http://www.ijpsr.com/V3I9/16%20Vol.%203%20(9),%20Sept.%202012,IJPSR-1623,Paper%2016.pdf
BIOMARKERS FOR THERAPEUTIC RESPONSE IN
CERVICAL CANCER
- INTRODUCTION
Cervical cancer is the most common gynecologic
malignancy in the world, and the second most frequently diagnosed cancer in
women worldwide. Hence it continues to present a significant challenge to the health care community with approximately
471, 000 new cases diagnosed each year [1]. The growing risk of cervical cancer
in women in India (aged 0-64 years) is 2.4% compared to 1.3% for the world [2].
As like any other cancers, stage of diagnosis is the most
important prognostic factor in cervical cancer. Women diagnosed with early
stage disease (IA-IB1) have survival rate of 85-100%. Stage IIA
tumors also have a good prognosis with a 5 year survival of approximately 90%.
However once the tumor spreads further, the prognosis is poor. About 35%
of women with cervical cancer will have persistent or recurrent disease [3].
Recurrences usually develop in the first 2 years after initial treatment. Hence
close monitoring is recommended in patients receiving any type of treatment [4].
Understanding the molecular basis of the biochemical
pathways involved in cervical cancer can facilitate the integration of
diagnosis, anticancer drug discovery, therapy for cancer and its
monitoring. Prognostication and the variability of tumor responses to
radio-/chemo-therapeutic agents is a topic of major interest in current cancer
research. A biomarker might be either a molecule secreted by tumor itself,
or it can be a specific response of the body to the presence of cancer.
Genetic, epigenetic, proteomic, glycomic, and imaging biomarkers can be used
for cancer diagnosis as well as prognosis. As an important biological
indicator of cancer status, progression and for the physiological state of the
cell at a specific time, biomarkers represent powerful tools for monitoring the
course of cancer and gauging the efficacy and safety of novel therapeutic
agents. They can have tremendous therapeutic impact in clinical oncology,
especially if the biomarker is detected before clinical symptoms or enable
real-time monitoring of drug response. The challenge in the field of
biomarkers is to develop methodology that will allow us to use the available
knowledge and tools to evaluate the usefulness of molecular abnormalities for
improving individual and population outcomes for cervical cancer [5].
Since cancer recurrence is the most common treatment
failure in patients with advanced tumor, increasing knowledge about new
biologic markers and better ability to predict risk of cancer recurrence is
very important for construction of more effective treatment
strategies. Biomarkers help to identify genetic variations or mutations as
well as changes in gene or protein expression or activity that can be linked to
a disease state or a response to a medical intervention [6].
The biomarkers, which indicate the response of cancer
patients to various therapy, predict response to particular therapies and
choose the drug that, most likely to yield a favorable response in a given
patient, Identify patients with a high probability of adverse effects of a
treatment and determine whether a therapy is having the intended effect on a
disease and whether adverse effects arise [7]. In this literature study we
have done a survey of the identified biomarkers.
Biomarkers can be either molecular biomarkers or imaging
biomarkers. Prognostic markers separate patients with good and bad prognosis to
aid the decision about how aggressive the therapy needs to be. It also provides
clues for the possible mechanism(s) responsible for the poor prognosis which
helps in identification of new targets for therapy. A predictive marker on the
other hand, identifies how likely an individual is to respond to a particular
therapy before initiation of treatment [8].
Prior to discussing individual biomarkers utilized, it is
important to note how the performance of biomarkers is evaluated. Biomarker
test performance is characterized by measures of sensitivity, specificity,
positive predictive value (PPV) and negative predictive value (NPV) with the
area under receiver operating characteristic curve (AUC). Detection of
biomarkers, either individually or as larger sets or patterns, can be
accomplished by a wide variety of methods, ranging from biochemical analysis of
blood or tissue samples to biomedical imaging, which include,
spectrophotometry, ELISA, DNA arrays, sequencing methods, mass spectrometry,
liquid chromatography and protein sequencing [6, 8].
