The actin cytoskeleton generates and senses forces. Here we report that branched actin networks from the cell cortex depend on ARPC1B-containing Arp2/3 complexes and that they are specifically monitored by type I coronins to control cell cycle progression in mammary epithelial cells. Cortical ARPC1B-dependent branched actin networks are regulated by the RAC1/WAVE/ARPIN pathway and drive lamellipodial protrusions. Accordingly, we uncover that the duration of the G1 phase scales with migration persistence in single migrating cells. Moreover, cortical branched actin more generally determines S-phase entry by integrating soluble stimuli such as growth factors and mechanotransduction signals, ensuing from substratum rigidity or stretching of epithelial monolayers. Many tumour cells lose this dependence for cortical branched actin. But the RAC1-transformed tumour cells stop cycling upon Arp2/3 inhibition. Among all genes encoding Arp2/3 subunits, ARPC1B overexpression in tumours is associated with the poorest metastasis-free survival in breast cancer patients. Arp2/3 specificity may thus provide diagnostic and therapeutic opportunities in cancer.

Tumor cell invasion and metastasis formation are the major cause of death in cancer patients. These processes rely on extracellular matrix (ECM) degradation mediated by organelles termed invadopodia, to which the transmembrane matrix metalloproteinase MT1-MMP (also known as MMP14) is delivered from its reservoir, the RAB7-containing endolysosomes. How MT1-MMP is targeted to endolysosomes remains to be elucidated. Flotillin-1 and -2 are upregulated in many invasive cancers. Here, we show that flotillin upregulation triggers a general mechanism, common to carcinoma and sarcoma, which promotes RAB5-dependent MT1-MMP endocytosis and its delivery to RAB7-positive endolysosomal reservoirs. Conversely, flotillin knockdown in invasive cancer cells greatly reduces MT1-MMP accumulation in endolysosomes, its subsequent exocytosis at invadopodia, ECM degradation and cell invasion. Our results demonstrate that flotillin upregulation is necessary and sufficient to promote epithelial and mesenchymal cancer cell invasion and ECM degradation by controlling MT1-MMP endocytosis and delivery to the endolysosomal recycling compartment.

Radionuclide molecular imaging of human epidermal growth factor type 2 (HER2) expression may enable a non-invasive discrimination between HER2-positive and HER2-negative breast cancers for stratification of patients for HER2-targeted treatments. DARPin G3 is a small (molecular weigh 14 kDa) scaffold protein with picomolar affinity to HER2. The aim of this firstin-human study was to evaluate the safety, biodistribution and dosimetry of 99m Tc-(HE)3-G3.Methods. Three cohorts of patients with primary breast cancer (each including at least 4 patients with HER2-negative and 5 patients with HER2-positive tumors) were injected with either 1000, 2000 or 3000 µg of 99m Tc-(HE)3-G3 (287±170 MBq). Whole-body planar imaging followed by SPECT was performed at 2, 4, 6 and 24 h after injection. Vital signs and possible side effects were monitored during imaging and up to 7 days after injection.Results. All injections were well tolerated. No side effects were observed. The results of blood and urine analyses did not differ before and after studies. 99m Tc-(HE)3-G3 cleared rapidly from the blood. The highest uptake was detected in the kidneys and liver followed by the lungs, breasts and small intestinal content. The hepatic uptake after injecting with 2000 or 3000 µg was significantly (p<0.05) lower than the uptake after injecting with 1000 µg. Effective doses did not differ significantly between cohorts (average 0.011± 0.004 mSv/MBq). Tumor-to-contralateral site ratios for HER-positive tumors were significantly (p< 0.05) higher than for HER2-negative at 2 and 4 h after injection. Conclusions.Imaging of HER2 expression using 99m Tc-(HE)3-G3 is safe, well-tolerated and provides a low absorbed dose burden on patients. This imaging enables discerning HER2-positive and HER2-negative breast cancer. Phase I study data justifies further clinical development of 99m Tc-(HE)3-G3.

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