1. Pharmacology of Antineoplastic Agents
Outline of Lecture Topics:
Background
Antineoplastic Agents: classification
a. Cell Cycle Specific (CCS) agents
b. Cell Cycle Non-Specific (CCNS) agents
c. Miscellaneous (e.g., antibodies) agents
Mechanisms of action
Side Effects
Drug Resistance
Overall, I have divided this lecture into two major parts. First, some general info. In the second part, we I will talk about cellular and molecular mechanisms of action of a few select
Antineoplastic compounds. Most important part of this lecture is the mechanism of action and side effects.
Dr.Rajarshi Patel, Ph.D.
Dept Chemistry

2. PART I Background Antineoplastic Agents
a. Cell Cycle Specific (CCS) agents
b. Cell Cycle Non-Specific (CCNS) agents
c. Miscellaneous (e.g., antibodies) agents
Before going into mechanisms of action, we will look at some basics.

3. Cancer
Definition:
Cancer* is a term used for diseases in which abnormal cells divide without control and are able to invade other tissues. Cancer cells can spread to other parts of the body through the blood and lymph systems, this process is called metastasis.
Categorized based on the functions/locations of the cells from which they originate:
Carcinoma - skin or in tissues that line or cover internal organs. E.g., Epithelial cells % reported cancer cases are carcinomas.
Sarcoma - bone, cartilage, fat, muscle, blood vessels, or other connective or supportive tissue.
Leukemia - White blood cells and their precursor cells such as the bone marrow cells, causes large numbers of abnormal blood cells to be produced and enter the blood.
Lymphoma - cells of the immune system that affects lymphatic system.
Myeloma - B-cells that produce antibodies- spreads through lymphatic system.
Central nervous system cancers - cancers that begin in the tissues of the brain and spinal cord.
In the next slide, we will look at the old and new ways to treat cancer.

4. Surgery Radiation Chemotherapy
Cancer Therapeutic Modalities (classical)
Surgery
Radiation
Chemotherapy
1/3 of patients without metastasis
Respond to surgery and radiation.
If diagnosed at early stage,
close to 50% cancer
could be cured.
50% patients will undergo chemotherapy,
to remove micrometastasis. However,
chemotherapy is able to cure only about 10-15%
of all cancer patients.
1. 30% of patients respond to surgery and radiation. 2. If diagnosed early, 50% cancer could be cured. 3. Although, 50% of the patients undergo chemotherapy, only 10-15% patients respond to chemotherapy.
In the next slide, we will look at the background of cancer chemotherapy.

5. New types of cancer treatment
Hormonal Treatments: These drugs are designed to prevent cancer cell growth by preventing the cells from receiving signals necessary for their continued growth and division. E.g., Breast cancer – tamoxifen after surgery and radiation
Specific Inhibitors: Drugs targeting specific proteins and processes that are limited primarily to cancer cells or that are much more prevalent in cancer cells.
Antibodies: The antibodies used in the treatment of cancer have been manufactured for use as drugs. E.g., Herceptin, avastin
Biological Response Modifiers: The use of naturally occuring, normal proteins to stimulate the body's own defenses against cancer. E.g., Abciximab, rituxmab
Vaccines: Stimulate the body's defenses against cancer. Vaccines usually contain proteins found on or produced by cancer cells. By administering these proteins, the treatment aims to increase the response of the body against the cancer cells.
This slide is for your information only. We are not going to cover these agents here. But I thought, you should be aware of the new developments.

6. Cancer Chemotherapy (Background)
Most of the recent progress using antineoplastic therapy is based on:
Development of new combination therapy of using existing drugs.
Better understanding of the mechanisms of antitumor activity.
Development of chemotherpeutic approaches to destroying micrometastases
Understanding the molecular mechanisms concerning the initiation of tumor growth and metastasis.
Recognition of the heterogeneity of tumors
B. Recently developed principles which have helped guide the treatment of neoplastic disease
A single clonogenic cell can produce enough progeny to kill the host.
2. Unless few malignant cells are present, host immune mechanisms do not play a significant role in therapy of neoplastic disease.
3. A given therapy results in destruction of a constant percentage as opposed to a constant number of cells, therefore, cell kill follows first order kinetics.