- APOPTOTIC
MARKERS IN CERVICAL CANCER
Apoptotic markers are indicators of the process of
apoptosis which is an essential mechanism to prevent the proliferation of cells
with a higher mutation rate, thus tempering malignant transformation.
2.1 ERCC 1 expression
Excision Repair Cross-Complementation group 1 enzyme
(ERCC1) is a molecular marker which can be used to rapidly evaluate the
cisplatin responsiveness of cervical tumors. Cisplatin is a valuable adjuvant
to radiotherapy for the treatment of cervical cancer. ERCCI encoded by
the ERCC1 gene, is a key enzyme in the
nucleotide excision repair pathway which is involved in the DNA repair
mechanisms in tumor cells damaged by treatment with platinum agents.
Preclinical and clinical data have suggested a potential use of ERCC1 as a
molecular predictor of clinical resistance to platinum-based
chemotherapy. Higher expression level of ERCC1 is significantly associated
with platinum drugs resistance [9] and it has been proved that ERCC1 mRNA
levels are also having significant correlation with cisplatin resistance [10].
Since the correlation between ERCC1 mRNA levels and cisplatin resistance in
single-cell-derived cervical carcinoma cell lines that exhibit a wide range of
inherent sensitivity to cisplatin is proved, pre-treatment ERCC1 mRNA level is
a useful predictive indicator for tumor-cell/clinical response to
cisplatin-based chemotherapy [11].
2.2 Protein 53 ( p 53)
p53 ( protein 53 or tumor protein 53),
is a tumor suppressor protein that in humans is encoded by
the TP53 gene. p53 is known to be a cell cycle checkpoint protein and thus
functions as a tumor suppressor playing a regulatory role in the
control of cell proliferation and apoptosis thereby preventing cancer. If
the p53 gene is damaged, tumor suppression is severely
reduced [12, 13]. The loss, or inactivation, of wild-type p53 has been
found to indirectly promote tumor angiogenesis by up-regulation of
angiogenesis-promoting protein, the vascular endothelial growth factor (VEGF)
[14].
2.3 Clusterin
Clusterin has been implicated in a variety of
activities including programmed cell death, regulation of complement mediated
cell lysis, membrane recycling, cell to cell adhesion and Src induced
transformation.Overexpression of clusterin, an anti-apoptotic molecule, has
been reported to induce resistance to chemotherapy in a variety of cancer cell
types. Clusterin induce resistance by physically binding to paclitaxel, which
may prevent paclitaxel from interacting with microtubules to induce apoptosis
[15]. The increased expression of clusterin in cervical cancer tissues than in
normal cervical tissues in different studies, suggests that clusterin may
confer paclitaxel resistance in cervical cancer cells [16]. Clusterin
expression could be a new molecular marker to predict response to
platinum-based chemotherapy and survival of patients with cervical cancer
treated with neo-adjuvant chemotherapy and radical hysterectomy.
2.4 Parathyroid Hormone Related Peptide (PTHrP)
Parathyroid hormone-related protein (or PTHrP) is
a protein member of the parathyroid hormone family.
It is occasionally secreted by cancer cells. It
regulates endochondral bone development by maintaining the endochondral
growth plate at a constant width. It also regulates epithelial-
2.5 Serum Nucleosomes
In
patients with cancer, there is often a correlation between tumor load and
amount of free DNA in circulation that at least part is present in the form of
oligo- and mononucleosomes, as a marker of cell death.Serum nucleosome level
determined in the third course of neoadjuvant chemotherapy in patients with
cervical cancer is marginally related with tumor response and survival. Serum
nucleosomes may have a predictive role for response and prognostic significance
in patients with cervical cancer patients treated with neoadjuvant chemotherapy
[18].
2.6 Cytokeratins
The level of cytokeratin-19 in cervical cancer cells will
be decreased. The reduction of cytokeratin-19 level has a killing effect on
cervical carcinoma SiHa and HeLa S3 cell lines. The apoptotic rate of cervical
carcinoma cells in response to cisplatin and vinblastin will be increased if
their cellular cytokeratin-19 level is reduced by specific antibody MAb Cx-99.
