1.
Understanding Cancer Drug Resistance by Developing and Studying Resistant Cell Line Models
Xavier CP, Pesic M, Vasconcelos MH.
Curr Cancer Drug Targets. 2016;16(3):
© 2016 Bentham Science Publishers
Chemotherapy Resistance Mechanisms and Requirement of Drug Resistant Cell Line Models
Presenter: Bhuvanesh Sukhlal Kalal,
PhD Scholar, Department of Biochemistry,
Yenepoya Medical College, Yenepoya University,
Mangaluru, India
Registration No. ACBICON/2016/549
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

2. INTRODUCTION
The development of chemotherapy resistance continues to be the main problem in the treatment of cancer patients
Key drug resistance mechanisms- intrinsic resistance and acquired resistance; Depends on- type of cancer and on the mode of action of the drug
Drug resistant cancer cell lines are useful tools for understanding the mechanisms of cancer drug resistance.
This review briefly discusses
The mechanisms of cancer drug resistance
Findings from different types of established cancer cell lines used
Validation methods in drug resistant cell lines to confirm the resistance mechanism
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

3. MOLECULAR MECHANISMS OF DRUG RESISTANCE
Causes:
Tumor heterogenicity,
Host factors (such as
pharmacokinetics of the drug, alterations in drug metabolism, drug absorption and intracellular drug transport) or
Tumor cell factors
Fig. (1). Drug resistance mechanisms developed by cancer cell lines. Cancer cell lines frequently acquire mechanisms of resistance during drug treatment, such as overexpression of drug efflux proteins, alteration in the drug target, acquired adaptive responses or deregulation of apoptosis. “Next-generation” drugs or combination of drugs may overcome such drug resistance.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

4. Table 1. Examples of mechanisms of resistance to anticancer drugs and possible alternative/additional treatments.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

5. Table 1. Examples of mechanisms of resistance to anticancer drugs and possible alternative/additional treatments. (conc.)
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

6. OTHER MECHANISMS OF RESISTANCE TO ANTICANCER DRUGS
Drug metabolism: second line of cellular defense
converted into metabolites that can be conjugated by enzymes such as Glutathione-S-Transferase
Drug can be trapped into subcellular organelles such as lysosomes and endosomes and then be expelled from the cell
Overexpression of one or more drug efflux pumps:
ABC) transporter family (best studied MDR1 or ABCB1; MRP1 or ABCC1; BCRP or ABCG2)
Difference in phases of the cell cycle:
In a mature solid tumor majority of quiescent cells (in the G0/G1) and a minority of proliferating cancer cells (in the S/G2/M)
Increased DNA damage repair and tolerance to DNA lesions
Deregulation of apoptosis: ex. Overexpression of the antiapoptotic protein.
Epithelial to mesenchymal transition.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

7. MODELS USED TO STUDYMECHANISMS OF DRUG RESISTANCE IN CANCER
Fig. (2). Experimental models of drug resistance. The different models that enable the study of cancer drug resistance include cancer cell lines, mouse models and human tumor samples. All of these models require to be analyzed before, during and after drug treatments. The in vitro studies engage different approaches, such as the establishment of isogenic pairs of resistant and sensitive cell lines, applied mutagenesis in sensitive cells, and gain or loss of function screens in sensitive or resistant cell lines. In vivo studies are performed using either xenografts or genetically engineered mice (GEM). In addition, patient tumor samples could be used together with samples from healthy donors.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

8. DEVELOPMENT OF DRUG RESITANT CANCER CELL LINES
Starting dose- Use IC20 or IC50 or, even a non-toxic concentration, to generating a resistant phenotype.
Continuous increase the concentration of the drug over time; maintain the cells at a final drug concentration.
Time- can take 3-10 months, depends on type cell line and drug target/s.
Fold resistance is usually calculated as the ratio between the IC50 of the resistant cell line and the IC50 of the corresponding parental cell line.
Fold resistance of ≥ 2.0 can be regarded as resistant.
Important issue: Stability (resistant phenotype may be reversible).
If drug resistant cells grow after drug withdrawal for 1-3 months and restored drug sensitivity to a level observed in parental cells, suggests- acquired resistance not arise due to mutations, since genetic alterations are irreversible events.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

9. ANALYSIS AND VALIDATION OF MECHANISM OF DRUG RESISTANCE
Drug resistant cancer cells
Drug sensitive cancer cells
Genetic mutation analysis- Sequencing the drug target gene or just the kinase domain of the gene
Introducing a mutation or alteration in the genes expected to be involved in the drug resistance mechanisms, using site-direct mutagenesis
Protein analysis-
Study the alternate signaling pathways or networks associated with drug resistance by Western blot, proteomic etc.
Silencing the gene/s thought to be responsible for the drug resistance by RNA interference (RNAi)
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

10. CONCLUSION
Development of drug resistant cancer cell lines- time-consuming process and the identification of the mechanism/s of resistance is a difficult.
Missed out factors- tumor microenvironment and tumor stroma interactions.
Useful tool for identifying drug resistance mechanisms and related targets, as well as to develop new approaches for overcoming drug resistance.
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)

11. Thank you all for patience and attention…
Acknowledgment
Organizer -ACBICON 2016
Bentham Science Publishers- for granting right to reproduce the copyrighted material
Dr. Vinitha R. Pai (Research guide)
Thank you all for patience and attention…
43rd Annual Conference of Association of Clinical Biochemists of India (ACBICON 2016)