Pre – clinical studies on oncolytic HSV1716

in hepatocellular carcinoma.

Lynne Braidwood

Origin of HSV1716 (Seprehvir)

Herpes simplex virus type 1 (HSV1) is a common human virus that naturally infects most of the population.

Our product HSV1716 is a modified version of HSV1 that lacks both copies of a single gene encoding a protein (ICP34.5) which is essential for virus replication and virulence in normal cells and tissues

In cancer cells, the requirement for ICP34.5 is negated by proteins and cellular pathways that are active only in cancer cells resulting in specific lysis of the tumour cells, whereas normal cells are unaffected.

Tumour selective mode of action

Clinical Studies Overview• Ongoing study in paediatric/young adults (Non-CNS tumours, 5

patients)

• Ongoing study in malignant pleural mesothelioma (2 patients)

• Completed safety / proof of concept studies in 72 patients

• 47 patients in brain tumour studies (4 studies)

• 5 patients in advanced metastatic melanoma study

• 20 patients in oral squamous cell carcinoma study

• No toxicity attributable to HSV1716 experienced by any patient

• Proof of Principle that HSV1716 virus replicates in tumours

Hepatocellular carcinoma(HCC)• Global health problem- HCC is the most common solid

organ malignancy 5th most common malignancy globally

3rd most common cause of death due to cancer

Increasing incidence (HCC rates tripled in US

between 1975 and 2005)

662,000 deaths annually

c1,500 deaths a year in the UK

• Treatment– Surgery

– Chemotherapy (eg doxorubicin)

• Dismal prognosis 90% of patients dead within six months

almost all patients dead with one year of diagnosis

HSV1716 synergizes with doxorubicin in vitro

• Doxorubicin – used frequently as single agent in advanced HCC and is frequently an active chemotherapeutic in TACE.

• HSV1716 in combination with doxorubicin assessed in vitro in HepG2-luc and HuH7

• Chou-Talalay analysis identifies strong synergy signals in both HCC cell lines

• Potential for enhanced efficacy with standard of care drugHuH7/Doxorubicin FaCI combination index plot.

0

0.5

1

1.5

2

2.5

3

3.5

4

0.00 0.20 0.40 0.60 0.80 1.00Fa

CI

moi 0.5

moi 0.05

HepG2-luc/Doxorubicin FaCI combination index plot

0

0.5

1

1.5

2

2.5

3

3.5

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0.00 0.20 0.40 0.60 0.80 1.00Fa

CI

moi 0.5

moi 0.05

HCC cell linesHepG2-luc •Adherent, epithelial-like cells derived from differentiated HCC •HepG2-luc2 (CaliperLS) is a luciferase expressing cell line derived from HepG2 •Fully permissive for HSV1716

HuH-7 •Epithelial-like tumorigenic cells from well-differentiated, hepatocyte-derived carcinoma male •Fully permissive for HSV1716

1 6 11 16 21 26 31 36 419.00E+04

9.00E+05

9.00E+06

9.00E+07Average radiance values for HepG2-luc xenografts

treated with 2e4pfu HSV1716

non treated xenograft"

average background

"2e4 pfu HSV1716

Days since cell injection

ROI v

alue

(rad

ianc

e)

HepG2-luc xenografts

Treatment Groups Number of mice Number of cures*

IT injection (2e6pfu) 13 12

IT injection (2e5 pfu) 5 5

IT injection (2e4 pfu) 7 6

IV injection (3x 2e6pfu)

5 4

No Virus 13 0

* cure = complete and permanent loss of light signal from HepG2-luc xenograft

0 2 4 6 8 10 12 140

250

500

750

1000

1250No virus1x10e71x10e6

day

Avera

ge t

um

ou

rvo

lum

e

• HSV1716 administered by IV on days 1 and 4• HuH7 tumours treated with HSV1716 at both doses have greatly reduced growth

compared to untreated controls • The difference in growth for untreated vs treated is highly significant by ANOVA (p<0.0001)

Growth of HuH-7 xenografts after 2x intravenous injectionof HSV1716

HuH7 xenograft growth until all no virus controls were sacrificed

• HSV1716 administered by IV on days 1 and 4 • Enhanced survival in both groups treated with

HSV1716 (p = 0.0008)

Survival of mice with HuH7 xenografts after 2x IV injection of HSV1716

0 10 20 30 40 500

102030405060708090

100110

no virus

1e7

1e6

day

Per

cen

t su

rviv

al

HuH-7 xenografts : Lower dose virus – spreading out the virus doses.

0 10 20 30 40 50 60 700

250

500

750

1000

12501x10e6pfu1x10e5pfuno virus

day

tum

ou

r v

olu

me

• HSV1716 administered by IV on days 1 and 14 and 29• HuH7 tumours treated with HSV1716 at both doses have greatly reduced growth compared to untreated controls • The difference in growth for untreated vs treated is highly significant by ANOVA (p<0.0001)

HuH-7 – Survival at lower treatment dose with more spread out virus

treatment

• HSV1716 administered by IV on days 1 and 14 and 29   • Enhanced survival in both groups treated with HSV1716 (p= 0.0157)• On day 66 when the experiment was stopped:-

• 2/6 mice treated with 1x105 pfu were cured• 4/6 mice treated with 1x106 pfu were cured

0 10 20 30 40 50 60 700

25

50

75

1001x10e5pfu1x10e6pfuno virus

day

Perc

en

t su

rviv

al

HSV1716 localisation to HuH7 xenografts

• Mice (n=2) were injected IV with increasing amounts of virus. • Tumours were harvested at 72hr and analyses by titration.• Virus only detected in tumours at titre > input at all doses

0

10000000

20000000

30000000

40000000

50000000

60000000

titr

e (

pfu

/ml)

input pfu

HSV1716 in pre-clinical HCC: Conclusions

72 hrs after systemic delivery the virus homes in exclusively on the tumour. Tumour growth is significantly reduced in the Huh7 model and survival lengthened.Giving virus treatment for a prolonged period results in a significant porportion of cures in the Huh7 model, even at lower doses of HSV1716. HSV1716 completely eliminated luc expressing HepG2 xenografts in 23 of 25 mice. HSV1716 may synergise with existing HCC treatments (doxorubicin). Results strongly supportive of progression to clinical studies of HSV1716 in HCC.

Acknowledgements

Virttu BiologicsDr Joe Conner & Kirsty Learmonth

Glasgow University Dept of Infection and ImmunityProf Shelia Graham

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