Dr Adeola Henry_Colorectal cancer book chapter 2014

Chapter 15
Prospects of ‘Omics Based Molecular Approaches in
Colorectal Cancer Diagnosis and Treatment in the
Developing World: A Case Study in Cape Town, South
Africa
Henry Adeola, Ryan William Goosen,
Paul Goldberg and Jonathan Blackburn
Additional information is available at the end of the chapter
http://dx.doi.org/10.5772/57485
1. Introduction
The emergence of the field of genomics, proteomics and more recently lipidomics in science
has advanced diagnostic and therapeutic medicine in no small measure. These fields typically
deal with the documentation of the identity, abundance and localization of DNA, RNA, protein
and lipid biomolecules in a given cell, tissue or organism. An in-depth knowledge of the
biologic and physiologic localization, chemistry, and methodology for isolation of these
essential biomolecules is key to a successful analysis and interpretation of information
retrieved in the ‘omics field.
The recent rapid development of these fields can be accounted for by the concurrent develop‐
ment of new state of the art, high throughput technologies such as real time qualitative
polymerase chain reaction (RTqPCR), microarrays, flow cytometry, mass spectrometry and
sequencing. These high throughput technologies have found extensive utility in diverse areas
of human biology, particularly following the completion of the human genome project (HGP)
in 2003. This project, which successfully documented the full complement of genes present
physiologically within the human cell, gave a scientific platform to newer experimental
initiatives thereafter.
Clinical application of ‘Omics based approaches have gained popularity and are believed to
be the future of medicine because of its inherent ability to determine disease-associated
changes in the human genome, transcriptome, proteome, lipidome and metabolome. Docu‐
© 2014 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons
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mentation of these changes in principle enables the identification of disease-associated
biomarkers for use in diagnostic tests, as well as with identifying molecular mechanisms of
disease.
Establishment of the HGP was then followed by the initiation of the Cancer Genome Atlas
project to create a repertoire of genomic profiles for 20 different types of cancers. This project
is designed to evaluate, across-the-board, the molecular and genomic depiction of different
cancer types and to delineate the networks and maps for disease pathogenesis; the first results
to be published from this project were for human glioblastomas [1], followed by ovarian
carcinomas. [2]
Most recently, human colon and rectal cancers have been added to the Cancer Genome Atlas
Project [3], the aim being to evaluate somatic modifications in such carcinomas using genome
level methodologies to assess variation in DNA copy number, methylation patterns, micro‐
RNA expression, exon utilization; and acquired mutations: To date, genetic mutations have
been found in 29 genes and amplifications of ERBB2 and IGF2 have also been observed. This
large scale identification of novel genetic changes in human colorectal cancer (CRC) may in
due course enable the identification of underlying molecular mechanisms of cancer develop‐
ment and new therapeutic targets, as well as the development of diagnostic and/or predictive
tests for CRC.
Current knowledge of cancer cell biology shows that the complex carcinogenesis process is
made up of several intricate molecular pathways and that different cancer cells express a
heterogeneous array of signals. Hence individualized administration of beneficial treatment
regimens to patients, rather than a ‘one-size-fits-all’ approach, is becoming more acceptable as
a generic principle to strive towards, not least since it would be similar to the use of specific
antibiotics based on microscopic culture and drug sensitivity testing instead of using a broad
spectrum/empirical antibiotic approach.
According to the GLOBOCAN cancer fact sheet of the International Agency for Research on
Cancer (IARC), over 12 new cancer cases and 7 million cancer deaths are reported worldwide
annually. Colorectal cancer is the third most common cancer in males (663,000 new cases
representing 10.0% of the total cancer cases) and the second most common cancer in females
(571,000 new cases representing 9.4% of the total cancer cases). [4] Roughly 600,000 cases of
colorectal cancer deaths are expected annually, accounting for almost a tenth of all cancer
deaths, making it the fourth leading cause of cancer death worldwide. In trend, mortality rates
are higher in males than in females except in the Caribbean regions.
Over half of the global CRC burden is occurs in developed countries and its incidence is known
to be lowest in third world countries. The lowest incidence is reported in Africa; however in
South Africa the incidence of CRC more closely resembles that of developed countries. Even
though the mortality and incidence figures are poorly reported and cannot be harmonized
reliably, reports from the currently defunct National Cancer Registry of South Africa revealed
that in 1999 CRC was the sixth commonest cancer in the general population, accounting for
about 2,367 cases of the total 26,606 new cancer cases reported. [5]
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Whilst research and health institutions in the developed world are replete with ongoing studies
and discoveries of candidate disease biomarkers, significant attendant challenges are associ‐
ated with ‘omics based studies in a developing world setting, inter alia: infrastructure, human
capacity and funding. Here we provide a synoptic overview of the prospects, challenges and
benefits of ‘omics based approaches in the diagnosis and treatment of CRC in a developing
world, using our Cape Town experience as a case example.
2. Diagnostic inaccuracies in colorectal cancer
The intricacies of CRC are due in part to its multiple implicated aetiologies and the heteroge‐
neity of the tumour. Important aetiologic considerations include; location (right sided or left
sided, surface or cryptic, colonic or rectal), whether sporadic or hereditary, de novo or sequel
to adenomatous polyp; intra-tumour heterogeneity (ITH) also plays an important role in the
accuracy of histopathologic and molecular diagnosis. The classical theory of ‘field cancerisa‐
tion’ [6] presupposes that cancer develops from multifocal areas of precancerous changes; this
concept is supported by latter studies which demonstrated that true cancer boundaries often
exceed the physically discernible margin taken out at surgery, with histological analyses
showing that these multiple abnormal tissues constituting premalignant cancer cell field
changes persist and surround the tumor and may form multiple independent lesions that
sometimes coalesce. The implications of such findings are profound when differentiating the
cancer field recurrence from second primary tumours [6-8]. Subsequently, others have
considered the concept of ‘sequential carcinogenesis’ which suggests that cancers develop as
a progression from patchy premalignant lesions which later show evidence of mild, then
moderate and then severe epithelial dysplasia. [7] These dysplastic lesions eventually progress
to carcinoma-in-situ (high grade intraepithelial neoplasm), then to microinvasive carcinoma
and advanced invasive carcinoma.
The literature is replete with the extensive application of the field concept of Slaughter in
surgical management of head and neck cancers and is gradually becoming a major consider‐
ation in the management of cancer of other organs such as the stomach, skin, cervix, lungs,
vulva, bladder, colon, breast, and ovaries. [9, 10] In addition, molecular investigations have
been used to assess correlation between phenotypic stages of cancer progression and expres‐
sion of cancer specific genes and mutations; these studies have demonstrated significant
correlations between both [7, [11], implying that molecular markers have the potential to
significantly enhance the precise diagnosis, staging and treatment monitoring of cancers.
