3D bio printing models of pharmacological research advantages and limitations

1. A SEMINAR ON
REVOLUTION OF 3D ORGAN MODEL IN
PHARMACOLOGICAL RESEARCH
PRESENTED BY:-
SYED DASTAGIR HUSSAIN
Mpharmacy I year I sem
UNDER THE GUIDANCE OF :-
Dr. M.V. KIRAN KUMAR
M.Pharm ., Ph.D., MBA.
HEAD OF DEPARTMENT OF
PHARMACOLOGY
C.M.R. COLLEGE OF PHARMACY
KANDLAKOYA(V), MEDCHAL ROAD, HYDERABAD,TELANGANA 1

2. CONTENT
2
01 Aim and Objective
02 Introduction
2.1 3D printing
2.2 3D Bioprinting
03 Types of 3D bioprinting and approaches
04 Creation of 3D Biostructures
05 Current Research on 3D Bioprinting
06 Concluding Remarks & Future
Perspectives
07 References

3. Aim and Objective:
 3D organ models have gained increasing attention as novel preclinical test systems and
alternatives to animal testing.
 Over the years, many excellent in vitro tissue models have been developed. In parallel,
microfluidic organ-on-a-chip tissue cultures have gained increasing interest for their ability
to house several organ models on a single device and interlink these within a human-like
environment.
 Human disease models have proven valuable for their ability to closely mimic disease
patterns in vitro, permitting the study of pathophysiological features and new treatment
options.
 Although animal studies remain the gold standard for preclinical testing, they have major
drawbacks such as high cost and ongoing controversy over their predictive value for several
human conditions.
3

4. 3D printing
3D printing, also known as additive manufacturing, was developed in the 1980’s as a process
used to make three dimensional objects. Additive manufacturing creates parts from the ground
up by fusing together layers of material. Its counterpart, subtractive manufacturing, begins
with material and removes excess until only the desired shape remains1
Methods
• There are several methods of 3D printing. The most commonly recognized is called Fused
Deposition Modeling (FDM).
• Other 3D printing processes, like Stereolithography (SLA)2
4
Introduction
Fig. No:01 figure of 3D printing process

5. 3D Bioprinting approaches
3D Bioprinting
advances have enabled 3D printing of biocompatible materials, cells and supporting components
into complex 3D functional living tissues. 3D bioprinting is being applied to regenerative
medicine to address the need for tissues and organs suitable for transplantation. Compared with
non-biological printing, 3D bioprinting involves additional complexities, such as the choice of
materials, cell types, growth and differentiation factors, and technical challenges related to the
sensitivities of living cells and the construction of tissues.3,4
Biomimicry:
 Biologically inspired engineering has been applied to many technological problems,
including flight5, materials research6, cell-culture methods6 and nanotechnology7.
 Its application to 3D bioprinting involves the manufacture of identical reproductions of the
cellular and extracellular components of a tissue or organ.8
Autonomous self-assembly:
 It an embryonic organ development as a guide,developing tissue produce their own ECM
components, appropriate cell signaling and autonomous organization and micro-
architecture and function9,10.
 A ‘scaffold-free’ version self-assembling cellular spheroids that undergo fusion and cellular
organization to mimic developing tissues.11
 Autonomous self-assembly relies on the cell as the primary driver of histogenesis, directing
the composition, localization, functional and structural properties of the tissue12.
5

6. 6
Types of 3D bioprinting and approaches
Fig. No:02 figure of Types 3D Bioprinting and approaches

7. Mini-tissues:
 Both of the above strategies for 3D bioprinting. Organs and tissues comprise smaller,
functional building blocks13,14 or mini-tissues.
 It is a smallest structural and functional component of a tissue, such as a kidney nephron.
Mini-tissues can be fabricated and assembled .
 There are two major strategies: first, self-assembling cell spheres (similar to mini-tissues)
are assembled into a macro-tissue using biologically inspired design and organization13,14
 Produce functional tissue units to create ‘organs-on-a-chip’, which are maintained and
connected by a microfluidic network for use in the screening of drugs and vaccines or as in
in vitro models of disease15,16.
Basic Principles:
The bases for reconstruction of tissues and organs through 3D Bioprinting consist of a set of
techniques that transfer biologically active material onto a substrate.17 The basic concept of
3DB allows the building process to be able to create cellular patterns, which are the
systematical organization of cell-to-cell interactions and produce mechanical and chemical
signaling. These patterns confined in a tridimensional structure hope to achieve cellular
functionality
and become a viable tissue or whole organ.17
7

