M6L2 ECM targeting therapies

  • Fibroblasts in the ECM:

    • Large secretome - cytokines, growth factors, ECM remodelling enzymes etc to maintain homeostasis in ECM

    • ECM production

  • Collagen - most abundant ECM protein

    • 28 types - collagen I (fibrillar, for support), collagen IV (basal lamina) most well known

    • Homo/heterotrimers

    • Fibrils and network forming collagens

    • Integrins can bind to collagen for structural support and signalling (proliferation, adhesion/migration, survival, differentiation pathways etc)

  • Fibronectin

    • ECM glycoprotein

    • Binds to other ECM proteins, eg fibrin

    • Contains RGD motifs which binds cell surface receptors

    • Important in TBFb deposition

  • Glycoproteins, proteoglycans, glycosaminoglycans (GAGs)

    • Proteoglycans - protein core with GAGs (negatively charged sugars) attached to it

    • 5 main GAGs - hyaluronic acid most important

    • Important role in lubrication, local sequestration of GFs and cytokines

    • Binds CD44

  • ECM modifying enzymes

    • MMPs - LOX, ADAMs

    • Degradation, modification, enzyme activation, stiffness/crosslinking, GF regulation

    • Can be secreted or be on cell surface

  • ECM in wound healing

    • Coagulation/inflammation - transient fibrin matrix, recruitment of neutrophils and later macrophages

    • Tissue formation - keratinocytes (re-epithelialisation), MMP production, myofiobroplasts, angiogenesis

      • Most relevant for cancer - cancer as the ‘never healing wound’, perpetual inflammation and extensive remodelling by MMPs

    • Remodeling - collagen formation, wound contraction, scar maturation

  • ECM in cancer

    • ECM becomes an active player in cancer, containing the niche and protecting the tumour from immune infiltration and drug delivery

    • Upon tissue invasion, a layer of ECM helps cancer to behave the way that it does

  • CAFs

    • Produce tumour ECM together with cancer cells

    • Very heterogenous (many progenitors - adipocytes, endothelial cells, epithelial cells, mesenchymal cells etc)

    • Produce cytokines and ECM remodelling enzymes

  • Collagen in cancer ECM

    • Cancer —> more collagen deposition by CAFs or cancer cells

    • Collagen fibres are crosslinked to a higher extent (stiffer)

    • Alignment of collegen fibres change

  • Tumour associated collagen signatures (TACS)

    • TACS1 - random alignment of collagen directly adjacent to tumour core

    • TACS2 - slightly more aligned

    • TACS3 - aligned collagen tracts which give cancer cells a path for invasion

  • Fibronectin in cancer

    • Higher expression of extra domain B fibronectin (EDB-FN)

    • Expressed in development (oncofoetal)

    • Vascularisation, angioghenesis

    • Can be used as predictive marker of cancer progression (more expression = more invasion, resistance…)

    • Not in all cancers, but about ~50%

  • Glycoproteins, proteoglycans, GAGs in cancer

    • Upregulated

    • Often coupled with upregulated proteoglycans

    • Correlates with poorer therapeutic penetration, eg Abs

  • Effects on therapies

  • Chemo/radioresistance

    • LOXL2 (collagen crosslinking enzyme) expression in spheroids significantly reduces doxorubicin penetration vs control

    • Increasing Gy has a greater killing effect in 2D cultures vs 3D environment

  • Immunosuppression

    • Covalent linkage from immunosuppressants to ECM

      • TGFb - master cytokine

      • Anti-tumour in early stages, pro-tumour as cancer evolves

      • Covalently linked into LTBP - LAP-dimer hides TBFb

      • Once it is released signalling is rapid due to proximity to cells

      • Hard to target TGFb as it is hidden

    • Association of GFs, GAGs, cytokines…

      • Binds to VEGF, PDGF, FGF, TGF, BMP, NGF, IL-10, IL-6 etc…

      • Hyaluronan (HA) in naked mole rat is x5 larger vs human or mouse

      • Introducing cancer cells into skin fibroblasts from naked mole rat does not develop into tumours

      • CD44 interaction with normal HA vs preventing CD44 interaction with high MW HA could be the reason

      • HA may be an immunosuppressant

    • ECM components

  • Most research is done in 2D - not representative of 3D architecture in vivo (though cheaper, less time consuming, less variable)

    • No ECM to mature ECM: 2D tissue culture —> spheroids —> organoids —> in vivo (subcutaneous) —> in vivo (orthotopic) —> patient derived tissue slices

  • ECM targeting

    • Targeting is difficult - expressed everywhere in the body

    • Conventional targeting/depletion of ECM proteins - transcriptional depletion, ECM component mimetics, destructiion of ECM producing cells, degradation of ECM

      • Collagen targeting with halofuginone - decreases alpha smooth muscle actin (marker for CAFs), but drug toxicity

      • GAG targeting with PEGPH20 - small TI, not much change in survival

      • Proteoglycan/GAG approach - PG545 (mimetic), activates TLR9 (immune activation), can combine with anti-PD1

      • Generally conventional approaches have not met goals or terminated due to severe AEs

    • ECM degradation as supportive strategy - combination with CART cells, OVs etc

      • OVs with transgene hyaluronidase - increaed spread and efficacy of OV in vivo

      • CAR-T cell + ECM degradation enzymes are more potent at infiltrating/clearing solid tumours, eg. CAR-T cell against HER2 expressing MMP9/12 and HPSE entering TME through synNotch system

    • Specific ECM targeting - specific ECM antigens/components

      • Targeting EDB-FN: EDB-FN x IL-2 could help clear renal cell carcinoma (some patients, not others)

      • Fibronum by Philogen 0 targeting antibody (L19) + TNF (immunostimulatory and cytotoxic payload)

      • Targeting tenascin C - anti-FBG + anti-PDL1

      • Targeting tenascin C with CAR-T cells - in GBM the ECM is very rich in tenascin C (increases with the grade)

      • PIGF-1 and PIGF-2: 21 AA motif which binds to ECM fused with anti-CTLA4/PDL1 rediuces tumour growth in mice