The possibility of a physically normal tissue adjacent to the tumour area and expressing cancer
specific genetic profiles has spurred interest in the origin of molecular level heterogeneity and
histologic variability in the immediate field surrounding the cancer area. Some researchers
have suggested that the heterogeneity in cancer growth follows a Darwinian evolutionary
theory of natural selection [12], whilst others have concluded that there are two plausible
origins of diversity in tumour cells expansion, described by the ‘clonal evolution’ theory and
the ‘cancer stem cell’ theory. [13-15] The clonal evolution theory (cancer monoclonality)
Prospects of ‘Omics Based Molecular Approaches in Colorectal Cancer Diagnosis and...
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presupposes that tumour cells result from a solitary clone of mitotically unstable cell that
differentiate into different offspring lineage clones that have developed additional unique
genetic damage down the lineage. [13] By contrast, the cancer stem cell theory (cancer
polyclonality) is premised on the possibility of cancer developing from multiple cancer stem
cells that proliferate concurrently and drive the expansion of the tumour. Both theories have
therapeutic implications in that only a fraction of the tumour bulk drives its expansion, hence
targeted molecular therapies at these ‘driver cells’ could in principle be established for cancer
treatment.
Phylogenetic evidence suggests that well-characterized subpopulations of tumour cells,
including annotations of genetic mutations, have been derived from sequential genetic events
[16] and mathematical models have been described to account for this, but to date have mostly
provided a one-dimensional insight into the complexities of ITH. [17, 18] The mechanistic
development of cancer is a multi- dimensional event and multiple factors have been estab‐
lished to govern its progression such as: the shape of the organ in which it occurs; blood supply;
surgical interference; the consistency of the surface on which it occurs; tumour microenviron‐
ment; and the genetic nature of the cell. Clearly ITH is a reality that affects tumour diagnosis,
classification, prognosis, and treatment; and requires further understanding.
2.1. Reliability of histopathology reports
Introduction: Histopathology is the science of utilising classical histological techniques to assess
micro- and macroscopic evidence of potential disease. Classical histology routinely utilises
microscopic observation of micrometer cross-sections of tissue, differential staining techni‐
ques, as well as immunohistochemical assays, to assess the tissue specimen in question. The
visual evidence provided by each technique, or combination thereof, allows a pathologist to
identify potential evidence of pathology, thereby providing a pathological diagnosis for a
given specimen.
Classical histological techniques are inherently limited in their scope for the detection of
pathology as they rely on a microscopically visible presentation of clear or strong evidence of
pathology, e.g. in the case of advanced disease. Furthermore, any aberrant change at the sub-
microscopic level, i.e. the molecular level, needs to have translated into morphological change
at the subcellular and/or overall cellular morphological level, or to have produced a variation
in the abundance of a particular protein, or set of proteins, that is detectable through immu‐
nohistochemistry.
Evidence of molecular changes that are not yet detectable at the histological level: In molecular studies
of tumour biopsies and adjacent “paired” normal samples, researchers often seek to identify
molecular signatures that can distinguish tumour samples compared to histologically normal
samples obtained from anatomical sites distal to the primary lesion. However, recent studies
have investigated the possibility of significant molecular differences between histologically
normal colorectal tissue from patients with polyps, or colorectal carcinoma, versus the same
tissue from healthy individuals [19], suggesting that significant aberrant molecular changes
occur in apparently normal colorectal tissue, despite being apparently distinctly located from
the original lesion. In turn, this suggests that classic histopathological techniques are limited

in their scope to identify such clinically relevant aberrant changes in tissue that appears
morphologically normal; interestingly, these molecular observations are entirely consistent
with Slaughter’s ‘field cancerisation’ theory. [6] However, whether or not such a field has a
gradually tapering aberrant effect as function of distance from the tumour remains incom‐
pletely answered. For instance, it has been found that there was no significant correlation
between the degree of aberrant gene expression perturbation and distance from a polyp or
tumour. [20] Irrespective, of the specific characteristics of any field effect, it is interesting to
note that supporting evidence of aberrant perturbations in histologically normal colorectal
mucosa appeared five decades after Slaughter’s original hypothesis as a direct result of modern
genomic, epigenomic, and proteomic research.
In a study of normal colonic mucosa from individuals with a family history of sporadic cancer
conducted by Hao et al [21], it was found that there was a significant difference in the expres‐
sion of several genes in these individuals’ normal mucosa relative to the same tissue from
healthy controls. In particular, the gene expression levels of PPAR-gamma, SAA1, and IL-8
were found to be significantly different in the morphologically normal rectosigmoid tissue
samples in the individuals with a family history of CRC. Furthermore, a follow-up study in
individuals with adenomatous polyps with or without familial history of colorectal carcinoma
again found that there was a difference between gene expression in normal rectosigmoid
mucosa from these individuals and healthy controls, regardless of the presence of a familial
history of cancer. [22]
Polley et al [19] observed significantly different proteomic signatures in morphologically
normal mucosa from patients with colorectal neoplasia compared to the same tissue from
healthy subjects. It therefore appears that a larger than anticipated field of tissue in the
colorectum may be affected by the presence of a neoplastic lesion, implying in turn that the
method used to determine clear margins estimated during surgical resection may need the
support of molecular assays in the future. However, whether or not the molecularly perturbed
normal mucosa will progress to disease remains to be determined.
In addition to genomic and proteomic perturbations, epigenetic changes have also been
reported in CRC tissues, one example being the hypomethylation of L1 promoter sequences
in colorectal tumours and in adjacent normal tissue of 6 out of 19 cancer patients, but not in
colonic mucosa of 14 healthy individuals. Furthermore, genomic CpG methylation appeared
to be lower in normal colorectal tissue from diseased patients, compared to healthy subjects,
and significantly lower in patients with hypomethylation of the L1 promoter sequences. [23]
The above examples at the genomic, proteomic and epigenetic level provide substantial
evidence of molecular aberrations in morphologically normal tissue sample adjacent to a
tumour. These changes might be subtle and may not effect a microscopically visible phenotype,
but could well represent significant perturbations that impart normal tissue samples with pre-
cancerous characteristics. It is therefore important that such findings are considered when
assessing individual biopsies by histopathology since these samples may in fact have under‐
lying molecular signals of disease that, if interpreted correctly, could provide insight into the
disease.
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Molecular Intratumour heterogeneity due to branched evolutionary clonal expansion: Intratumour
heterogeneity in terms of cellular morphology and types of cells, is a well-established obser‐
vation in anatomical pathology. However, it is only recently that a further layer of complexity
has been introduced through the discovery of an added layer of heterogeneity at the genomic
and epigenomic level. This molecular heterogeneity further complicates biopsy selection and
subsequent histopathological assessment because a specific lineage of clones, with a distinct
genomic profile, may occur in physical clusters on a given tumour and thus be physically
separated from other clonal populations. Therefore, each biopsy taken may present a uniform
or slightly variable morphological appearance but might be distinct at the molecular level with
unique functional potential and characteristics. This has fundamental ramifications when it
comes to assessing the severity of disease and the prognosis, as well as deciding the appropriate
course of treatment. Furthermore, intra-tumour heterogeneity governs how each unique
tumour lineage evolves and adapts to therapeutic interventions and should therefore ideally
be understood when developing new chemo- and biological treatments.
