Background
Introduction: from the theory of angiogenesis to an orchestra of alternatively spliced angiogenic genes
Alternative splicing
Main text
Genome-wide AS changes in ECs
AS isoforms acting on the extracellular space during physiological and tumor angiogenesis
GENE | AS variant | Relevance in cancer angiogenesis |
---|---|---|
VEGF-A | VEGF-Axxxa | Expression/function Overexpressed by a wide variety of human tumors. Pro-angiogenic function, produced by both cancer cells and ECs [46]. |
VEGF-Axxxb | Expression/function Anti-angiogenic function, generally downregulated in cancer [46]; not detected in normal or tumor ECs [47]. Examples of potential use for therapy | |
VEGF receptors (VEGFRs) | sVEGFR1 | Expression/function Anti-angiogenic function, inhibits VEGF signalling in ECs [51]. Controversial role in cancer [52‐54]. Examples of potential use for therapy Morpholino oligonucleotides to promote AS into sVEGFR1 [55]. |
sVEGFR2 | Expression/function Decreases lymphangiogenesis. Downregulated in neuroblastoma patients [56]. | |
Neuropilins (NRPs) | sNRP1 | Expression/function Examples of potential use for therapy Overexpression of sNRP1 to prevent VEGF signalling [60]. |
NRP1-∆7 | Expression/function Altered glycosylation. Anti-angiogenic function [61]. | |
NRP1-∆E4, NRP1-∆E5 | Expression/function Altered glycosylation and endocytic trafficking [62]. | |
s9NRP2 | Expression/function Decoy function [63]. | |
Membrane-bound NRP2 variants | Expression/function Differentially activate signalling pathways [58]. | |
Fibroblasts growth factor receptors (FGFRs) | FGFRIIIb | Expression/function Examples of potential use for therapy Anti-FGFR2-IIIb–Specific Antibody (GP369) [67]. |
FGFRIIIc | Expression/function | |
sFGFRs | Expression/function Possible decoy function [69]. | |
Deletion of auto-inhibitory domain | Expression/function Hyper-activation of the signalling pathway [69]. | |
C-term FGFRs AS variants C1, C2, C3 | Expression/function Differential impact on receptor internalization and downstream signalling. C3 implicated in oncogenesis [70]. | |
Deletion of VT motif | Expression/function Deletion affects downstream signalling [71]. | |
Vasohibins (VASHs) | VASH1A | Expression/function Anti-angiogenic-function. Expressed by ECs [72]. Examples of potential use for therapy Overexpression of VASH1A [72]. |
VASH1B | Expression/function Expressed by ECs. Promotes the normalization of tumor blood vessels [72]. Examples of potential use for therapy Overexpression of VASH1B [72]. | |
VASH2-355aa | Expression/function Expressed by ECs [73]; unknown function. | |
VASH2-290aa | Expression/function Anti-angiogenic function [73]. | |
Angiopoietins (ANGs) | ANG1–0.7, − 0.9 and − 1.3 kb | Expression/function Differentially activates TIE2 pathway [74]. |
ANG2443 | Expression/function Expressed in primary ECs and non-endothelial tumor cell lines. It antagonizes TIE2 signalling during tumorigenesis and inflammation [75]. | |
ANG2B | Expression/function Differentially activates TIE2 signalling [76]. | |
Fibronectin (FN) | EDA/EDB-FN | Expression/function Expressed during embryonic and tumor angiogenesis. EDA-FN plays a role in vascular remodelling and prevents vascular oxidative stress in diabetic conditions [77, 78]. Examples of potential use for therapy Drug delivery [79]. |
Tenascin C (TNC) | Large TNC variants | Expression/function Expressed in pathological tissues undergoing active remodelling. Favour cell migration [80]. Specific spliced variants or single AS domains are associated with different tumor types [80] types; FNIII C-bearing TNC isoform is highly expressed in brain and lung tumors, associated with tumor stroma [81]. Examples of potential use for therapy TNC antibodies to deliver cytotoxic molecules, recognizing the AS domains A1 to D of the large isoform of TNC. Aptamer TTA1 [82]. |
SLIT2 | Slit2-WT | Expression/function Expressed and released by tumor cells. Reduces EC permeability [83]. |
Slit2-ΔE15 | Expression/function Expressed and released by normal cells. Reduces EC permeability and plays a role in vessel normalization [83]. | |
PECAM1 | PECAM1-FL, Δ12, Δ13, Δ14, Δ15, Δ14&15 | Expression/function |
