R1 서자희

Sonic hedgehog signaling pathway

• Cell proliferation and differentiation dur-ing embryonic development– Paracrine manner

– Loss of signaling activity congenital anom-aly • holoprosencephaly, polydactly, craniofacial de-

fects, and skeletal malformations

–Mutation, misregulation cancer • Medulloblastoma (MBL)

• Basal cell carcinoma (BCC)

• Rhabdomyosarcoma

Role of Hh signaling in cancer

• 1st identified in Gorlin syndrome– Loss of function : PTCH1

– Numerous BCCs, MBL and rhabdomyosarcoma

• Most spordic BCCs & 1/3 of spordic MBLs– Inactivating mutation in

PTCH1

– Activating mutation in SMO (more rarely)

Role of Hh signaling in cancer

• Tumor cell produce and respond to Hh ligand in an autocrine manner– In the absence of mutations in compo-

nents of the Hh pathway, Hh signaling has been described to play a role in the growth of a variety of epithelial cancer types

– Small cell lung cancer, prostate cancer, pancreatic and other GI tract malignacies

• Inhibition of Hh pathway therapeutic effect in tumors – Mutation level : PTCH1 or SMO

– Hh ligand : overexpression in tumors

Natural compound cyclopamine

Small molecule drug optimized Hh-Antag• Inhibition of SMO : interference of tumorogenesis

• GDC-0499 treatment : Tumor regression reported in advanced BCCs & MBLs

• Alternative model in three recent articles– Tumor-produced Hh ligand trigger signaling

in the stromal environment in a paracrine manner

Hh

Hh

Hh

GLI1, PTCH, cyclin D, E, Myc etc

relocalization

Positive & negative feedback loop

Epithelial cell

Mesenchymal cell

De-repressing

Hh pathway in paracrine manner

• Hh ligand-expressing cancer cells of epithelial origin are refractory to ligand, whereas surrounding stroma cells are ligand respon-sive

• Smo-antagonist-mediated growth inhibition not correlate with Hh target gene expression – in vitro growth repression d/t off-target effects– compounds are used at high concentrations

• IC50* of Hh Antag to inhibit ptw in mesenchymal cell line >> IC 50 required to inhibit cell growth x 20

• High drug concentration not correlate with regulation of Hh target

genes (GLI1, PTCH1)• Studies using high concentrations of some

of the SMO-inhibitors need to be interpreted with caution

*half-maximal inhibitory concentration

Nature vol. 455, 2008

Hh-Antag-mediated growth inhibition

• Colorectal, pancreatic, ovarian tumor– Hh ligand expression in particular SHH and/or

IHH

– levels of Hh ligand mRNA expression in tumor cells correlated with increased Gli1 and Ptch1 mRNA levels in the stroma and not the tumor compartment

• Hh ligands activate Hh signaling in the surrounding stroma and not the tumor epithelium

Therapeutic value of inhibition of paracrine signal

• Specific inhibition of Hh signaling in the stroma with either Hh-Antag or a neutralizing anti-Hh antibody

growth inhibition of Hh ligand-expressing xenografts

• Genetic ablation of Smo in the murine stroma

growth inhibition of human tumor xenografts contribution of Hh-activated stroma in tumorigenesis

• Provide important clinical biomarker for selection of Hh-inhibitor responsive tumors– Hh ligand expression in tumor cells

– Hh pathway activation in the stroma

Studies on mouse models of pancreatic ductal adenocarcinoma (P-

DAC)

• Genetic deletion of Smo from pan-creatic epithelial cells – NOT affect the development and or pro-

gression of KrasG12D-driven PDAC– NOT affect Ptch1 and Gli1 expression

levels– Smo expression was dispensable for the

initiation and progression of PDAC

• Epithelial expression of an oncogenic allele of Smoothened (SmoM2 *) – Not induce neoplastic transformation in murine

pancreatic epithelium

– Not affect tumor development and progression in KrasG12D-driven PDAC models

* Smoothened (SmoM2) : triggers ligand-independent activation of Hh pathway constitutively

• SmoM2 failed to activate the Hh pathway in epithelial cells as measured – by measuring Gli1 mRNA levels using quantita-

tive PCR following laser capture microdissection of the tumor and stromal compartments

GENES & DEVELOPMENT 23:24–36 2009

• Epithelial expression of activated Gli2

cooperate with activated KrasG12D to induce U/D pancreatic tumors– expression of truncated form of Gli2 : resistant to key

posttranscriptional regulatory mechanism

– not reflect ability of cell to transduce signal

– recent identification of mutations in the Gli transcrip-tion factors in human pancreatic cancer cell lines

• However, effect of these mutations on Gli activ-ity needs to be determined before concluding – downstream Hh signaling components can contribute

cell autonomously to adenocarcinoma formation

• Gli activity in pancreatic tumor cells

stimulation of Hh independent manner by TGF-β and KRAS

• TGF- β treatment upregulation of Gli1 & Gli3– Smo expression status in PDAC cells

– depletion of Kras expression

downregulation of Gli1 and Ptch1 mRNA

• Gli1 PDAC cancer cell survival KRAS-dependent malignant cellular phenotype

Limitation

• In vitro cell culture

• Levels of Gli activity in the tumor cell compart-ment : much lower than in adjacent stromal cells – by quantitative mRNA analysis(RT-PCR) of microdis-

sected cells from mouse or human PDAC and col-orectal tumor cell culture

• It will be interesting to extend these results to in vivo models– i.e ; by specific deletion of Gli1 from epithelial com-

partment in the mouse PDAC model

Proc Natl Acad Sci USA 2009:106;4254-9

Hh pathway in paracrine manner

• Hh ligands secreted by the epithelial tumor cells activate Hh signaling in the surrounding stroma– Hh signaling expression of soluble

factors & ECM components • acting upon tumor epithelium or other cell

types• promoting tumor growth

• Sonic hedgehog pathway

contribute to fibrotic tissue formation in tumor microenvironment of orthotopic model of human pancreatic cancer

indirect angiogenic factor – induce expression of vascular endothelial

growth factors and angiopoietins

– inhibition of stromal Hh signaling

expression of components of the Wnt and insulin-like growth factor signaling path-ways • insulin-like growth factor 2 : medulloblastoma

• Wnt/β-catenin signaling : skin hamartoma

Requirement for future stud-ies

How these and other factors affect tumor growth

Which factors can be targeted for therapy

– Hh signaling in the self-renewal and survival of var-ious epithelial and nonepithelial cancer stem cells

– Breast, multiple myeloma, chronic myelogenous leukemia stem cells

• Paracrine Hh signaling in stromal cells might also be involved in the establishment and maintenance of a cancer stem cell

In conclusion…

• Tumor-stromal interactions : critical to tumor establishment and growth, and the devel-opment of anti-angiogenic drugs– inhibition of signaling in the relatively normal

stromal cell– less likely to acquire mechanisms of therapeutic

resistance

• Challenging in a clinical setting– GDC-0449

• Ovarian cancer : increase the time until tumor recur-rence

• Metastatic colorectal cancer : improve response to therapy or increase the time until disease recurs

• In BCCs…– Hh signaling in mutation-driven tumors–more difficult to characterize the role of

paracrine signaling of Hh in cancer

• It may be possible that both paracrine and autocrine Hedgehog signaling cooperate in some tumors

Thank You !

Nikon F80 by Ja Hee