In a landmark study by Gerlinger et al [14], multiple spatially separated biopsy samples
obtained from primary renal carcinomas and associated metastatic sites showed evidence of
branched evolutionally growth by unique clonal population lineages. Furthermore, the
majority of all somatic mutations (63 – 69%) were not found to be present across all tumour
biopsies. One clinically relevant consequence of these findings was the identification of both
good and poor prognostic signatures in different biopsies of the same tumour. This study
highlights the inherent complexity of utilising a single biopsy for biomarker development,
prognostic or predictive measures based on molecular markers to guide treatment regimen. It
is clear that future biopsy collection - for the purpose of treatment planning or for biomarker
research - should ensure collection and molecular assessment of multiple biopsies from
physically separate sites on the same tumour.
A recent study by Kreso et al [24] has provided supporting evidence, in the context of colorectal
carcinoma, that the distinct genetic profile of each intratumoural clonal subpopulation has
implications in growth potential and chemotherapy tolerance and therefore has an impact on
treatment outcomes. This appears to be due to Darwinian style natural selection, with certain
subpopulations becoming more dominant by leveraging their intrinsic tolerance to selective
pressure such as chemotherapeutic intervention. These findings highlight the fact that different
tumour subpopulations have distinct proliferative potentials and chemotherapy tolerance
mechanisms; as such, treatment regimens in the future my need to target each type of cellular
population individually in order to prevent disease recurrence.
Future directions for modern pathological assessments of cancer tissue samples: Given the recent
developments in the field of intratumour heterogeneity, and the evidence of significant
molecular aberrations in histologically normal mucosa, it is clear that classical histopatholog‐
ical techniques are limited in their ability to assess the underlying clinically relevant hetero‐
geneityintumourandnormaltissuesamples,suggestingthatmodern,validated,cost-effective
molecular assays should be integrated into histopathological assessments. Amongst others,
this would ensure that clinically relevant phenotypic or functional characteristics - whether
dormant or active - which may directly govern a tumours response to therapeutic intervention

canbe identified.Thisenhancedapproachshouldfacilitatethedevelopmentofnoveltreatment
regimens that efficaciously target all heterogeneous subpopulations for a given tumour and,
in doing so, potentially also thwart aberrant histologically normal tissue from progressing to
a malignant lesion.
The possibility that distinct subpopulation of cells and/ or genetic signals exist in different
areas of a particular biopsy specimen poses a major challenge in implementing personalized
therapy. [25] ‘Omics based research can in principle generate a more robust predictor of
therapeutic benefits, but this often involve an extensive sample collection for discovery and
validation, adequate funding and availability of appropriate manpower: an example of one
such initiative is the ‘Personalized RNA Interference to Enhance the Delivery of Individualized
Cytotoxic and Targeted Therapeutics’ (PREDICT) consortium [26] that aims to identify reliable
biomarkers of different cancer types.
2.2. Tumour classification and staging
The earliest classification system for CRC by Dukes [27], modified by Astler [28], considered
the staging of CRC severity to be based on its depth of invasion. More recently, a standardized
widely accepted staging system of the American Joint Committee on Cancer (AJCC) also
known as the TNM system which incorporated the tumour size, nodal involvement and
presence of distant metastasis was introduced [29]: These three classifications are largely based
on morphologic evidence and can be subject to inaccuracies. To minimize errors in diagnosis,
these phenotypically driven systems of staging colorectal tumour could now in principle be
usefully complemented with a validated molecular diagnostic method.
Newer and often more reliable molecular based assessment of tumour staging and prognosis
are beginning to emerge for CRC based on new molecular cancer knowledge and ‘omics based
techniques and complement existing orthodox morphology based methods. One of such
molecular classification is based on CpG island methylator phenotype (CIMP), as well as
microsatellite instability (MSI), and is scored as type 1-5 based on different combinations of
these molecular features. [11] Chromosomal instability (CIN) has also been identified as an
important global molecular classifiers of CRC. [30] In addition, KRAS mutations have emerged
as the most common prognostic biomarker of CRC for anti-EGFR therapy patients and many
more candidate markers for prediction of therapeutic outcomes in colorectal cancer are being
discovered now using ‘omics based techniques, including loss of PTEN signals, PI3KCA
mutation and BRAF gene mutation. [31]
Identification of cancer specific genes, proteins, lipids and metabolites is increasingly regarded
as a promising route to early diagnosis, treatment and monitoring of disease progress. For
instance, the following genes have been recognized as associated with the risk of developing
colorectal cancer; TP53, MLH1, MSH2, MSH6, MYH, EPCAM, KIT, BLM, SMAD4, PGDFRA,
BMPR1A, APC, AXIN2, and STK11. Vogelstein et al [32] In a recent meta-analysis of 25 different
genetic expression studies of colorectal cancer, a statistically significant down-regulation of
carbonic anhydrase II (CAII) and CEACAM1 was observed, while there was up-regulation of
TGFβ1, IFITM1, SPARC, GDF15 and MYC genes. [33] Based on these molecular patterns, a
novel staging and classification system that is devoid of errors can now potentially be evolved.
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2.3. Role of surgical pathology
The rationale behind the histopathologic use of slide sections from biopsy samples has been
to evaluate for diagnostic purposes a representative microcosm of disease from a routine‐
ly processed, waxed, and miniaturized specimen blocks. However this has sometimes led
to mis- or under- diagnosis of tumour depending on the exact site from which the biopsy
was taken from. In principle, the goal of surgical resection is to take adequate ‘tumour-
free’ margin; however achievement of this goal in practice is at best an estimated blind
procedure, since cancer specific molecular alterations in the ‘apparently’ tumour free regions
are largely unknown. This makes it difficult to readily determine the adequacy of a
surgically resected margin. Not least, pathologists have had to take multiple biopsies from
different sites in a resected tumour mass, trying to maximize the chances of locating the
accurate tumour areas. This blind sampling procedure is a major potential source of
diagnostic inaccuracies in practice and it is therefore important to detect colorectal cancer
early to improve the chances of getting an adequate ‘tumour free’ surgical margin. Prompt
surgical intervention, coupled with accurate determination of the most beneficial adju‐
vant therapy for the specific patient, appears to hold the key. Surgical pathology - which
is the interface between surgery and pathologic specimen processing - is thus a vital control
point for the eradication of diagnostic inaccuracies. Given the current molecular eviden‐
ces on intratumour and interbiopsy heterogeneity, an exclusive morphology based sampling
technique is a potential minefield for diagnostic errors, even with the best mastery.