This indicates that elevation of cytokeratin-19 expression could associate with
the apoptotic resistance and malignant progression of cervical carcinoma [19].
The LD80 values were at least 15-fold reduced when cancer cells were treated
with cisplatin or vinblastine in the presence of MAb Cx-99. These results
suggest that the functional role of cytokeratin-19 is associated with the
apoptosis resistance and thus leads to drug resistance of cervical cancer cells
[20].
2.7 Bax Gene
Bax can function as an effector of p53 in
chemotherapy-induced apoptosis and contributes to a p53 pathway to
suppress oncogenic transformation thus becoming a determinant of p53-dependent
chemosensitivity [21]. HPV-positive cervical tumors were shown to undergo
apoptosis upon expression of Bax, a molecule directly regulated by functional
p53 proving its role in apoptosis and sensitivity to chemotherapy [22,
23]. Patients in whom BOAI (cisplatin-based cyclic balloon-occluded
arterial infusion chemotherapy) is effective will show significantly higher
expression of the bax protein and gene after BOAI, and cancer cell apoptosis
will be accelerated. On the other hand, patients in whom BOAI is ineffective
will show significantly higher expression of the bcl-xL protein and gene
after BOAI. Thus bax/bcl-xL expression can be used as an indicator of the
effectiveness of BOAI therapy[24]. In conclusion, increased Bax expression
results in good response to chemotherapy.
2.8 Survivin
Survivin is a structurally unique member of the inhibitors
of apoptosis protein (IAP) family that is involved in both control of cell
division and inhibition of apoptosis. The survivin protein is expressed highly
in most human tumors and fetal tissue, but is completely absent in terminally
differentiated cells. Invitro and in vivo studies targeting survivin with
antisense oligonucleotides have shown that negative mutants or ribozymes induce
apoptosis, reduce tumour-growth potential and sensitise tumour cells to
chemotherapeutic drugs such as taxol, cisplatin, etoposide, γ-irradiation, and
immunotherapy. Study of the markers of survival demonstrated that survivin
expression correlates with drug resistance in CaSki and SiHa cells [25].
2.9 Bcl – xL (B-cell lymphoma-extra-large)
It is one of several anti-apoptotic proteins which are
members of the Bcl-2 family of proteins. It has been implicated in the
survival of cancer cells. Increased expression of antiapoptotic BAG-1, p50, p33
and Bcl-xL may cause resistance to apoptosis through reduction of caspase-3
activity in human cervical cells having an MDR phenotype [26]. Increased levels
of Bcl-xL in the HPV16-immortalized and the CSC-transformed HEN, correlated
with progressively increased resistance of these cells to apoptosis induced by
staurosporine or cisplatin [27]. The ability of bcl-xL to prevent apoptotic
cell death in response to chemotherapy-induced DNA damage and cell-cycle
arrest may contribute to the accumulation of chromosomal aberrations
within tumors. The expression of bcl-xL in tumor cells is likely to be an
important indicator of chemotherapeutic efficacy [28].
2.10 mTOR
The mammalian target of rapamycin (mTOR) also
known as FK506 binding protein 12-rapamycin associated protein
1 (FRAP1) is a protein which in humans is encoded by the
FRAP1 gene. The expression of phosphorylated mTOR may have a role as
a marker to predict response to chemotherapy and survival of cervical cancer
patients who are treated with cisplatin-based neoadjuvant chemotherapy. In
vitro studies have shown that pretreatment with rapamycin inhibited activation
of mTOR signaling and significantly enhanced the sensitivity of CaSki cells to
paclitaxel by increasing apoptotic cell death. Thus mTOR inhibitors enhance
chemosensitivity of paclitaxel and cisplatin in ovarian and cervical cancer
cells [29]. The mTOR cascade may be a promising target for therapeutic
intervention in cervical cancer. These studies address potential of
targeting mTOR protein in the enhancement of therapeutic efficacy of
chemotherapy in human cervical cancer [30].
