Targeting angiogenesis and - PowerPoint Presentation

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Targeting angiogenesis and the tumour microenvironment in cancer

Last updated: April 2020

IntroductionThis chapter provides:An overview of the regulatory processes involved in angiogenesis in normal physiology and cancer, and explains the role of abnormal vasculature

in tumour progressionA discussion of angiogenesis and immunosuppression in the TME and how the key angiogenic factors VEGF and Ang2 can both promote immunosuppressionA rationale for vascular normalisation as a therapeutic strategy, including dual targeting of VEGF and Ang2, as well as combination therapy with PD-1 blockade

Ang2, angiopoietin 2

;

PD-1, programmed

cell death protein

1; TME

, tumour microenvironment;

VEGF, vascular

endothelial growth

factor.

Regulation of angiogenesis in normal physiology and cancer

Tip cell

Stalk cell

Physiological angiogenesis occurs in embryonic development and wound healing

Angiogenesis is the

formation of new blood vessels from pre-existing

vessels and is controlled by a

balance of endogenous stimulators and inhibitors

. During embryonic development and wound healing, various stimuli can

tip the balance

in favour of angiogenesis

1,2

1. Ramjiawan RR, et al. Angiogenesis 2017;20(2):185–204. 2.

Albini

A. et al.

Nat Rev Clin Oncol

2012;9:498–509;

3. Lugano R, et al. Cell

Mol Life

Sci 2019 [Epub

ahead of print

].

Sprouting angiogenesis

3

Intussusceptive

angiogenesis

3

Stimulators

of angiogenesis

1,2

VEGF; PIGF; Ang1/2; Tie2; FGF; PDGF; EGF; MMP-2; MMP-9; COX-2;

mTOR

; ROS; calories; glucose

,

fat; pH

, oxygen

levels

Inhibitors of angiogenisis

1,2

TSP-1; PF-4; angiostatin; endostatin; tumstatin; interferon

α

,

β

,

γ

; TIMPs; AMPK; tight junctions; integrins (ligated)

AMPK, 5' AMP-activated protein

kinase; Ang1/2, angiopoietin 1/2;

COX-2,

cyclooxygenase 2;

EGF, epidermal growth

factor; FGF

, fibroblast growth

factor

;

MMP-2/9, matrix metallopeptidase 2/9; mTOR,

mammalian target of

rapamycin;

PD-1, programmed cell death protein 1; PDGF,

platelet-derived

growth

factor; PF-4

,

platelet

factor

4;

PIGF, placental growth factor; ROS,

reactive

oxygen

species;

TIMP, tissue inhibitor of

metalloproteinase; TME, tumour microenvironment

; TSP-1, thrombospondin

1; VEGF, vascular endothelial growth factor.

In

sprouting angiogenesis

, outgrowing tip cells fuse with an existing vessel or newly formed sprout, whereas in

intussusceptive

angiogenesis

a pre-existing vessel splits into two

3

In cancer, angiogenesis is critical for tumour growth and metastasis1. Zimna A, Kurpisz M. Biomed Res Int 2015;2015:549412; 2. Tocris Bioscience. Angiogenesis. https://www.tocris.com/cell-biology/angiogenesis (Accessed: February 2020).

1. Hypoxia induces the release of pro-angiogenic factors 2. Hypoxia also upregulates protease expression, leading to basement membrane degradation and pericyte detachment3. Tip cells migrate along the angiogenic factor gradient4. Endothelial cells differentiate into proliferative stalk cells

5

. VEGF stimulates DLL4 secretion, which binds to Notch-1 receptors, thereby downregulating VEGFR suppressing proliferation

As

a tumour develops, its size is limited by the diffusion of nutrients, oxygen and metabolites from existing blood

vessels.