A delicate balance needs to be achieved during sampling of surgical specimens for the
assessment of tumour heterogeneity. The most prominent, average pattern of expression is
most likely to be identified during sampling of a large tumour biopsy sample, but this
approach risks masking the less prominent but potentially equally important information
about sub-populations of tumour cells in the biopsy sample. By contrast, signal-to-noise
ratios have to be carefully balanced when analysing smaller biopsy samples since this
approach presents a smaller sampling frame within which it may be impractical to identify
all possible biomarker signals a tumour could express. [34] Surgeons and pathologists also
need to bear in mind the downstream requirements and applications of ‘omics based
research when surgical biopsy samples are taken, especially since mining the molecular
archives of formalin fixed paraffin embedded (FFPE) specimens through genomics,
proteomics and lipidomics research is beginning to gain traction now. However, factors
such as age of tissue, condition of storage, tissue sample size, fixation time, pH influen‐
ces and buffers are known to influence the outcome of ‘omics based analyses on surgical
specimens. [35] In particular, RNA degrades rapidly at room temperature, whilst formal‐
in damages nucleic acids within the specimen by forming sclerotic crosslinks of DNA and
RNA via methylol adducts and methylene bridges. [35] Thus, immediate snap freezing of
fresh sections in liquid nitrogen or dry ice is good practice, whilst use of newer alcohol-
based kits [36], or of phosphate buffered formalin [37, 38], provide useful alternatives to
standard formalin for rapid fixation of surgical specimens prior transport to the lab.

2.4. Patient evaluation and therapeutic loopholes
Biomarker discovery research will in principle enable accurate stratification of patients into
appropriate risk categories. The orthodox clinical approach of prescribing a common thera‐
peutic cancer regimen to all colorectal cancer patients is fast becoming a subject of evidence-
based debates. Certain differences exist between patients who come from demographically,
geographically and genetically divergent backgrounds. Such inter-patient variability may also
present significant differences in tumour phenotype, behavior and natural histories across a
population, an important observation that is referred to as single nucleotide polymorphism
(SNP) noise. [39] Patient stratification based on these natural clusters can enable meaningful
biomarker discovery using ‘omics based techniques. For instance, in a culturally heterogene‐
ous South African population composed of Caucasians, Indigenous African, Indian, and Mixed
Ancestries, the a priori expectation is that different racial groups may express different
biomarkers of disease. In the same light, candidate biomarkers discovered from studies on
developed world patient cohorts may not necessarily be effective for the management of
colorectal cancer in a developing world situation due to differing ethnicities, which has clear
implications for the planning and execution of ‘omics based discovery and validation of
biomarkers.
In principle, it is possible to predict response to specific targeted therapies (e.g. Herceptin)
using ‘omics based approaches, with patients who would not respond to specific targeted
therapies being identified at the outset and given alternative therapies. Patient dependent
source of variation for biomarker discovery includes individual genetic make-up, metabolism,
stage of disease, health, immunocompetence, nutritional factors, and environmental factors.
Careful patient selection to eliminate confounders must be carried out prior to experiments
and biomarkers must be validated in a standardized acceptable manner.
Genetic profiling of patients for KRAS mutation, BRAF genes, Microsatellite instability (MSI)
and CpG island methylator phenotypes (CIMP) is now being increasingly carried out for
colorectal cancer patients in the developed world and this has contributed significantly to
treatment planning.PatientsthatexhibitMSIhavebeenfoundtopossessbetteroverall survival
than those with chromosomal instability and are less affected by p53 mutations [40], whilst
CRC with p53 +, MSI + profiles are usually more aggressive than those with p53-, MSI + profiles.
[41] Biomarkers such as telomerase and survivin have been used to assess the long term risk
of CRC development [42] whilst morphologic biomarkers in patients include neoplastic
colorectal polyposis and the presence of aberrant cryptic foci (AFC); the presence of adenomas
with Intraepithelial Neoplasm in the colorectal region can also be used in surveillance as
surrogate endpoint biomarkers. [43] Paradoxically, most anti-cancer agents do not have well-
established single predictors of individualized response, however with the advances in ‘omics
based approaches it should be possible to provide such molecular predictors. For example,
MSI has been documented to be an effective predictor of response to fluoropyrimidine therapy,
whilst ERCC1 was found to be beneficial in patients using platinum containing anticancer
regimen. [31]
‘Omics based techniques relevant to colorectal cancer management
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The management of CRC has to date been based on fairly invasive techniques for diagnosis
and treatment. In contrast, ‘omics based techniques in most instances are non- or minimally
invasive, thereby improving patient compliance, eliminating surgical morbidities and
ultimately reducing the burden of disease through early diagnosis and effective treatment
monitoring. The potential utility of increasingly common-place ‘omics based techniques in the
diagnosis, surveillance, treatment, and prevention of colorectal cancer is thus discussed below.
2.5. Genomics and epigenomics
Introduction: In genomic investigations, high-throughput technologies such as microarray
platforms or DNA/ RNA sequencing are now commonplace. These technologies are now
routinely applied to large sample collections, with complete clinical annotations, and aim to
produce profound insights into disease at the resolution of single nucleotide polymorphisms,
gene expression, and the status of epigenetic regulatory mechanisms such as hypo- and
hypermethylation.
Extensive bioinformatic analysis of these high-throughput, multi-level, datasets has provided
in-depth insights into the mechanisms of disease and treatment resistance. These findings have
been translated into biomedical research aimed at addressing the significant challenge in
treating cancer of the colorectum. While it is well established that cure rate for treating TNM
stage I colorectal cancer is over 90%, the clinical management of stage II CRC is more compli‐
cated. [44, 45] Furthermore, improved chemo- or biological treatment regimens are needed to
address the high rates of recurrent disease observed with stage II and III CRC disease, as well
as for TNM stage IV disease which is generally considered incurable at present. [46, 47]
The overall disease-free survival statistics for TNM stage II disease, treated with surgery alone,
are as high as 75%. [44] However, certain sub-populations of patients experience a worse
prognosis and have clinical outcomes more similar to TNM stage III disease. Therefore, much
focus has been on finding genetic markers to guide treatment regimen selection in stage II and
III disease, the goal being to improve overall efficacy, decrease treatment failures, reduce the
incidence of recurrent disease, whilst at the same time lowering the cost of treatment by
limiting costly chemotherapeutics to those patients with predicted benefit.
High-throughput genomic studies today provide a comprehensive means to analyse amongst
others the expression level of every individual gene, as well as to assess chromosomal segment
copy number- and DNA methylation pattern variations and to determine single nucleotide
polymorphism and mutation frequencies, all in a genome-wide manner. Biomarker research
can thereafter be carried out to identify and validate distinct signatures derived from inte‐
grated value measurements associated with each gene, as a prelude to translating such findings
into a clinical setting, as either biomarkers or novel therapeutic targets.
3. Transcriptomics
The field of transcriptomic research, within the context of prognosis and treatment outcome
prediction, has seen much attention in recent years. High-volume real-time gene expression

assays, gene expression microarrays and, more recently, ‘next generation sequencing’ based
methods have provided the necessary platforms to investigate putative prognostic and
predictive genetic markers. These platforms, combined with known clinical outcomes, enable
panels of genes to be significantly correlated with prognosis, or the outcome of treatment with
various chemotherapeutic agents. However, genetic association studies require large datasets
in order to identify putative prognostic and predictive markers with significance worthy of
clinical utility. As such, there are to date a limited number of landmark publications that
present such multi-gene panel lists associated with prognosis and treatment outcome predic‐
tion in TNM stage II and III colon and rectal cancers.