3. ANGIOGENIC MARKERS
Angiogenesis is essential for the growth of solid tumors.
Tumor hypoxia and progression of cervical cancer seems to correlate with the
angiogenic potential of the tumor. High blood vessel density predicts improved
survival with radiotherapy and chemotherapy.
3.1. Cycloxygenase (cox)-2
Cyclooxygenase (COX) is an enzyme that is responsible for
the formation of prostanoids. Two cyclooxygenase isoforms that
have been identified are COX-1 which is produced constitutively (i.e., gastric
mucosa) and COX-2 which is inducible (i.e., sites of inflammation) by the
action of macrophages. Cycloxygenase 2 (COX-2) is an important marker for
predicting response of cervical cancer to adjuvant chemotherapy. COX-2
levels significantly correlates with VEGF levels in uterine cervical cancers.
VEGF associated with COX-2 might work in the advancement of angiogenesis
[31]. The expression of COX-2 has been shown to be induced by
proinflammatory cytokines, and suggestions have been made that overexpression
of COX-2 suppresses apoptosis and is directly related to tumor growth. The
expression of COX-2 in stage IB cervical cancer may down regulate apoptotic
processes and thus enhances tumor invasion and metastasis [32].
3.2. Thymidine Phosphorylase (dThdPase or TP)
dThdPase gene and protein expression levels in cervical
carcinoma cell lines were closely related to the number of cells that migrated
and invaded. Studies also suggest that dThdPase expression may be closely
related to tumor invasion and metastasis[33]. Immunohistochemical expression of
TP in tumor cells has been suggested as a useful prognostic factor for uterine
cervical squamous cell carcinomas treated with radiotherapy. Choosing therapy
for individual cases by referring to factors including TP expression should
contribute to an improved prognosis[34].
3.3. Major Vault Protein (MVP)
Hypoxia induces drug resistance in clinical tumours. Vaults
are multi sub unit structures which mediate bidirectional nucleocytoplasmic
transport of a wide range of substrates, including cytotoxic drugs. Chemo-resistance
would be mediated by up-regulation of Major Vault Protein (MVP) through the
Hypoxia-inducible factor 1 (HIF-1). Increased levels of MVP have been reported
in numerous cell lines after treatment with a wide panel of cytostatic drugs
like doxorubicin, methotrexate, vincristine or cisplatin [35]. Thus
over-expression of MVP has been associated with chemotherapy resistance. MVP
over-expression was associated with reduced long-term local control in patients
who achieved clinical complete response to radio-chemotherapy [36].
Vascular endothelial growth factor (VEGF) is a
chemical signal produced by cells that stimulates the growth of new blood
vessels. VEGF production is considered essential for angiogenesis
and cancer metastasis, with high titres being indicative of a poor prognosis
[37]. Patients with cervical cancer who are positive for VEGF
expression are less likely to benefit from neoadjuvant chemotherapy.
Pretreatment assessment of VEGF expression may provide additional information
for identification of patients with cervical cancer who had a low likelihood of
response to neoadjuvant chemotherapy and an unfavorable prognosis.
3.5. CD31 (PECAM-1)
CD31 is a member of the adhesion molecule family and is
also known as platelet endothelial cell adhesion molecule-1 (PECAM-1) or
endothelial cell adhesion molecule (endoCAM-1). It plays a key role
in removing aged neutrophils from the body. CD31 has been used to measure
angiogenesis, which reportedly predicts tumor recurrence. Micro Vascular
Density (MVD) is assumed to reflect the intensity of tumor angiogenesis;
indeed, it has been established as a good indicator of prognosis in
several cancer types. If validated, CD31 MVD has the potential to identify
a group of women who are less likely to benefit from standard adjuvant
chemoradiotherapy and might be better served with neoadjuvantchemotherapy and
surgery, with a new radiosensitizing regimen like cisplatin and tirapazamine or
an anti-angiogenesis drug like bevacizumab which exhibited single-agent
activity in recurrent cervical cancer [35, 38].
