You are on page 1of 66

THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

FACULTY OF SCIENCE
MODULAR DEGREE SCHEME

BSc HONOURS DEGREE


IN
PHARMACOLOGY

Kudakwashe Emmanuel Mupamhanga

K0433939

The Biological and Clinical Significance


of
P-Glycoprotein in cancer
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

ABSTRACT & BACKGROUND

CANCER

– BIOLOGY OF CANCER

– CANCER CAUSES

– CANCER TYPES

– GENES IN CANCER

– TREATMENT SCHEMES

P-GLYCOPROTEIN

– HISTORY

– STRUCTURE

– LOCALISATION OF Pgp IN NORMAL TISSUES

Pgp SIGNIFICANCE ON THE THERAPY OF CANCER

– DRUG RESISTENCE

– P-GP EXPRESSION IN CANCER CELLS

– PHARMACOLOGICAL TARGETING OF Pgp

– 1ST TO 3RD GENERATION Pgp INHIBITORS

– STEM CELLS AND CANCER GENE THERAPY

CONCLUSION

REFERENCES
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
ABSTRACT

Cancer is a leading cause of death worldwide. It is characterized by rapid and

unregulated growth of the body’s cells as a result of a mutation in a proto-oncogene

or a tumor suppressor gene or both. It is however resultant of multiple mechanism

leading to its development ranging from environmental factors to hereditary

influence. The membrane bound MDR1 gene product P-glycoprotein (Pgp) has been

reported to be associated with drug resistance incidence in cancer to chemotherapy.

Pgp is thought to have an evolutionary role as a protective mechanism against toxins

ingested or inhaled from the environment. Pharmacological and gene therapy

research has strived to modulate the effects of Pgp or more recently make use of this

drug resistant characteristic for chemoprotection respectively.

Background
According to the World Health Organisation (WHO), cancer is a leading cause of

death worldwide and accounts for an estimated 7.4 million deaths (2004 statistics),

13% of all deaths worldwide. Cancer can affect any part of the body and there are

approximately 200 different types of cancer. (Cancer Research UK)

In the U.K alone there are approximately 285,000 new cases of cancer diagnosed

each year and it is estimated that 1 in 3 people will develop some form of cancer in

their lifetime. It is non-discriminate of age however it is most prevalent in older people

with 75% of cases developing in people at the age of 60 and above. Cancers in

children, teenagers and young adults account for approximately 1% of all cases.

Although cancer incidence has remained relatively stable over the last decade there

has been an overall increase in incidence rates in the U.K constituting a rise of one

quarter since 1975.

Table 1: Main types of cancer leading to mortality each year


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Cancer type Number of

deaths per

year
Lung 1.3 million

Stomach 803 000

Colorectal 639 000

Liver 610 000

Breast 519 000

WHO FACT SHEET No. 297

The WHO projects deaths from cancer worldwide to continue rising with an estimated

12 million deaths worldwide in 2030. Both men and women are affected however

variations exist between genders for the most frequent types of cancer.

Among men lung cancer is the most prevalent whereas breast cancer dominates

cancer incidences among women (Table 1). In the U.K the overall cancer death rates

have fallen by about 10% however a staggering 150,000 deaths, 1 in 4 of all deaths

are as a result of cancer (Figure 1). Despite declines in death caused by uterine,

oesophageal and male skin cancer 1 in 5 cancer fatalities are attributed to lung

cancer.

Figure 1: All deaths caused by cancer 2006


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Cancer Research UK (2007)


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
These figures and statistics give insight into the global burden of cancer. The

projected increases (12 million by 2030) place an ever mounting challenge on the

scientific community to understand the process of carcinogenesis and tofind effective

treatment strategies to combat the disease.

Biology of Cancer
“Tumors destroy man in a unique and appalling way, as flesh of his own flesh which

has somehow been rendered proliferative, rampant, predatory, and ungovernable . . .

Yet, despite more than 70 years of experimental study, they remain the least

understood. . . What can be the why for these happenings?”

Peyton Rous, in his acceptance lecture for the Nobel Prize in Physiology or Medicine

(1966)

Cancer refers to group of diseases that develop across time and are predominantly

characterised by uncontrolled division of the body’s cells. In the case of normal cells

external growth factors are required to instruct the cell to divide. Normal cell

regulation inhibits these growth factors accordingly and halts further division. Cancer

cells operate independently of these positive growth factors and thus divide in their

presence or absence. National Institutes for Health (1999)

Cancerous cells begin to dictate their own agenda for proliferation. Not only does this

ancestral cell display inappropriate proliferation all of its subsequent progeny operate

in this manner a mass of cells formed of these abnormal cells are referred to as a

tumour which can either stay in the tissue it originated in ( in situ cancer) or it may

begin to invade nearby tissue (invasive cancer). Invasive tumors are said to be
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
malignant. National Institutes for Health., (1999)., Normal cells can divide to fill in a

gap but a soon as there are a sufficient number of cells to fill the gap they cease to

divide. Cancer cells show no contact inhibition and continue to divide after they touch

other cells and consequentially form this large mass of cells -tumour. Blackburn et

al.,

Cells have a lifespan and this entails ageing and death via apoptosis which is the

normal, regulated programmed death of cells. Their ability to replicate their DNA is

limited to approximately 50 times due to the fact that each time a chromosome

replicates the (ends) shorten. Growing cells utilise telomerase enzymes to replace

lost cells whereas maturet cells lack this enzyme resulting in their limited replications.

Cancerous cells have the ability to activate telomerase in adult cells and this allows

for an unlimited number of cell divisions. Columbia Encyclopaedia 6th edition., (2007)

; National Institutes for Health., (1999); Blackburn et al.,

In addition when DNA is damaged or replicates abnormally, normal cells cease to

divide and since cancerous cells divide regardless of DNA damage or abnormal cell

replication they manage to accumulate increasing amounts of damaged DNA.

Blackburn et al.,

Figure 2: Stages of Tumour development

National Cancer Institutes (1999)


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Malignant tissue can cause secondary effects whereby expanding abnormal growth

puts pressure on surrounding organs and tissue or cancer cells metastasize and

invade other organs by shedding into the blood or the lymph.

It is believed that tumour development and growth is preceded a mutation within a

cell(s) leading to hyperplasia (figure 2) mentioned above which creates an

environment conducive to further mutation. This applies to virtually all of the body’s

tissues as all of them are susceptible to mutations. As a result of these mutations

cancerous cells attain a selective advantage over normal proliferating cells and thus

multiply much more rapidly.

The general consensus within modern science is that cancer is a disease of

molecules and genes. It is a multistep process developing across time as long

succession of genetic changes. Through these changes precancerous cells manage

to acquire the traits together and manifest into a malignant growth of cells. National

Institutes for Health., (1999)The main functioning genes are subdivided into three

categories. Proto-oncogenes produce proteins that enhance cell growth and division.

When mutated these genes are referred to as oncogenes. The second group are the

tumour suppressor genes and these create proteins that terminate cell division and

induce apoptosis. The third group are genes coding for DNA repair mechanisms

which help repair DNA from molecular changes that lead to cancer. National

Institutes for Health., (1999)

Mutations in any three of the groups of genes potentiate the development of cancer

(Table 2).
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Table 2: Gene mutations and their implications

Mutation Present Effects potentiating Cancer

Proto-oncogenes Results in oncogenes-

accelerated and

uncontrolled growth
Tumour suppressor genes Limited ability of signalling

inhibitory messages

DNA repair genes Loss of ability to repair

alterations in DNA

Genes in Cancer
Modern science views on cancer centre around it being a multistep process

developing across time as long succession of genetic changes. Through these

changes precancerous cells manage to acquire the traits together and manifest into

a malignant growth of cells. National Institutes for Health., (1999)

Mutations to proto-oncogenes forms oncogenes and these genes stimulate

excessive division whilst mutations in tumour suppressor genes (figure 4), results in
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
their inactivation. Consequentially the ability to inhibit excessive growth is lost and

collectively mutations in these groups of cancers account for most human cancers.