8. Creation of 3D Biostructures:
steps of creating a tridimensional biostructure18
1. Preprocessing
2. Processing
(a) Layer of hydrogel or hydrogel container
(b) Bioink
(c) Hydrogel
(d) Bioink dispensation process
3. Maturation
4. Application 8

9. True 3D Bioprinting Advances:
1)Vascular:
 Bioprinting for organ creation is the establishment of vascular networks that are essential to the
organ, the desired tissue cannot survive. described a method for fabricating .
 3D biostructures with vasculature, multiple types of cells, and ECM, with the use of four
different bioinks in vitro,19
 A layer-by-layer printing technique with the use of multicellular spheroids containing smooth
cells and fibroblasts along withagarose rods, resulting in single- and double-layer small diameter
vascular structures.
Applications
 Meso-rex bypass left portal vein for the treatment of portal vein thrombosis,
 Living donor liver transplantation vasculature for the drainage of segments V and VIII of
the liver to the middle hepatic vein or vena cava to prevent outflow issues in right lobe grafts20
9
Current Research on 3D Bioprinting
Fig. No:03 figure of Biostructure of vascular

10. 2) Liver:
 Drug metabolism has fueled multiple interests around the world to develop liver
biostructures to test the biopharmacology of many drugs.
 Research to develop 3D liver structures will eventually benefit the field of transplantation.
Till a date, only reports of early data in in vitro models are available. Robbins et al.21
 3D filament networks made of carbohydrate glass in a cylindrical shape that were lined
with endothelial cells and perfused with blood.
 This model was tested in a rat hepatocyte model, maintaining the metabolic function of
the cells.22
Applications :
 Hepatotoxic effects of the antibiotics levofloxacin and trovafloxacin.23
10
Fig. No:04 figure of Biostructure of liver

11. 3) Intestine:
 Although GI tract models have been developed with diverse bioengineering techniques.
 3D bioprinting world has not focused on the development of GI tract models. The
complexity of creating an intestine requires the creation of vasculature, neural, and
lymphoid tissue along with epithelial tissue with absorptive and secretory functions.
 At our institution, efforts are being focused on the creation of a muscular graft onto which
all the functions previously mentioned are added.24
11
Fig. No:05 figure of Biostructure of Intestine

12. 4) Kidney:
 Drug nephrotoxicity is estimated to cause 25% of acute renal failure.
 3D-bioprinted kidney models is being fueled by the need to understand better the
interaction between the kidney and multiple drugs.
 Created an in vitro model of multicellular, 3D-bioprinted proximal tubules.25
In these model, the interface between tubular epithelium and renal interstitial cells was
observed within an extracellular matrix and housed in perfusable tissue chips that allowed
the model to survive for more than 2 months.
Applications:
 Their model exhibits enhanced epithelial morphology and functional properties.
Cyclosporine demonstrated disruption of the epithelial barrier in a dose-dependent
manner, proving the utility of the in vitro model.26
12Fig. No:06 figure of bioprinted kidney model

13. 5) Heart:
 The complexity of the heart tissue poses a barrier that only few researchers are willing to
confront. 27 created a hybrid strategy based on 3D bioprinting and scaffolding 28.
 First with the use of bioink containing endothelial cells, they injected microfibrous hydrogel
scaffolds.
 This endothelial layer was then seeded with cardiomyocytes in order to generate aligned
myocardium capable of spontaneous and synchronous contractions.
 Lack of structure and functionality. This is one of the earliest demonstrations of 3D-
bioprinted heart tissue29
13Fig. No:07 figure of Biostructure of cardiomyocytes

14. 6) Neural tissue:
 Engineering nervous system tissues limited work done in the context of bioprinting for
neural tissue fabrication.
 Studied the effect of vascular endothelial growth factor (VEGF) release on proliferation and
migration of murine neural stem cells .30
 Proliferated successfully in contrast to the cells that could not proliferate within the
collagen matrix.31
7) Pancreas tissue :
 Pancreatic β-cells do not easily survive in vitro and only a very few attempts have taken
place to differentiate β-cells from human stem cells.
 Regeneration of pancreas tissue is primarily embodied to the extent that β- cells from
mouse lines or insulinoma cells have been used to fabricate pancreatic islets.32
 Encapsulated human and mouse islets as well as rat insulinoma INS1E β-cells within
hydrogels and bioprinted them in dual layer scaffolds 33.
 The scaffolds were later implanted in diabetic mice and explanted 7 days thereafter.
14

