EMATOLOG
`Basic Principles and Practice
`
`Edited by
`Ronald Hoffman, M.D.
`Bruce Kenneth Wiseman Professor of Medicine, Professor of Pathology, and
`Chief, Hematology/Oncology Section, Department of Medicine,
`Indiana University School of Medicine,
`Indianapolis, Indiana
`
`Edward J. Benz, Jr., M.D.
`
`Chief, Section of Hematology, Associate Chainnan for Academic Affairs,
`Department of Internal Medicine, Professor, Departments of Internal Medicine
`and Human Genetics, Yale University School of Medicine;
`Attending Physician, Yale-New Haven Hospital,
`New Haven, Connecticut
`
`Sanford J. Shattil, M.D .
`
`Professor, Departments of Medicine and Pathology and Laboratory Medicine,
`University of Pennsylvania School of Medicine;
`Chief, Hematology-Oncology Section, Hospital of the University of Pennsylvania,
`Philadelphia, Pennsylvania
`
`Bruce Furie, M.D.
`Professor, Departments of Medicine and Biochemistry,
`Tufts University School of Medicine;
`Chief, Division of Hematology-Oncology, Department of Medicine,
`New England Medical Center Hospital,
`Boston, Massachusetts
`
`Harvey J. Cohen, M.D., Ph.D.
`
`Professor of Pediatrics, and Chief, Di vision of Pediatric Hematology /Oncology
`Department of Pediatrics and the Cancer Center,
`'
`University of Rochester School of Medicine and Dentistry,
`Rochester, New York
`
`(cid:141)-(cid:141) •••
`IIIIIUIII _,.
`Churchill Livingstone
`New York, Edinburgh, London Melbourn T k
`e, o yo
`'
`
`NOVARTIS EXHIBIT 2096
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`

`Library of Congress Cataloging-in-Publication Data
`
`Hematology : basic principles and practice / edited by Ronald Hoffman
`... [et al.].
`p. cm.
`Includes bibliographical references and index.
`ISBN 0-443-08643-5
`1. Hematology. I. Hoffman, Ronald, date.
`[DNLM: 1. Hematologic Diseases--diagnosis. 2. Hematologic
`Diseases-therapy. WH 100 H48745]
`RC633.H434 1991
`616. l '5--dc20
`DNLM/DLC
`for Library of Congress
`
`90-15112
`CIP
`
`© Churchill Livingstone Inc. 1991
`
`All rights reseived. No part of this publication may be reproduced, stored in a retrieval sys(cid:173)
`tem, or transmitted in any form or by any means, electronic, mechanical, photocopying,
`recording, or otherwise, without prior permission of the publisher (Churchill Livingstone
`Inc., 650 Avenue of the Americas, New York, NY 10011 ).
`
`Distributed in the United Kingdom by Churchill Livingstone, Robert Stevenson House, 1-3
`Baxter 's Place, Leith Walk, Edinburgh EHi 3AF, and by associated companies, branches,
`and representatives throughout the world.
`
`Accurate indications, adverse reactions, and dosage schedules for drugs are provided in this
`book, but it is possible that they may change. The reader is urged to review the package
`information data of the manufacturers of the medications mentioned.
`
`The Publishers have made every effort to trace the copyright holders for borrowed material.
`If they have inadvertently overlooked any, they will be pleased to make the necessary
`arrangements at the first opportunity.
`
`Acquisitions Editor: Beth Kaufman Barry
`Assistant Editor: Leslie Burgess
`Copy Editor: Kamely Dahir
`Production Designer: Charlie Lebeda
`Production Supervisor: Sharon Tuder
`
`Printed in the United States of America
`
`First published in 1991
`
`7 6 5 4 3
`
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`

`pharmacology of
`Antineo~lastic Agents and
`IVle~hamsms of Multidrug
`Resistance
`
`Antonio C. Buzaid and Ed Cadman
`
`52
`
`~Go
`
`HISTORICAL ASPECTS
`
`The vesicant properties of sulfur mustard have been known for
`more than 100 years. It was only in 1919, however, that Krumb(cid:173)
`haar and Krumbha~r observed that poisoning by sulfur mustard
`produc~d leukop~m~, aplasia of the bone marrow, marked de(cid:173)
`crease m lymph~,d tissue, and ulceration of the gastrointestinal
`tract. These fmdmgs prompted Goodman and associates to test
`wh~ther nitrogen m~stard could be used therapeutically. 1 Fol(cid:173)
`lowing successful ammal studies, clinical trials were launched in
`1942, which ushered in the modern era of chemotherapy.
