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									www.valentiabiopharma.com
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Valentia Biopharma - In vivo Drug Discovery


      Research and development of new drugs for human
                         diseases


  Currently focused on finding lead
  compounds to achieve an effective
  treatment on Myotonic Dystrophy
  Type 1 (DM1) disease
Valentia Biopharma - In vivo Drug Discovery


     Drosophila melanogaster, a new technology for Drug
                        Discovery


Valentia Biopharma R&D



Generation of disease models               • Genetic diseases
Generation of genetic/biochemical models   • Other disease
                                             mechanisms



In vivo High Throughput Screening          • Phenotypic assays
                                           • Biochemical assays
Why Drosophila?

                 75% of human disease causing genes are conserved in
                 Drosophila

                 Short time frame from drug dosing to results

                 No ethical issues.

                 Large number of genetic tools available

                 Low variability at low costs
                 (Flies are CHEAP)


Not only Drosophila:
Drug delivery can be problematic and not all human systems can be easily modeled. (pulmonary
disease…)
So we always validate any positive result with mouse models and human cell lines
Drosophila models. Screening experimental approaches


(1) Phenotypic assay (viability)

                                                                   n=191
                                                        200
                             Viability model            160
                                                        120
                           Toxic in mushroom bodies      80                n=67
                                (neuronal cells)         40
                                                          0
                                                                  Control DM1 fly




(2) Biochemical assay (gene-reporter) by fusing known human
    genes involved in DM1 pathologies to a reporter
                  PROMOTER-CONSTRUCT                                TRANSGENIC FLY



                    Gene         Reporter

                                                          FLY
                                                      INJECTION
In vivo automated HTS platform


Our high-throughput screen (HTS) platform allows testing
thousands of compounds per week with the advantage of working
in in vivo conditions.



TOXICITY AND ACTIVITY EVALUATION
   IN THE SAME EXPERIMENTAL
           APPROACH
In vivo automated HTS platform                              Screening platform - Steps in red are automated




     1. FLY CROSSES                                                                5. READING
                             3. SEEDING           4. HOMOGENIZATION            (Envision Reader
     2. DRUG PLATES
                             (Sorter Cytometry)   (Robot with stackers)               /Scanner)
        PREPARATION
     (Robot with stackers)
                                                                                 6. ANALYSIS




     F0:Adults                         F1:Embryo/Larvae                        F1:Adults

 By genetic modification we have been able to develop transgenic flies models that reproduces some
 aspects of the genetic human disease Myotonic Dystrophy. This has been the first model we developed in
 and now it is being used with our High Throughput Screening (HTS) technology. Our HTS platform allows
 the testing of thousands of compounds per week, with the differential advantage of working with in vivo
 models. Large scale in vivo compound testing on Drosophila provides important early information on
 multiple key parameters of drug discovery
DM1 Drug Discovery Pipeline
                                                     > 15,000 small molecules

                                             screening
                                                                  >8,000
                                            screening analyisis
                                               and validation
                                                                        30 candidates
                                                     Secondary assays
                                                        validation
                                                                           4 hits

                  Research                                                    Development
                   Exploratory         Hit to lead       Optimization           Pre-clinic          Clinic

 *VLT001
 (ABP1)

 VLT002

 VLT003

 VLT004

 VLT005
*ABP1 was discovered by an academic group performing manual screening in the DM1 Drosophila model
The disease: Myotonic Dystrophy Type 1 (Steiner disease)

Rare Disease. Overall worldwide prevalence: 1 / 8,000. Higher in
some populations like in Quebec (Canada).


High penetrance

Multifactorial disease. Mainly a muscular disorder:
Myotonia, progressive muscular wasting and weakness.


