The NICB (National Institute for Cellular Biotechnology) is located on the Dublin City University (DCU) campus in Dublin, Ireland. It is a leading multidisciplinary centre of translational research in fundamental and applied cellular biotechnology, molecular cell Biology, ocular diseases and biological chemistry. It includes a multidisciplinary team of Cell and Molecular Biologists, Biotechnologists, Chemists and Informatics specialists. The NICB prioritises translational research involving collaborations with industry and with clinicians, and is committed to educating people from all backgrounds in the area of Biomedical Science. This slideshare summarises the main research areas of the NICB, including: Molecular basis for biopharmaceutical production by animal cells Cancer – drug resistance, invasion and biomarkers Tissue Engineering/Stem Cell Therapy – ocular diseases, diabetes Using animal cells as research tools and models for disease research

1. N a t i o n a l I n s t i t u t e F o r C e l l u l a r B i o t e c h n o l o g y
Research Overview

2. NICB – Dublin City University
• Government-designated National Centre of Expertise in Basic
and Applied Molecular Cell Biotechnology since 1987
• New 3,200m2 building (opened 2006)

3. Main Areas of Research
• Molecular basis for biopharmaceutical production by animal
cells
• Tissue Engineering/Stem Cell Therapy – ocular diseases,
diabetes
• Cells as research tools and models for disease research
• Cancer – drug resistance, invasion and biomarkers

4. Cells as products
Stem cell therapy/ Tissue engineering

5. Tissue Engineering/
Stem Cell Therapy
• Twin focus – basic research and clinical application
• Corneal stem cell transplant – entering clinical phase
• Pancreatic islet transplant – currently technology
transfer stage (Partner: University of Oxford)

6. • The Corneal surface consists of an
epithelial layer that, in the healthy eye,
is constantly renewed.
• Located at the limbus a narrow ring of
stem cells surrounding the cornea.
Stem Cell Treatments For
Eye Injuries 1

7. Stem Cell Treatments For
Eye Injuries 2
• Patients with limbal deficiencies are unable to maintain a stable
cornea. e.g. Chemical and thermal burns, Stevens-Johnson syndrome
• Cornea transplantation in these patients typically fail due to
conjunctival invasion, vascularization, and persistent epithelial defect.
• The NICB in collaboration with its partners, proposes to introduce a
stem cell therapy for these patients.

8. • Isolate and culture stem cells from the limbal ring.
These cultured cells are then transplanted on to the
surface of the injured eye, repopulating the
damaged cornea
• Research to refine techniques, find better
differentiation markers and investigate biology of
the process, e.g. using microarrays and proteomics
Stem Cell Treatments For Eye
Injuries - 3

9. Diabetes Research
at the NICB

10. 5-10% of these have severe
recurring hypoglycemia
23,000 adults in Ireland
suffer from Type 1 Diabetes
(T1DM)

11. Significant sustained decrease in
severe hypoglycaemic events and
improved quality of life.
Islet
transplantation

12. Islets from the Donor’s pancreas are
infused into the recipient’s hepatic
portal vein and produce insulin

13. Islet
Transplantation
Islet Isolation &
Purification
Donated
Pancreas

14. DRIVE – Horizon 2020
funded project
Diabetes
Reversing
Implants with enhanced
Viability and long-term
Efficacy

15. Mitochondrial
Function
Oroboros oxygraph
Measures oxygen consumption
in cells and tissues in real time

16. Cells as factories
Manufacture of recombinant
proteins

17. Research Related to Biopharmaceutical
Production
Our goal is to work closely with biopharmaceutical companies to improve basic
understanding of the molecular basis for recombinant protein production by
mammalian cells (including CHO and hybridomas) and
• to translate this understanding into improved biopharmaceutical
production processes
• Special focus on miRNA

18. Why do we need improved biopharmaceutical
production processes?
• Increasing number of products
• Current high unit cost
• Reduced production costs will contribute to
(a) Access for patients to treatments they need
(b) Long-term health of the biopharmaceutical industry

19. Invited talks

20. Cells as research tools
& as models for
disease research

21. Cell culture
Access to living human material
- primary cultures
- cell lines

22. Cell culture systems may not always
completely reflect complex multi-cell type
and spatial organisation in the body

23. C o r e t e c h n o l o g i e s :
M o l e c u l a r p r of i l i n g of c e l l u l a r r e s p o n s e
e . g . t o d r u g s ; t o t e m p e r a t u r e c h a n g e : t o o x i d a t i v e s t r e s s
• mRNA (Affymetrix)
• miRNA (ABI)
• Proteins
• Importance of bioinformatics analysis