`In 1948 methotrexate became available, and shortly thereafter
`5-fluoro~racil was synthesized. In 1955 the first ch~motherapeutic
`cure usmg methotrexate as a treatment for m:::tastatic: chorio(cid:173)
`carcinoma was recorded. During the 1960s investiga.tion 0f cer(cid:173)
`tain medicinal plants resulted in the development of the Vinca
`alkaloids and podophyllotoxin derivatives. Late ir; that decade
`DeVita and colleagues showed that combinations of chemother(cid:173)
`apeutic drugs produced better results than single-agent therapy.2
`Since then the incorporation o f daunorubicin, doxorubicin, and
`cytosine arabinoside in the oncologic armamentarium has re(cid:173)
`sulted in a significant impact in the curability of many non-Hodg(cid:173)
`kin's lymphomas and acute leukemias. Clinical trials since about
`1970 have produced a number of useful chemotherapeutic regi(cid:173)
`mens.
`In this chapter we review the pharmacology of antineoplast_ic
`drugs, with special focus on the cytotoxic agents employed in
`the treatment of hematologic malignancies. In addition, we will
`provide an overview of their mechanisms of action as well as
`potential strategies designed to overcome drug resistance.
`
`CELLULAR KINETICS AND TUMOR GROWTH
`
`~t any given time only a portion of the cells in a tumor. are
`actively dividing; this subset of cells is called the growth fract10~.
`~.en a malignancy first arises, most of the tumor cells are d1-
`Vtd1ng, and the growth fraction is hi~h. A_s the tumor grows, a
`~arger proportion of the cells become macbve and assume a re~t(cid:173)
`'?g state. The decline in growth fraction may be due to r~stnc(cid:173)
`t,ons of space, nutrient availability, and blood supply: This pat(cid:173)
`tern of growth does not follow a classical exponentta! growth
`curve and is best described by the Gompertz equa_t,on. The
`growth fraction depends also on the type of tumor w!th values
`ranging from less than 10 percent for some ad~noc?I'cmomas t_o.
`greater than 90 percent for some lymphomas. This concept is
`
`Fig. 52-1. Phases of the cell cycle. Go. resting phase (nonproliferation of cells!;
`G1. pre-ONA synthetic phase (12 hours to a few days!; S, ONA synthesis (usually
`2 to 4 hours!; G2, post-ONA synthesis (2 to 4 hours; cells are tetraploid in this
`stage!; M. mitosis l 1 to 2 hours!.
`
`of great importance since most chemotherapeutic agents are
`more effective against dividing cells than against resting cells.
`A schematic presentation of the events occurring during the
`cell cycle is shown in Figure 52-l. Cytotoxic agents can be divided
`into phase-specific and phase-nonspecific according to their pre(cid:173)
`dominant effect on the cell cycle.
`I. Phase-nonspecific agents are effective in any phase of the cell
`cycle. Agents that fall into this category usuaJ!y have a linear
`dose-response curve (i.e., the greater the dose administered,
`the greater the fraction of cell kill). They are divided into two
`subgroups:
`a. Cycle-specific agents kill cells that are proceeding through
`the cell cycle independently of whether the cell is in GI, G2,
`S, or M phase (e.g., alkylating agents, cisplatin).
`b. Cycle nonspecific agents kill nondividing cells (e.g., steroids
`and antitumor antibiotics except bleomycin).
`2. Phase-specific agents are effective only if present during acer(cid:173)
`tain phase of the cell cycle. Within a certain dose range agents
`of this category show no increase in cell kill with further in(cid:173)
`crease in dose. If the drug is maintained over a period of time,
`however, more cells will enter the specific lethal phase of the
`cycle arid ~e killed_. Examples include L-asparaginase (GI
`phase), antimetabohtes (S phase), and Vinca alkaloids (M
`phase).
`Chemotherapeutic agents are not completely specific and af(cid:173)
`fect normal as well as neoplastic cells. This effect is most pro(cid:173)
`nounced with rapidly proliferating cells, such as the mucosa of
`the gastrointestinal tract and the bone marrow. This limits dose
`escalation and usually determines the maximum tolerated dose.