7. Cancer Chemotherapy
C. Malignancies which respond favorably to chemotherapy:
choriocarcinoma,
Acute leukemia,
Hodgkin's disease,
Burkitt's lymphoma,
Wilms' tumor,
Testicular carcinoma,
Ewing's sarcoma,
Retinoblastoma in children,
Diffuse histiocytic lymphoma and
Rhabdomyosarcoma.
D. Antineoplastic drugs are most effective against rapidly dividing tumor cells.

8. LD50 ----- Therapeutic Index = ED50
E. The Main Goal of Antineoplastic Agents
IS to eliminate the cancer cells without affecting normal tissues (the concept of differential sensitivity).  In reality, all cytotoxic drugs affect normal tissues as well as malignancies - aim for a favorable therapeutic index (aka therapeutic ratio).
Therapeutic Index =
LD50
-----
ED50
A therapeutic index is the lethal dose of a drug for 50% of the population (LD50) divided by the minimum effective dose for 50% of the population (ED50).

9. Infrequent scheduling of
treatment courses.
Prolongs survival but does not cure.
More intensive and
frequent treatment.
Kill rate > growth rate.
Untreated patients
F. The effects of tumor burden, scheduling, dosing, and initiation/duration of treatment on patient survival.
Early surgical removal of the primary tumor decreases the tumor burden. Chemotherapy will remove persistant secondary tumors.

10. General rules of chemotherapy
Combination of several drugs with different mechanisms of action, different resistance mechanisms, different dose-limiting toxicities.
Adjuvant therapy: Additional cancer treatment given after the primary treatment to lower the risk that the cancer will come back. Adjuvant therapy may include chemotherapy, radiation therapy, hormone therapy, targeted therapy, or biological therapy.
Neoadjuvant therapy: Treatment given as a first step to shrink a tumor before the main treatment, which is usually surgery, is given. Examples of neoadjuvant therapy include chemotherapy, radiation therapy, and hormone therapy. It is a type of induction therapy.

11. Antineoplastic Agents
Alkylating agents
Topoisomerase inhibitors
Antimetabolites
Molecularly targeted
busulfan
dactinomycin
cytarabine
erlotinib
carboplatin
daunomycin
clofarabine
imatinib
carmustine
doxorubicin
fludarabine
sorafenib
cisplatin
etoposide
gemcitabine
sunitinib
cyclophosphamide
etoposide phosphate
mercaptopurine
tretinoin
dacarbazine
idarubicin
methotrexate
Herceptin
ifosfamide
irinotecan
nelarabine
Miscellaneous
lomustine
liposomal daunomycin
thioguanine
arsenic trioxide
mechlorethamine
liposomal doxorubicin
Tubulin binders
asparaginase
melphalan
mitoxantrone
docetaxel
bleomycin
oxaliplatin
teniposide
ixabepilone
dexamethasone
procarbazine
topotecan
vinblastine
hydroxyurea
temozolomide
vincristine
mitotane
thiotepa
vinorelbine
PEG-asparaginase
paclitaxel
prednisone
Here is a list of antineoplastic agents. By no means it is complete list. In the next slide, we will look at how these compounds may inside tumor cells.

12. Chemotherapy: classification based on the
mechanism of action
Antimetabolites: Drugs that interfere with the formation of key biomolecules including nucleotides, the building blocks of DNA.
Genotoxic Drugs: Drugs that alkylate or intercalate the DNA causing the loss of its function.
Plant-derived inhibitors of mitosis: These agents prevent proper cell division by interfering with the cytoskeletal components that enable the cell to divide.
Other Chemotherapy Agents: These agents inhibit cell division by mechanisms that are not covered in the categories listed above.
All these agents could be either cell cycle specific or cell cycle non specific anti-neoplastic agents.