But also cataracts, hypogonadism, ECG changes,
infertility, cognitive dysfunction, mental retardation…



Source:http://omim.org/entry/160900


Harper 2001
DM1 DNA mutation: repeat expansion
                          (A)                                         (B)                 (C)

                            Size of CUG repeat
             DM1 Phenotype        50 - 400
“Premutation” Asymptomatic          38 - 49

                      Normal         5 - 37


CAP                                  (CUG)n
      5’                                         3’ (AAA)
                                                  n
  5’ UTR DMPK Gene (Coding region) 3’ UTR


 (A) In DM1, the repeat involved is a CTG tract located in the 3´UTR region of the DMPK gene. In normal
 population we have two alleles between 5-37 CTG repeats. In patients the length of one allele is
 expanded from more than 50 repeats up to even thousands of repeats. (B) Mutant transcripts form
 stable CUG hairpins that avoid their normal transportation to cytoplasm, with the ability of forming
 nuclear aggregates and sequester RNA-binding factors such as Muscleblind-like-1 protein (MBLN1),
 which plays an important role in alternative splicing and gene expression regulation. (C) Thus, in DM1
 patients several aberrant splicing events in many genes (spliceopathy) have been characterized, most of
 them dependent of MBLN1. A few of these aberrant splicing event have already been linked to distinct
 DM1 clinical symptoms.
VLT001 (ABP1) active compound

           In vivo discovery of a peptide that prevents CUG–RNA hairpin formation
                   and reverses RNA toxicity in myotonic dystrophy models
          Amparo García-López, Beatriz Llamusía Mar Orzáez, Enrique Pérez-Payá, and Ruben D. Artero
                                 PNAS U S A. 2011 Jul 19;108(29):11866-71.


         Summary Results:

         1. Orally administered ABP1 increased adult viability and reversed muscle
         degeneration phenotypes in DM model flies in a dose-dependent manner.

         2. Aberrant ribonuclear CUG foci diminished and Muscleblind (the
         Drosophila functional homolog of human MBNL1) misdistribution in CUG foci
         was improved in model flies taking ABP1 orally.

         3. Intramuscular administration of ABP1 suppressed muscle histopathology
         signs and reversed missplicing events in DM model mice up to one month
         after administration. Expression of Clcn1, which is low in DM1 model mice,
         recovered in ABP1 injected muscle.

         4. Binding and destabilizing CUG repeat RNA hairpins in vitro suggest a
         mechanism of action of ABP1 located very upstream of the disease pathway
         suggesting the molecule should be therapeutically active against many of the
         clinical signs.
  ABP1 suggested mechanism of action
   MBNL1




                        CUG
                       G    C                                CUG
                        U U                                 G    C
                         CG                                  U U
                                                              CG
       MBNL1             GC
                       U U
                         CG
                                                              GC
                                                             U U
                         GC                                   CG
    sequestration      U U                                    GC
                         CG
                         GC              ABP1                U U
                                                             G C
                       U U
                         CG                               C       G
                         GC
                       U U
                         CG
                                                         U           U              MBNL1
                         GC                             G           C
                       U U
                         CG                                C       G               released
                         GC                                  U U
                       U U                                    CG
   Splicing              CG
                         GC
                       U U
                                                              GC
                                                             U U
                         CG                                   CG
misregulation            GC
                       U U
                                                              GC
                                                             U U
                                                              CG
                         CG
                         GC                                   GC
                       U U                                   U U
                         CG                                   CG
                         GC                                   GC
                       U U                                   U U
                         CG
                         GC
                                                              CG
                                                              GC                 Normal Splicing
                       UC GU                                 U U
                         GC                                C     G
                       U U                              G          C
                         CG                            U             U
                                   A)n          CUGCUGC                GCUGCUG
                    DMPK 3’ UTR
                                                        DMPK 3’ UTR


      ABP1 binds to CUG repeat RNA and induce a switch to a single-stranded conformation,
      releasing MBLN1 and decreasing CUG toxicity.
Business case for DM

   Among rare diseases, Myotonic Dystrophy presents an attractive
            commercial opportunity on multiple fronts:

 Clinical state (urgent unmet medical need, lack of suitable
  treatments)

 Scientific rationale (solid understanding of disease pathology,
  druggable targets, and proof of concept for therapeutic
  intervention leading to clinically meaningful benefit in animal
  models)

 Logistics/marketing considerations (significant disease
  prevalence/potential market size, accessibility of patients, well-
  organized global research community)



Source: 2010 Marigold therapeutic strategies for myotonic dystrophy

								
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