15. 8) Pharmaceutics and high-throughput screening:
 Improving the ability to predict the efficacy and toxicity of drug candidates earlier in the
drug discovery process will speed up the translation of new drugs into clinics.
 Recent attempts in 3D in vitro assay systems is an ideal way to resolve this bottleneck
because 3D tissue models can closely mimic the native tissue and have the capability to be
used in high-throughput .
 Among various methods for engineering 3D in vitro systems34, bioprinting has superiorities
such as controllability on size and microarchitecture, high-throughput capability, co culture
ability and low-risk of cross-contamination.
15
 Bioprinted tissue and organ models have been increasingly considered for the potential of
pharmaceutics use such as drug toxicology and high throughput screening.35
Fig. No:08 figure of extracellular matrix and perfusable tissue chips

16.  Organ models hold the potential to revolutionize preclinical research, although there
is still a long road ahead and the predictivity of most organ models has yet to be
proven.
 Pharmaceutical companies and regulatory authorities have recognized the importance
of this technology and are highly engaged with them.
 Scientists must learn from previous mistakes and combine efforts to improve
preclinical-to-clinical translation.
 In this regard political and financial support is pivotal. Furthermore, joint efforts of
expert groups in academia, pharmaceutical industry, and regulatory authorities are
now needed to approach and overcome current bottlenecks.
 Accordingly, governments initiatives are required to promote not only the
development of organ (disease) models but also their implementation in preclinical
drug testing, otherwise the full potential of organ models may never be realized.
16
Concluding Remarks & Future Perspectives:

17. References:
1. Nakamura, M., Iwanaga, S., Henmi, C., Arai, K. & Nishiyama, Y. Biomatrices and
biomaterials for future developments of bioprinting and biofabrication. Biofabrication 2,
014110 (2010).
2. Mironov, V. et al. Organ printing: tissue spheroids as building blocks. Biomaterials
30,2164–2174 (2014)
3. Elsevier; mini-tissue image is reprinted from Norotte, C. et al. Scaffold-free vascular tissue
engineering using bioprinting. Biomaterials 30, 5910–5917 (2009)
4. Zopf, D.A., Hollister, S.J., Nelson, M.E., Ohye, R.G. & Green, G.E. Bioresorbable airwa
splint created with a three-dimensional printer. N. Engl. J. Med. 368, 2043–2045 (2013).
5. Michelson, R.C. Novel approaches to miniature flight platforms. Proc. Inst. Mech. Eng.
Part G J. Aerosp. Eng. 218, 363–373 (2004).
6. Reed, E.J., Klumb, L., Koobatian, M. & Viney, C. Biomimicry as a route to new materials:
what kinds of lessons are useful? Philos Trans A Math Phys. Eng. Sci. 367, 1571–1585
(2009).
7. Huh, D., Torisawa, Y.S., Hamilton, G.A., Kim, H.J. & Ingber, D.E. Microengineered
physiological biomimicry: organs-on-chips. Lab Chip 12, 2156–2164 (2012).
8. Ingber, D.E. et al. Tissue engineering and developmental biology: going biomimetic. Tissue
Eng. 12, 3265–3283 (2006).
9. Marga, F., Neagu, A., Kosztin, I. & Forgacs, G. Developmental biology and tissue
engineering. Birth Defects Res. C Embryo Today 81, 320–328 (2007).
10. Steer, D.L. & Nigam, S.K. Developmental approaches to kidney tissue engineering. Am. J.
Physiol. Renal Physiol. 286, F1–F7 (2004).
17

