MINERVAL:

A BREAKTHROUGH IN THE TREATMENT OF CANCER

MECHANISM OF ACTION

Minerval +SMS1 SM K/H-Ras MAPKs PI3K/Akt/mTOR Selective

inactivation inhibition Cancer cell death

Minerval (2OHOA) is an orally bioavailable synthetic derivative of oleic acid that crosses the Blood Brain Barrier and activates sphingomyelin
synthase 1 (SMS1), a key enzyme that catalyses the reversible conversion of PC, PE or PS into SM and DAG, leading to an increased concentration of
sphingomyelin (SM) in the cancer cell membrane. This mislocalizes K/H-Ras from the Plasma Membrane and inhibits its nanoclustering, short-cutting Ras-
associated proliferative signalling pathways, which exhibit an aberrant activity in at least one third of all human cancers

HES1 mRNA (relative values)

150
Right. The beginning and the end of the mechanism of action. The anti-tumoral effect SNB19

of Minerval (IC50) on a variety of cancer cell lines strongly correlates with basal levels of MRC5

SMS1 mRNA expression (the beginning) and with a decrease of HES1 mRNA expression 100
SNB75
U87MG
after 24h treatment, expressed in % of basal levels (untreated controls). Regulation of
HES1 (transcription factor at the end of Ras/Notch/autophagic antiproliferative signal 50
SF268
U251
transduction pathways) confirms the modulation of Ras signalling by Minerval SF295
U118

0
0 500 1000
IC50 (M)

MOA overview: SMS1 activation

Increases the levels of SM and DAG
products in cancer cell membranes,
accompanied with reductions of PC, PE
and PS levels, which mislocalizes K/H-
Ras from its active domain in the plasma
membrane and inhibits its nanoclustering,
short-cutting key Ras-dependent
proliferation pathways (like Ras/MAPK,
Top. The activation of SMS1 accumulates sphingomyelin in the cell membrane, Pi3K/AKT/mTOR or PKC/Cyclin CDK),
translocating K-Ras to non-active cytosolic domains (left) in human glioma U118 cells. causing cell cycle arrest followed by
As a consequence Ras-dependent proliferative signalling is inhibited and autophagy selective death of cancer cells
is selectively induced as determined by fluorescence autophagosome presence in non-
cancer (MRC5) and human glioma (1321N1) cell lines (right)

EFFICACY IN PRECLINICAL STUDIES

Minerval has greater efficacy than temozolomide (TMZ) and no tumour relapse was observed after treatment termination

Top. Minervals (2OHOA) efficacy against human glioma in nude mice bearing human gliomas

Minerval crosses the Blood Brain Barrier (BBB)

Left. Immunocytochemical analysis of brains Top and left. Minerval has demonstrated
from nude mice in an orthotopic model a marked anticancer effect in xenograft
inoculated with human glioma cells (SF767) animal models, outperforming
and treated (p.o.) during 42 days with temozolomide (TMZ) in 50-day treatment
vehicle (control, C), Minerval (2OHOA) or (upper right). Combinatory regime with
temozolomide (TMZ) TMZ showed strong synergistic results
Bars: 200 mm (1, 2, 3) and 50 mm (4, 5, 6) after 60-day treatment (upper left)
In mouse #2 (treated with 2OHOA) almost Animals treated with Minerval do not
no traces of tumour were found. In mouse show tumour relapse after treatment
#3 (representative also of #4 and #5) no termination, as it happens with animals
lesions were detected (right panels) treated with TMZ (bottom left)

Minerval Min+Gem Minerval

Top. Minervals efficacy in xenograft models of human lung cancer (A549), pancreatic cancer (BXPC3), prostate cancer (PC3) and leukaemia (Jurkat), all with low SMS1 levels
Supported by:
 MINERVAL:
A BREAKTHROUGH IN THE TREATMENT OF GLIOMA

MIN-001-1203: A phase I/IIa open-label dose escalation study of Minerval in subjects with advanced
solid tumors including malignant glioma
Study design
Case study 1: response in GBM patient 010202 (54y old male)
Part A. Dose escalating study. Up to 30 patients.
Apr 2012: Partial debulking surgery followed by radical chemo-radiotherapy
21-day cycles. Glioma and other solid tumours
Aug-Sep 2012: Adjuvant Temozolomide, with PD after 3 cycles
Part B. Exploratory study. Up to 20 patients in
Nov-Feb 2013: PCV chemotherapy with PD after 4 cycles
two groups. 21-day cycles. 1st group with glioma
Aug 2013: enrolled in MIN-001-1203 trial (2nd cohort, 1g/day BID)
patients. 2nd group with biopsiable solid-tumours
patients for biomarker and efficacy evaluation.