Proto-oncogenes code for proteins that play a part in pathways that process and

receive growth signals (figure 3) from other cells within a particular tissue. When

growth factors

Figure 3: Control of gene expression, stimulatory signals

Blackburn et al,.

are produced they move into the gaps between cells attaching to specific receptors

on their membranes. Upon activation these receptors transmit stimulatory signals to

proteins within the cell cytoplasm. These proteins in turn convey these messages

stimulating other proteins all the way to the nucleus and activate genes that facilitate

the movement of the cell through its growth cycle. The mutated form of these genes

(oncogenes) cause proteins involved in growth signalling pathways to become

overactive and as a result cell proliferation progresses at much faster rate than in

comparison to the rate before the mutation. Oncogenes also produce deviating

signals to receptor proteins leading to the release of stimulatory signals in the


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
presence or absence of growth factors. This continuous disruption of the signalling

cascade results in the nucleus receiving constant stimulatory signals instructing it to

divide. Blackburn et al,.

Figure 4: Some Genes implicated in Cancer development

Blackburn et al,.

When DNA repair mechanisms involved in maintenance of the chromosome are

damaged errors in the DNA go unattended. Without DNA mechanisms mutations are

allowed to accumulate within the cell. These mutations increase the cancerous

changes within a cell. Blackburn et al,.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

CANCER CAUSES
There maybe multiple mechanisms leading to the development of cancer (figure 5).

Although disputed the prevailing model centres around mutations occurring within

tumour suppressor and oncogenes which result in cancer. Alternative models

indicate mutations occurring in “master genes” which control the cell cycle and that

once a mutation occurs in these genes inappropriate gene dosing occurs. As a result

of this cells produce too much or too little of particular proteins required for proper

cell growth and an imbalance of this sort leads to cancer. Blackburn et al,.; National

Institutes for Health (1999)

Viruses that affect humans may also promote cancer; this is the case with viruses

which insert their DNA into the chromosome at the same point in which proto-

oncogenes are located thus inactivating them (converting them into oncogenes).

Virus DNA located close to genes involved in the regulation of cell growth may take

over host machinery resulting in increased transcription of those genes

(inappropriately) which provides the possibility of a cancer occurring. The table below

shows a few viruses that have been implicated in cancer. Blackburn et al,.
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Table 3: Viruses that can potentiate cancer

Viruses associated with increased risk of Cancer type caused

cancer
Human papillomavirus Genital Carcinomas

Hepatitis B Liver Carcinoma

Epstein-Barr virus Burkitts lymphoma

Human T-cell leukaemia virus T-cell lymphoma

Herpes Virus Kaposi’s sarcoma, B

cell lymphoma

Cancer research over the years had lead to knowledge that environmental factors

contribute to an individual’s chances of getting cancer. In 1775 Percival Pott found

that there was an unusually high incidence of scrotal cancer amongst men who

worked as chimney sweep boys. Hawes et al (1775) When there is a significant

correlation between exposure to an environmental factor and occurrence of a specific

cancer the factor is referred to as a carcinogenic agent. Carcinogenic agents range

from X-rays, UV light, Tobacco smoke, industrial solvents. Some cancers associated

with these factors are not associated with cancer genes furthermore some are

preventable. Blackburn et al,.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Figure 5:The Genesis of cancer 4 Theories

Padilla-Nash Hesed M and Reid Thomas (2003)


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
CANCER TYPES
Cancers are classified according to the type of tumour that develops and this is

based on where the original alteration occurred (See Table 4 below). Blackburn et al,.

Table 4: Origin of mutation and cancer

Type Origin
Carcinoma Epithelial cells (most common)

Sarcoma Muscle, bone, fat and

connective tissue

leukaemia White blood cells


Lymphoma Bone marrow (lymphatic

system)
Myeloma Specialised anti-body

producing white blood cells

Pgp SIGNIFICANCE IN THE THERAPY OF CANCER


The ideal cancer therapy aims at being highly efficacious with high tumour specificity

and maintaining minimum levels of toxicity. Roylance (2007)

TREATMENT SCHEMES
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
The basis of medical cancer therapy falls under the categories of chemotherapy,

radiotherapy, endocrine therapy and biological therapy. Although their mechanisms

may differ, treatment strategies tend to involve a combination of separate therapies.

Table 5: Mainline Therapies in cancer

Therapy type Treatment strategy


Chemotherapy The use of cytotoxic agents

Radiotherapy Ionising gamma radiation

Endocrine Blocking hormonal action

Biological Monoclonal antibodies

Surgery -ectomy

Chemotherapy is one of the mainline treatments in cancer and involves the use of

alkylating agents, platinum compounds, anthracyclines, antimicrotubuleagnets,

antimetabolites and topoisomerase II inhibitors. These drugs target rapidly dividing

cells consistent with tumour growth with a 90% efficacy on 10% of all cancers

Roylance 2007. They are however non-specific to tumour cells thus normal cells my

affected if they exhibit similar characteristics of division.

Figure 6: Chemotherapeutic effects on the cell cycle

VINCA
PHASE NON-SPECIFIC
ALKALOIDS
Alkylating agents
TAXANES
Cisplatin
Nitrosoureas
Antibiotics

M
G0

G2 G1

S
METHOTREXATE
HYDOXYUREA
CYTOSINE ARABINOSIDE
ANTHRACYCLINES
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Roylance 2007
Combination chemotherapy involves using more than 1 class of chemotherapeutic

agent at optimal dose and schedule. This enables synergistic action of the drugs as

different drug classes will affect different points within the cell cycle (figure 6).

Radiotherapy much like chemotherapy is non-specific to cancer cells thus normal

cells are damaged in the process. Unlike chemotherapy these cells have the ability to

repair themselves after. Cancer Research UK

It involves the use of ionising radiation in measured doses (X-rays). This radiation

damages cells hindering their growth and division. It is usually given before and after

surgery to reduce tumour size and after to improve treatment results. Palliative

treatment of cancer by radiotherapy buys times for other treatment strategies to be

implemented. Cancer Research UK

Hormonal therapy is usually associated with breast and prostate cancer and it is a

relatively specific form of treatment with minimal toxicity. Beaston 1896 postulated a

link existed between the ovaries and the proliferation of breast cells after removing

the ovaries of a woman with advanced metastatic breast cancer, she responded

drastically. When tamoxifen a selective oestrogen modulator was developed as a

contraceptive its uses in breast cancer soon superseded its relatively low efficacy as

a contraceptive (it even induced ovulation in some cases). In 1973 it was licensed for

use in breast cancer and its mechanism of action (Jordan 1974) involved the

blockade of oestradiol to oestrogen receptor (OER) in human breast and rat

mammary tumour. Hormonal therapy currently includes anti-oestrogen agent

(fulvesant) and aromatase inhibitors both steroidal (exemestane) and non-steroidal

(arimidex). Roylance (2007).

Biological therapy involves the use of monoclonal antibodies and small molecule

inhibitors. This strategy is efficacious however the mechanism of action is not fully

understood. The treatment is specific and of minimal toxicity (e.g. herceptin). This is
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
a relatively novel treatment strategy in cancer thus many of the treatment regimes

are not implemented unless conventional treatment has failed. Roylance (2007)

P-GLYCOPROTEIN

P-glycoprotein (Pgp) is the most characterised member of the human ATP Binding

Cassette (ABC)-transporter family. It is the gene product of the MDR1 gene and

serves as an integral efflux membrane protein. Pgp and members of this superfamily

are characterised by the ATP driven active transport of substances out of the cell. It
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
is believed to have evolved as a protective mechanism against harmful toxins

Gottesman et al (1995). Evidence of this is based on the highly conserved protein

regions that can be observed in prokaryotes, archea and eukaryotes.Seelig and

Landwojtowicz (2000)

The ABC efflux transporters are primarily located in plasma membranes; here they

extrude a variety of structurally diverse drugs, drug conjugates and metabolites

Schinkel and Jonker (2002). The human genome codes approximately 48 ABC

proteins, which are subdivided into subfamilies via sequence alignments (From A to

G). For a transporter to be considered functional it must contain at least 2 ABC

subunits. These proteins are membrane bound consisting of various domains and

specialised structures. (Table 6)

Table 6: ABC transporters

Szakacs et al 2008

Pgp transports a variety of chemically diverse substances across the cell membrane

thus protecting the cell from passively transported drugs. It plays a role in the
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
bioavailability of substances that enter the cell (including drugs, metabolites, and

xenobiotics). This wide substrate specificity limits compounds crossing tissues with a

protective barrier function which include; blood-brain barrier, liver, intestines, kidney

and testis/placenta (Figure 4). Most Pgp substrates are hydrophobic and many of

them contain aromatic ring structures Endicott and Ling (1989); Gottesman and

Pastan (1993). Its substrate specificity extends to non cytotoxic compounds as well;

calcium channel blockers, immunosuppressants, steroid hormones and neuroplectic

drugs.

