Cultured Autologous Oral Mucosal Epithelial Cell-Sheet for Corneal Epithelial Reconstruction (CAOMECS)
One-year follow-up for 17 patients/25 included
ASCRS, Boston, april 2010
C.Burillon, O.Damour
Tissus et Cells HCL Bank
The authors have no financial interest in the subject matter of this poster
Ophthalmology department
Limbal epithelial deficiency (LSCD)
Population Consequences
Thermal or chemical Burns
Stevens Johnson and Lyell Syndrom
Ocular pemphigoid
Aniridia
Severe dry eye
Limbal stem-cell deficiency after cornea transplantations
Other bilateral disorders of the ocular surface
Ulcers
pain
Dryness
Neovascularisation
Conjunctivalisation
Symblepharon
Total bilateral LSCD gives rise to blindness
What do we have to do in LSCD?
To Bring an Epithelium
Autologous limbus graftCultured Autologous
limbal EpitheliumOral Mucosa Graft
For unilateral deficiencyAutograft from controlateral
safe eyeBrings stem cells to the
corneal epithelium
1-3 mm² Biopsy
For unilateral deficiency
For bilateral deficiencyOral mucosa very closed to cornea epithelium +++
PurposeHypothesis of CAOMECS use:
1 – To restore the ocular surface
2 – To obtain sufficient Cornea transparency when stroma is healthy.
3 – To allow secondary donor cornea graft when stroma is opaque.
Materials and methodsWe use a specific support for the culture: an UpCell ®
Insert which is a thermosensitive polymer
- The temperature responsive polymer (PIPAAm) is immobilized by covalent binding on the surface.
- The surface change between hydrophobic and hydrophilic is temperature-dependent and reversible.
- At the temperature 20~25 , cells can't keep attachment because the surface turned to ℃hydrophilic to liberate hydrophobic interaction.
- The surface antigen and extra cellular matrix (ECM) of harvested cells are intact, because trypsin or dispase is not necessary to harvest the cells.
37°C 20°C
UpCell Basal membrane
hydrophobic hydrophilic
Biopsy and Culture using UpCell Insert ®
UpCell®-Insert
Biopsy
Culture on Upcell Insert at 37°C: 3 weeks
Isolation
NIH/3T3 in storage
Temperature reduced to 20ºC
PVDF doughnut ring as a white ring
After three weeks, we obtain in 100% cases: • Robust, viable, multilayered epithelial cell sheets, • Transparent to read a letter Arial size 10.• Holoclones is garant of their functionality. • Keratin 3, p63 and b1 integrin and Laminin 5 expression on the sheet
Final Product Validation
Cytokeratin 3
Laminin 5 Integrin beta-1
p63
EfficiencyCFE > 2.1 ± 0.9%
Qualitykeratin 3/76, p63, β1 integrin
Microbiological SafetyOn biopsy transport medium On the culture medium4 days before harvest the sheet Graft day: on the medium just before harvest the sheetTransport medium
Clinical Trial Hospices Civils de Lyon, France prospective, open, non comparative and monocentric study
Stage I/II study by Gehan study design
Results: safety and efficacy at 12 monthsGrade changes improvement in blue worsening in red
# Primary endpoint Secondary endpoint
1
Corneal burn / 4 keratoplasties cataract surgery, Trabeculectomy + Mitomycine
No Ulcer anymore No more vessels
Comfort (dryness or watering, pain and photophobia)
2Neuroparalytic keratitis 1 lamellar keratoplasty
PEK (2-level improvement)No vessels anymore(3 to 0)
visual acuity, Comfort (pain, photophobia, dryness watering)
3Rosacea keratitis / no surgical treatment
No Ulcer anymore,vascularisationRecurrence of few localized vessels
Visual acuity Comfort (dryness or watering and pain)
4 Corneal burn /1 amniotic membrane
Blood vessels’ activitiesRecurrence Localized conjunctivalisation since at 6 months, due to surgery or contact lens
