biopharmaceuticals - current fda & emas regulations on ... · galactosylation, sialylation...
TRANSCRIPT
![Page 1: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/1.jpg)
©2013 Waters Corporation 1
Jayesh Kattla, PhD March 2015
Gothenburg & Copenhagen
Biopharmaceuticals - Current FDA & EMAs Regulations on glycan analysis
![Page 2: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/2.jpg)
©2013 Waters Corporation 2
Hope and Risk
![Page 3: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/3.jpg)
©2013 Waters Corporation 3
4,288
6,340
13,600
24,000
45,000 50,000
E. coli Yeast Fruit fly Human Rice Corn
Num
ber
of G
enes
The total number of genes does not define complexity
![Page 4: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/4.jpg)
©2013 Waters Corporation 4
Post translational modifications explains the complexitity of protein function
![Page 5: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/5.jpg)
©2013 Waters Corporation 5
Glycan Biosynthesis is a complex process
Complext process No genetic code for glycosylation
Glycan sysntiesis occurs in ER and Golgi Main types are N linked and O Linked
glycans All proteins need not go thru the whole
pathway makes the glycans highly Heterogeneous
Macro heterogeniety Micro heterogeniety Size of the glycans can vary Glycan are involved in protein fuction in
terms of binding, safety, Efficacy, immune response, half life,
![Page 6: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/6.jpg)
©2013 Waters Corporation 6
EPO glycosylation
![Page 7: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/7.jpg)
©2013 Waters Corporation 7
Glycan micro heterogeneity affects protein function
Asn Desialylation of IVIg abrogates anti-inflammatory properties in K/N mice Kaneko et al (2006). Science; 313(5787): 670-673
Involved with placental transport of IgG IgG galactosylation increased in pregnant women Kibe et al (1996). J Clin Biochem Nutr; 21(1): 57-63
Asn
IgG G0 interacts with MBL to activate complement Malhotra et al (1995). Nat Medicine; 1(3): 237-243
Asn
Loss of core α(1,6) fucose on IgG results in enhanced ADCC activity Okazaki et al (2004). J Mol Biol; 336(5): 1239-1249
Asn
Sialylation increases the half life of protein
Highly mannosylated proteins can be cleared out the system within minutes by mannose binding lectin
![Page 8: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/8.jpg)
©2013 Waters Corporation 8
Glycosylation Functions: Risks and Regulatory Concerns
Structure based on PDB file 1H3Y from Krapp S et al. J Mol Biol, 2003, 325(5): 979-989
Mediates biological activity
Glycans impact safety and efficacy
– Correct and consistent structure of the glycans
– Obtaining the desired medical effect
– Avoid adverse immunological reaction
Alteration in glycans may eliminate or alter
activity
Immune response triggered by unrecognized
glycans
Consistent glycan distribution indicates
process stability
![Page 9: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/9.jpg)
©2013 Waters Corporation 9
Biopharmaceuticals
$163 billion dollar market 20% are biopharmaceuticals 8% Annual Growth 50% of top drugs are
biomolecules Was 28% in 2008 Biopharmaceuticals are more
complex in structures
![Page 10: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/10.jpg)
©2013 Waters Corporation 10
The Biologics Market
Blood Proteins
Hormones
Growth Factors
Cytokines
Vaccines
mabs
Bone Proteins
Blood Proteins
Hormones
Growth Factors
Cytokines
Vaccines
Monoclonal Antibodies
Bone Proteins
Other
190 EMA/FDA Approved
Walsh (2010). Nature Biotech; 28(9):917-924
![Page 11: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/11.jpg)
©2013 Waters Corporation 11
The Biologics Market
Blood Proteins
Hormones
Growth Factors
Cytokines
Vaccines
mabs
Bone Proteins
Blood Proteins
Hormones
Growth Factors
Cytokines
Vaccines
Monoclonal Antibodies
Bone Proteins
Other
127 are Glycoproteins (>66%)
Walsh (2010). Nature Biotech; 28(9):917-924
![Page 12: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/12.jpg)
©2013 Waters Corporation 12
Biopharma Attempts to Replicate Human Glycosylation
Struwe WB, Cosgrave EFJ, and Rudd PM. (2011). Glycoproteomics in Health and Disease. Functional and Structural Proteomics of Glycoproteins.