Angiogenesis

acts as a checkpoint

that permits further tumour

growth

1,2

Pathological

angiogenesis

provides the

vascular supply

for proliferating

cells1,2

Steps involved in tumour vascularisation

2

In cancer, angiogenesis is driven by hypoxia

2

DLL4,

delta-like canonical Notch ligand 4; PDGFβ, platelet-derived growth factor beta; VEGF, vascular endothelial growth factor; VEGFR, vascular endothelial growth factor receptor.Endothelial cellPericyte detachmentBasement membrane degradationEndothelialprogenitor cellBasementmembraneHypoxicenvironmentstabilisesexpressionof HIF-1VEGFR2, EGFR and FGFRVEGFEGFFGFTGFβIGF1MMPTip cellVEGFR2Stalk cellVEGFR2Notch 1DLL4PDGFPericyte recruitmentHypersprouting6. PDGFβ stimulates pericyte attachment and reduces proliferation and VEGF sensitivity. The blood supply stimulates further tumour growth

Angiogenesis

Oncogenic angiogenesis and the resulting abnormal vasculature is one of the hallmarks of cancer

1

1. Hanahan D, et al. Cell 2011;144(5);646–74; 2. Ramjiawan RR, et al. Angiogenesis 2017;20(2):185–204

.

Pathological angiogenesis leads to vasculature with

abnormal structure and function

2

Blood vessels supplying solid tumours are often

tortuous and

disorganised

,

and are often

excessively leaky

2

This leads to

changes in the TME

2

TME

, tumour

microenvironment

.

↑ Adhesion

molecules

TGF

β

, adenosine

IL-10

↓ Phagocytosis

↓

Antitumour

M1

phenotype

↑ Pro-tumour

M2

phenotype

↑ T

reg

Cell

EC

DC

TAM

↓ Maturation

↓ Antigen

presentation↑ PD-L1↑ PD-L1↑ PD-L1↓ T-cell Infiltration↑ FASLLow O2Low pHTumour↑ PD-L1CTLAbnormal vasculature promotes tumour progressionAbnormal vasculature contributes to tumour progression by impairing perfusion, resulting in tumour hypoxia and low intratumoural pH1The leaky nature of tumour blood vessels in combination with dysfunctional lymphatic drainage also leads to elevated interstitial fluid pressure in the TME1Abnormal vessels and impaired perfusion can restrict entry of cytotoxic drugs and immune cells from the circulation into tumours1Factors in the TME can also impair the function of CD8+ cytotoxic lymphocytes, and limit their penetration into the tumour1Furthermore, abnormal and leaky tumour blood vessels facilitate the intravasation of cancer cells into the systemic circulation, promoting metastasis11. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40.Abnormal vasculature↑ Immunity suppression↓ Adaptive immunityCD, cluster of differentiation; CTL, cytotoxic T lymphocyte; DC, dendritic cell; EC, endothelial cell; FASL, Fas ligand; IL-10, interleukin-10; PD-1, programmed cell death protein 1; PD-L1; programmed death ligand 1; TAM, tumour-associated macrophage; TGFβ, transforming growth factor beta; TME, tumour microenvironment; Treg, regulatory T cell.

The tumour microenvironment contributes to resistance to immunotherapy1. Jenkins RW, et al. Br. J. Cancer 2018;118:9–16; 2.Topalian

SL, et al. Cancer Cell 2015;27(4):450–61.

Arg1, arginase-1; CD, cluster of differentiation; CTLA-4, cytotoxic T lymphocyte-associated protein 4; IDO, indoleamine-pyrrole 2,3-dioxygenase; JAK1,2; Janus kinase 1, 2; LAG3, lymphocyte-activation gene 3; MDSC, myeloid-derived suppressor cell; PD-L1; programmed death ligand 1; PD-1, programmed cell death protein 1; PGE

2

, prostaglandin

E

2

;

TCR, T-cell receptor; TIM3, T-cell immunoglobulin and mucin domain 3; TME, tumour microenvironment; T

reg

, regulatory T cell; VISTA, V-domain Ig suppressor of T-cell activation;

α

PD-L1, anti-PD-L1 antibody;

α

PD-1, anti-PD-1 antibody;

Β2

M, beta-2 microglobulin.