`
`TUMOR HETEROGENEITY
`Cancer has been shown to be a clonal d isease, (i.e., the cancer
`cells descend from a single progenitor cell). However, as cancers
`progress they become markedly heterogeneous. Within a single
`
`669
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`670 Part V / Hematolo ic Mali nancies
`
`A
`
`Fi1. 52-2 . .. Growth trees .. of stem cell compartments up to eight cells for _IAI a
`death rate of 0 and (Bl a death rate of 0.6. A circle that branches into two circles
`indicates the division of a stem cell to form two new stem cells: the gray circles
`indicate that the division produced two differentiated cells. Dotted hnes indicate
`ancestries in which all the cells have subsequently become d1lferentIated lor d1edl,
`and solid lines indicate continued stem cell proliferation. Resistance (pink circles)
`occlr.i at the fifth branch !line connecting two viable s:em cells! in cells that are
`sensitive lnot resistant!. All cells resulting from the drv1s1on of a resistant cell
`are themselves resistant. In A it can be seen that this leads to three cells being
`resistant:; in B, the comparatively greater number of branches leads to five of the
`eight cells being resistant. This illustrates the enhancement of resistance caused
`by cellular differentiation [or death!. Note: resistance is assumed to occur de(cid:173)
`terministically rather than randomly in order to simplify illustration, but it would
`not be expected to do so in reality. !Modified from Goldie and Coldman, 6 with
`permission.I
`
`tumor this heterogeneity is expressed as variations in histopa(cid:173)
`thology. cytogenetics, expression of surface antigens, growth
`rate, metastatic potential, and more importantly, sensitivity to
`cytotoxic agents. The major factor responsible for this hetero(cid:173)
`geneity is spontaneous mutation. Tumors are also heterogeneous
`in their supply of nutrients and oxygen, factors that may further
`increase their genetic instability.4
`5 The overall growth of a tumor
`•
`is dictated by the number of cell doublings. its growth fraction,
`and the death rate of the cancer cells. For example, in a given
`tumor (Fig. 52-2A) for which a constant mutation rate and a death
`rate of zero are assumed, the higher the number of cell doublings,
`the larger the tumor, the larger the number of mutations. and
`thus the higher the chance of having chemotherapy-resistant
`clones. If a constant mutation rate is assumed in a tumor with a
`high death rate (Fig. 52-28), many more cell doublings and there(cid:173)
`fore many more mutations must occur for the tumor to reach the
`same size.6 The latter situation applies to tumors that are slow(cid:173)
`growing, apparently because the rate of cell loss is high. Thus,
`at the time that these slow-growing tumors an,: clinically de(cid:173)
`tectable, they have already undergone multiple mutations and
`have a large numbe.r of cells that are resistant to virtually all
`available anticancer agents. In simple terms, one can envision a
`patient with a bulky and slow-growing tumor as actually having
`multiple different cancers. Consequently, current chemotherapy
`is seldom successful in patients with this type of condition.
`The mathematical model of Goldie and Coldman6 allows one
`to develop a better intuitive understanding of the events that
`occur during the treatment of cancer. This model substantiates
`the concept of dose intensity developed by Hryniuk and Bush7
`and also validates the importance of employing multiple cyto(cid:173)
`toxic drugs instead of single drugs to decrease the development
`of resistance. In addition, this hypothesis has resulted in the use
`of alternating non-cross-resistant regimens. This approach has
`recently gained wide popularity. However, since most chemo(cid:173)
`therapy regimens presently available are at best only partially
`non-cross-resistant-as, for example MOPP (mechlorethamine,
`oncovin, procarbazine, and prednisone) alternating with ABVD
`(Adriamycin, bleomycin, vinblastine, and dacarbazine) the ther(cid:173)
`apeutic value of this strategy has not been properly tested.
`
`DEVELOPMENT Of CHEMOTHERAPEUTIC AlitNrs
`Chemotherapeutic ag~nts m~y be developed (I) by s
`edures using new b1ochem1cal and pharmacologic YllU~ti
`pr~c structure-activity relationships; ( 2) from natura1conc%
`an
`1 nt extracts, microbial fermentation, and rnar- SOurtes
`(e.g .. Pa
`. .
`th t·
`ine or
`isms); and (3) by exammmg new syn e ic compounds lllad:a~.
`other purposes.
`fo1
`
`Screening for Antitumor Activity
`The concept of screening agen~ for a~titumor activity is b
`on the rationale that an appropnate b1oassay rnay reliabl ii.I~
`dicate activity against human_ ca~cers. At the National c;~n(cid:173)
`lnstitute the algorithm shown m Figure ~2-3 has been used si er
`1972. Modifications are frequent!}'. made m an attempt to incr:
`the predictive value of the screen mg panel. After drug acquis•,·
`.
`.
`th
`.
`l t<ll)
`and formulation, the firs! ~tep m. e screening process is evai.
`uation of anticancer act1v1ty agamst the mouse P388 leukern·
`model. Active drugs are subjected to a broader testing ~
`which includes many ~ouse tumo~s. as _well as_ human tumors
`xenografted in nude mice. If no act1v1ty ts seen m the P388 ieu.
`kemia model, the drug is cross-checked against hurnan turnors
`grown in soft agar and by the subr~nal capsule assay. Drugs that
`are not active in the P388 leukemia model but show activity in
`other biologic or biochemical systems will also undergo evalu(cid:173)
`ation in the broader testing panel. If anticancer activity is ob(cid:173)
`served either in the broader panel or in the soft agar or subrena(
`capsule assay. the drug enters preclinical toxicologic testing. (k.