13. In next slides we are going to look at mechanism of action of alkylating agents.

14. Cell cycle specificity of Anti-Neoplastic Agents
Vincristine,
Vinblastine
Paclitaxel, Docetaxel
Cyclophosphamide
Bleomycin
Actinomycin D M G
resting G G
Hydrocortisone 2 1
G0 = resting phase
G1 = pre-replicative phase
G2 = post-replicative phase
S = DNA synthesis
M = mitosis or cell division
In next slides we are going to look at mechanism of action of alkylating agents. S
Actinomycin D
Purine antagonists
5-Fluorouracil
Methotrexate
Cytosine arabinoside
Cyclophosphamide
Methotrexate
5-Fluorouracil
6-Mercaptopurine
Cytosine arabinoside
6-Thioguanine
Daunomycin

15. PART II Mechanisms of action Side Effects Drug Resistance
Pharmacology of Antineoplastic Agents
PART II
Mechanisms of action
Side Effects
Drug Resistance
In next slides we are going to look at mechanism of action of alkylating agents.

16. Chemotherapy: Mechanisms of Action1
Asparaginase
Tubulin binders
Alkylating agents
Topoisomerase Inh.
Antimetabolites
DNA
Purines and
Pyrimidines
RNA
In next slides we are going to look at mechanisms of action of alkylating agents.
Protein
tubulin

17. Major Clinically Useful Alkylating Agents
Cancer Chemotherapy
Chapter 55. B.G. Katzung
Bis(mechloroethyl)amines
Nitrosoureas
Aziridines

18. An Example of DNA Crosslinking
Crosslinking: Joining two or more molecules by a covalent bond. This can either occur in the same strand (intrastrand crosslink) or in the opposite strands of the DNA (interstrand crosslink). Crosslinks also occur between DNA and protein. DNA replication is blocked by crosslinks, which causes replication arrest and cell death if the crosslink is not repaired.

19. Alkylating Agents (Covalent DNA binding drugs)
The first class of chemotherapy agents used.
They stop tumour growth by cross-linking guanine nucleobases in DNA double-helix strands - directly attacking DNA.
This makes the strands unable to uncoil and separate.
As this is necessary in DNA replication, the cells can no longer divide.
Cell-cycle nonspecific effect
Alkylating agents are also mutagenic and carcinogenic

20. E.g., Mechlorethamine (Nitrogen Mustards)
Cancer Chemotherapy
Dr.Rajarshi N. Patel

21. Cyclophosphamide
Cyclophosphamide is an alkylating agent. It is a widely used as a DNA crosslinking and cytotoxic chemotherapeutic agent.
It is given orally as well as intravenously with efficacy.
It is inactive in parent form, and must be activated to cytotoxic form by liver CYT450 liver microsomaal system to 4-Hydroxycyclophamide and Aldophosphamide.
4-Hydroxycyclophamide and Aldophosphamide are delivered to the dividing normal and tumor cells.
Aldophosphamide is converted into acrolein and phosphoramide mustard.
They crosslink DNAs resulting in inhibition of DNA synthesis .

22. Cyclophosphamide Metabolism
Inactive
Cyclophosphamide is a widely used alkylating agent. It is given orally as well as intravenously. It is inactive in parent form, and must be activated to cytotoxic form by liver CYT Hydroxycyclophamide and Aldophosphamide are delivered to the normal as well as tumor cells. Aldophosphamide is converted into acrolein and phosphoramide mustard. They crosslink DNAs resulting in inhibition of DNA synthesis.

23. Clinical Applications: Breast Cancer Ovarian Cancer
Cyclophosphamide
Clinical Applications:
Breast Cancer
Ovarian Cancer
Non-Hodgkin’s Lymphoma
Chronic Lymphocytic Leukemia (CLL)
Soft tissue sarcoma
Neuroblastoma
Wilms’ tumor
Rhabdomyosarcoma
Cyclophosphamide is a widely used alkylating agent. It is given orally as well as intravenously. It is inactive in parent form, and must be activated to cytotoxic form by liver CYT Hydroxycyclophamide and Aldophosphamide are delivered to the normal as well as tumor cells. Aldophosphamide is converted into acrolein and phosphoramide mustard. They crosslink DNAs resulting in inhibition of DNA synthesis.