Inadequate tumour-specific T-cell function is

a putative

mechanism

of resistance to

immunotherapy that is driven by the TME

1

An immunosuppressive TME is characterised by:1,2High levels of immune-suppressing cytokines and/or metabolitesRecruitment of immune suppressive cells such as MDSCs and TregsMutations in key effector pathwaysHigh levels of PD-L1 expressionPreclinical models have shown an association between elevation of immune-suppressive cell types (including Tregs, MDSCs, Th2 CD4+ T cells, and M2-polarised tumour-associated macrophages) in the TME and impaired immunotherapy efficacy1These cell types promote an immunosuppressive TME that inhibits antitumour cytotoxic and Th1-directed T-cell activities, primarily through the release of cytokines, chemokines, and other soluble mediators1

Angiogenesis and immunosuppression in the TME

Angiogenesis and immunosuppression are interconnected processesPreclinical and clinical evidence suggests that angiogenesis and immunosuppression have shared

regulators1Molecules that regulate angiogenesis can affect immune cells and their interaction with tumours through direct effects on immune cells, as well as indirect effects on the endothelium and through vascular normalisation2Pro-angiogenic molecules are associated with immunosuppressive effects on antigen presentation, T-cell priming, T-cell trafficking and T-cell tumour infiltration21. Campesato LF, Merghoub T. Ann Transl Med 2017;5(24):

497;

2.

Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40

.

VEGF

Ang2

(others)

Angiogenic factors

Angiogenesis

Immunosuppression

Ang2, angiopoietin 2

;

VEGF, vascular

endothelial growth

factor.

Excessive VEGF in the TME promotes immunosuppression

In addition to its key role in angiogenesis, excessive

VEGF in the TME induces immunosuppression via the following

mechanisms:

1

Increased VEGF directly

inhibits CTL trafficking, proliferation and effector function

VEGF

inhibits dendritic cell maturation and antigen presentation

, hampering

T-cell

activation and reducing the T cell-mediated immune response

VEGF increases the

number and

enhances the function

of

immunosuppressive

T

reg

cells

,

TAMs and/or

monocytesVEGF promotes angiogenesis, resulting in abnormal tumour vasculature, hypoxia, and low pH in the TME1. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40.Ang2, angiopoietin 2; CTL, cytotoxic T lymphocyte; DC, dendritic cell; MDSC, myeloid-derived suppressor cell; TAM, tumour-associated macrophage; TME, tumour microenvironment; Treg, regulatory T cell; VEGF, vascular endothelial growth factor. ↓ Adaptive immunity↓ Innate immunity TumourCells known to produce VEGFCells known to produce ANG2

Ang2 signalling can also induce immunosuppressionThe immuno-modulatory role of Ang2, another regulator of angiogenesis, is not as well understood as that of VEGF. However, activated Ang2 promotes

immunosuppression in tumours through the following mechanisms:11. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40.Increasing leukocyte–endothelial interactions via upregulation of adhesion molecules, facilitating recruitment of immunosuppressive cells, e.g. MDSCs, Tregs and Tie2-expressing monocytes

Disrupting

EC–pericyte

contacts, facilitating

migration of immune cells

out of the vasculature and into the TME

Modulating the function

of

monocytes

by suppressing the secretion

of

TNF, thus restricting their anticancer activity

Ang2, angiopoietin 2

; EC, endothelial

cell; MDSC

, myeloid-derived suppressor

cell

; TAM, tumour-associated

macrophage;

TME

, tumour microenvironment;

TNF, tumour

necrosis factor; Treg, regulatory T cell; VEGF, vascular endothelial growth factor.

The VEGF/Ang2 signalling axis facilitates tumour cell migration and metastasis1. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40

.Overexpression of VEGF and Ang2 may mediate metastasis through a variety of mechanisms:1

Furthermore,

high

levels of Ang2 expression

in patients with breast cancer

have been shown to correlate with

the

presence of metastatic

disease

1

Inducing structural

and biochemical

abnormality of

tumour-associated

blood vessels

In animal models, VEGF also exerts pro-metastatic effects through activation of the

calcineurin–nuclear

factor of activated cell

dissemination

Preclinical evidence

in models of breast cancer suggests

that a high level of Ang2 pathway activation results in lymph node invasion and metastasis

Ang2, angiopoietin 2; VEGF, vascular endothelial growth factor.