`spite many modifications, it is not known how well this current
`system identifies potentially active antineoplastic agents.
`
`Clinical Trials
`Following extensive toxicologic studies, the new drug usual~
`enters clinical trials in humans, which are divided into low
`phases. Phase I trials are designed to determine the toxicity pro(cid:173)
`file, the maximum tolerated dose(MTD), and pharmacologicdata;
`the determination of anticancer activity is a secondary goal. Pa•
`tients with refractory cancers are candidates for phase I trials.
`The initial dose employed is usually IO percent of the lethal dose
`(LD,o) found in rodents during the preclinical studies. This dose
`is progressively escalated, generally according to a modified Fi·
`bonacci scale (Table 52-1 ). At least three patients are usual~
`enrolled and evaluated at each dose level. Once the M1l> and
`tox_icity profile are determined, the drug enters phase II studies.
`which are designed primarily to determine the efficacy of the new
`compound in different types of cancer generally the 7 to 10 most
`common ones. Phase Ill trials generally compare the efficacy 01
`the new drug in a randomized fashion with the existing "standard
`therapy." In phase IV trials, usually conducted after the drug has
`
`T(cid:127)
`
`lllle 52-1 Modified Fibonacci Dose Escalation
`Scheme Used in Phase I Trials
`Drug Dose
`Percent Increment
`lmg/m2l
`above Prior Dose level
`n
`[Initial dose levell
`
`2n
`
`3.3n
`Sn
`7n
`Sn
`
`12n
`
`16n
`
`1OD
`65
`51
`40
`28
`33
`33
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`Synthesis
`of
`new compounds
`
`or
`
`isolation of
`natural
`products
`
`Cha ter 52 / Pharmacolo of Antineo lastic A ents and Mechanisms of Multidru Resistance 671
`
`~ ....
`
`Other biologic
`or biochemical
`systems
`
`active(cid:141)
`
`•
`
`•
`
`Broader Panel
`
`Testing in other
`mouse tumors
`(Leukemia L 1210,
`B16 melanoma,
`M5076 sarcoma)
`
`Human tumors
`xenogratted In
`nude mice
`
`(cid:141)
`
`P388 mouse
`leukemia
`model
`
`I not
`
`• active
`
`Cross-checked in
`human tumors grown
`In soft agar and
`subrenal capsule assay
`
`active (cid:141)
`
`• Mammary xenogratt
`MX-1
`
`aciive
`
`Pre-clinical toxicologic
`studies
`
`/
`
`not aciive
`
`/
`
`no further study
`Fig. 52-3. Algorithm used for drug screening at the National Cancer Institute.
`
`been marketed, the drug is combined with other treatment mo(cid:173)
`dalities (e.g., radiation and/or s urgery) and c ompared in a ran(cid:173)
`domized study with the standard therapy.
`
`PHARMACOLOGY OF CHEMOTHERAPEUTIC AGENTS
`A number of fundamental molecular processes must take place
`for cells to proliferate. DNA must be replicated without error, a
`~r.ocess that requires a n appropriate supply of purine and pyrim(cid:173)
`idine nucleotides and multiple enzymes such as DNA polymer(cid:173)
`ases. These enzymes are produced from complementary RNA ~yn(cid:173)
`lhesized from an intact DNA template. The RNA is translated mto
`proteins through a complex polymerization reaction, which takes
`place on the ribosomes in the cell cytoplasm. After the cells have
`replicated their DNA, they undergo mitosis. Cytotoxic agents i.n(cid:173)
`terfere with one or more of these essential cellular processes (Fig.
`52-4).
`Cytotoxic agents have been classically divided i~to alkyla.ting
`agents, plant a lkaloids, antitumor antibiotics, ant1metabolltes,
`and a miscellaneous group. There are presently mor~ than 40
`standard (i.e., commercially available) chemotherapeutic agents.
`The Pharmacology of those agents used in the treatment of he(cid:173)
`lllatologic malignancies is presented below.
`
`Alkylating Agents
`1:;;i
`Wi~kylating agents conta in alkyl groups that bonr~t~f;:
`ciat dnu~leophilic s ubs tances of the D~A a ndl~r f structure the
`e with the DNA. On the basis of the ir chemica
`
`alkylating agents are divided into the five groups shown in Table
`52-2.