18. 11. Derby, B. Printing and prototyping of tissues and scaffolds. Science 338, 921–926 (2012).
12. Kasza, K.E. et al. The cell as a material. Curr. Opin. Cell Biol. 19, 101–107 (2007).
13. Mironov, V. et al. Organ printing: tissue spheroids as building blocks. Biomaterials 30,
2164–2174 (2009).
14. Kelm, J.M. et al. A novel concept for scaffold-free vessel tissue engineering: self-assembly
of microtissue building blocks. J. Biotechnol. 148, 46–55 (2010).
15. Huh, D. et al. Reconstituting organ-level lung functions on a chip. Science 328, 1662–1668
(2010).
16. Gunther, A. et al. A microfluidic platform for probing small artery structure and function.
Lab Chip 10, 2341–2349 (2010).
17. Mironov, V. (2003). Printing technology to produce living tissue. Expert Opinion on
Biological Therapy, 3, 701–704.
18. Catros, S., Fricain, J. C., Guillotin, B., Pippenger, B., Bareille, R., Remy, M., Lebraud, E.,
Desbat, B., Amedee, J., & Guillemot, F. (2011). Laser-assisted bioprinting for creating on-
demand patterns of human osteoprogenitor cells and nano-hydroxyapatite. Biofabrication,
3, 025001.
19. Kolesky, D. B., Truby, R. L., Gladman, A. S., Busbee, T. A., Homan, K. A., & Lewis, J. A.
(2014). 3D bioprinting of vascularized, heterogeneous cell-laden tissue constructs.
Advanced Materials, 26, 3124–3130.
20. Norotte, C., Marga, F. S., Niklason, L. E., & Forgacs, G. (2009). Scaffold-free vascular
tissue engineering using bioprinting. Biomaterials, 30, 5910–5917.
21. Robbins, J. B., Gorgen, V., Min, P., Shepherd, B. R., & Presnell, S. C. (2013). A novel
in vitro three-dimensional bioprinted liver tissue system for drug development. FASEB
Journal,
18

19. 22. Miller, J. S., Stevens, K. R., Yang, M. T., Baker, B. M., Nguyen, D. H., Cohen, D. M., Toro,
E., Chen, A. A., Galie, P. A., Yu, X., Chaturvedi, R., Bhatia, S. N., & Chen, C. S. (2012).
Rapid casting of patterned vascular networks for perfusable engineered three-dimensional
tissues. Nature Materials, 11, 768–774.
23. Ma, X., X. Qu, et al., Deterministically patterned biomimetic human iPSC-derived hepatic
model via rapid 3D bioprinting. Proc Natl Acad Sci U S A, 2016. 113: 22062211.
24. Wengerter, B. C., Emre, G., Park, J. Y., & Geibel, J. (2016). Three-dimensional printing in
the intestine. Clinical Gastroenterology and Hepatology, 14, 1081–1085.
25. King, S., Creasey, O., Presnell, S., & Nguyen, D. (2015). Design and characterization of a
multicellular, three-dimensional (3D) tissue model of the human kidney proximal tubule. The
FASEB Journal, 29(1 Supplement), LB426.
26. Homan, K. A., Kolesky, D. B., Skylar-Scott, M. A., Herrmann, J., Obuobi, H., Moisan, A., &
Lewis, J. A. (2016). Bioprinting of 3D convoluted renal proximal tubules on Perfusable
chips. Scientific Reports, 6, 34845.
27. Zhang, X. Y., & Zhang, Y. D. (2015). Tissue engineering applications of three-dimensional
Bioprinting. Cell Biochemistry and Biophysics, 72, 777–782.
28. Organovo. 2015. The bioprinting process.
29. Zhang, Y. S., Arneri, A., Bersini, S., Shin, S. R., Zhu, K., Goli-Malekabadi, Z., Aleman, J.,
Colosi, C., Busignani, F., Dell'erba, V., Bishop, C., Shupe, T., Demarchi, D., Moretti, M.,
Rasponi, M., Dokmeci, M. R., Atala, A., & Khademhosseini, A. (2016). Bioprinting 3D
microfibrous scaffolds for engineering endothelialized myocardium and heart-on-a-chip.
Biomaterials, 110, 45–59. 19

20. 30. Lee, Y-B. et al. (2010) Bio-printing of collagen and VEGF-releasing fibrin gel scaffolds for
neural stem cell culture. Exp. Neurol. 223, 645–652.
31. Hsieh, F-Y. et al (2015) 3D bioprinting of neural stem cell-laden thermoresponsive
biodegradable polyurethane hydrogel and potential in central nervous system repair.
Biomaterials 71, 48–57.
32. Pagliuca, F.W. et al. (2015) Generation of functional human pancreatic β cells in vitro. Cell
159,428–439.
33. Marchioli, G. (2015) Fabrication of three-dimensional bioplotted hydrogel scaffolds for
islets ofLangerhans transplantation. Biofabrication 7, 25009.
34. Xu, F. et al. (2011) Microengineering methods for cell-based microarrays and high-
throughput drug-screening applications. Biofabrication 3, 34101.
35. Ozbolat, I.T. and Hospodiuk, M. (2016) Current advances and future perspectives in
extrusionbased bioprinting. Biomaterials 76, 321–343
20

21. THANK YOU
21