A study by O’Connell et al [48] involved the combined analyses of four independent studies
of colorectal cancer patients. Samples were obtained from 1851 CRC patients in the United
States, with stage II or III disease, who participated in the National Surgical Adjuvant Breast
and Bowel Project (C-01/C-02 and Cleveland Clinic (CC); C04; C-06) and 1136 candidate genes
(761 genes assessed in C-01/C-02; 375 genes in CC/C-06) were evaluated. The aim of the study
was to establish a panel of markers that could be associated with the risk of disease recurrence
and that could determine the likelihood of patients benefitting from adjuvant 5-fluorouracil/
leucovorin adjuvant therapy. The analyses resulted in the identification of 48 genes signifi‐
cantly associated with the risk of disease recurrence and 66 genes significantly associated with
5-FU/LV benefit (with four genes in common between the two sets). For practical reasons a
gene panel of 7 genes predictive of disease recurrence, 6 predicted of 5-FU/LV benefit, and 5
reference genes were selected. The clinical utility of this predictive panel was then independ‐
ently evaluated in the Quick and Simple and Reliable (QUASAR) study. [49] The aforemen‐
tioned study validated the use of this gene panel, and it’s consequent recurrence score, as an
independent predictor of the risk of recurrent disease in stage II colon cancer patients who had
undergone surgery. This gene panel was then commercialized by Genomic Health by the
production of a multi-gene panel called the Oncotype DX® Colon Cancer Assay.
In separate work, Salazar et al [50] described a gene expression signature - named ColoPrint -
in early 2011 that allowed for improved prediction of prognosis in TNM stage II and III
colorectal cancer. The gene signature consists of 18 genes that were identified from analysis of
188 frozen tumour samples (TNM stage I-IV; 78.7% not treated with adjuvant chemotherapy),
and a cross-validated on 206 independent tumour samples (TNM stage I-III; 60.7% not treated
with adjuvant chemotherapy) originating from three sites in the Netherlands. This panel
showed better predictive accuracy, in comparison to the American Society of Clinical Oncology
criteria for assessing the risk of cancer recurrence without prescreening for microsatellite
instability (MSI), with a hazard ratio of 2.69 (95% CI, 1.41 to 5.13; P = 0.003) for patients with
stage II disease. Results such as these are encouraging, and provide strong supporting evidence
for the utility of molecular approaches over purely clinical markers.
In a related study, a panel of 13 genes - referred to as ColoGuideEx - was reported by Ågesen
et al [51] to be significantly associated with predicting prognosis for stage II colorectal cancer
patients. This study was conducted on an initial dataset obtained from 207 colorectal samples
originating from three independent Norwegian patient series, and validated on a 108-sample
gene expression dataset originating from the USA and Australia. The independent prognostic
373

value of the panel of genes was confirmed by multivariate Cox regression analyses (p≤0.004),
which included various clinicopathological variables and all three-sample series.
O’Connell et al (2010)
O’Connell et al (2010)
Oncotype DX® Colon Assay
Salazar et al (2011)
ColoPrint®
Ågesen et al (2012)
ColoGuideEx
5FU benefit Disease recurrence Disease recurrence Disease recurrence
ATP5E ATP5E CA4388O2 AZGP1
AXIN2 BGN CTSC BNIP3
BIK C-MYC CYFIP2 CXCL10
EFNB2 FAP EDEM1 CXCL13
GPX1 GADD45B HSD3B1 DSC3
HSPE1 GPX1 IL2RA ENPP3
MAD2L1 INHBA IL2RB EPHA7
PGK1 Ki-67 LAMA3 KLK6
RUNX1 MYBL2 LIF MMP3
UBB PGK1 MCTP1 PIGR
VDAC2 UBB PIM3 SEMA3A
VDAC2 PLIN3 SESN1
PPARA TUBA1B
PYROX D1
SLC6A11
THNSL2
ZBED4
ZNF697
Table 1. A list of multi-gene panels that are significantly associated with disease recurrence, or benefit from adjuvant
5-fluorouracil (5-FU) and leucovorin (LV) chemotherapy, as published in three independent studies.
Table 1 illustrates the various gene panels from the four studies described above and it is
noteworthy that there are no genes in common between these gene panels. Furthermore, the
biological relevance the genes utilised in each panel has yet to be fully explained and may
represent an opportunity to identify pathologically associated genes and pathways when
molecular enrichment analyses are applied across multiple gene panel studies in the context
of prognostic and predictive biomarkers used in recurrent colorectal cancer. For example, there
are genes across the lists in Table 1 that share a relationship by virtue of their KEGG pathway
associations: (1) Cytokine-cytokine receptor interaction (n=6 genes; CXCL10 and CXCL13 [Che‐
mokines; CXC subfamily] from Ågesen et al (2012), LIF [gpl30 shared] from Salazar et al (2011),
IL2RA and IL2RB [IL2RG shared] from the Hematopoietins from Salazar et al (2011), and

INHBA [TGF-ß family] O’Connell et al (2010); (2) Axon guidance (n=3 genes; EPHA7 from
Ågesen et al (2012), EFNB2 from O’Connell et al (2010), and SEMA3A from Ågesen et al (2012);
(3) Pathways in Cancer (n=3 genes; LAMA3 from Salazar et al (2011), AXIN2 from O’Connell et
al (2010), RUNX1 from O’Connell et al (2010)); (4) p53 signaling pathway – target genes (n=2 genes;
SESN1 from Ågesen et al (2012), and GADD45B from O’Connell et al (2010)). These KEGG
pathway association mappings were adapted from outputs generated by GeneCodis. [52-54]
In a novel study that utilised eight published prognostic and predictive gene expression
signatures, Shi et al [55] combined the datasets and integrated them with publically available
protein-protein interaction network data in order to identify candidate molecular markers
associated directly with the recurrent colorectal cancer phenotype. As a result, they were able
to not only infer pathophysiological mechanisms underpinning the recurrent disease pheno‐
type, but also used the augmented and cross-study integrated signature to identify both a
prognostic signature and a multi-gene signature to predict treatment outcome. The resultant
gene signature consists of 487 genes and is referred to as the NEM signature (as it integrates
information from Network, Expression, and Mutation datasets).
It is clear therefore that gene expression panels are developing to the point where they are
close to translation into a clinical setting and adoption into routine clinical practice. It remains
to be seen though how many of the published gene panels will make it into the clinic after
appropriate validation studies have been carried out to assess performance across larger and
more ethnically diverse patient populations. In this context, it is important to note that such
signatures might yet be inherently compromised by the likely existence of multiple, possibly
opposing, signatures in the same tissue sample (vide supra). Therefore, testing of multiple
biopsies from the same tumour may be necessary in order to generate a holistic prognosis and
to provide guidance on treatment strategy. Furthermore, in developing nations where the
incidence of colorectal cancer is not yet decreasing and cases consistently present at more
advanced stages of disease, it remains to be seen whether the cost of such diagnostic or
prognostic panels will be affordable in the public sector where there is the most need and where
the greatest diversity exists.