`
`General Mechanism of Action
`The cytotoxic as well as the mutagenic effects of the alkylating
`agents are directly related to the alkylation a nd disruption of
`DNA. Figure 52-5 shows the various mechanisms by which mech(cid:173)
`lorethamine (nitrogen mustard) may alkylate the DNA. While
`mechlorethamine is used here to illustrate the effects of the al(cid:173)
`kylating agents on the DNA, the same basic mechanisms apply
`
`Table 52-2. Alkylating Agents
`Nitrogen mustards
`Mechlorethamine
`Cyclophosphamide
`lfosfamide
`Chlorambucil
`Melphalan
`Ethylenirnines
`Thiotepa
`Hexamethylmelamine•
`Alkylsulfonates
`Busulfan
`Nitrosoureas
`Carmustine
`Lomustine
`Streptozocin
`Triazines
`Oacarbazine !DTICl
`" lnvestigational drug.
`
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`

`!6~7!2~£P!!art~VJ/'.,_H~e~m~a~to~log~ic1Ma~ligil!:na!nc~ie!s _____________________________________ ~~~~~~~~----~-
`,,...... ~
`I Purine Synthesis ]
`
`Pyrimidine synthesis
`
`Inhibition of purine '
`biosynthesis
`(e.g., 6MP, 6TG, MTX)
`
`Inhibition of adenosine
`deaminase
`(e.g., pentostatin)
`
`Topolsomerase-11 mediated
`ONA cleavage via
`stabilization of DNA
`cleavable complex
`(e.g., anthracyclines,
`mitoxantrone, VP16, VM26,
`m-AMSA, dactinomycin)
`
`/
`
`.
`. of pyrimidine sysnthesis
`tnh1blt1on PALA. pyrazoturin)
`(e.g.,
`
`Rlbonucleotldes
`
`Inhibition of ribonucleotide
`reductase
`- --- (e.g., hydroxyurea,
`tludarabine)
`
`Inhibition of dTMP synthesis
`(cid:141) _ -
`(e.g., SFU, MTX)
`
`Inhibition of DNA polymerase alpha
`~ ___ -
`(e.g., Ara-C)
`
`Deoxyrlbonucleotld••
`
`! (cid:141)
`+ !
`
`Topo II
`
`JI
`/
`
`Incorporation into DNA
`.. ,
`.
`(e g Ara-C fludarablne)
`~~...._,..-j......_
`
`Single strand binding and
`intra and interstrand cross(cid:173)
`linkage of DNA
`(e.g., alkylating agents, heavy
`metals, mltomycin, OTIC,
`? procarbazine)
`
`,
`
`,
`
`! "-
`(e.g., vincristine, vinblastine) , l ',,,,
`'@~
`' Hydrolysis cl extracallular
`
`Scission of DNA
`(e.g., bleomycin)
`
`+
`
`drug
`
`~
`
`Binding to tubulin preventing
`microtubule assembly
`
`e;odi"9: :.--.blooklog DNA
`
`,
`
`and RNA production
`r:::::::l '-
`(e.g., anthracyclines, mitoxantrono,
`~ '
`dactinomycin, mithramycin)
`
`L-asparagine
`(e.g., asparai;inase)
`
`fil- 52--'. Overview of sites and mechanism of act10n of the most useful chem<Xherapeutic agents.
`
`to the other alkylating agents. The nitrogen atom at the 7 posi~ion
`in guanine is particularly susceptible to covalent bond formation,
`but other atoms in the purine and pyrimidine bases (e.g., the
`nitrogens at the I and 3 positions in adenine) as well as phosphate
`moieties of the DNA and proteins associated with DNA also may
`be aJkylated.8 The pharmacology of these agents is discussed in
`Appendix 52-1, and the metabolism of cyclophosphamide is
`illustrated in Figure 52-6.
`
`fractionation of this extract by other investigators led to ~ pu(cid:173)
`rification of vincristine, vinblastine, and other Vinca aikal01~
`Podophyllotoxin is extracted from the mandrake plant. P Eto(cid:173)
`phyllum peltatum L., which was also used as a folk reme<iY•
`poside and teniposide are semisynthetic derivatives 01 podO(cid:173)
`phyllotoxin. The mechanism of action of these drugs haS not~
`co~pletely elucidated, but it appears to be related to theirabilt~
`to m~u~e stabilization of a topoisomerase II-DNA cleavage C()l1I'
`plex m its putative cleavable states (Fig. 52-7).
`
`Plant Alkaloids
`
`The plant alkaloids include the vincas and the epipodophyl(cid:173)
`lotoxins (Table 52-3 and Appendix 52-2).
`The periwinkle plant, Vinca rosea L., has been described for
`many years in medicinal folklore as having beneficial properties.