Targeting angiogenesis and the TME in cancerTherapeutic strategies

­­­­Tumour perfusion

and oxygenation

Vasogenic oedema

Interstitial pressure

Drug delivery

Cancer cell shedding

Invasiveness

Metastasis

Drug and/ or radiation

sensitivity

Immune regulatory cells

Immune effector cells

Anti-angiogenic treatment

Vascular

normalisation

Progression

Response

Immunosuppressive

microenvironment

Immunosupportive

microenvironment

TAM

CTL

Vascular normalisation converts an immunosuppressive TME to an immunosupportive one

Treatment with anti-angiogenic agents can

renormalise tumour vasculature

. Vascular normalisation

converts an immunosuppressive microenvironment to an immunosupportive one by improving blood perfusion and oxygenation, thereby enabling increased infiltration of immune effector cells1Vascular normalisation also enhances the delivery and effectiveness of chemotherapy and immunotherapy1A challenge with anti-VEGF therapies is that vascular normalisation may only be transient (i.e. lasting days to weeks), and not sustained1Preclinical evidence suggests that dual inhibition of Ang2 and VEGF may extend the window of normalisation compared with inhibition of either pathway alone11. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40.Ang2, angiopoietin 2; CTL, cytotoxic T lymphocyte; TAM, tumour-associated macrophage; TME, tumour microenvironment; VEGF, vascular endothelial growth factor.

Endothelial

cell membrane

Inhibition of

angiogenesis and

metastasis

Tie2

Ang2

VEGFR2

VEGF

Dual blockade

Dual targeting of VEGF and Ang2 may be more effective than targeting either pathway alone

VEGF

and Ang2 signalling have different but

complementary functions

in tumour

angiogenesis:

1–3

Ang2

destabilises established blood vessels

through interruption of vascular tyrosine protein kinase receptor Tie2 signalling, which promotes vessel remodelling – a prerequisite for sprouting angiogenesis

Signalling via VEGF

regulates endothelial cell proliferation and migration, and vessel

sprouting

Tumours might escape anti-VEGF therapies through alternative modes of vascularisation, e.g. upregulation of alternative angiogenic pathways such as Ang2–Tie2 signalling1Preclinical studies in mouse models of various solid tumours have shown that combined VEGF/Ang2 blockade prolongs survival compared with blockade of either pathway alone1Dual VEGF/Ang2 inhibition was also associated with a reduced frequency of metastatic dissemination compared with anti-VEGF therapy alone in models of breast cancer11. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40; 2. Gerald D, et al. Cancer Res 2013;73(6):1649–57; 3. Huang H, et al. Nat Rev Cancer 2010;10(8):575–85. Targeting angiogenesis by inhibiting VEGF and Ang2 signallingAng2, angiopoietin 2; VEGF, vascular endothelial growth factor; VEGFR(2), vascular endothelial growth factor receptor (2).

VEGF/Ang2 inhibition in combination with PD-1 blockade could further improve therapeutic efficacyDual inhibition of VEGF and Ang2 enhances the TME to support T-cell trafficking and function

in the tumour, providing a rationale for their use in combination with a PD-1 inhibitor1‒5The triple combination of VEGF, Ang2 and PD-1 inhibition may further drive T cell-mediated tumour cell death1‒51. Hofmann I, et al. Poster presentation at the 8th Euro Global Summit on Cancer Therapy 2015; 2. Fukumura D, et al. Nat Rev Clin Oncol 2018;15(5):325‒40; 3. Gerald D, et al. Cancer Res 2013;73(6):1649–57; 4. Huang H, et al. Nat Rev Cancer 2010;10(8):575–85; 5. Boehringer Ingelheim. Data on file.

Ang2, angiopoietin 2

; CD, cluster of differentiation; PD-1, programmed cell death protein 1; TME, tumour

microenvironment; VEGF, vascular

endothelial growth

factor

, VEGFR(2), vascular endothelial growth factor receptor (2).

Summary

SummaryPathological angiogenesis in cancer can result in abnormal vasculature and an immunosuppressed TME, which can fuel tumour progression

Angiogenesis and immunosuppression are interconnected processes, both regulated by VEGF and Ang2Vascular normalisation with antiangiogenic agents can convert an immunosuppressive TME to an immunosupportive oneVEGF/Ang2 inhibition in combination with PD-1 blockade represents a rational therapeutic strategy to normalise tumour vasculature, reprogramme the TME and maximise the effectiveness of immunotherapy

Ang2, angiopoietin 2

;

PD-1, programmed

cell death protein

1; TME

, tumour microenvironment;

VEGF, vascular

endothelial growth

factor.