High CD31 MVD is not only associated with prolonged Progression-free survival (PFS)
and Overall survival (OS)but is an independent prognostic factor in women with
high-risk, early stage cervical cancer treated with radiation therapy alone or
chemoradiation. High CD31 MVD is a surrogate marker for improved tumor blood
flow and oxygenation, resulting in a better response to adjuvant radiotherapy
with chemotherapy [38].
4. OTHER PROGNOSTIC MARKERS
4.1. P- Glycoprotein
Many cancers fail to respond to chemotherapy by acquiring
multi drug resistance (MDR) to which it has been attributed the failure of
treatment in over 90% of patients with metastatic cancer. Although MDR can have
several causes, one major cause of multi drug resistance is the presence of
molecular "pumps" that transport drugs out of the cell. The most
prevalent of these MDR transporters is P-glycoprotein, a particularly
"promiscuous" molecule that transports distinct types of molecules
ranging from peptides to steroids as well as chemotherapy drugs. Once a cancer
cell starts to produce P-gp it becomes resistant to chemotherapy drugs
and it becomes much less likely that the patient will recover [39].
P-glycoprotein was found to be increased in radical
hysterectomy samples from locally advanced or bulky cervical carcinoma treated
with two courses of intraarterial infusion of cisplatin, doxorubicin, mitomycin
C, and 5-fluorouracil (5-FU), when compared with pretreatment biopsy.
Assessment of the expression of P-glycoprotein is thus potentially useful for the
prediction of tumor response to neoadjuvant chemotherapy for cervical
carcinomas [40].
4.2. Carcinoembryonic Antigen (CEA)
Carcinoembryonic Antigen (CEA) is a glycoprotein which is
produced in significant amounts by the large intestine during fetal
development. Chemotherapy and radiation therapy can cause a temporary rise in
CEA due to the death of tumor cells and release of CEA into the blood stream.
Benign disease does not usually result in an increase above 10 ng/ml. CEA is a
valuable tumor marker to predict the prognosis of squamous cell carcinoma of
the uterine cervix and to foresee a clinical response to subsequent neoadjuvant
chemotherapy [41]. Kjorstad and Orjasaester have reported that all the patients
with pre-treatment levels over 15ug/l died during the follow up. In the range
between 5 – 15 ug/l, two third of them developed recurrence [42].
4.3. Squamous Cell Carcinoma Antigen (SSCA)
SCCA belongs to the family of serine and cysteine protease
inhibitors. It is a protein that in humans is encoded by the SERPINB4 gene.
This antigen is present in normal cervix epithelium with an increased
expression in proportion to dysplastic lesion and cervical squamous cell
carcinoma. Approximately 60% of patients with cervical cancer are detected with
elevated levels of serum SCCA at initial diagnosis, when all stages are
included. Several studies have concluded that serum SCCA is useful in
monitoring the course of squamous cell cervical cancer following primary
therapy. Persistently elevated serum SCCA levels after and/or during treatment
suggest tumor persistence or progressive disease especially in distant
metastatic sites [43]. Levels of antigen prior to chemotherapy may not predict
the response to treatment, but patients whose levels decrease during
chemotherapy are more likely to show responses to treatment and the antigen
levels are more predictive of response after two cycles of chemotherapy than
one [44].
SCC assay may provide useful information to improve the
prognostic characterization and disease monitoring of patients with locally
advanced cervical cancer undergoing neoadjuvant chemotherapy [45].
4.4. Thymidylate Synthase (TS)
Thymidylate synthase (EC2.1.1.45) is the enzyme used to
generate thymidine monophosphate (dTMP), which is subsequently phosphorylated
to thymidine triphosphate for use in DNA synthesis and repair. Certain germline
polymorphisms in TS have been shown to influence the level of TS expression
which supports the hypothesis that genotyping patients for TS polymorphisms may
also serve as a useful predictive marker of fluoropyrimidine response and
toxicity [46] and unlike the determination of intratumoral gene expression
which requires available tumor tissue, genotyping can be performed on blood,
which is more readily available. Those tumors with high TS gene expression are
generally non-responsive to protocols that include the TS-directed combination
of 5-FU and leucovorin [47].