24. Depression of blood cell counts Bleeding Alopecia (hair loss)
Cyclophosphamide
Major Side effects
Nausea and vomiting
Decrease in PBL count
Depression of blood cell counts
Bleeding
Alopecia (hair loss)
Skin pigmentation
Pulmonary fibrosis
Nausea and vomiting, decrease in PBL count, BM depression, bleeding, alopecia, skin pigmentation, pulmonary fibrosis

25. Ifosphamide Mechanisms of Action Similar to cyclophosphamide
Application
Germ cell cancer,
Cervical carcinoma,
Lung cancer
Hodgkins and non-Hodgkins lymphoma
Sarcomas
Major Side Effects
Nausea and vomiting, decrease in PBL count, BM depression, bleeding, alopecia, skin pigmentation, pulmonary fibrosis

26. A. Alkylating agents 1. Mechanism of Action 2. Clinical application
3. Route
4. Side effects
a. Nitrogen Mustards
A. Mechlorethamine
DNA cross-links, resulting in inhibition of DNA synthesis and function
Hodgkin’s and non-Hodgkin’s lymphoma
Must be given Orally
Nausea and vomiting, decrease in
PBL count, BM depression, bleeding, alopecia, skin pigmentation, pulmonary fibrosis
B. Cyclophosphamide
Same as above
Breast, ovarian, CLL, soft tissue sarcoma, WT, neuroblastoma
Orally and I.V.
C. Chlorambucil
Chronic lymphocytic leukemia
Orally effective
D. Melphalan
Multiple myeloma, breast, ovarian
E. Ifosfamide
Germ cell cancer, cervical carcinoma, lung, Hodgkins and non-Hodgkins lymphoma, sarcomas
Orally effective

27. A. Alkylating agents 1. Mechanism of Action 2. Clinical application
3. Route
4. Side effects
b. Alkyl Sulfonates
A. Busulfan
Atypical alkylating agent.
Chronic granulocytic leukemia
Orally effective
Bone marrow depression, pulmonary fibrosis, and hyperuricemia
c. Nitrosoureas
1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
A. Carmustine
DNA damage, it can
cross blood-brain barrier
Hodgkins and non-Hodgkins lymphoma, brain tumors, G.I. carcinoma
Given I.V. must be given slowly.
Bone marrow depression,
CNS depression, renal toxicity
B. Lomustine
Lomustine alkylates and crosslinks DNA, thereby inhibiting DNA and RNA synthesis. Also carbamoylates DNA and proteins, resulting in inhibition of DNA and RNA synthesis and disruption of RNA processing. Lomustine is lipophilic and crosses the blood-brain barrier
Hodgkins and non-Hodgkins lymphoma, malignant melanoma and epidermoid carcinoma of lung
Orally effective
Nausea and vomiting, Nephrotoxicity, nerve dysfunction
C. Streptozotocin
DNA damage
pancreatic cancer
Given I.V.
Nausea and vomiting, nephrotoxicity, liver toxicity

28. A. Alkylating agents d. Ethylenimines 1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
A. Triethylene thiophosphoramide (Thio-TEPA)
DNA damage, Cytochrome
P450
Bladder cancer
Given I.V.
Nausea and vomiting, fatigue
B. Hexamethylmelamine
(HMM)
DNA damage
Advanced ovarian tumor
Given orally after food
Nausea and vomiting, low blood counts, diarrhea
d. Triazenes
1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
A. Dacarbazine (DTIC)
Blocks, DNA, RNA and protein synthesis
Malignant Melanoma, Hodgkins and non-Hodgkins lymphoma
Given I.V.
Bone marrow depression, hepatotoxicity, neurotoxicity, bleeding, bruising, blood clots, sore mouths.

29. Summary

30. Dihydrofolate reductase (DHFR);
C. Antimetabolites
Folic acid is a growth factor that provides single carbons to the precursors used to form the nucleotides used in the synthesis of DNA and RNA. To function as a cofactor folate must be reduced by DHFR to THF.
Folic acid
Tetrahydrofolate (THF)
Methyl group and related carbon provider
Dihydrofolate reductase (DHFR);
Key factor for the synthesis of DNA & RNA
Methotrexate (MTX) is a folic acid analog that binds with active site of DHFR, interfering with synthesis of tetrahydrofolate (THF), which serves as the key one-carbon carrier for enzymatic processes for involved in de novo synthesis of thymidylate, purine nucleotides, and amino acid serine and methionine.

