected target genes predicts human breast cancer outcome. We conclude that therapies, which rely on senescence induction by inhibiting Shp2 or controlling its target gene products, may be useful in blocking breast cancer.

Cell differentiation mediated by co-culture of human umbilical cord blood stem cells with murine hepatic cells.

In the present study, purified human cord blood stem cells were co-cultivated with murine hepatic alpha mouse liver 12 (AML12) cells to compare the effect on endodermal stem cell differentiation by either direct cell-cell interaction or by soluble factors in conditioned hepatic cell medium. With that approach, we want to mimic in vitro the situation of preclinical transplantation experiments using human cells in mice. Cord blood stem cells, cultivated with hepatic conditioned medium, showed a low endodermal differentiation but an increased connexin 32 (Cx32) and Cx43, and cytokeratin 8 (CK8) and CK19 expression was monitored by reverse transcription polymerase chain reaction (RT-PCR). Microarray profiling indicated that in cultivated cord blood cells, 604 genes were upregulated 2-fold, with the highest expression for epithelial CK19 and epithelial cadherin (E-cadherin). On ultrastructural level, there were no major changes in the cellular morphology, except a higher presence of phago(ly)some-like structures observed. Direct co-culture of AML12 cells with cord blood cells led to less incisive differentiation with increased sex-determining region Y-box 17 (SOX17), Cx32 and Cx43, as well as epithelial CK8 and CK19 expressions. On ultrastructural level, tight cell contacts along the plasma membranes were revealed. FACS analysis in co-cultivated cells quantified dye exchange on low level, as also proved by time relapse video-imaging of labelled cells. Modulators of gap junction formation influenced dye transfer between the co-cultured cells, whereby retinoic acid increased and 3-heptanol reduced the dye transfer. The study indicated that the cell-co-cultured model of human umbilical cord blood cells and murine AML12 cells may be a suitable approach to study some aspects of endodermal/hepatic cell differentiation induction.

A comprehensively characterized large panel of head and neck cancer patient-derived xenografts identifies the mTOR inhibitor everolimus as potential new treatment option.

Patient-derived xenograft (PDX) models have shown to reflect original patient tumors better than any other preclinical model. We embarked in a study establishing a large panel of head and neck squamous cell carcinomas PDX for biomarker analysis and evaluation of established and novel compounds. Out of 115 transplanted specimens 52 models were established of which 29 were characterized for response to docetaxel, cetuximab, methotrexate, carboplatin, 5-fluorouracil and everolimus. Further, tumors were subjected to sequencing analysis and gene expression profiling of selected mTOR pathway members. Most frequent response was observed for docetaxel and cetuximab. Responses to carboplatin, 5-fluorouracil and methotrexate were moderate. Everolimus revealed activity in the majority of PDX. Mutational profiling and gene expression analysis did not reveal a predictive biomarker for everolimus even though by trend RPS6KB1 mRNA expression was associated with response. In conclusion we demonstrate a comprehensively characterized panel of head and neck cancer PDX models, which represent a valuable and renewable tissue resource for evaluation of novel compounds and associated biomarkers.

Activation of MAPK overrides the termination of myelin growth and replaces Nrg1/ErbB3 signals during Schwann cell development and myelination.

Myelination depends on the synthesis of large amounts of myelin transcripts and proteins and is controlled by Nrg1/ErbB/Shp2 signaling. We developed a novel pulse labeling strategy based on stable isotope labeling with amino acids in cell culture (SILAC) to measure the dynamics of myelin protein production in mice. We found that protein synthesis is dampened in the maturing postnatal peripheral nervous system, and myelination then slows down. Remarkably, sustained activation of MAPK signaling by expression of the Mek1DD allele in mice overcomes the signals that end myelination, resulting in continuous myelin growth. MAPK activation leads to minor changes in transcript levels but massively up-regulates protein production. Pharmacological interference in vivo demonstrates that the effects of activated MAPK signaling on translation are mediated by mTOR-independent mechanisms but in part also by mTOR-dependent mechanisms. Previous work demonstrated that loss of ErbB3/Shp2 signaling impairs Schwann cell development and disrupts the myelination program. We found that activated MAPK signaling strikingly compensates for the absence of ErbB3 or Shp2 during Schwann cell development and myelination.

Bace1 and Neuregulin-1 cooperate to control formation and maintenance of muscle spindles.

The protease ß-secretase 1 (Bace1) was identified through its critical role in production of amyloid-ß peptides (Aß), the major component of amyloid plaques in Alzheimer's disease. Bace1 is considered a promising target for the treatment of this pathology, but processes additional substrates, among them Neuregulin-1 (Nrg1). Our biochemical analysis indicates that Bace1 processes the Ig-containing ß1 Nrg1 (IgNrg1ß1) isoform. We find that a graded reduction in IgNrg1 signal strength in vivo results in increasingly severe deficits in formation and maturation of muscle spindles, a proprioceptive organ critical for muscle coordination. Further, we show that Bace1 is required for formation and maturation of the muscle spindle. Finally, pharmacological inhibition and conditional mutagenesis in adult animals demonstrate that Bace1 and Nrg1 are essential to sustain muscle spindles and to maintain motor coordination. Our results assign to Bace1 a role in the control of coordinated movement through its regulation of muscle spindle physiology, and implicate IgNrg1-dependent processing as a molecular mechanism.