`The observation of bone marrow suppression by Noble et al. in
`J 958 Jed to the isolation of an active alkaloid extract.1 Further
`
`Tait 52-3. Plant Alkaloids
`Vinca alkaklids
`Vinblastine
`Vincristine
`
`EoiDOdophyllotoxin
`Etoposide CVP1 61
`T eniposide IVM26J•
`• 1nvestigational drug.
`
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`

`'
`
`Chapter 52 / Pharmacolo of Antineoplastic Agents and Mechanisms of Multidru Resistance
`
`673
`
`p
`
`""'------'------,, 3'
`Guanine
`
`p
`
`/
`
`HO
`
`oo✓✓✓: O
`
`N~~ H2N
`o ✓ ✓
`
`lnterstrand crosslinking
`between guanines
`
`Abnormal base pairing
`with thymine
`
`/
`
`p
`
`L - . - -- ~-_J
`
`Ring cleavage
`
`p
`
`Depurination
`
`•
`
`Excision of guanine residue
`
`Fig. 52-5. Meclianisr:1 cf 3ction of alkylating agents, illustrated by mechlorethamine.
`
`C;yclophosphamide
`
`Hepatic cytochrome
`P-450 system
`
`4-Hydroxycyclophosphamide
`
`Eo,yma1;, j
`
`·-1/
`
`Spontaneous
`
`Hepatic
`aldehyde
`oxidase
`
`Aldophosphamide
`
`Nonenzymatic in
`peripheral tissue
`
`I
`.
`~ - - - - - - - -
`I
`I
`Carboxyphospham,de,
`4-Ketocyclophosphamide
`L ____________________ I
`
`1nact1ve metaboliW:;,
`
`I - - - - - - - - - - - ,
`I
`I
`I
`I
`
`Phosphamide mustard +
`acrolein (toxic to urothelium)
`
`'------------'
`
`1 If,') .
`
`Fig. 52.&. Metabolism of cyclophosphamide.
`
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`

`674
`
`ancies
`
`•somerase II
`ro~~:entlY bound
`co
`ONA:
`to
`lex
`oleavable comP
`
`5'
`
`p ........_3. Stabilization of the
`cfeavable complex by
`antineoplastic drug
`
`3·
`
`Development of
`DNA strandbreaks
`( - - - )
`
`Fig. 52-l . Topoisomerase II-mediated ONA cleavage.
`
`Antitumor Antibiotics
`The antitumor antibiotics are natural products obtained from
`the culture broth of various species of streptomyces. By direct
`inhibition of DNA and/or RNA synthesis, they affect different
`phases of the cell cycle. Table 52-4 summarizes the most clinically
`useful antitumor antibiotics (see also Appendix 52-3). Oactino(cid:173)
`mycin and mitomycin are of limited value to the hematologist
`and will not be discussed here.
`
`Anti metabolites
`. ~ti~etabolites are compounds that because of their strocturil
`similarity to physiol?gic intermediates are incorporated as Ira~
`ulent substrates, ultimately interfering with vital processes oft
`lular metabolism. The antimetabolites can be divided into t}lree
`groups: (I) folic acid analogues (Fig. 52-8), (2) pyrimidine_~(cid:173)
`logues, and (3) purine analogues (Table 52-5 and Appendix 5
`
`T1bl1 52-4. Antitumor Antibiotics
`Anthracyclines
`Daunorubicin
`Ooxorubicin
`ldarubicin'
`Mit.oxantrone
`Dactinomycin
`Plicamycin
`Mitomycin
`Bleomycin
`'lnvestigational drug.
`
`T1bl1 52-5. Antimetabolites
`Folic acid analogues
`Methotrexate
`Pyrimidine analogues
`Cytarabine Iara-CJ
`Fludarabine•
`Azacytidine•
`Fluorouracil (5FUJ
`Floxuridine lFUORJ
`Purine analogues
`Mercaptopurine (6MP)
`Thioguanine (6TGl
`-.Jentostatin•
`slnvestigational drug.
`
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`
`

`

`SFU
`
`Cha ter 52 1
`
`Pharmacol
`
`of Antineoplast ic A ents and Mechanisms of Multidru Resistance &75
`
`FUMP ~
`
`UDP
`
`~
`
`dUOp ~
`
`Methotrexate
`
`dUDP ~dUTP
`
`dUMP ~
`
`l
`
`dUMp
`
`- - - - 1:._ _ _ ~(cid:141) dTMP
`Thymidylate Synthase
`
`Formyl FH(cid:157)
`
`N
`5
`(leucovorin)
`
`NS, 1 ~
`
`NS 10 MethyleneFH4
`
`t "'-•H• ~
`
`ethenyl FH4
`
`/
`
`.