24. Proteomics
• Proteomics Infrastructure at the NICB
• 2D DIGE (Difference Gel Electrophoresis)
• Mass Spectrometry for Protein
Identification/Characterisation, Including
phosphoproteomics
• Quantitative LC-MS approaches (i.e. gel-free)

25. Mass Spectrometry Suite
• LTQ Orbitrap XL (Thermo Fisher Scientific)
High mass accuracy
Interfaced with Dionex Ultimate 3000
1D and 2D LC capabilities
• LTQ XL with ETD
Electron Transfer Dissociation (ETD)
PTM analysis (e.g. phosphorylation)
• MALDI TOF-TOF 4800 (Applied Biosystems)
High throughput MALDI MS/MS
LC-MALDI

26. Using proteomic profiling to identify
new cancer markers in blood

27. Cancer Biomarkers
for What?
• Early detection
• Monitoring of tumour burden (response to
therapy)
• Detection of relapse
• Predicting response to specific therapy

28. Advanced Lung
Cancer
Haptoglobin ELISA
0
1000
2000
3000
4000
5000
6000
ug/mL
Control
Squamous
Adenocarcinoma
Small cell carcinoma

29. Ultimate aims of our
Cancer Research Programmes
Paired diagnostic and therapeutic approaches to
treatment of individual cancer patients

30. Toxicity Testing
Drug Resistance in Cancer
Mechanisms of Cancer Cell Invasion

31. CHEMOTHERAPY WORKS,
BUT ONLY SOMETIMES:
WHY?
One reason is drug resistance
(acquired or intrinsic)

32. Adriamycin Distribution in Resistant Cancer Cells
DLKP-A resistant cancer cells
after exposure to Adriamycin.
Fluorescent view
DLKP sensitive cancer cells after
exposure to Adriamycin
Fluorescent view

33. Immunodetection of MRP-1
MRP-1 Negative Breast Carcinoma MRP-1 Positive Breast Carcinoma

34. Cancer invasion & Metastasis
• Cell models, including clonal variants
• In vitro properties and investigation of relevance in human cancer
• Molecular profiling
• New antigens/monoclonal antibodies

35. Therapeutic Targeting Using ADCs

36. Identification and Investigation of novel
membrane protein targets with high cancer
specific expression for potential therapeutic
targeting using Antibody Drug Conjugates (ADCs)

37. Colon Cancer
The 9E1 target antigen is highly
expressed in colon cancer and show
limited expression in normal colon

38. Pancreatic Cancer
Novel membrane target is highly
expressed in Pancreatic Cancer

39. Breast Cancer
Novel Membrane Target is highly
expressed in triple negative breast
cancer (TNBC) and shows very
limited expression in normal breast

40. Development of functional anti-invasive
MAbs to identify potential drug targets
associated with cancer invasion/metastasis.

41. MAb 7B7 targeting the Ku70/80 heterodimer
significantly blocks the invasion of
MiaPaCa-2 pancreatic cancer cells

42. Effect of LY6G6F siRNA on 2D colony formation
of Mia Paca-2 cells
Cell number seeded:
100 200 400
Control
Negative
siRNA
LY6G6F
siRNA #2

43. Uveal Melanoma

44. Proteomic Analysis of uveal melanoma
to understand metastatic disease

45. Enucleation & Plaque
Radiotherapy
Treatments for
Primary Tumor

46. metastasis in
50% of cases
Primarily to the liver with
average survival of
5-8 months

47. Loss of chromosome 3
Class I v Class 2 gene signature
Loss of BAP1
GNAQ/GNA11 mutation
Current
Biomarkers

48. Aims of t h e St u dy :
Iden tif y differen tially expressed protein s in primar y
u veal melan oma t issu es of pat ien t s wh o developed
met ast at ic disease ver su s t h ose wh o did n ot .
Un der st an d t h e biology of t h e disease
Iden tif y n ew th erapeu tic targets

49. Cancer Biotherapeutics

50. Background
• NSABP-B31 (Paik et al. 2007) and N9831 (Perez et al.
2010)
• Could a strong immune response to trastuzumab in the
adjuvant setting be responsible for the response in HER2
low patients?
• Trastuzumab has two mechanisms of action 1) it inhibits
HER2 signaling and 2) engages the immune system
through ADCC.

