Simultaneously targeting extracellular vesicle trafficking and TGF-ß receptor kinase activity blocks signaling hyperactivation and metastasis.

Metastasis is the leading cause of cancer-related deaths. Transforming growth factor beta (TGF-ß) signaling drives metastasis and is strongly enhanced during cancer progression. Yet, the use of on-target TGF-ß signaling inhibitors in the treatment of cancer patients remains unsuccessful, highlighting a gap in the understanding of TGF-ß biology that limits the establishment of efficient anti-metastatic therapies. Here, we show that TGF-ß signaling hyperactivation in breast cancer cells is required for metastasis and relies on increased small extracellular vesicle (sEV) secretion. Demonstrating sEV's unique role, TGF-ß signaling levels induced by sEVs exceed the activity of matching concentrations of soluble ligand TGF-ß. Further, genetic disruption of sEV secretion in highly-metastatic breast cancer cells impairs cancer cell aggressiveness by reducing TGF-ß signaling to nearly-normal levels. Otherwise, TGF-ß signaling activity in non-invasive breast cancer cells is inherently low, but can be amplified by sEVs, enabling invasion and metastasis of poorly-metastatic breast cancer cells. Underscoring the translational potential of inhibiting sEV trafficking in advanced breast cancers, treatment with dimethyl amiloride (DMA) decreases sEV secretion, TGF-ß signaling activity, and breast cancer progression in vivo. Targeting both the sEV trafficking and TGF-ß signaling by combining DMA and SB431542 at suboptimal doses potentiated this effect, normalizing the TGF-ß signaling in primary tumors to potently reduce circulating tumor cells, metastasis, and tumor self-seeding. Collectively, this study establishes sEVs as critical elements in TGF-ß biology, demonstrating the feasibility of inhibiting sEV trafficking as a new therapeutic approach to impair metastasis by normalizing TGF-ß signaling levels in breast cancer cells.

Cancer associated-fibroblast-derived exosomes in cancer progression.

To identify novel cancer therapies, the tumor microenvironment (TME) has received a lot of attention in recent years in particular with the advent of clinical successes achieved by targeting immune checkpoint inhibitors (ICIs). The TME consists of multiple cell types that are embedded in the extracellular matrix (ECM), including immune cells, endothelial cells and cancer associated fibroblasts (CAFs), which communicate with cancer cells and each other during tumor progression. CAFs are a dominant and heterogeneous cell type within the TME with a pivotal role in controlling cancer cell invasion and metastasis, immune evasion, angiogenesis and chemotherapy resistance. CAFs mediate their effects in part by remodeling the ECM and by secreting soluble factors and extracellular vesicles. Exosomes are a subtype of extracellular vesicles (EVs), which contain various biomolecules such as nucleic acids, lipids, and proteins. The biomolecules in exosomes can be transmitted from one to another cell, and thereby affect the behavior of the receiving cell. As exosomes are also present in circulation, their contents can also be explored as biomarkers for the diagnosis and prognosis of cancer patients. In this review, we concentrate on the role of CAFs-derived exosomes in the communication between CAFs and cancer cells and other cells of the TME. First, we introduce the multiple roles of CAFs in tumorigenesis. Thereafter, we discuss the ways CAFs communicate with cancer cells and interplay with other cells of the TME, and focus in particular on the role of exosomes. Then, we elaborate on the mechanisms by which CAFs-derived exosomes contribute to cancer progression, as well as and the clinical impact of exosomes. We conclude by discussing aspects of exosomes that deserve further investigation, including emerging insights into making treatment with immune checkpoint inhibitor blockade more efficient.

On-Target Anti-TGF-ß Therapies Are Not Succeeding in Clinical Cancer Treatments: What Are Remaining Challenges?

Metastasis is the leading cause of death for cancer patients. During cancer progression, the initial detachment of cells from the primary tumor and the later colonization of a secondary organ are characterized as limiting steps for metastasis. Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) are opposite dynamic multistep processes that enable these critical events in metastasis by altering the phenotype of cancer cells and improving their ability to migrate, invade and seed at distant organs. Among the molecular pathways that promote tumorigenesis in late-stage cancers, transforming growth factor-ß (TGF-ß) is described as an EMT master inducer by controlling different genes and proteins related to cytoskeleton assembly, cell-cell attachment and extracellular matrix remodeling. Still, despite the successful outcomes of different TGF-ß pharmacological inhibitors in cell culture (in vitro) and animal models (in vivo), results in cancer clinical trials are poor or inconsistent at least, highlighting the existence of crucial components in human cancers that have not been properly explored. Here we review most recent findings to provide perspectives bridging the gap between on-target anti-TGF-ß therapies in vitro and in pre-clinical models and the poor clinical outcomes in treating cancer patients. Specifically, we focus on (i) the dual roles of TGF-ß signaling in cancer metastasis; (ii) dynamic signaling; (iii) functional differences of TGF-ß free in solution vs. in exosomes; (iv) the regulatory effects of tumor microenvironment (TME) - particularly by cancer-associated fibroblasts - on TGF-ß signaling pathway. Clearly identifying and establishing those missing links may provide strategies to revitalize and clinically improve the efficacy of TGF-ß targeted therapies.