Objectives
Dose-escalation, multicentre phase I clinical trial
of 2-OHOA in advanced solid tumours including
refractory malignant glioma

Primary objective: To determine safety,

tolerability and recommended phase 2 dose of
of 2-OHOA

Secondary objectives: To determine

pharmacokinetics (PK) and pharmacodynamics
(PD) profile and preliminary anti-tumoral activity

Exploratory objectives:

To evaluate the effect of 2-OHOA on glial fibrillary
acidic protein (GFAP) in glioma pts

To study miRNA as a potential response biomarker
22AUG13 22OCT13 17MAR14 27MAR15 01FEB16
Main results
Part A highlights (completed): Top. Pt 010202. Partial Response (PR) by RANO (tumour shrinkage of 93% on primary lesion lasting for almost 3 years)

7 cohorts completed. 32 pts (28 evaluable) treated Minerval in recurrent GBM patients in MIN-001-1203 PI study
No relevant safety issues (only GI effects 200% Recurrent glioblastoma patients in MIN-001-1203 study (n=13)
Target lesion. Best response 010202* 145,4

(diarrhea, nausea,..) in some pts at the higher doses) 175%

* -> New non-target lesion detected 013006 30,1

% change of tumour volume. Best response Vs baseline assessment

150% # -> Clinical deterioration

MTD established as 12g/day (4g TDS) 125%
A -> previously treated with Avastin 010606* 26,9

030603 26,7

Part B highlights (completed):

100%
013012 6,1
75%

22 pts (18 evaluable) treated with 12g/day. 50%

033007A 6,0
Patients with recurrent glioblastoma treated for
030203 6,0 at least 1 cycle in MIN-001-1203 (n = 13)
No DLTs. No safety/tolerability issues 25%
030301A 5,9 A -> previously treated with Avastin

MTD confirmed as 12g/day (4g TDS) 0%

013001 5,0
-25%

Anti-tumour activity -50%

010103# 4,0

010302# 3,6
Encouraging sings of efficacy, specially in rGBM pts: -75%
030201A 3,1

46 patients evaluable (21 with glioma) -100% -93%

033002# 2,4
-125%
50% (4/8) of rGBM pts treated for 2 cycles had 030201A# 033007A 013001 013012 030203 030301A 010103# 033002# 013006 030603 010302# 010606* 010202*
0 6 20 26 40 60 80
Weeks on treatment
100 120 140 160

objective clinical benefit (6 months) by RANO 13 rGBM patients completed at least 1 cycle of treatment with Minerval. Most pts had received 2 or more previous lines of
Almost all (4/5) of rGBM pts treated for 2 cycles, (experimental) chemotherapy treatment. 4/13 rGBM patients treated had overall clinical benefit. Of the 9 rGBM pts eligible for
not treated before with Avastin, had clinical benefit DLT assessment that did not experienced clinical benefit, at least 5 had previously failed another experimental treatment (4 of them
with bevacizumab) 4/9 rGBM pts not previously treated with bevacizumab had clinical benefit with Minerval
1 rGBM pt PR lasting for almost 3 years 8 rGBM patients completed at least 2 cycles of treatment and had their planned reassessment scan at 6 weeks. 4 of them (50%)

1 oligodendroglioma pt SD for 9 months had objective clinical benefit by RANO. Of the 4 rGBM pts treated with Minerval for at least 2 cycles with no clinical benefit, 3 of
1 rGBM pt SD for 7 months them had previously failed another experimental treatment (2 of them with bevacizumab)
2 rGBM pts SD for 6 months 4/6 rGBM pts treated for 2 or more cycles with Minerval and that were bevacizumab naive had objective clinical benefit

1 mesothelioma pt SD for 10 months

Bottom. PK curves for cohorts #01 - #06 MIN-001-1203. Safety summary
1 colon ADK pt SD for 3 months
1 lung ADK pt SD for 5 months 21 Serious Adverse Events (SAE)
reported in all cases assessed as not
Bottom. Dose escalation process related or unlikely related to the study
7th Cohort: BID drug
16g/day N=7 3 DLT +
1 non-evaluable pt No treatment-related deaths
6th Cohort: TDS Dose limiting toxicities (DLTs) were
12g/day (MTD)
12g/day N=8 1 DLT + Grade 3 diarrhoea (n=3) and vomiting
2 non-evaluable pts
(n=1) despite medical optimization
8g/day 5th Cohort: BID Of the 202 drug-related Adverse Events
N=3 NO DLT (AE) reported in the 54 patients treated in
4th Cohort: BID the study, 97% (195/202) were grade 1-
4g/day N=4 NO DLT 2 and 3% (7/202) were grade 3
1 non-evaluable pt
The majority of the adverse events
3rd Cohort: BID related to the treatment were of
2g/day
N=3 NO DLT Pharmacokinetic profile: gastrointestinal (GI) nature, including all

The PK profile was dose-proportional up to 12g/day. Grade 3 AEs
1g/day 2nd Cohort: BID
Accumulation ratios on Cmax near 1.5 (D21/D1) starting at 1g/day and close to 2 for doses No safety or tolerability issues reported in
N=4 NO DLT greater or equal to 12 g/day. Saturation of absorption starting at 8g/day the safety expansion cohorts: 22

No food interactions were observed patients treated with 12000mg/day,
0.5g/day 1st Cohort: BID
On Day 1, half-life derived from the 8h post-dose concentrations was between 1,4 and 4,6 h at
n=3 NO DLT
several of them treated for >6 months
doses up to 8g/day, increasing to 7,4 hr at 12g/day.
Supported by: