It has long been known that endothelial Ca²⁺ signals drive angiogenesis by recruiting multiple Ca²⁺-sensitive decoders in response to pro-angiogenic cues, such as vascular endothelial growth factor, basic fibroblast growth factor, stromal derived factor-1α and angiopoietins. Recently, it was shown that intracellular Ca²⁺ signaling also drives vasculogenesis by stimulation proliferation, tube formation and neovessel formation in endothelial progenitor cells. Herein, we survey how growth factors, chemokines and angiogenic modulators use endothelial Ca²⁺ signaling to regulate angiogenesis and vasculogenesis. The endothelial Ca²⁺ response to pro-angiogenic cues may adopt different waveforms, ranging from Ca²⁺ transients or biphasic Ca²⁺ signals to repetitive Ca²⁺ oscillations, and is mainly driven by endogenous Ca²⁺ release through inositol-1,4,5-trisphosphate receptors and by store-operated Ca²⁺ entry through Orai1 channels. Lysosomal Ca²⁺ release through nicotinic acid adenine dinucleotide phosphate-gated two-pore channels is, however, emerging as a crucial pro-angiogenic pathway, which sustains intracellular Ca²⁺ mobilization. Understanding how endothelial Ca²⁺ signaling regulates angiogenesis and vasculogenesis could shed light on alternative strategies to induce therapeutic angiogenesis or interfere with the aberrant vascularization featuring cancer and intraocular disorders.

CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-associated proteins) has undergone marked advancements since its discovery as an adaptive immune system in bacteria and archaea, emerged as a potent gene-editing tool after the successful engineering of its synthetic guide RNA (sgRNA) toward the targeting of specific DNA sequences with high accuracy. Besides its DNA editing ability, further-developed Cas variants can also edit the epigenome, rendering the CRISPR-Cas system a versatile tool for genome and epigenome manipulation and a pioneering force in precision medicine. This review explores the latest advancements in CRISPR-Cas technology and its therapeutic and biomedical applications, highlighting its transformative impact on precision medicine. Moreover, the current status of CRISPR therapeutics in clinical trials is discussed. Finally, we address the persisting challenges and prospects of CRISPR-Cas technology.

Nicotinic acid adenine dinucleotide phosphate (NAADP) gates two-pore channels 1 and 2 (TPC1 and TPC2) to elicit endo-lysosomal (EL) Ca²⁺ release. NAADP-induced EL Ca²⁺ signals may be amplified by the endoplasmic reticulum (ER) through the Ca²⁺-induced Ca²⁺ release mechanism (CICR). Herein, we aimed at assessing for the first time the role of EL Ca²⁺ signaling in primary cultures of human metastatic colorectal carcinoma (mCRC) by exploiting Ca²⁺ imaging and molecular biology techniques. The lysosomotropic agent, Gly-Phe β-naphthylamide (GPN), and nigericin, which dissipates the ΔpH which drives Ca²⁺ refilling of acidic organelles, caused massive Ca²⁺ release in the presence of a functional inositol-1,4,5-trisphosphate (InsP₃)-sensitive ER Ca²⁺ store. Liposomal delivery of NAADP induced a transient Ca²⁺ release that was reduced by GPN and NED-19, a selective TPC antagonist. Pharmacological and genetic manipulations revealed that the Ca²⁺ response to NAADP was triggered by TPC1, the most expressed TPC isoform in mCRC cells, and required ER-embedded InsP₃ receptors. Finally, NED-19 and genetic silencing of TPC1 reduced fetal calf serum-induced Ca²⁺ signals, proliferation, and extracellular signal-regulated kinase and Akt phoshorylation in mCRC cells. These data demonstrate that NAADP-gated TPC1 could be regarded as a novel target for alternative therapies to treat mCRC.

The acidic vesicles of the endolysosomal (EL) system are emerging as an intracellular Ca²⁺ store implicated in the regulation of multiple cellular functions. The EL Ca²⁺ store releases Ca²⁺ through a variety of Ca²⁺-permeable channels, including Transient Receptor Potential (TRP) Mucolipin 1-3 (TRPML1-3) and two-pore channels 1-2 (TPC1-2), whereas EL Ca²⁺ refilling is sustained by the proton gradient across the EL membrane and/or by the endoplasmic reticulum (ER). EL Ca²⁺ signals may be either spatially restricted to control vesicle trafficking, autophagy and membrane repair or may be amplified into a global Ca²⁺ signal through the Ca²⁺-dependent recruitment of ER-embedded channels. Emerging evidence suggested that nicotinic acid adenine dinucleotide phosphate (NAADP)-gated TPCs sustain multiple cancer hallmarks, such as migration, invasiveness and angiogenesis. Herein, we first survey the EL Ca²⁺ refilling and release mechanisms and then focus on the oncogenic role of EL Ca²⁺ signaling. While the evidence in favor of TRPML1 involvement in neoplastic transformation is yet to be clearly provided, TPCs are emerging as an alternative target for anticancer therapies.