Low-power laser alters mRNA levels from DNA repair genes in acute lung injury induced by sepsis in Wistar rats.

Acute lung injury (ALI) is defined as respiratory failure syndrome, in which the pathogenesis could occur from sepsis making it a life-threatening disease by uncontrolled hyperinflammatory responses. A possible treatment for ALI is the use of low-power infrared lasers (LPIL), whose therapeutical effects depend on wavelength, power, fluence, and emission mode. The evaluation mRNA levels of repair gene related to oxidative damage after exposure to LPIL could provide important information about the modulation of genes as treatment for ALI. Thus, the aim of this study was to evaluate the mRNA levels from OGG1, APEX1, ERCC2, and ERCC1 genes in lung tissue from Wistar rats affected by ALI and after exposure to LPIL (808 nm; 100 mW). Adult male Wistar rats (n = 30) were randomized into six groups (n = 5, for each group): control, 10 J/cm2 (2 J), 20 J/cm2 (5 J), ALI, ALI + LPIL 10 J/cm2 and ALI + LPIL 20 J/cm2. ALI was induced by intraperitoneal E. coli lipopolysaccharide injection (10 mg/kg). Lungs were removed, and samples were withdrawn for total RNA extraction, cDNA synthesis, and mRNA levels were evaluated by RT-qPCR. Data normality was verified by Kolmogorov-Smirnov, comparisons among groups were by Student's t test, Mann-Whitney test, one-way ANOVA, Kruskal-Wallis followed by post-tests. Data showed that OGG1 (0.39 ± 0.10), ERCC2 (0.67 ± 0.24), and ERCC1 (0.60 ± 0.19) mRNA levels are reduced in ALI group when compared with the control group (1.00 ± 0.07, 1.03 ± 0.25, 1.01 ± 0.16, respectively) and, after LPIL, mRNA relative levels from DNA repair genes are altered when compared to non-exposed ALI group. Our research shows that ALI alter mRNA levels from genes related to base and nucleotide excision repair genes, suggesting that DNA repair is part of cell response to sepsis, and that photobiomodulation could modulate the mRNA levels from these genes in lung tissue.

Low power blue LED exposure increases effects of doxorubicin on MDA-MB-231 breast cancer cells.

Patients with triple negative breast cancer can develop side effects as a result of chemotherapy. Photodynamic therapy may reduce these side effects if the chemotherapy agent could also act as a photosensitizer. Thus, the aim of this work was to evaluate cytotoxicity and reactive oxygen species production induced by doxorubicin and low power blue LED in breast cancer cultures. Cell viability and reactive oxygen species (ROS) in MDA-MB-231 cultures were evaluated in response to different doxorubicin concentrations and blue LED fluences. Compared with control, cell cultures only incubated with doxorubicin at 25 nM showed 23% of cell viability reduction while its combination with blue LED at 640 J/cm2 reduced 40% of cell viability after 24 h. After 48 h, reduction of cell viability raises to 40% in cell cultures only incubated with doxorubicin and 55% when combined with blue LED. Evaluation 30 min after treatment showed that cells incubated with doxorubicin and exposed to blue LED generated 22% more ROS than controls. Those results show that incubation with doxorubicin combined with exposure to low power blue LED is more cytotoxic and more effective to increase ROS levels in MDA-MB-231 cultures than incubation with doxorubicin alone.

Photobiomodulation effects on mRNA levels from genomic and chromosome stabilization genes in injured muscle.

Muscle injuries are the most prevalent type of injury in sports. A great number of athletes have relapsed in muscle injuries not being treated properly. Photobiomodulation therapy is an inexpensive and safe technique with many benefits in muscle injury treatment. However, little has been explored about the infrared laser effects on DNA and telomeres in muscle injuries. Thus, the aim of this study was to evaluate photobiomodulation effects on mRNA relative levels from genes related to telomere and genomic stabilization in injured muscle. Wistar male rats were randomly divided into six groups: control, laser 25 mW, laser 75 mW, injury, injury laser 25 mW, and injury laser 75 mW. Photobiomodulation was performed with 904 nm, 3 J/cm2 at 25 or 75 mW. Cryoinjury was induced by two applications of a metal probe cooled in liquid nitrogen directly on the tibialis anterior muscle. After euthanasia, skeletal muscle samples were withdrawn and total RNA extracted for evaluation of mRNA levels from genomic (ATM and p53) and chromosome stabilization (TRF1 and TRF2) genes by real-time quantitative polymerization chain reaction. Data show that photobiomodulation reduces the mRNA levels from ATM and p53, as well reduces mRNA levels from TRF1 and TRF2 at 25 and 75 mW in injured skeletal muscle. In conclusion, photobiomodulation alters mRNA relative levels from genes related to genomic and telomere stabilization in injured skeletal muscle.