P-gp History
Juliano and Ling (1976) first characterised P-gp while working with Chinese hamster

ovary (CHO) clonal cell lines where they reported the expression of a protein (P-gp)

in colchicine resistance. This followed previous work by Ling and Thompson in

(1974) (figure7) in which they isolated a series of related colchicine resistant CHO

clonal cell lines using single step selections. They also found that increasing

colchicine resistance correlated to increased resistance to other drugs and reduced

uptake.

figure 7: Development of Pgp understanding


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Mcdevitt and Callaghan (2007)

The isolation of clones using Chinese hamster ovary (CHO) cells (Juliano and Ling

1976) was useful as a method of investigating somatic cell mutations and the

phenotypic expression in cultured mammalian cells. Isolation also aided in identifying

environmental mutations.

Researchers attempted to investigate genetically linked mutations and specialised

only in genuine mutations (that is heritable alterations in particular genes). Heritable

mutations are valuable as they can also serve as genetic markers. Single step

selection methods were implemented and analyses were undertaken to determine

whether the appearance of variants was consistent with spontaneous mutation rates.

In 1970 while assessing the resistance of murine leukaemia sublines (L5178Y and

L5178Y/D) to actinomycin D Kessel and Bosmann (1970) found that administration of

50µg/kg actinomycin D inhibited uridine incorporation into RNA in L5178Y but not in

L5178Y/D. Following enzymatic studies (using galactosyl transferases, fucosyl

transferases, glucosyl transferases) they managed to identify an altered cell surface

glycoprotein. These alterations to the membrane composition where found to have

attributed to the changes in actinomycin D permeability.

These findings concurred with those of other researchers at the time and during the

isolation of colchicine-resistant CHO cells in which Juliano and Thompson (1974)

found that the colchicine-resistant cells characterised had pleiotropic cross-

resistance to other drugs (daunomycin and puromycin) furthermore reduced uptake

of colchicine was proportionate to the degree of drug resistance. This led to the

conclusion that colchicine resistance was bought about by alterations to membrane

permeability.
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Juliano and Ling (1976) attributed the multi-drug resistant characteristic of tumor

cells as being brought about by the overexpression of this surface glycoprotein and

named it Plasma glycoprotein.

STRUCTURE OF P-GP
The structure of Pgp like other functional entities of ABC transporters consists of 4

domains.2 membrane domains (MDs) and 2 nucleotide binding domains (NBDs also

referred to as the ATP-binding cassettes). The NBDs are responsible for the

generation of motional force while the MDs provide a translocation pathway for

substrates bound to the protein. Pgp shares a number of conserved sequence motifs

with other ABC transporters (Walker A, Walker B and the ABC signature) due to ATP

their common substrate. MDs however are more diverse and this diversity is

reflected by the large diversity of substrates that are transported. Seeger and W. van

Veen ( 2008)
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Figure 8a: P glycoprotein structure
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Schinkel and Jonker 2003

MDs (Figure 8a) consist of 6 putative transmembrane segments with heavy

glycosylation occurring on the first loop. The NBDs or similarly the ATP binding

cassettes are located intracellularly (Figures 8a and 8b). In vitro studies on Pgp

structure Schinkel et al (1993) showed that N-glycosylation was not necessary for

basic transport function. Up to 3 sites present in the mdra1 protein, they also deleted

a stretch of 20 amino acids containing 2 out of the 3 glycosylation sites following

transfection into drug-sensitive cells the effects of the mutations were analysed. It

was found that the absence of N-glycosylation did not alter the level or pattern of

cross resistance however drastically reduced the efficiency in which drug-resistant

clones where generated. As a result of these findings it was postulated that the

Glycosylation of the transmembrane loops contributed to the stability of Pgp (within

the plasma membrane) but not to drug transportation.

Figure 8b: Pgp structure showing mechanism of action

Modified from Gottesman et al,.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Photoaffinity labelling experiments Cornwall et al (1986); Bruggeman et (1992)

show that following the passive diffusion of chemical agents into the cell

cytoplasm they are bond by Pgp and exported out into the extracellular space

ATP hydrolysis or GTP in certain scenarios Ambudkar et al 1992

LOCALISATION OF Pgp IN NORMAL TISSUES


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Figure 9: localisation of Pgp as detected by MRK16 a monoclonal antibody
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Theibaut et al 1987

Thiebaut et al ‘s findings support the hypothesis of the MDR1 gene product (P-gp) as

having a role as a pump against physiological metabolites and chemotherapeutic

agents.
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

DRUG RESISTANCE
“The first successful chemotherapy of human cancer led soon to the realization that

drug resistance was going to be a major impediment to cure or long-term palliation”

Gottesman and Ling (2006)

In the 1940s Sidney Farber administered aminopterin in patients who where

diagnosed with leukaemia in what was the beginnings of modern age treatment to

caner. This also symbolised the beginnings of the understandings of drug resistance

and its clinical consequences as the treated children experienced an initial response

to therapy followed by remissions that were resistant to further treatment.

Drug resistance is a phenomenon associated with the chemotherapy of metastatic

cancers. Conventional chemotherapy of most common cancers (see table 1) can be

expected to reduce tumour size in 50% of all cases. In almost all cases drug

resistance develops and is the major cause of fatalities Baird and Kaye (2003).

Resistance falls into 2 categories; intrinsic or acquired. Chemotherapy has limited

efficacy in patients with intrinsic resistance which is present at the time of diagnosis

whereas acquired resistance is born of tumours which are initially responsive to

chemotherapy however reoccurrence of the tumour expresses a completely different

phenotype which is non-responsive to previous therapy. In some cases “multidrug

resistance”(MDR) occurs and this refers to pleiotropic cross-resistance to a range of

structurally unrelated compounds as a result of increased expression of the

transporter protein.
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Drug resistance is either as a result of alterations in the cancer cells or insufficient

drug exposure. Insufficient dosing, low bioavailabiulity, increased metabolism,

excretion are some of the factors that lead to drug resistance. Alterations in cancer

cells can lead to increased drug efflux (ABC transporters; Pgp), decreased drug

influx or activation of detoxification systems. Biard and Kaye (2003)

Pgp EXPRESSION IN CANCER CELLS


In vitro studies of multidrug resistance reveal that application of a single

chemotherapeutic agent, to a selected cancerous cell line can in fact confer


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
resistance, to other unrelated compounds. Salmon et al (1989) while investigating

multiple myeloma found that the frequency of Pgp expression increased after

chemotherapy (Fig 6 ).

(See Clinical Studies table 7).

The MDR1 gene is reported to be overexpressed in up to 50% of clinical tumour

specimens Goldstein et al (1989): Gottesman et al (1995) and the overexpression of

MDR1 transcripts, is associated with lack of treatment efficacy in a number of

cancers Baldini et al 1996; Chan et al 1990. Furthermore studies carried out by

Campos et al 1992; Pirker et al in the same year identified Pgp expression to be an

independent risk factor for treatment failure.