4.6. Dihydropyramidine Dehydrogenase (DPD)
Dihydropyrimidine dehydrogenase (DPD) is an enzyme that is
involved in pyrimidine degradation. It is also involved in the degradation of
the chemotherapeutic drugs like 5-fluorouracil and Tegafur-uracil. Individuals
with Dihydropyrimidine dehydrogenase deficiency may develop
life-threatening toxicity following exposure to 5-fluorouracil (5-FU) or
oral fluoropyrimidine capecitabine (Xeloda) because the drugs are not at all
degraded. Patients with low expression levels of DPD are more likely to respond
to 5-FU therapy [48]. Study on Thymidine phosphorylase/
4.7. X-Ray Repair Cross Complementing Protein 1(XRCC1)
The protein encoded by this gene is involved in the
efficient repair of DNA single-strand breaks formed by exposure to ionizing
radiation and alkylating agents. This protein interacts with DNA ligase III,
polymerase beta and poly (ADP-ribose) polymerase to participate in the base
excision repair pathway. Genetic polymorphism of XRCC1 R399Q is associated with
response to platinum-based NAC in bulky cervical cancer, and MDR analysis
documented association between gene-gene interaction of XRCC1 R399Q and treatment
response [50].
4.8. NF-κB (Nuclear Factor Kappa-Light-Chain-Enhancer Of
Activated B Cells)
NF-κB is a protein complex that controls the transcription
of DNA and plays a key role in regulating the immune response to infection.
Conversely, incorrect regulation of NF-κB has been linked to cancer,
inflammatory and autoimmune diseases. If treated with a small molecule
inhibitor towards aurora kinases, (which helps in cellular division by
controlling chromatid segregation), the NF-κB activity will be down regulated
and the efficacy of cytotoxic drugs will be enhanced. This shows that there is
an association between cell resistance to chemotherapeutic agents and NF-κB
activation [51].
4.9. c-erbB-2 Oncoprotein
c-erbB-2 oncoprotein is a 185 KDa membrane bound glycoprotein.
It is a receptor on the cytoplasmic membrane which is homologous to the
epidermal growth factor receptor (c-erbB-1). c-erbB-2 oncoprotein is associated
with a reduced response to neo-adjuvant chemotherapy in the primary treatment
of invasive cervical cancer. Thus over expression of c-erbB-2 can be used as a
useful marker to identify patients who are likely to benefit from high doses of
adjuvant chemotherapy [52].
4.10. Cadherin Methylation
Cadherins (Calcium dependent adhesion molecules) are a class
of type-1 transmembrane proteins. They play important roles in cell adhesion,
ensuring that cells within tissues are bound together. Inactivation of the
cadherin-mediated cell adhesion system, caused by aberrant methylation, is a
common finding in human cancers. Detection of aberrant cadherin methylation may
be of potential use as a marker for selecting cervical cancer patients at high
risk for relapse who could benefit from additional systemic therapy [53].
5. CONCLUSION
Biomarker research has become a sign of the times, and the
identified biomarkers can be used for relating patient’s response to drugs. The
drug-related biomarkers indicate whether a drug will be effective in a specific
patient and how the patient’s body will process it. Prognostic biomarkers help
devising an optimal therapeutic treatment plan for different patient subsets
and to monitor the effect of treatment. It is becoming clear that mapping the
entire networks rather than individual markers may be necessary for robust
diagnostics and tailoring of therapy.
Genomics offers the opportunity to examine gene expression
or the variation in gene sequence, whereas proteomics encompasses evaluation of
protein expression, activation, modification, degradation, and ambitiously
targets protein function. In years to come, a serum or urine test for every
phase of cancer may drive clinical decision making, supplementing or replacing
currently existing invasive techniques.
Advances in the field of biomarkers will enable techniques
to be developed that can profile tumor cells for their genetic background,
allowing selection of anticancer agents on an individual basis. The next
generation of anticancer treatments might therefore be tailored according to
the molecular alterations identified in tumor cells of individual patients.
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