Single nucleotide polymorphisms: Colorectal cancer has classically been treated by 5-fluorouracil
(Capecitabine; Xeloda®; Hoffmann-La Roche Inc.), combined with either Oxaliplatin (Eloxa‐
tin®; Sanofi-Synthélabo Inc.) or Irinotecan (Camptosar® or CPT-11; Pfizer Pharmaceuticals
Inc.). Over the past decade these regimens have been augmented by the addition of biological
therapeutic agents, such as the monoclonal antibodies Cetuximab (Erbitux®; ImClone Systems
Inc.), Panitumumab (Vectibix®; Amgen Inc.), and Bevacizumab (Avastin®; Genentech Inc).
Each drug has a different mechanism of action and a unique set of molecular targets, as well
as distinct sets of enzymes responsible for its metabolism; furthermore, particularly in the case
of biological agents, specific enzymes that are functionally important to the pathways targeted
by the biologic are major determinants of response. It is well-established that polymorphic
variations in enzymes involved in drug metabolism can alter drug availability, thereby causing
a variation in clinical response, by altering the rate and specificity of drug metabolism. Such
variant drug metabolizing enzymes are generally encoded by single nucleotide polymor‐
phisms (SNPs) that result in changes in the amino acid composition of the relevant gene
375

product (i.e. protein), resulting in altered enzymatic activity. As such, the biomedical com‐
munity has utilised an arsenal of molecular techniques, including high-throughput genomic
platforms, to identify such SNPs and to quantify their frequency in populations in order to
correlate with treatment outcomes and thereby to identify biomarkers that are predictive of
therapeutic response.
This area of research has been comprehensively reviewed recently by Asghar et al [56], Bandrés
et al [57], Benheim et al [58], Coate et al [59], De Roock et al [60], and Ross et al [61] and forms
the basis of the rich pharmacogenetic resource, PharmGKB® (http://www.pharmgkb.org)
which documents each therapeutic agent together with an aggregated list of SNPs reported in
the literature to be associated with treatment outcome.
4. Pharmacogenomics
4.1. Markers of treatment outcomes when treated with classical chemotherapeutics
5-Fluorouracil: 5-Fluorouracil (5-FU) was introduced more than 50 years ago by Heidelberger
et al [62] and is the foundational cytotoxic agent used in the treatment of colorectal neoplasia.
5-FU can be administered in three different physical forms: either through an intravenous
solution, or as an oral compound (Capecitabine, and Tegafur). The metabolism of either of
these compounds results in the formation of fluoronucleotides. A subset of these molecules,
fluorouridine triphosphate (FUTP) and fluorodeoxyuridine monophosphate (FdUMP) are
misincorporation into DNA and RNA during their in vivo biosynthesis. In addition, FdUMP
inhibits thymidylate, synthase thereby resulting in a nucleotide imbalance due to a depleted
intracellular reserve of thymidine for DNA synthesis. The enzymes responsible for producing
the active metabolites have been studied for polymorphic variation and analysed for their
correlation to treatment response (as seen in Table 2).
Gene SNP Molecular effect
Associated treatment
outcome
Reference
TP Increased expression Increased response Bandrés et al (2007)
TS TSER2R Decreased expression Increased response Bandrés et al (2007)
DPD DYPD*2A Decreased activity
Increased response
Increased toxicity
Bandrés et al (2007)
MTHFR 677T
Increased production of
CH2FH4
Increased response Bandrés et al (2007)
Table 2. A list of single nucleotide polymorphism (SNP) markers associated with treatment outcome, as reported by
Bandrés et al (2007), when using a 5-fluorouracil-based regimen.
Oxaliplatin: Oxaliplatin is a platinum analog that results in inter- and intra-molecular DNA
cross-links, resulting in the inhibition of DNA synthesis, transcription and repair processes.

This compound has been extensively utilised in combination with 5-FU and Leucovorin, a
regimen referred to as FOLFOX and commonly prescribed for treatment of advanced colorectal
cancer.
There are two groups of genes that are primarily responsible for altered response to Oxalipla‐
tin, namely genes involved in DNA repair and in glutathione conjugation reactions. In the
former group, a polymorphism in the X-ray repair cross-complementing group 1 enzyme
(XRCC1) - which is part of the base excision repair system - has been associated with variable
initiation of DNA repair. [57] In addition, a polymorphism in a component of the ubiquitous
nucleotide excision repair pathway - the excision repair cross complementing group 2 (ERCC2)
gene - has been significantly associated with a clinical response to platinum-based chemo‐
therapy. [57] However, there are no known polymorphisms in the DNA mismatch repair
pathway associated with variation in treatment response with Oxaliplatin. In the second
group, it has been reported that platinum compounds are inactivated by glutathione conju‐
gation and therefore the enzymes responsible for catalyzing this reaction have been investi‐
gated. In particular, SNPs in several glutathione-S-transferase (GST) genes have been
implicated in conferring resistance to Oxaliplatin. [57]
outcome
Reference
GSTP1 613G Decreased activity Increased response Bandrés et al (2007)
GSTT1 Deletion Decreased activity Increased response Bandrés et al (2007)
GSTM1 Deletion Decreased activity Increased response Bandrés et al (2007)
XRCC1 388Gln Decreased activity Increased response Bandrés et al (2007)
ERCC2 751Gln Decreased activity Increased response Bandrés et al (2007)
Table 3. A list of single nucleotide polymorphism (SNP) markers associated with treatment outcome, as reported by
Bandrés et al (2007), when using a chemotherapeutic regimen that includes Oxaliplatin.
It is noteworthy that in a recent study by Fernandez-Rozadilla et al [63], seven SNPs
(rs16857540, rs2465403, rs10876844, rs10784749, rs17626122, rs7325568, rs4243761) were found
to be significantly associated with adverse drug reactions in the context of singular 5-FU or
FOLFOX treatment. Given the relatively large sample size of 221 CRC patients and a validation
set of 791 patients, these results hold strong statistical significance and provide potential
predictive capacity for toxicity response on an individual patient basis if validated through a
larger cohort.
Irinotecan (CPT-11): Irinotecan is a camptothecin analogue with well-established anti-neoplasia
activity exerted though stabilization of the ordinarily transient DNA topoisomerase I-DNA
complex, thus preventing the repair of temporary single stranded breaks during DNA
replication and leading to cell death. [64] There are two primary pathways that have been
implicated in variable response to Irinotecan treatment: drug transport into the extracellular
377

environment, and metabolism of Irinotecan into its active (SN-38 and SN-38G) and inactive
metabolites.