Despite the extensive cataloguing of Pgp expression in many cancer types a clear

relationship between Pgp detection and its implications on prognosis and response

to therapy is still debated. Poor design of clinical trials and a general lack of

consensus on detection methods, low patient numbers significantly hampered

progress Pgp understanding. In 1996 Beck et al attempted to standardise the

detection of Pgp expression making recommendations for future studies which

included:

(a) Although detection of Pgp and MDR1 is at present likely to be more reliable in

leukemias and lymphomas than in solid tumors, accurate measurement of low levels

of Pgp expression under most conditions remains an elusive goal;

(b) Tissue-specific controls, antibody controls, and standardized MDR cell lines are

essential for calibrating any detection method and for subsequent analyses of clinical

samples;

(c) Use of two or more vendor-standardized anti-Pgp antibody reagents that

recognize different epitope improves the reliability of immunological detection of Pgp;

(d) Sample fixation and antigen preservation must be carefully controlled;

(e) Multiparameter analysis is useful in clinical assays of MDR1/Pgp expression


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
(f) Immunostaining data are best reported as staining intensity and the percentage of

positive cells; and

(g) Arbitrary minimal cut-off points for analysis compromise the reliability of

conclusions

Beck et al (1996)
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Table 6: clinical studies implicating Pgp expression in cancer.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Figure 9: Cancers reprted to have high MDR1 expression post-chemotherapy

Gotetesman and Pastan et al 1991

PHARMACOLOGICAL TARGETING OF Pgp


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Drug resistance presents the greatest challenge to cancer treatment by

chemotherapy and is the main reason for failure of treatment Aouali et al (2005). One

of the first compounds to be identified as being able to circumvent resistance

(vincristine in particular) was the calcium channel blocker Verapamil . This was

followed up in 1986 by Slater et al in which cyclosporine was found to have

modulating ability on resistance. Early clinical trials Salmon et al (1991) on verapamil

assessed it clinical feasibility as a modulator. Salmon et al (1991) first evaluated the

resistance patterns from bone marrows from 59 myeloma patients and found that

verapamil was capable of sensitising myeloma cells exhibiting resistance to

doxorubicin and vincristine in vitro but did not enhance sensitivity of cells that were

drug sensitive (P>.001). Clinical trials were then conducted on 22 patients with

myeloma refractory to vincristine-adriamycin-dexamethasone (VAD) which was

administered with i.v verapamil. The clinical efficacy of the trials prompted the

supposition that clinical reversal of MDR could be achieved by verapamil. Cardiac

toxicity however was observed in some patients (fig 9), an unacceptable routine for

cancer treatment. Licht et al (1998)

Figure 9: Toxicities of VAD Plus High-Dose IV Verapamil in 22 Patients


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Salmon et al 1991
The use of verapamil, cyclosporine Yahanda et al (1992) and other 1stgeneration Pgp

inhibitors was plagued by an inability to reach sufficient plasma concentrations to

block Pgp activity and by clinically significant toxicity profiles. These findings

prompted research using available in vitro assays to investigate any common

pharmacophoric elements on substrates of


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Pgp. In 2003 Wang et al utilised quantitative structure activity relationships (QSAR)

to describe criteria that must be fulfilled in order for a substance to be a substrate or

modulator for Pgp. Their findings proposed that a Pgp modulator candidate should

have

i. a log P value of at least 2.92

ii. 18-atom-long molecular axis and

iii. a high Ehomo value

iv. at least 1 tertiary base nitrogen atom

(P version 4.0 QSAR software and HyperChem version 5.0 program) (Wang et al

2003)

Figure 10: 1st -3rd Generation Pgp Modulators

Mcdevitt and Callaghan et al 2007


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
The low binding affinities of the 1st generation Pgp inhibitors necessitated the use of

high doses subsequently leading to unacceptable toxicity. Despite the works of

Zamora et al 1988 and Wang et al 2003 the pharmacophoric parameters lacked the

needed stringency to facilitate drug development significantly. Moreover 1st

generation modulators had shown efficacy and thus they formed the template of

further drug development. 2nd generation Pgp modulators (Fig 10)

1ST -3RD GENERATION Pgp MODULATORS


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

For a compound to eligible as a modulator of Pgp it must fulfil at least one of the

following criteria:

i. Increase the potency of cytotoxic drugs in resistant cells

ii. Increase the accumulation of cytotoxic drugs intracellularly

iii. Interfere with photo affinity labelling of Pgp

As mentioned before (pharmacological targeting of Pgp) 1st generation modulators

resulted in high levels of toxicity and a major contributing factor to this was the low

binding affinities observed which prompted higher doses. The clinical application of

cyclosporine has been extensively investigated Yahanda et al (1992); Sonneveld et

al (1992).

Manetta et al 1993 conducted phase I trials in which the potential clinical use of

cyclosporine A modulation of cisplatin was investigated, they also set out to identify a

tolerable dose of Cyclosporin A when combined with a standard dose of cisplatin

(75mg/m2 ). Their sample size consisted of 20 patients with refractory gynaecologic

cancer received 81 courses of therapy.

20% of patients developed nephrotoxicity with 25% of the patients being partial (3

patients) and complete (2 patients) responders. Although there was evidence of

chemosensitising of MDR the overall results showed cyclosporine A achieves this

with considerable toxicity levels.

The toxicity of 1st generation chemosensitising agents prompted the development of

new compounds. Analogues of these agents where developed and these included

dexverapamil, dexniguldipine, PSC 833 and VX-710. Wilson et al 1995 reported 10-

fold decrease in cardiac toxicity at equal chemosensitising levels. The most

characterised of these compounds is PSC 833 a derivative of cyclosporine that was


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
reported Boekhorst et al 1992; Twentyman and Bleehen 1991 to inhibit Pgp by 10-

20fold greater activity. Many studies show it to have high reversing potency Boesch

et al 1991; Gaveriaux et al 1991. While findings by Coley et al 2000 on fresh tumour

material from patients with soft tissue sarcoma recorded modest on accumulation of

ancthracycline (20%) when given at 1nM. Animal models of 2nd Pgp generation

modulators show significant results Watanabe et al 1996 however clinical trials show

evidence of limited success. VX710 like PSC 833 has been extensively studied. It

directly interferes with the efflux of substances due to its affinity to the Pgp pump as

well as other related ABC transporters (namely MRP1). The coadministration of this

compound with chemotherapeutic agents has had limited success in the treatment of

refractory cancers. Gandhi et al 2007 conducted a phase II trial in order to evaluate

the efficacy of VX710 when co administered with doxorubicin and vincristine patients.

The patients suffered from inoperable, local advanced or metastatic sarcoma of the

soft tissue. In addition the patients had anthracycline-resistance. The study was on

36 patients who enrolled over a two year period and of varying demographic. VX710

was reported as no significantly enhancing anti-tumour activity or survival.

Neutropenia was also found to be the major toxicity occurring in 26/30 patients in

trials. There was however partial responders 7/36 patients as detected by radiology

however accurate calculation of response duration was hindered by censoring of the

results and a halt to the trials. Similarly earlier studies conducted by Bramwell et al

2002 revealed objective responses for the drug with disease stabilisation in partial

responders lasting duration of only 3-4 months.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Ghandi et al 2007

2nd generation inhibitors have a better pharmacologic profile than 1st generation

inhibitors however they also posses similar characteristics which limits their chemical

profile. By limiting the metabolism and clearance of chemotherapeutic agents they

facilitate the incidence of unacceptable toxicity that necessitates the reduction in

doses administered within the trials.

Cytochrome P450 enzymes are also induced along with ABC transporters leading to

suppositions that regulatory elements of these genes overlap Lum and Gosland

1995. The P450 3A4 isoenyme shares many substrates with Pgp thus substances

affected by MDR development are also liable to metabolism by P450 3A4 (PSC 833

and VX710) and thus results in many of the reported unpredictable pharmacokinetic

interactions. These agents inhibit the P450 3A4 mediated metabolism and is the

main cause of toxicity associated with modulators off Pgp. Dose reductions had to be

implemented by researchers due to safety however achieving therapy at a safer

dosing regime limits the efficacy of many of these 2nd generation modulators.

Furthermore intrinsic activity of some 2nd generation modulators (VX710, PSC 833)

suggests they have affinities for other ABC transporters as well as Pgp and this

inhibitory activity of non-target transporters may contribute to the limited efficacy and

raised toxicity of these agents Yanagisawa et al 1999;Rowinsky et al 1998.

3rd generation Pgp modulators (fig 6 pharmacological targeting of Pgp) specifically

and potently inhibit Pgp function. Their development occurred as a result of structure

activity relationships and combinational chemistry Thomas and Coley 2003. At

relevant concentrations they do not show activity at P450 3A4 enzymes Dantzig et al

1999 where the selectivity of LY335979’s selectivity for Pgp was evaluated as well as

its effects of P450 activities. It was found to have a significantly lower affinity for
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
CYP3A than for Pgp. Furthermore it was characterised as a potent modulator of Pgp

and not other members within the ABC transporter family (namely MPR1 and MPR2).