`
`contnbutes second
`Carbon of purine ring
`synthesis
`
`~ JI'
`t ro
`N 1 O Formyl FH4
`J 7
`
`O
`
`a
`
`2
`
`4
`
`3
`
`II
`
`FH2
`
`;hyd,o~I"'
`reductase
`
`Methotrexate i . . - -- - - - '
`
`tural
`aud(cid:173)
`cel-
`1ree
`1na·
`52·
`
`Fig. 52-8. Sites of action of methotrexate and 5-fluorouracil.
`
`Ta.Ille 5!-&. M iscellaneous Aoents
`Heavy metals
`Cisplatin
`Carboplation
`Hydroxyurea
`Procarbazine
`Asparaginase
`Amsacrine•
`Gallium nitrate•
`Hormones and antihormones
`'lnvestigational drug.
`
`41 In contrast to the alkylating agents, most antimetabolit~ are
`~ ·specific and present minimal risk in terms of potenti_al for
`carcinogenesis, leukemogenesis and infertility. Fluorouracil and
`Uoxuridine are of limited value to the hematologist and will not
`be discUSsed here.
`
`Miscellaneous Agents
`rn ~e agents included in the miscellaneous category are sum(cid:173)
`rn~zed in Table 52-6 and Appendix 52-5. Of the~e, only the h~vy
`tals, _hydroxyurea, procarbazine, L-asparagmase, amsc1:c~me
`:gallium nitrate, and glucocorticoids are currently of clinical
`ance to the hematologist.
`
`MULTIDRUG RESISTANCE
`PY may be defined
`th
`.
`Oinicat
`. ·r a t response
`resistance to cytotoxic chemo era
`as lack
`followec1of initial response to chemotherapy or mi~any causes,
`by Progression after chemotherapy. It has
`
`which can relate to either the tumor and/or the host Table 52-
`7 lists factors that may adversely influence the efficacy of chem(cid:173)
`otherapeutic agents and result in clinical resistance, of which
`intrinsic or acquired resistance to chemotherapeutic drugs is
`probably the most important. In a broad sense multidrug resis(cid:173)
`tance (MDR) can be defined as resistance to dissimilar agents.9
`Cancer cells may develop resistance to the chemotherapeutic
`agents by various mechanisms, an overview of the most common
`of these is shown in Figure 52-9. ln most cases these mechanisms
`have been primarily characterized in cell lines, and their clinical
`relevance has yet to be defined.
`It is important to emphasize that MOR implies the likelihood
`of more than one mechanism being operative in vivo, not only
`within the same tumor but also within the same cancer cell. Each
`of the mechanisms for MOR will be discussed.
`
`The P-Glycoprotein System
`Cancer cells selected for resistance to one natural product such
`as doxorubicin are frequently cross-resistant to structurally dis(cid:173)
`similar agents, including the vinca alkaloids and the podophyl(cid:173)
`lotoxin derivatives. This pattern of drug resistance has been
`termed pleiotropic resistance or MOR. 10 In 1976 Juliano and Ling
`reported a pleiotropically resistant cell line derived from Chinese
`hamster ovary cells which accumulated less colchicine than its
`drug-sensi~ve parent.11 Believing changes in cell permeability to
`be responsible lor decreased drug accumulation, their group in(cid:173)
`vestigated the cell membranes of these drug-resistant cells and
`identified a highly amplified 170-kDa phosphoglycoprotein.11 Sub(cid:173)
`sequent work demonstrated that this phosphoglycoprotein
`termed P-glycoprotein, actually functioned as an adenosine tri:
`phosphate (ATP} dependent drug efflux pump capable of extrud(cid:173)
`ing plant alkaloids and antitumor antibiotics fro m the cancer cells
`rather than decreasing drug uptake.10
`
`NOVARTIS EXHIBIT 2096
`Breckenridge v. Novartis, IPR 2017-01592
`Page 9 of 17
`
`

`

`676 Part V I Hematologic Mali nancies
`Tabla 52-7. Factors That May Adveraely Influence the Efficacy of
`_ _ _ _ _ _ _ Chem_ otherapeuticAgen~~::'.--- - - - -
`Host Factors
`1 •n al•
`.
`Alteration of drug pharmacokinetics su;h that a g,ven dose resu ~ 1
`Failure of the drug to undergo activation
`to
`tcred distribution of metabolism (e.g., increased catabolrsml
`Strategic location of the tumor, resulting 1n poor eccc~s of the drug
`the tumor site (e.g .. testis, central remus system)
`Tumor Factors
`Intrinsic or acquired chemotherapy drug resistance
`Alteration of blood flow to the tumor, resulting in an inadequate dose
`.
`reaching the tumor
`Changes in the tumor environment, reducing the effecuveness of the dn,g
`.
`.
`.