Hydrolysis of Irinotecan by carboxylesterases: CES1 and CES2 results in the production of its
active metabolite, SN-38. This metabolite then undergoes detoxification via the process of
glucuronidation, catalyzed by uridine diphosphate-glucurono-syltransferase 1A (UGT1A), to
produce a metabolite called SN-38G which then interacts directly with the DNA topoisomerase
I enzyme. However, Irinotecan can also be oxidized by members of the Cytochrome P450 3A
subfamily (CYP3A) to produce inactivate metabolites. Each of these metabolites is transported
out of the cell by the adenosine-triphosphate (ATP) binding cassette (ABC) transporter
transmembrane proteins. As such, polymorphic variation in these enzymes has been associ‐
ated with altered metabolism, transport and therapeutic efficacy of this drug and its metabo‐
lites. A selection of these SNPs is detailed in the Table 4.
outcome
Reference
CES2 IVS10-88 Decreased expression Decreased response Bandrés et al (2007)
CYP3A ? Decreased activity Decreased response Bandrés et al (2007)
UGT1A1 UGT1A1*28 Decreased activity
Increased response
Increased toxicity
ABCB1 1236C"/>T Decreased activity
Increased response
Increased toxicity
ABCB1 3435C"/>T Decreased activity
Increased response
Increased toxicity
ABCC2 3792T Increased response Bandrés et al (2007)
ABCG2 421A Decreased activity Increased response Bandrés et al (2007)
Table 4. A list of the single nucleotide polymorphism (SNP) markers associated with treatment outcome, as reported
by Bandrés et al (2007), when using a chemotherapeutic regimen that includes Irinotecan (CPT-11).
Markers of treatment outcomes when treated with biological agents: In recent years there has been
an emergence of a new class of antibody-based therapeutics that directly and specifically target
molecules belonging to processes fundamental to cancer pathophysiology. Comprehensive
reviews of the hallmarks of the cancer phenotype have been updated recently by Hanahan and
Weinberg [65, 66]; in addition, comprehensive reviews of biomarkers associated with mono‐
clonal antibody therapies that are currently in being used in the treatment of this disease have
also been published recently. [56, 60]
Vascular endothelial growth factor as a target: It is well understood that tumours have angiogenic
potential, i.e. they possess the ability to induce the production of new blood vessels, and
thereby increase their supply of oxygen and nutrients; this characteristic provides an oppor‐
tunity for therapeutic intervention, targeting the vascular endothelial growth factor (VEGF)
and associated VEGF receptors (VEGFR-1, -2, and -3). In particular, overexpression of VEGF
has been associated with vascularity, endothelial cell migration and invasion, poor prognosis

and aggressiveness in most malignancies [67]; mAbs have thus been designed to target VEGF,
thereby reducing the amount of free VEGF, reducing VEGF receptor activation, and ultimately
reducing angiogenesis. [68] A recombinant humanized IgG1 monoclonal antibody, Bevacizu‐
mab, that specifically targets VEGF is currently indicated as part of combination therapy in
metastatic colorectal cancer; however, a biomarker that is predictive of response to anti-VEGF
therapy is yet to be discovered.
Epidermal growth factor receptor as a target: Another fundamental hallmark of the cancer
phenotype is the ability for tumours to alter their response to growth factors through differ‐
ential gene expression of growth factor receptors. This too presents the opportunity interrupt
aberrant growth mechanisms. In colorectal cancer, it has been shown that there is an abnormal
activation of epidermal growth factor receptor (EGFR). [69] As such, a humanized monoclonal
antibody, Cetuximab, targeted at the extracellular domain of EGFR has been evaluated in
clinical trials. This mAb blocks specific EGF-mediated signal transduction events [70], thereby
inhibiting cellular proliferation and inducing apoptosis. [71] Treatment with Cetuximab also
leads to increased responses to classic chemotherapeutic agents and radiotherapy, as well as
inhibiting cellular proliferation, angiogenesis and metastasis. [72] Another recombinant
human IgG2 monoclonal antibody, Panitumumab, which also targets EGFR is currently
indicated in metastatic colorectal carcinoma where there has been resistance to fluoropyrimi‐
dine, Oxaliplatin, and Irinotecan containing regimens.
To date there have been a small but useful number of genes harboring mutations that provide
insight into the outcome of anti-EGFR therapy. For example, the mutational status of the
Kirsten-ras (KRAS) gene is currently being used in predicting treatment benefit in the context
of anti-EGFR therapy for patients with metastatic disease. Other common genes with muta‐
tions relevant to anti-EGFR mAb therapy include BRAF and PIK3CA, as well as loss of
expression of PTEN.
Conclusion: It is clear that SNP biomarkers associated with treatment outcome are generally
found in genes with specific characteristics of cancer pathophysiology, or with drug metabo‐
lism and transport, and numerous low-throughput, focused, yet fruitful studies have resulted
in clinically translatable SNP biomarkers being identified. This contrasts with the high-
throughput transcriptomic approach to identification of biomarker gene panels that is typically
initially blind to the functional significance of each individual gene. As such, these simple SNP
markers present a cost-effective option to predicting the efficacy of a specific therapeutic agent,
or combination thereof, prior to prescription, enabling design of individualized therapy that
will result in increased efficacy and improved treatment outcomes.
5. Epigenomics and epigenetics
Molecular studies of cancer have revealed the presence of not only of genetic mutations, copy
number alterations, altered gene expression, but also of aberrant epigenetic changes. Intense
investigation in recent years has shown that epigenetic regulation of gene expression plays a
crucial role in embryonic development, imprinting, and tissue differentiation. [73] Therefore,
379

deregulation of such a fundamental mechanism might offer insight into the driving molecular
mechanisms associated with the development, and regulation, of a carcinoma phenotype,
particularly in cases where other genomic perspectives have not yet provided an answer.
In cancer studies, the field of epigenomics has encompassed investigations into aberrant DNA
methylation, post-translational modification of histones that affects chromatin structure,
altered expression of microRNAs and non-coding RNAs, and nucleosome positioning. [73-75]
For complete reviews of this specific topic see Ballestar et al [76], Hatziapostolou et al [77],
Khare et al [74], Lao et al [75], Liu et al [78], Sawan et al [79], Sharma et al [73], Ting et al [80],
and van Engeland et al [81].
DNA methylation studies have focused particularly on the methylation patterns of cytosine
and guanine (CpG)-rich DNA sequences. In particular, regions of the genome that have a
higher than expected number of CpG nucleotides compared to the rest of the genome – so-
called “CpG islands” – are of particular interest because they have been shown to overlap the
promoter regions of 60-70% of genes. [75] An extension of this concept has been the discovery
of CpG sites outside of promoter regions, referred to as “CpG Island shores”, that are within
two kilobases of the 5’ end of a CpG island. [75] Methylation of CpG islands is largely associated
with transcriptional repression [75, 82] and, in general, CpG islands are protected from
aberrant methylation but in cancer this does not appear to be the case. Since methylation
changes can significantly alter gene expression profiles and thereby deregulate important
biological pathways, a sound understanding of which genes and which associated pathways
are affected in which individual patients might in the future be applied in the clinic to guide
the selection of appropriate treatment regimens.
The first report of epigenetic alterations in tumours of the colon revealed an extensive loss of
5’-methylcytosine when compared to normal colon tissue. [83] However, it is only recently
that this area of research has gained increased attention. Today, high-throughput methylation-
specific microarray and sequencing technologies, together with a well-established array of
commercially available methylation assay kits, have facilitated large-scale epigenomic
investigations and contributed to an increased understanding of the methylome on a multi-
gene scale.
