Production of erythropoietin MCQs With Answer

Production of erythropoietin MCQs With Answer

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Introduction

Production of erythropoietin MCQs With Answer is a focused quiz resource designed for M.Pharm students studying Advanced Pharmaceutical Biotechnology. This collection explores the molecular biology, upstream cell-culture strategies, downstream purification, glycosylation requirements, analytical characterization and regulatory considerations specific to recombinant erythropoietin (rEPO) manufacture. Questions probe practical aspects such as host cell selection, expression constructs, fermentation modes, glycoengineering and potency assays, as well as common purification challenges and formulation stability. Intended to deepen conceptual understanding and exam preparation, the MCQs emphasize critical determinants of rEPO quality, activity and safety, preparing students for research or industry roles in biopharmaceutical production.

Q1. Which glycosylation feature of erythropoietin is most critical for its in vivo serum half-life?

  • High-mannose oligosaccharides
  • Oligosaccharide branching without sialic acid
  • Sialylation of terminal glycans
  • Absence of N-linked glycans

Correct Answer: Sialylation of terminal glycans

Q2. Which mammalian cell line is most commonly used for industrial-scale recombinant human erythropoietin production?

  • Escherichia coli
  • Chinese hamster ovary (CHO) cells
  • Saccharomyces cerevisiae
  • Pichia pastoris

Correct Answer: Chinese hamster ovary (CHO) cells

Q3. What is the main reason E. coli is not suitable for producing therapeutic erythropoietin?

  • Low plasmid stability
  • Inability to perform human-like N-linked glycosylation
  • Poor protein folding in the periplasm
  • High endotoxin production only

Correct Answer: Inability to perform human-like N-linked glycosylation

Q4. Which gene-engineering strategy increases secreted rEPO yield from mammalian cells?

  • Removal of native signal peptide without replacement
  • Incorporation of a strong secretion signal peptide (e.g., IgG signal)
  • Use of a bacterial ribosome binding site
  • Deletion of all N-glycosylation sites

Correct Answer: Incorporation of a strong secretion signal peptide (e.g., IgG signal)

Q5. During clone selection for high rEPO producers using CHO-DHFR system, what is the role of methotrexate (MTX)?

  • MTX acts as a glycosylation enhancer
  • MTX selects for clones with amplified DHFR and co-amplified EPO gene copies
  • MTX increases cell growth rate making selection faster
  • MTX stabilizes secreted protein in culture medium

Correct Answer: MTX selects for clones with amplified DHFR and co-amplified EPO gene copies

Q6. Which upstream culture strategy is commonly used to maximize volumetric productivity of rEPO while maintaining quality?

  • Continuous batch culture without feeding
  • Fed-batch with controlled nutrient feeds
  • Static culture in flasks only
  • Repeated freeze-thaw cycles of culture

Correct Answer: Fed-batch with controlled nutrient feeds

Q7. Which parameter manipulation is often used to reduce proteolytic cleavage and improve rEPO stability during culture?

  • Raising culture temperature by 5–8°C
  • Lowering culture temperature mildly (temperature shift)
  • Increasing shear stress by intense agitation
  • Adding high concentrations of proteases

Correct Answer: Lowering culture temperature mildly (temperature shift)

Q8. In downstream processing of rEPO, which chromatography is commonly used as a capture step for secreted glycoproteins lacking Fc regions?

  • Protein A affinity chromatography
  • Ion-exchange chromatography (IEX)
  • Lectin affinity chromatography specific to mannose only
  • Reverse phase chromatography as first step

Correct Answer: Ion-exchange chromatography (IEX)

Q9. Which analytical method is most appropriate for assessing rEPO glycoform heterogeneity and sialylation state?