`(e.g .. low oxygen pressure. low pH
`Presence of u.nor cells predominantly in Go phase (k1net1c res1stanc~ _
`
`Genetics of P-glycoprotein
`P-glycoprotein, also known as PGY-1 , is the product of the
`MOR-I gene, 12 which is located on chromosome 7. 13 At least one
`other homologous MOR gene, termed MDR-3, has been found in
`humans, but its function remains to be determined. 14 Examina(cid:173)
`tion of the chromosomes of highly resistant cell lines often re(cid:173)
`veals the presence of many amplified sequences of DNA, which
`either may exist as small extrachromosomal circles, termed min(cid:173)
`utes or double minutes, or may be integrated into the chromo(cid:173)
`some as a _homogeneously staining region. These regions are
`kno:,vn to arise whe~ ?ene amplification occurs and contain many
`copies of the amphfted gene. Most often, amplification of the
`
`ears to be the basis for increased P. 1
`MOR-I ~en~ a~t hly resistant cell lines. Gene arnpJifi~ Y~Pt~
`in some P-gl ycoprotein-positive atton i,..._
`,g
`expression in
`observed. howev~~ls of resistance. Thus, it appearCe)J Ii~~
`lected f?r lo~v ~~glycoprotein is n ot always depend! that !'Ir. "t
`0 nd may also be affected at levels su nt on g "·
`expr~~•0 ~
`amph(1cat1on t:anslation. 'o Ampl ification of other clih as ~~
`scnpt1on or lasses such as the cytoplasmic protein seJy i~
`posed eg~n~e~ected in P-glyc<;>protein-p_o~itive cells, aJ~orcin ~
`of this protein is not sulflc1en t, nor is it oU&Ji ,.__
`n~e "'·
`also b e .
`typ 1s Th
`ssa,.
`e in vitro d
`h
`erexpressIon
`e.
`. •t ·,on of the MDR p eno
`emo ~,.
`·
`M·DR I
`• gene mt? sensitive tel ns_tri.
`for acquIsI
`tion that transfection of the
`expression of both the P-glycoprotem and the l ,0 I h"s
`'" R
`·d
`t
`t
`notype represen ts the most "'!1~or an ev, ence that the pr P~.
`-
`f
`con ers
`f th MDR-l ,,ene is a sufhc1ent and necessary cond· ~lite
`Ition
`e 16
`e
`o
`for
`o
`expression of the MDR phenotyp ·
`Structure and Function of P-glycopratein
`The cloning and sequencin~ o f th_e MDR-1 gene have allo
`prediction of the correct ammo acid sequence of the p I we.i
`protein. Computer modeling i~dicates that the protein rn~pco(cid:173)
`contains two homologous regions, each consisting ol ap ecu~
`duplication 10 (Fig. 52-10). ThE:re ar e 1.280 amin? acids and r
`the protein, which suggests a history 0t 0Xi(cid:173)
`mately ha.Ii
`helical transmemb~ane ~omams (shown as cylinders in Fig. 5;_
`JO) in P-glycopro tem. It Is not clear, h owever, how many Of these
`transmembrane domains are required for channel fonnati
`There are two nucleotide (ATP)-binding domains localized in :
`
`Increased efflux
`(e.g., P-glycoprotein
`in typical MOR)
`
`Decreased influx
`(e.g., decreased
`membrane)
`
`Decreased transformation
`from inactive to active drug
`(e.g., mitomycin resistance) ~~ • .,......,
`
`'
`
`Increased intracellular
`drug binding
`j(
`(e.g., glutathione system)
`,._.,D . ..,,..,.
`
`Increased ONA repair ~
`(e.g., increased 0
`_
`6
`methylguanine DNA
`. methyltransferase
`m BCNU resistance)
`
`Altered intracelluar
`drug distribution
`(e.g. , to lysosomes)
`
`Increased metabolism to
`Alteration of 5
`. .
`_inactive drug
`pec1flc
`target
`(e.g., increased aldehyd
`(e.g., altered toen~Ymes
`dehydrogenase in e
`etoposid PD•s?merase II in
`cyclophosphamide resistance)
`and increas e resistance
`.
`F"
`resfia~HF)R in MTX
`••· 52-9. Overvtew of potential sites for d
`ce
`rug resista
`rce at th
`e cellular level
`
`Fit. 52-1
`as a cha
`Produc:i
`and are
`1ndirectI
`Protein.
`Bradley
`
`()'top
`Porti<
`lllell\l
`as tr;
`extt\J
`folio,
`tide)
`glYcc
`Prat
`
`De1
`Nu,
`l'
`clet,
`cl)~
`r~
`the
`Pa..,
`
`NOVARTIS EXHIBIT 2096
`Breckenridge v. Novartis, IPR 201