Comfort (dryness or watering) Visual acuity
5 Lyell syndrome /1 lamellar keratoplasty
SAECorneal Perforation at 6 month, but not related with CAOMECS
6Corneal burn / 1 lamellar keratoplasty, 1 amniotic membrane
No more PEKCorneal vascularisation (uncountable to 1), blood vessels activity
Comfort (dryness and pain) 1 level decrease of visualAcuity
7Corneal burn /1 keratoplasty1 amniotic membrance
No more UlcerNo more PEKCorneal vascularisation (5 to 1)
Comfort (Photophobia, dryness, pain)
Follow up of safety and efficacy at 12 monthsGrade changes improvement in blue worsening in red
# Primary endpoint Secondary endpoint
8Trachoma / Several corneal grafts
No Ulcer anymore neither PEKNo vascularisation anymore(20 to 0)
Visual acuityComfort (no dryness Photophobia,)
9 Lyell syndrome / 1 lamellar keratoplasty
No more ulcerPEKCorneal vascularisation (20 to 2)
Visual acuityComfort (no dryness, no photophobia)
11Corneal burn /1 keratoplasty
No Ulcer anymore PEKNo vascularisation anymore(12 to 0)
Visual acuityBut Corneal opacification Comfort (no photophobia, no dryness)
12
Corneal burn / 1 amniotic membrane, 1 lamellar keratoplasty
No Ulcer anymoreRecurrence Vessel (firstly decrease 20 to 0, but shown recurrence to 5 at 2-month and 7 at 12 month post-graftRecurrence PEK (firstly decreased grade to 0, but shown recurrence to + since 3 –month post-graft)
Comfort (no photophobia and no pain)but Comfort (dryness was appeared after 1 month.)
Patient No 13 could not come for the follow-up assessments since 2-month time.
Follow up of safety and efficacy at 12 months Grade changes improvement in blue worsening in red
# Primary endpoint Secondary endpoint
10
Rosacea keratitis/3 keratoplasties, Cataract surgery
No Ulcer anymore PEK Corneal vascularisation (15 to 3), vessels activity
visual acuity, Comfort (no photophobia, no dryness)
14
Corneal burn/1 Palpebral reconstruction1 amniotic membrance
KPS de ++ to 0Recurrence of
neovasculatisation at 8months
visual acuity, Comfort (no pain)
15Aphakia post-congenital cataract + corneal anoxia induce by contact lens/no surgical treatment
PEK Corneal vascularisation(40 to 2), Blood vessels activity
visual acuity, Comfort (no pain, photophobia, no dryness)
16
Corneal burn/1 keratoplasty, 1 amniotic membrane
No more Ulcer PEK Corneal vascularisation (30 to 3
visual acuity, Comfort (no pain, no dryness, photophobia)
17Lyell syndrome/1 amniotic membrane
PEK Corneal vascularisation (30 to 8) , vessels activity
Comfort (dryness +++ to +)
Without fluorescein with fluorescein
Inclusion One year follow up Inclusion One year follow up
Some examples:Primary criteria : Ocular Surface observed by Slit Lamp Examination
After corneal graft
Conclusions• Significant regenerated epithelium on the ocular surface:
– new good functional epithelium– less vascularisation and vessels activities
• When stroma and endothelium are healthy, visual acuity can be improved
• When stroma and endothelium are damaged, visual acuity is difficult to be improved with only CAOMECS treatment but it allows a conventional donor graft without rejections and visual acuity is improved (5 cases up to day)
Histology of corneaAfter CAOMECS + corneal graft
Aknowlegments
GCS/CTC tissus and cells bankOdile DamourChantal HéloireEric VenetPascale PascalCéline Auxenfans
Laboratoire ImmunocytologySerge Nataf
CellSeedYukio HasegawaNao NakamuraAyumi MurayamaVirginie Justin
OphthalmologyCarole Burillon
Preliminary Human StudyNishida et al , N. Engl.J.Med, 2004, 351,1187-1196