31. inhibits formation of FH4 (tetrahydrofolate) from folic
C. Antimetabolites
1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
1. Methotrexate
inhibits formation of FH4 (tetrahydrofolate) from folic
acid by inhibiting the enzyme dihydrofolate reductase (DHFR); since FH4 transfers
methyl groups essential to DNA synthesis and hence DNA synthesis blocked.
Choriocarcinoma, acute lymphoblastic leukemia (children), osteogenic sarcoma, Burkitt's and other non-Hodgkin‘s lymphomas, cancer of breast, ovary, bladder, head & neck
Orally effective as well as given I.V.
bone marrow depression, intestinal lesions and interference with embryogenesis.
Drug interaction: aspirin and sulfonamides displace methotrexate
from plasma proteins.
Methotrexate (MTX) is a folic acid analog that binds with active site of DHFR, interfering with synthesis of tetrahydrofolate (THF), which serves as the key one-carbon carrier for enzymatic processes for involved in de novo synthesis of thymidylate, purine nucleotides, and amino acid serine and methionine.

32. 1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
2 Pyrimidine Analogs: Cytosine Arabinoside
inhibits DNA synthesis
most effective agent for induction of remission in acute myelocytic
leukemia; also used for induction of remission acute lymphoblastic leukemia,
non-Hodgkin's lymphomas; usually used in combination chemotherapy
Orally effective
bone marrow depression
1. Mechanism of Action
2. Clinical application
3. Route
4. Side effects
2 Purine analogs:
6-Mercaptopurine (6-MP) and Thioguanine
Blocks DNA synthesis by inhibiting conversion of
IMP to AMPS and to XMP as well as blocking conversion of AMP to
ADP; also blocks first step in purine synthesis.
Feedback inhibition
blocks DNA synthesis by inhibiting conversion of IMP to
XMP as well as GMP to GDP; also blocks first step in purine synthesis by
feedback inhibition
most effective agent for induction of remission in acute myelocytic
leukemia; also used for induction of remission acute lymphoblastic leukemia,
non-Hodgkin's lymphomas; usually used in combination chemotherapy
Orally effective
bone marrow depression,

33. 6. Drug Resistance
One of the fundamental issue in cancer chemotherapy is the development of cellular drug resistance. It means, tumor cells are no longer respond to chemotherapeutic agents. For example, melanoma, renal cell cancer,
brain cancer often become resistant to chemo.
A few known reasons:
Mutation in p53 tumor suppressor gene occurs in 50% of all tumors. This leads to resistance to radiation therapy and wide range of chemotherapy.
Defects or loss in mismatch repair (MMR) enzyme family. E.g., colon cancer no longer respond to fluoropyrimidines, the thiopurines, and cisplatins.
Increased expression of multidrug resistance MDR1 gene which encodes P-glycoprotein resulting in enhanced drug efflux and reduced intracellular accumulation. Drugs such as athracyclines, vinca alkaloids, taxanes, campothecins, even antibody such as imatinib.

34. Summary
The main goal of anti-neoplastic drug is to eliminate the cancer cells without affecting normal tissues.
Log-Kill Hypothesis states that a given therapy kills a percentage of cells, rather then a constant number, therefore, it follows first order kinetics. Aim for a favorable therapeutic index.
Early diagnosis is the key.
Combination therapy and adjuvant chemotherapy are effective for small tumor burden.
Two major classes of antineoplastic agents are:
a. Cell Cycle Specific and
b. Cell Cycle Non-Specific agents
Because chemotherapeutic agents target not only tumor cells, but also affect normal dividing cells including bone marrow, hematopoietic, and GI epithelium. Know what the side effects are.
Drug resistance is often associated with loss of p53 function, DNA mismatch repair system, and increased MDR1 gene expression.
Surgery then chemotherapy is adjuvant