Other 3rd generation modulators; XR9576, R101933,ONT-093,GF120918 are of

varying chemical structures but are common in their level of potency on Pgp. This

has also been observed in clinical trials as well the most promising agent of

3rdgeneration inhibitors is XR9576 which is believed to achieve it modulation by

binding to the ATP binding sites of Pgp. Martin et al 1996; roe et al 1999 findings

identified XR9576 as a potent inhibitor of Pgp in vivo and in vivo. Martin et al 1999

used drug resistant CHO ovary cell lines and demonstrated XR9576 showing greater

selectivity, duration of inhibition and potency of interaction at Pgp than with any other

reported modulators.

Furthermore it was shown to increase accumulation of [H3]-Vinblastine and [3H]-

paclitaxel transport as high as in cell lines that were not overexpressing Pgp

(EC50=487+50 nM). By binding to the ATP sites means that Xr9576 like most 3rd

generation modulators are not substrates themselves. This induces conformational

changes which prevent hydrolysis of ATP a requirement in the efflux mechanism

MODULATION OF Pgp; A GENETIC APPROACH

Molecular strategies directed at modulating Pgp activity have focused on interfering

with the synthesis of Pgp. This has brought into use antisense oligonuleotides,

ribozymes and protein that regulate differentiation of cancer cells Corrias et al 1992;

Efferth et al 1993. These strategies focus on cleavage of MDR1 mRNA and have the

ability to restore chemosensitivity in MDR cells Kobayashi et al 1999. They

developed 2 anti-MDR1 hammerhead ribozymes driven by the beta-lactin promoter.

They transduced these ribozymes into MDR1 expressing cells which were designed
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
to target specific predetermined sites (fig11) in addition a retroviral vector containing

polymerase III promoter was used to improve ribozyme activity.

Figure 11: MDR1 gene and anti-MDR1 hammerhead ribozymes

Kobayashi et al 1999

Human leukaemia cell lines (MOLT-3 where MDR subline used was MOLT-3/TMQ800)

where used and these were co-cultured with virus producer cells. Ribozyme efficacy

was then determined by G418 selection and ribozyme transduced cells became

vincristine-sensitive. Original MOLT-3/TMQ800 cells(conrol) were reported to be 600-

fold more resistant to vincristine whereas stably transduced cells showed 92 to 296-

fold resistance. Partial restoration of vincristine sensitivity and doxorubicin activity

was accompanied by decreased expression MDR1, decrease in Pgp amount and

function (fig 10). Kobayashi et al (1999)

iMDR1-sRz one of the ribozymes used targeted the translation-initiation site and

pooled transformants that occurred as a resulted of its introduction to original MOLT-

3/TMQ800 cells exhibited 10% of MDR1 mRNA compared to controls.

Further studies by Scanlon et al (1993) complimented these findings and revealed

that in addition to MDR1-directed ribozymes, c-fos interacting ribozymes could also

decrease expression of Pgp. Hammerhead ribozymes designed to cleave c-fos


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
mRNA cloned into pMAMneo were transfected into drug sensitive human ovarian

carcinoma (A2780) cell lines. Cells exhibiting MDR also had elevated levels of the c-

fos furthermore cells transfected with c-fos also exhibited MDR and the anti-fos

ribozyme reversed the MDR phenotype in A2780AD cells. These studies reveal

potential pharmacological targets in particular Fos which was previously believed to

play a role in resistance to agents not within the MDR family such as cisplatin, AZT

and 5-Flourouracil Scanlon et al (1993). Transcription factors are involved in the

regulation of expression in drug resistance genes and thus appear to be suitable

targets for strategies to circumvent drug resistance.

STEM CELLS AND MDR1 EXPRESSION


The prospect of transferring drug resistance genes to chemosensitive cells like stem

cells has been explored to protect them from the adverse effects of chemotherapy.

The drug resistance genes may serve as selectable markers in vivo facilitating gene

expression in transduced cells after exposure to drugs.

Licht et al (1998) transferred MDR1 cDNA to mouse bone marrow cells that lacked

expression of lineage –specific antigens nor MHC II antigen Ia. These cell had high

expression of Ly6A/E. Isolated cells were expanded ex vivo using growth factors.

Gene transfer was achieved via coculture containing retrovirus. Functional Pgp was

detected in 60% of expanded cells. The recipient animals expressed Pgp in high

proportion compared to those observed in MDR cancers in the clinic.

In 1994 Ward et al conducted research using human CD34+ progenitor cells a

reflection of the feasibility of gene transfer to stem cells as demonstrated in previous

animal models. In normal and also in tumour tissue MDR expression varies greatly
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
whereas in bone marrow cells it remains consistently low. This renders stem cell

particularly sensitive to many chemotherapeutic agents; taxol, anthracylcines,

etoposide, vinca alkaloids all of which are effect against many cancer forms.

They reported the successful transfer and expression of the human MDR1into

human bone marrow cells (via amphotropic retroviral supernatant) and the resistance

to taxol exposure gained thereafter. PCR analysis of transduced, colchicine resistant,

amphotoric cells indicated 17/20 clones possessing th MDR cDNA

The main MDR producer A12M1 had the highest titer (5 x 1014 viral particles/ mL).

FCAS analysis showed a 1to 2 log increase in MDR expression compared t

untransduced cells.

Taxol was introduced to MDR-transduced cells and their level of MDR expression

compared to non-transduced cells after exposure to 1 x 10-6 mol/L on day 10 post

transduction. Analysis took place on day 12 and 12.5% of cells showed increase in

Pgp compared to non-transduced cells.

The levels of taxol resistance achieved by MDR-transduced cells and non-

transduced cells and preferential survival was observed in transduced cells at doses

of 5 x 10-8 mol/L. This ability to select for progenitor cells resistant to taxol bears

significant perspective on its potential use in providing drug resistant marrow cells for

patients undergoing cancer therapy. Alternatively it could be useful in enriching

marrow populations of cells with the MDR gene and a non-selectable gene (e.g beta-

globin) Ward et al (1994).

Be it promising the first published clinical trials conducted on MDR1 levels reported

to be disappointingly low. Hanania et al (1996). It is believed that low and unstable

expression of these transferred genes is the major cause for failure of gene therapy

Licht et al (1998). As a result trials conducting high-dose chemotherapy with

autologous hematopoietic stem cell transplantation report insufficient function of

reconstituted bone marrow limiting the efficacy of transplantation chemotherapy.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
Efforts by Takahashi et al 2007 (Fig 12) to increase transduction efficiency of MDR1

gene transfer yielded limited success.

Figure 12: MDR gene therapy

Takahashi et al 2007

Peripheral blood was stimulated using 5 cytokines; SCF, TPO, IL-6, FL-Ligand & sIL-

6r. They achieved transduction efficiencies of 8-17% by Pgp expression.

Transplantation of induced peripheral blood stem cells (PBSC) (1/3) and untreated

PBSC (2/3) yielded 3-5% increases in the ratio of peripheral white blood cells which

corresponded to the ratio of induced MDR1-tranduced CD34+ cells. However despite

this after 6 months Pgp expressing cells decreased to an undetectable level. These

limited successes in clinical scenarios may indicate low survival chances of MDR1-

transduced stem cells in the bone marrow or may lie in transduction strategies which

still seem to lack sufficient efficiency.


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Discussion/ Conclusion
Extensive studies have been carried out on the MDR gene and it gene product P-

glycoprotein. It has been characterised as a ATP-dependant efflux protein whose

main role is the expulsion of toxic substances that enter the cells of the body.

The use of radiolabelled dyes monoclonal antibodies have elucidated its distribution

within the body’s organs leading to researchers suggesting evolutionary significance

pertaining to its protective characteristics. This is further substantiated by it

occurrence in many different species.

More than 50% of cancer relapses show an overexpression of MDR1 gene as a

result of conventional chemotherapy. Furthermore the increased expression of Pgp

has been associated with multidrug resistance to a number of unrelated

chemotherapeutic agents.

This greater understanding of Pgp gave rise to many pharmacological studies aimed

at modulating Pgp function as a means of increasing drug accumulation of

chemotherapeutic agents. As a result drug development on Pgp inhibitors advanced

rapidly. Many agents proved promising however clinical trials yielded limited

successes. A significant trend in these studies was that although Pgp modulators

have the potential to increase drug accumulation of cytotoxic agents a significant

number of studies resulted in toxicities associated with this increase accumulation of

drug. Alterations of doses and more potent drugs were designed slightly increasing

the number of responders to therapy.