  • SDS-PAGE without further analysis
  • Mass spectrometry of released glycans and HPLC or CE for sialic acid profiling
  • UV spectrophotometry at 280 nm
  • Simple Bradford protein assay

Correct Answer: Mass spectrometry of released glycans and HPLC or CE for sialic acid profiling

Q10. Darbepoetin alfa differs from recombinant human EPO primarily because it:

  • Is produced in bacteria without glycosylation
  • Has additional N-linked glycosylation sites to extend half-life
  • Lacks any glycosylation to increase potency
  • Is a pegylated version of epoetin alfa

Correct Answer: Has additional N-linked glycosylation sites to extend half-life

Q11. Which potency assay is classically used to measure biological activity of erythropoietin?

  • TF-1 cell proliferation assay or in vivo reticulocyte count assay
  • Hemagglutination assay
  • ELISA measuring total protein only
  • Antimicrobial activity assay

Correct Answer: TF-1 cell proliferation assay or in vivo reticulocyte count assay

Q12. Why is sialic acid removal (desialylation) from rEPO detrimental to its pharmacokinetics?

  • It improves receptor binding but reduces clearance
  • It exposes galactose residues leading to rapid hepatic clearance via asialoglycoprotein receptor
  • It increases half-life by reducing renal filtration
  • It prevents dimerization and aggregation only

Correct Answer: It exposes galactose residues leading to rapid hepatic clearance via asialoglycoprotein receptor

Q13. Which quality attribute is particularly critical to control to avoid immunogenic responses to therapeutic rEPO?

  • Residual host cell DNA amount only
  • Glycosylation profile and aggregation level
  • Color of the final solution
  • Presence of bacterial LPS only

Correct Answer: Glycosylation profile and aggregation level

Q14. During scale-up of rEPO production from shake flasks to bioreactors, which factor must be carefully matched to maintain cell physiology?

  • Plasmid copy number only
  • Oxygen transfer rate (kLa) and shear environment
  • Incubator brand
  • Color of plasticware

Correct Answer: Oxygen transfer rate (kLa) and shear environment

Q15. Which regulatory consideration is specifically important for biosimilar erythropoietin approval compared to small-molecule generics?

  • Exact matching of sequence is unnecessary
  • Comprehensive comparability studies on glycosylation, potency and immunogenicity
  • No clinical data needed if in vitro assays match
  • Only pharmacokinetic studies are required

Correct Answer: Comprehensive comparability studies on glycosylation, potency and immunogenicity

Q16. A common impurity in mammalian cell-derived rEPO preparations that must be reduced is:

  • Plasmid-coded antibiotic molecules
  • Host cell proteins (HCPs) and DNA
  • Peptidoglycan from cell wall fragments
  • Yeast-derived mannans

Correct Answer: Host cell proteins (HCPs) and DNA

Q17. Which feed strategy can maintain high cell density and continuous product removal to potentially improve rEPO productivity?

  • Batch culture with single harvest
  • Perfusion culture with cell retention
  • Static standing culture without mixing
  • Repeated small-volume bolus harvests with no replenishment

Correct Answer: Perfusion culture with cell retention

Q18. For assessing charge variants of erythropoietin resulting from glycan or sialic acid differences, which analytical technique is commonly used?

  • Size-exclusion chromatography only
  • Isoelectric focusing or ion-exchange chromatography
  • Western blot without focusing
  • Simple UV absorbance at 214 nm

Correct Answer: Isoelectric focusing or ion-exchange chromatography

Q19. Codon optimization of the human EPO gene for CHO expression primarily aims to:

  • Alter the amino acid sequence to increase activity
  • Improve translation efficiency by matching host-preferred codons
  • Reduce mRNA stability to lower expression
  • Introduce bacterial promoters for higher expression

Correct Answer: Improve translation efficiency by matching host-preferred codons

Q20. Viral safety in rEPO production is addressed during manufacturing primarily by which approaches?

  • Not applicable since mammalian cells never carry viruses
  • Use of robust donor-screened cell banks, validated viral inactivation/removal steps and testing
  • Only final sterile filtration is sufficient
  • Relying solely on antibiotics in culture medium

Correct Answer: Use of robust donor-screened cell banks, validated viral inactivation/removal steps and testing

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