![Page 13: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/13.jpg)
©2013 Waters Corporation 13
First Plant made antibody
![Page 14: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/14.jpg)
©2013 Waters Corporation 14
Negative Attributes to Mammalian Cell Culture Glycosylation
Asn
Asn
Asn
Asn Presence of gal-α(1,3)-gal can induce anaphylaxis Chung et al (2006). N Engl J Med; 358(11): 1109-1117
50% of non-allergic blood donors contain antibodies against β(1,2)-xylose and α(1,3)-core fucose Bardor et al (1995). Glycobiology; 13(6): 427-434
N-glycolylneuraminic acid is an oncofetal antigen in humans Muchmore et al (1989). J Biol Chem; 264(34): 20216-20223
![Page 15: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/15.jpg)
©2013 Waters Corporation 15
Genetically engineered expression system /QBD
The concept promotes industry's understanding of the product and manufacturing process starting with product development, basically building quality in, not testing it. Under the concept of QbD, when designing and developing a product, a company needs to define desired product performance and identify CQAs.
![Page 16: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/16.jpg)
©2013 Waters Corporation 16
Bioprocessing Conditions Can Influence Glycosylation
Cell Line Critical in affecting
glycosylation (Raju et al 2000)
Manufacturing Mode Perfusion increases sialylation
over fed-batch (Lipscomb et al 2005)
Dissolved O2 Variable effect, cell line specific
(Restelli et al 2006)
Temperature Low temps (30°C) decrease sialylation
(Trummer et al 2006)
Ammonia High concns affect
terminal glycosylation (Yang and Butler 2000, 2002)
pH Galactosylation, sialylation microheterogeneity vary
(Muthing et al 2003)
![Page 17: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/17.jpg)
©2013 Waters Corporation 17
Analysis of critical quality attributes during the manufacturing process
Process analytical technology (PAT) has been defined by the United States Food and Drug Administration (FDA) as a mechanism to design, analyze, and control pharmaceutical manufacturing processes through the measurement of Critical Process Parameters (CPP) which affect Critical Quality Attributes (CQA).
It is absolutely essential to tightly control the levels of glycan critical quality attributes during the manufacturing process. Eg your Desired glycoforms that is involved in activitating ADCC Maintain the levels of immunogenenic epitopes Mannose containing glycans and so on
![Page 18: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/18.jpg)
©2013 Waters Corporation 18
Changes in Bioprocess Can Affect Glycosylation
![Page 19: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/19.jpg)
©2013 Waters Corporation 19
Regulatory Agencies Are Demanding More Detailed Glycan Analysis
![Page 20: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/20.jpg)
©2013 Waters Corporation 20
FDA Drug application pathway
IND BLA Developmental discussion
![Page 21: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/21.jpg)
©2013 Waters Corporation 21 keith webber, FDA
![Page 22: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/22.jpg)
©2013 Waters Corporation 22 keith webber, FDA
![Page 23: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/23.jpg)
©2013 Waters Corporation 23 keith webber, FDA
![Page 24: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/24.jpg)
©2013 Waters Corporation 24 keith webber, FDA
![Page 25: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/25.jpg)
©2013 Waters Corporation 25 keith webber, FDA
![Page 26: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/26.jpg)
©2013 Waters Corporation 26 keith webber, FDA
![Page 27: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/27.jpg)
©2013 Waters Corporation 27 keith webber, FDA
![Page 28: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/28.jpg)
©2013 Waters Corporation 28
Micro & Macro Heterogeniety
keith webber, FDA
![Page 29: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/29.jpg)
©2013 Waters Corporation 29 keith webber, FDA
![Page 30: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/30.jpg)
©2013 Waters Corporation 30 keith webber, FDA
![Page 31: Biopharmaceuticals - Current FDA & EMAs Regulations on ... · Galactosylation, sialylation microheterogeneity vary (Muthing et al 2003) ©2013 Waters Corporation 17 Analysis of critical](https://reader034.vdocument.in/reader034/viewer/2022050716/5e41eae4305b7c446f466738/html5/thumbnails/31.jpg)
©2013 Waters Corporation 31
Summary
Biopharmaceuticals are complex molecules Glycan synthesis is a complex process Glycosylation plays a critical role in protein function Glycan complexity increases due to its micro and macro
heterogeneity Glycans are in involved in safety and efficacy of the protein Company are genetically engg several expression systems to
produce proteins with more human like glycosylation Glycosylation is an important aspect of QbD Glycosylation important in PAT Companies are required to do indepth Glycosylation analysis
before regulatory submission