51. Antibody-Dependent Cell-
mediated Cytotoxicity
ADCC

52. NK cells and peripheral blood
mononuclear cells (PBMCs)
Immune cells from
healthy Volunteers

53. Immunoblotting and high
content analysis.
HER2 Levels
quantified

54. Examine the effects of TKIs on
HER2 expression in a HER2-
positive cell line (SKBR3)
Laser scanning
Confocal
Microscopy

55. Determining the effect of PD-1/PD-L1
inhibition on response to
trastuzumab in preclinical HER2-
expressing breast cancer models.

56. Assessing the expression of
immunomodulatory proteins in
HER2-targeted therapy-resistant
tumour cells.

57. Pancreatic Cancer Programme

58. • St Lukes Radiation Oncology
Network, Rathgar
• University of Buffalo
Collaboration

59. Generation of primary pancreatic
cancer and stromal cell lines

60. With pancreatic cancer
cells and how they impact
treatment success
Stromal Cancer
Cell interactions

61. L e v e l o f c o l o ny fo r m a t i o n i n Pa n c - 1 c e l l s p o s t c o - c u l t u r e
w i t h p a n c re a t i c p a t i e n t - d e r i v e d f i b ro b l a st s .

62. Medicinal Inorganic Chemistry

63. artificial
metallonuceases
• First reported “self-active” oxidative system
capable of inducing single-stranded DNA scission
in the absence of exogenous reductant or oxidant.
• Copper based complex, di-nuclear structure.
• Displays excellent in vitro chemotherapeutic
activity toward cisplatin-resistant ovarian cancer.

64. • Detection and quantification DNA
oxidation by synthetic artificial
metallonucleases using capillary
electrophoresis.
• Analysis completed using Agilent
Bioanalyzer 2100 located within the
molecular biological laboratory at NICB.
Chemical nuclease detection
using microfluidics

65. • Potent non-covalent DNA binding agents where nucleic
acid recognition is achieved through use of the
“phosphate clamp”.
• Phosphate clamp-DNA interactions result in
condensation of superhelical and B-DNA.
• Triplatin-DNA binding inhibits endonuclease activity by
type II restriction enzymes.
• High chemotherapeutic potential for human cancer.
Novel chemotherapeutic
platinum(II) complexes

66. • Detection of apoptosis induced mitochondrial
depolarization through exposure to ROS active
copper developmental therapeutics.
• Flow cytometry and confocal imaging analyses.
• Broad spectrum of activity identified using the
National Cancer Institute (NCI) 60-human cancer
cell panel.
• Unique mechanism compared to marketed
therapeutics.
Copper chemotherapeutic
drug development

67. • Individual morphine molecules and derivatives
thereof lack nucleic acid recognition properties.
• In the triplet drug form unique properties
emerge with this alkaloid substructure interacting
with dsDNA and condensing superhelical DNA.
Tripodal opioids as
unique DNA interacting
agents

68. Phenazine based
biomaterials
• Enhanced high affinity DNA binding
• Distinctive nucleotide binding
specificity.
• High intercalative capacity
• In vitro chemotherapeutic potential

69. • Metal catalyzed reactive oxygen species (ROS)
in biological systems can cause a wide variety of
pathological conditions including cancer.
• The extent of DNA damage owing to these
radicals can be quantified through 8-oxo-2’-
deoxyguanosine (8-oxo-dG) lesion detection
using both ELISA or LC-MS/MS analysis.
Detection of free radical
DNA damage

70. Metallodrug
topoisomerase
inhibitors
• Development of intercalating metal complexes to
unwind and relax negatively supercoiled DNA.
• Applications as unique topoisomerase inhibitors.

71. Collaboration with Industry
• Cells as factories
• Cells as products
• Cells as Research Tools
• Cells for toxicity assay and bioassay
• Making monoclonal antibodies
• Whole genome mRNA and miRNA profiling
• Proteomic profiling
• Pharmacokinetics/drug analysis

72. Some past & present industry collaborators

73. Questions?
N a t i o n a l I n s t i t u t e f o r C e l l u l a r
B i o t e c h n o l o g y
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