Research has generally shifted to stem cell biology and gene therapy and significant

progress has been made since the early days of introducing foreign genes into

murine cells via retroviral vectors. Gene therapy directed at transferring resistance to

stem cell as a means of protection from chemotherapy was anticipated to


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
revolutionize cancer treatment drastically however optimism in these ambitions have

rapidly declined. This largely due to the limited positive results obtained in clinical

trails to a variety of gene therapy approaches in which low efficiency was reported.

P-glycoprotein’s significance in cancer relates to multidrug resistance and its

overexpression leads to the failure of many conventional chemotherapeutic

strategies. Despite advances in pharmacotherapy and stem cell research many

challenges lie ahead in the translation of recent advances into reproducible clinical

benefit.

A major factor to be considered is that resistance to chemotherapeutics is

multicausative. A number of genes have been identified that are associated with

chemoresistance and despite strategies having being developed to circumvent Pgp

mediated resistance in cancer various other approaches involving other genes are

still at a preclinical stage.

Any major advances in Pgp modulation or application will have to encompass other

related ABC transporters and other factors contributing to the MDR phenomenon this

project recognises the feasibility of pharmacological Pgp modulation and its

application in gene therapy which is still to be validated as a means of cancer

therapy.
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

REFERENCES
Ambudkar S V, Lelong I H and Zhang J P. (1992). Partial purification

and reconstitution of tthe human multidrug-resistance pump-

Characterisation of drug stimulable ATP hydrolysis. Proc Natl Acad Sci

USA; 89: 8472-8476

Alan Milburn. (2000). The NHS cancer plan: A plan for investment A

plan for reform. www.doh.gov.uk/cancer

Aouali Nassera, Eddebra Lahcen, Macadre Jerome and Morjani Hamid.

(2005). Immunosuppressors and reversion of multidrug-resistance.

Critical Reviews in Oncology/Hematology; 56: 61-70

Baird R D and Kaye S B. (2003). Drug resistance reversal-are we

getting closer? European Journal of Cancer; 39: 2450-2461

Baldini Nicola, Scotlandi Katia, Barbanti-Brodano, Manara Maria

Cristina, Maurici Daniela, Bacci Gaetano, Bertoni Franco, Picci Piero,

Sottili Sandra, Campanacci Mario and Massimo Serra. (1995).

Expression of P-Glycoprotein in High-Grade Osteosarcomas in

Relation to Clinical Outcome. The New England Journal Of Medicine;

333: 1380-1385

Bates Susan E, Mickley Lyn A, Chen Yi-Nan, Richert Nancy, Rudick

Jonathan, Biedler June L and Fojo Antonio T. (1989). Expression of

Drug Resistance Gene in Human Neuroblastoma Cell Lines:

Modulation by Retinoic Acid-Induced Differentiation. Molecular and

Cellualr Biology; 9: 4337-4344


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Bech-Hansen N T, Till J E and Ling V.(1975). Pleiotropic Phenotype of

Colchicine-Resistant CHO Cells Cross-Resistance and Collateral

Sensitivity. J. Cell, Pysiol; 88: 23-32

Beck W T, Grogan T M, William C L, Cordon-Cardo C, Parham D M and

Kuttesch J F. (1996). Methods to detect P-glycoprotein-associated

multidrug resistance in patients’s tumors: Consensus

recommendations. Cancer Research; 56: 3010-3020

Bertino Joseph R.( 2008).Transfer of drug resistance into

Hematopoietic Stem Cells for Marrow Protection. The

Oncologist;13:1036-1042

Blackburn Elizabeth, Druker Brian, Hartwell Leland, King Mary-Claire,

Weinberg Robert. (2009). Rediscovering Biology: Cell Biology and

Cancer. www.learner.org/courses/biology/index.html

Bodine DM, seidel NE, Gale MS, Nienhuis AW and Orlic D.( 1994).

Efficient retrovirus transduction of mouse pluripotent hematopoietic

stem cells mobilized into peripheral blood by treatment with granulocyte

colony-stimulating factor and stem cell factor. Blood; 84:1482-1491

Bramwell Vivien H C, Morris Donald, Ernst Scott D, Hings Ingrid,

Blackstein Martin, Venner Peter M, Ette Ene I, Harding Matthew W,

Waxman Allison and Demetri George D. (2002). Safety and Efficacy of

the Multidrug-Resistance Inhibitor Biricodar (VX-710) with Concurrent

Doxorubicin in Patients with Anthracycline-resistant Advanced Soft

Tissue Sarcoma. Clinical Cancer Research; 8: 383-393


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Bruggemann E P, Currier S J, Gottesman M M, Pastan I. (1992).

Characterisation of the azidopine and vinblastine binding site of P-

glycoprotein. J Biol Chem; 264: 15483-15488

Campos L, Guyotat D and Achimbaud E. (1992). Clinical significance of

multidrug resistance P-glycoprotein expression on acute

nonlymphoblastic leukemia cells at diagnosis. Blood; 79: 473-476

Cancer Research UK. (2009). Cancer Stats Key Facts, All cancers.

info.cancerresearchuk.org/cancerstats/incidence/

http://www.cancerhelp.org.uk/help/default.asp?page=91

Chaudhary P M, Mechetner E B and Roninson I B. (1992) Expression

and activity of the multidrug resistance P-glycoprotein in human

peripheral blood lymphocytes. Blood Journal; 80: 2735-2739

Chan H S L, Thorner P S and Haddad G. (1990). Immunohistochemical

detection of P-glycoprotein. Prognostic correlation in soft tissue

sarcoma of childhood. Journal of Clinical Oncology; 8: 689-704

Coley H M, Verrill M W, Gregson S E, Odell D E, Fisher C and Judson I

R. (2000). Incidence of P-glycoprotein overexpression and multidrug

resistance (MDR) reversal in adult soft tissue sarcoma. European

Journal of Cancer; 36: 881-888

Cordon-Cardo C, O’brien J P, Boccia J, Casals D, Bertino J R and

Melamed M R. (1990). Expression of the Multidrug resistance Gene

Product (P-Glycoprotein) in Human Normal and Tumor Tissues. The

Journal of Histochemistry and Cytochemistry; 38: 1277-1287


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Cordon-Cardo Carlos, O’brien James P, Casals Dolors, Rittman-Grauer

Lana, Biedler June L, Melamed Myron R and Bertino Joseph R. (1989).

Multidrug-resistance gene (P-glycoprotein) is expressed by endothelial

cells at blood-brain barrier sites. Medical Sciences; 86: 695-698

Corrias M V and Tonini G P. (1992). An oligomer complimentary to the

5’ region of the MDR1 gene decreases resistance to doxorubicin of

human ardenocrcinoma multidrug resistance cells. Anticancer

Research; 12: 1431-1438

Cornwell M M, Safa A R and Felsted R L. (1986). Membrane vesicle

from multidrug resistant human cancer cells contain a specific 150-170

kDa protein detected by photoaffinity labelling. Proc Natl Acad Sci USA;

83: 3847-3890

Dantzig A H, Shepard R L, Law K L, Tabas L, Pratt S, Gillespies J S,

Binkley S N, Kuhfel M T, Starling J J and Wrighton S A. (1999) .

Selectivity of the Multidrug Resistance Modulator, LY335979, for P-

Glycoprotein and Effect on Cytochrome P-450 Activities. The journal of

Pharmacology and Experimental Theraputics; 290: 554-562

Debenham Paul G, Kartner Norboert, Siminovitch Louis, Riordan John

R and Ling Victor. (1982). DNA-Mediated Transfer of Multiple Drug


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Resistance and Plasma Membrane Glycoprotein Expression. Molecular

and Cell Biology; 2: 881-889

Efferth T and Volm M. (1993). Modulation of P-glycoprotein mediated

multidrug resistance by monoclonal antibodies, immunotoxins or

antisense oligodeoxynucleotides in kidney carcinoma and normal

kidney cells. Oncology; 50: 303-308

Endicott Jane A and Ling Victor.(1989) The Biochemistry of P-

Glycoprotein-Mediated Multidrug Resistance. Annual Reviews

Biochemistry; 58: 137-171

Fenneteau Frederique, Turgeon Jacques, Couture Lucie, Michaud

Veronique, Li Jun, Nekka Fahima. (2009). Assesing drug distribution in

tissues expressing P-glycoprotein through physiologically based

pharmacokinetics modelling: model structure and parameters

determination. Theoretical Biology and Medical Modelling; 6: 1742-

4682

Fromm Martin F. (2002). Genetically determined differences in P-

glycoprotein function: implications for disease risk. Toxicology;181-

182:299-303

Frredenberg William R, Salzaman Sherry A, Phan Sonja M and

Burmester James K. (1999). Transforming growth factor-β and


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

multidrug resistance in chronic lymphocytic leukemia. Medical

Oncology;16:110-118

Ghandhi Leena, Harding Matthew W, Neubauer Marcus, Langer Corey

L, Moore Melvin, Ross Helen J, Johnson Bruce E and Lynch Thomas J.