sPECAM1 | Expression/function Possible function in regulating PECAM1-mediated cellular interactions [87]. | |
CD146 | shCD146 | Expression/function Promotes EC proliferation, migration and adhesion [88]. |
lgCD146 | Expression/function Promotes EC tube formation and stabilization [88]. | |
CD44 | CD44v6 | Expression/function Controls EC migration, sprouting and tube formation, acting as a VEGFR2 co-receptor for VEGF-A [89]. Examples of potential use for therapy |
Endoglin (ENG) | L-endoglin | Expression/function |
S-endoglin | Expression/function Interacts with TGFβ type I receptors ALK5, stimulating ALK5 pathway. Associated with altered pulmonary angiogenesis [98]. It is induced by senescence and able to contribute to NO-dependent vascular homeostasis. | |
Insulin receptor (IR) | IR-A | Expression/function Pro-proliferative function; overexpressed in tumor vasculature [99]. |
Tissue factor (TF) | asTF | Expression/function Soluble factor, highly expressed in advanced stages of several human cancers [100, 101]. Stimulates tumor growth, angiogenesis and metastasis [102]. Examples of potential use for therapy Antibody drug conjugate of TF and monomethyl auristatin E [103]. |
flTF | Expression/function Highly expressed in several types of cancer. Involved in cancer-related thrombosis, tumor growth and metastasis [104]. Examples of potential use for therapy Anti-flTF antibody 10H10 [105]. | |
L1CAM (L1) | L1-ΔTM | Expression/function Soluble form of L1CAM, released by ECs. Promotes EC tube formation and neovascularization. Overexpressed in the ovarian cancer vasculature; associated with tumor vascularization [106]. |
L1-FL | Expression/function Highly expressed in tumor vasculature several types of cancer. Pro-angiogenic function [107]. |
VEGF-A
VEGF receptors (VEGFRs)
Neuropilins (NRPs)
Fibroblasts growth factor receptors (FGFRs)
Vasohibins
Angiopoietins
Fibronectin (FN)
Tenascin C (TNC)
SLIT guidance ligand 2 (SLIT2)
PECAM1
CD146
CD44
Endoglin (ENG)
Insulin receptor (IR)
Tissue factor (TF)
Cell adhesion molecule L1 (L1CAM)
SRFs regulating EC functions
PTBP1
SRSF1
NOVA2
MBNLs
ELAVL1
RBFOX2
Therapeutic strategies exploiting AS of angiogenic factors in cancer
Therapeutic approach | Examples | Pros and cons |
---|---|---|
Controlling the activity of splicing factor regulators | Poor specificity, resulting in AS modification of multiple genes besides VEGFA. | |
Inhibiting the assembly of the spliceosome machinery | - Compounds binding to the spliceosome component SF3b: FR901464 and its methylated derivative, spliceostatin A [199]. | Poor specificity, affecting AS of multiple genes; partial understanding of mechanism of action. |
Interfering with splicing sites | - Morpholino oligonucleotides targeting the exon 13/intron 13 junction of the VEGFR1 pre-mRNA, favoring the production of the anti-angiogenic, soluble form of VEGFR1 [55]. | Possibility to target one single gene; off-target effects due to either the presence of the targeted sequence in other portions of the genome or tolerance toward mismatches. |
Blocking pro-angiogenic isoforms | - Intravenous delivery of autologous T cells, modified to recognize CD44v6 on the surface of cancer cells (ClinicalTrials.gov: NCT04427449 [95]). | High specificity with minimal side effects; cumbersome and expensive design and production. |
Overexpressing anti-angiogenic isoforms | - Overexpression of sNRP1 to prevent VEGF signalling [60]. - Overexpression of either VASH1B or VASH1A [72]. | Delivery requiring either gene therapy or production of recombinant proteins; no effect on the level of pro-angiogenic isoforms. |
Exploiting cancer-specific isoforms for drug delivery | - Monoclonal antibodies and aptides targeting EDA/EDB domains of FN: F8 fused to IL-2 [205, 206]; L19 fused to either IL-2 or IL-12 [207, 208]; EDB-targeting aptides conjugated with doxorubicin-containing liposomes [209, 210]. - Monoclonal antibodies (F16 fused to IL-2) and aptamers targeting domains A1-D of TNC [211]. | High specificity for cancer cells; cumbersome and expensive design and production; toxicity related to the chemotherapeutic agent. |