(2007). A Phase II Study of the Safety and Efficacy of the Multidrug

Resistant Inhibitor VX-710 Combined With Doxorubicin and Vincristine

in Patients With Recurrent Small Cell Lung Cancer. American Cancer

Society; 109: 5

Gibbs Wayt W. (2003). Untangling the roots of cancer. Scientific

American.

Gottesman Michael M. (1993). How Cancer Cells Evade

Chemotherapy: Sixteenth Richard and Hinda Rosenthal Foundation

Award Lecture. Cancer Research; 53: 747-754

Gottesman Michael M. (2009). Learning some new tricks from a

multidrug transporter. National Health Institute;

Gottesman Michael M and Ling Victor. (2006). The molecular basis of

multidrug resistance in cancer: The early years of P-glycoprotein

research. FEBS Letters; 580: 998-1009

Goldstein L, Galsi H and Fojo A. (1989). Expression of multidrug

resistance gene in human cancers. J Natl Cancer Inst; 81: 116-124

Hanania E G, Giles R E, Kavanagh J, Ellerson D, Zu Z, Wang T , Su Y,

Kudelka A, Rahman Z, Holmes F, Hortobagyi G, Claxton D, Bachier P,

Thall P, Cheng S, Hester J, Ostrove J M, Bird R E, Chang A Korbling M,

Seong D, Cote R, Holzmayer T, Merchetner E, Heimfeld S, Berenson


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

R, Burtness B, Edwards C, Bast R, Andreeff M, Champlin R and

Deisseroth A B. (1996). Results of MDR-1 vector modification trial

indicate that granulocyte/macrophage colony-forming unit cells do not

contribute to post transplant hematopoietic recovery following intensive

systemic therapy. Medical Sciences; 93: 15346-15351

Israeli David, Ziaei Simin, Gonin Patrick and Garcia Luis. (2005). A

proposal for the physiological significance of mdr1 and bcrp1/abcg2

gene expression in normal tissue regeneration and after cancer

therapy. Journal of Theoretical Biology; 232: 41-45

Jones M E, Thomas S M and Rogers A. (1993). Luria-Delbruck

Fluctuation Experiments: Design and Analysis. Genetics Society of

America*

Jones Craig V. (2006). Tamoxifen (ICI46,474) as a targeted therapy to

treat and prevent breast cancer. British Journal of Pharmacology; 147:

269-276

Juliano R L, Ling V. (1976). A suface glycoprotein modulating drug

permeability in Chinese hamster ovary cell mutants. Biochim Biophys

Acta; 455: 152-162


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Juliano R L. (1978). Adhesion and Detachment Characteristics of

Chinese Hamster Cell Membrane Mutants. Journal of Cell Biology; 76:

43-49

Kang Hyumin, Fisher Michael, Xu Dong, Miyamoto Yuko J, Marchand

Arnaud, Van Aerchot Arthur, Herdewijn Piet and Juliano Rudolph L.

(2004). Inhibition of MDR1 gene expression by chimeric HNA antisense

oligonucleotides. Nucleic Acids Research; 32: 441-4419

Kessel David and Bosmann Bruce. (1970). On the Characteristics of

Actinomycin D Resistance in L5178Y Cells. Cancer Research; 30:

2695-2701

Kobayashi Hiroyuki, Takemura Yuzuru, Wang Fu-Sheng, Oka Tetsuo,

Ohnuma Takao.(1999). Retrovirus-mediated transfer of anti-MDR1

hammerhead ribozymes into multidrug-resistant human leukemia cells:

screening for effective target sites. Int. J. Cancer; 81: 944-950

Kuroda Yukiaki. (1974). Mutagenesis in cultured human diploid cells I.

Effects of some mutagens and a selective agent on cell survival. Japan

Journal Of Genetics; 49: 379-387

Kuwazuru Yasuo, Yoshimura Akihiko, Hanada Shuichi, tsunomiyaAtae

U, Makino Torahiko, Ishibashi Kazuaki, Kodama Masahiko, Iwahashi

Masato, Arima Terukatsu, and Akiyama Shin-Ichi. Expression of the

Multidrug Transporter P-glycoprotein, in Acute Leukemia Cells and

Correlation to Clinical Drug Resistance. Cancer; 66: 868-873


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Kuo Macus Tien, Bednarski Patrick, Kohno Kimitoshi, Salerno Milena,

Tarasuik Jolanta and Zunino Franco. (2009). Redox Regulation of

Multidrug Resistance in Cancer Chemotherapy. Antioxid Redox Signal;

1: 99-134

Lagas Jurjen S, Sparidans Rolf W, van Waterschoot Robert A B,

Wagenaar Els, Beijen Jos H and Schinkel Alfred H. (2008). P-

Glycoprotein Limits Oral Availabity, Brain Penetration and Toxicity of an

Anionic Drug, the Antibiotic Salinomycin. Antimicrobial Angents and

Chemotherapy; 52: 1034-1039

Lee Kyung-Yung, Prak Jae-Gahb, Gazdar Adi F, Goldstein Lori J,

Hwang Ee-Sook and Kim Jin-Pok. (1990). Correlation between MDR1

Gene Expression and In Vitro Drug Sensitivity Testing (DST) of Human

Cancer Cell Lines. Journal of Korean Cancer Association;

Li Yan-Hong, Wang Yong-Hua, Li Yan, Yang Ling. (2006). Gene

Polymorphisms and Clinical Relevance. Acta Genetica Sinca; 33: 93-

104

Li Qing-Quan, Wang Wen-Juan, Xu Jing-Da, Cao Xi-Xi, Chen Qi, Yang

Jin-Ming and Xu Zu-De. (2007). Involvement of CD147 in regulation of

multidrug resistance to Pgp substrate drugs and in vitro invasion in

breast cancer cells. Cancer Sci; 98: 1064-1069

Licht Thomas, Gottesman Michael M, Pastan Ira. (1998). Clinical

applications of gene therapy in cancer: modification of sensitivity to

therapeutic agents. Stem cell biology and gene therapy


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Ling Victor and Thompson Larry H. (1974). Reduced permeability in

CHO cells as a mechanism of resistance to colchicine. J. Cell. Physiol;

83: 103-116

Loscher Wolfgang and Potschka Heidrum. (2005). Progress in

Neurobiology; 76: 22-76

Lown Kenneth S, Mayo Robert R, Leichtman Alan B, Hsiao Hsiu-ling,

Turgeon Kim, Schmiedlin Phyllissa, Brown Morton B, Guo Wesheng

and Rossi Stephan J. (1997). Role of intestinal P-glycoprotein (mdr1) in

interpatient variation in the oral bioavailability of cyclosporine. Clinical

Pharmacology Therapy; 62: 248-260

Lum Bert L, Fisher George A, Brophy Nathalie A, Yahanda Anne M,

Adler Kent M, Kaubisch Sonja, Halsey Joanne R N and Sikic Branimir I.

(1993). Clinical Trials of Modulation of Multidrug Resistance. Cancer

Supplement; 72: 11

Martin Catherine, Berridge Georgina, Mistry Prakash, Higgins

Chistopher, Charlton Peter, Callaghan Richard. (1999). The molecular

interaction of the high affinity reversal agent XR9576 with P-

glycoprotein. British Journal of Pharmacology; 128: 403-411

Manetta Alberto, Boyle Julie, Berman Michael L, Disaia Philip J, Lentz

Samuel, Yuan Lia Shu, Mutch David, Slater Lewis. Cyclosporin

Enhancement of Cisplatin Chemotherapy in Patients with Refractory

Gynecologic Cancer. Cancer; 73: 1

Marcus Menashe, Fainsod Abraham and Diamond Gill. (1985). The

Genetic Analysis of Mammalian Cell-Cycle Mutants. Annual Review of

Genetics; 19: 389-421


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

McDevitt Christopher A and Callaghan Richard. (2007). How can we

best use structural information on P-glycoprotein to design inhibitors?

Pharmacology & Therapeutics; 113: 429-441

Moscow Jeffrey A and Cowan Kenneth H. (1988). Multidrug

Resistance. Journal of the National Cancer Institute; 80: 14-20

National Institutes of Health. (1999). NIH Publication No. 99-4646

Pastan Ira and Gottesman. (1991) Multidrug Resistance. Annual

Reviews in Medicine; 42: 277-86

Pirker R, Wallner J and Gotzl M. (1992). MDR1 RNA expression is an

independent prognostic factor in acute myeloid leukemia. Blood; 80:

557-559

Riehm Hansjorg and Biedler June L. (1971). Cellular Resistance to

Daunomycin in Chinese Hamster Cells in Vitro. Cancer Research; 31:

409-412

Roe Michael, Folkes Adrian, Ashworth Philip, Brumwell Julie, Chima

Lal, Hunjan Sukhjit, Pretswell Ian, Dangerfield Wendy, Ryder Hamish

and Charlton Peter. (1999).Reversal of P-Glycoprotein mediated

Multidrug Resistance by Novel Anthranilamide Derivatives. Bioorganic

& Medicinal Chemistry Letters; 9: 595-600

Roylance Rebecca. (2007) Basis of medical cancer therapy.

www.smd.qmul.ac.uk/morbidanatomy/medica~1.ppt

Saad Maha, Garbuzenko Olga B and Minko Tamara. (2008). Co-

delivery of siRNA and an Anticancer Drug for Treatment of Multidrug-

resistant Cancer. Nanomedicine; 6: 761-776


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Salmon S E, Dalton W S, Grogan T M, Plezia P, Lenhert M, Roe D J

and Miller T P. (1991). Multidrug-resistant myeloma: laboratory and

clinical effects of verapamil as a chemosensitizer. Blood Journal; 78:

44-50

Scanlon Kevin J, Ishida Hironori, Kashani-Sabet Mohammed.(1994).

Ribozyme-mediated reversal of the multidrug-resistant phenotype.

Biochemistry; 91: 11123-11127

Schinkel Alfred H and Jonker Johan W. (2003). Mammalian drug efflux

traansporters of the ATP binding cassette (ABC) family: an overview.

Advanced Drug Delivery Reviews; 55: 3-29

Schinkel Alfred H, Wagenaar Els, van Deemter Liesbeth, Mol Carla A A

M and Borst Piet. (1995). Absence of the mdr1a P-Glycoprotein in Mice

Affects Tissue Distribution and Pharmacokinetics of Dexamethasone,

Digoxin and Cyclosporin A. Journal of Clinical Investigations; 96: 1698-

1705

Schinkel Alfred H, Kemp Stephan, Dolle Martijn, Rudenko Gabrielle and

Wagnaar Els. (1993). N-glycosilation and Deletion Mutants of the

Human MDR1 P-glycoprotein. The Journal of Biological Chemistry;

268: 7474-7481

Scripture Charity D and Figg William D. (2006). Drug Interactions in

Cancer Therapy. Nat Rev Cancer; 7:546-558

Seeger Markus A and van Veen Hendrik. (2008). Molecular basis of multidrug

transport by ABC transporters. Biochimica et Biophysica Acta; uncorrected

version
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Sheps Jonathan A and Ling Victor. (2007). Perface: the concept and

consequences of multidrug resistance. European Journal of

Physiology; 453: 545-553

Sonneveld P, Suciu S, Weijermans P, Beksac R, Neuwirtova G

Lokhorst H, Van Der Lelie J, Dohner H, Gerhartz H, Sergeren M,

Willemze R and Lowenberg B. (2001). Cyclosporin A combined with

vincristine, doxorubicin and dexamethasone (VAD) compared with VAD

alone in payients with advanced refractory multiple myeloma: an

EORTC-HOVON randomised phase III study (06914). British Journal of

Haematology; 115: 985-902

Sotomayor Maria G, Yu Hua, Antonia Scott, Sotomayor Eduardo M and

Pardoll Drew M.( 2002). Advances in Gene Therapy for Malignant

Melanoma. Cancer Control; 9: 1

Stein U, Shoemaker R H and Schlag P M. (1996). MDR1 Gene

Expression: Evaluation of its Use as a Molecular Marker for Prognosis

and Chemotherapy of Bone and Soft Tissue Sarcomas. European

Journal of Cancer; 32: 86-92

Styczynnski Jan and Wysocki Mariusz. (2007). Cyclosporine increases

in vitro sensitivity to prednisolone in childhood acute lymphoblastic

leukemia. Cancer Therapy; 5: 213-218

Szakacs Gergely, Varadi Andras, Ozvegy-Laczka and Sarkadi Balazs.

(2008). The role of ABC transporters in drug absorption, distribution,

metabolism, excretion and toxicity (ADME-Tox). Drug Discovery Today;

13: 1359-6446
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

Takahashi Shunji, Aiba Keisuke, Ito Yoshinori, Hatake Kiyohiko, Nakane

Minoru, Kakayashi Takayuuki, Minowa Sayuri, Shibata Harumi,

Mitsuhashi Junko, Tsukahara Satomi, Ishikawa Estuko, Suzuki Rieko,

Tsuruo Takashi and Sugimoto Yoshikazu. (2007). Pilot study of MDR1

gene transfer into hematopoietic stem cells and chemoprotection in

metastatic breast cancer patients. Cancer Sci;98:1069-1616

Thiebaut Franz, Tsuruo Takashi, Hamada Hirofumi, Gottesman Michael

M, Pastan Ira and Willingham Mark C. (1987). Cellular localization of

the multidrug-resistance gene product P-glycoprotein in normal human

tissues. Medical Sciences; 84: 7735-7738

Thomas Hilary and Coley Helen M.(2003). Overcoming Multidrug

resistance in Cancer: An Update on the Cinical Strategy of Inhibiting P-

Glycoprotein. Cancer Control; 10: 2

Trivedi Meghana, Budihardjo Imawati, Loureiro Kirsten, Reid Tony R

and Ma Joseph D. (2008). Epothilones: A Novel Class of Microtuble-

Stabalizing Drugs for the Treatment of Cancer. Future Oncology; 4:

483-500

Visvader Jane E and Lindeman Geoffrey J. (2008). Cancer Stem Cells

in Solid Tumours: Accumulating Evidence and Unresolved Questions.

Nature Review Cancer; 10: 755-768

Wang R B, Kuo C L, Lien L L and Lien E J. (2003). Structure-activity

relationships: analyses of p-glycoprotein substrates and inhibitors.

Journal of Clinical Pharmacy and Therapeutics; 28: 203-228

Ward M, Richardson C, Pioli P, Smith L, Podda S, Goff S, Hesdorffer C

and Bank A. (1994).Transfer and expression of the human multiple


THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

drug resistance gene in human CD34+ cells. Blood Journal; 84: 1408-

1414

Watanabe Toru, Naito Mikihiko, Oh-hara Tomoko, Itoh Yohjiro, Cohen

Dalia and Tsuruo Takashi. (1996). Modulation of Multidrug Resistance

by SDZ PSC 833 in leukemic and Solid-tumor-bearing Mouse Models.

Jpn. J. Cancer Res; 87: 184-193

World Health Organisation. (2009). Cancer. Factsheet No.297.

www.who.int/mediacentre/factsheets/fs297/en.hmtl

Yahanda A M, Adler K M and Fisher G A. (1992). Phase I trial of

etopiside with cyclosporine as a modulator of multidrugresistance.

Journal Clinical Oncology; 10: 1624-1634

Acknowledgements
THE BIOLOGICAL AND CLINICAL SIGNIFICANCE OF P-GLYCOPROTEIN IN CANCER

I would like to thank Dr. Helmout Modjtahedi For all his assistance. He

managed to create a positive environment conducive to successful

completion of this poroject. Many thanks.

You might also like