Arguably the most important epigenetic finding to date, in the context of colorectal cancer, has
been the identification of a unique molecular subtype characterized by a high frequency of
gene methylation. Colorectal tumours of this variety are now referred to as having the CpG
island Methylator Phenotype (CIMP). The exact panel for diagnosing CIMP varies from study
to study, but the panel of genes proposed by Weisenberger et al [84] has been commonly used,
i.e. NEUROG1, SOCS1, RUNX3, IGF2, and CACNA1G. In general, if a panel has more than 60%
of its genes methylated, then is considered to be CIMP positive. Despite, the lack of a stand‐
ardized CIMP diagnostic panel, this type of tumour is reported to be predominantly associated
with right-sided colon cancer, and tends to be more common in woman. [84, 85]
Diagnosing CIMP tumours is clinically relevant because approximately 20% of colorectal
tumours have this phenotype and generally share a high frequency of the BRAF c.1799T>A (p.
V600E) mutation [75] that has been reported to negatively impact treatment outcomes in anti-

EGFR mAb-mediated therapy in patients with KRAS wild-type tumours and metastatic
disease. [60, 86, 87] As such, classifying a patient’s tumour as having the CIMP phenotype
could be used in the future to guide the selection of biological therapeutics to ensure that the
most efficacious treatment regimen is utilised.
The current lack of a standardized panel of genes from which to assess the status of genomic
methylation highlights the fact that this is an emerging field of study. As such, the translation
of these panels into the clinic as diagnostic or prognostic biomarkers is still premature.
However, as the field progresses and more literature-based evidence is published in support
and validation of a particular panel, the full potential of such epigenomic biomarkers may
ultimately be translated into patient benefit.
While the field is currently in its infancy, there are a number of encouraging studies that
represent good examples of novel discoveries of methylation markers that have highly
significant associations. For example, Wang et al [88] examined the methylation status of five
tumour suppressor genes in eighty-five paired colorectal tumours and normal mucosa samples
from Chinese patients and showed that the methylation status of two genes, CDH13 and
FLBN3, were significantly associated with stage of disease and prognosis. In particular, CDH13
was significantly associated with poor differentiation (p=0.019) and had a relatively strong
association with advanced stage of disease (p=0.084). In a similar fashion, FLBN3 was signif‐
icantly associated with advanced disease (p=0.027) and with the presence of lymph node
metastasis (p=0.029). Furthermore, CDH13 and/or FLBN3 methylation status was found to be
predictive of a poor overall survival (p=0.001) and conversely the presence of methylated
hMHL1 indicated a better chance of survival (p=0.046).
In a clinical setting, samples that are obtained in a non- or minimally invasive manner are
preferred and there have been a number of studies based on stool and blood plasma samples
to assess aberrant methylation patterns. One example is the clinically validated methylation
status of the Vimentin gene (VIM) - which has been found in the majority of colorectal tumours
(53 – 84%; Lao et al 2011) – with assays conducted on stool samples, thus providing a non-
invasive mechanism for early detection of colorectal cancer. This assay is currently commer‐
cially available in the United States as the ColoGuard assay (LabCorp), and reports a sensitivity
of 83% and a specificity of 82%. [89, 90] Outside of the United States, in Europe and the Middle
East there is now an additional non-invasive test for early detection of colorectal cancer by
assessing the methylation status of SEPT9. This assay is currently commercially available as
Epi proColon (Epigenomics AG).
The field of epigenomics, and epigenetics, applied to colorectal cancer is providing valuable
insights into underlying mechanisms of this disease and seems to be a promising avenue of
research. It is clear that assessing the status of DNA methylation alone holds prognostic and
predictive value, and as such we would expect this field to gain much momentum towards
translating these findings into a routine clinical setting. As this field of study can be conducted
on a broad spectrum of sample types, along with commercially available kits to assess
methylation, this field is likely to see an increased number of significant findings in the near
future. However, as in the case of gene expression-based biomarkers, each epigenetic finding
will need to be assessed in patients of diverse ethnicities. A recent study by Nieminen et al [91]
381

highlighted this in observing that colorectal carcinomas in an Egyptian cohort had a signifi‐
cantly higher state of methylation, in microsatellite stable tumours, compared with sporadic
colorectal cancers in a Finnish cohort.
5.1. Proteomics
The field of proteomics deals with identification of the total complement of peptides and
proteins expressed in a cell, tissue or an organism and is in principle more directly related to
phenotypic changes associated with disease pathogenesis. Proteomic studies are able to define:
the functional state of protein activities; protein-ligand interactions; protein-protein interac‐
tions; and a host of dynamic post-translational modifications such as glycosylation, phos‐
phorylation, ubiquitinylation, SUMOylation, proteolytic cleavage, lipoylation and acetylation
of proteins. The proteome of a cell may vary from one time point to another and in different
states of health and disease so proteomics techniques have been employed to identify cancer
specific proteomes as a means to identify candidate biomarkers of early disease. A huge
amount of data is generated from single proteomic experiments and these are typically
analyzed to understand the mechanistic pathways of pathologic events in protein networks
using various databases, workflows and algorithms. For proteomics analysis, complex
mixtures of proteins derived from a given biological sample are typically rendered into a set
of peptides via proteolysis, most commonly using the enzyme trypsin; direct liquid chroma‐
tography-tandem mass spectrometry (LC-MS/MS) based methods are then typically employed
to separate, quantify and identify thousands of individual tryptic peptides in a sample [92-94],
from which the identity and quantity of the parent proteins in the original biological sample
can be inferred. For example, one notable recent study identified and quantified >7,000 unique
proteins from FFPE tissue blocks from individual colorectal cancer patients [95], amply
demonstrating the potential of the technology.
Common sources of current proteomics biomarkers for CRC include stool, blood, biopsy and
urine samples. A common fecal proteomics biomarker in use today is hemoglobin [96], while
carcinoembryonic antigen (CEA) is a common blood-based biomarker currently in use. [97]
Tissue inhibitor of metalloproteinase 1 (TIMP-1) has also been used and has been found useful
in the early detection of cancer although, paradoxically, other studies using multivariate
analysis have described TIMP-1 as independent of the stage of cancer. [98] Proteasome
activator complex subunit-3 (PSME-3), nicotinamide N-methyltransferase (NNMT), collapsin
response mediator protein-2 (CRMP-2), MIF, M2-PK, M-CSF, HNP 1-3, CCSA-2, CCSA-3,
CCSA-4, laminin, MMP-9, MMP-7, and a host of other serum proteomics biomarkers are being
developed and optimized for clinical use. For example, surface enhanced laser desorption-
ionisation (SELDI) mass spectrometry was used to analyze the serum of 62 CRC patients
compared to 31 controls, and four proteomic markers were found in detectable levels in the
serum of CRC patients: apolipoprotein C1, alpha-1-antitrypsin, C3a-desArg and transferrin.
[99] A separate study carried out at the Mayo Clinic revealed elevated levels of 5 serum
biomarkers in CRC: DcR3, TRAIL-2, spondin-1, MIC 1 and Reg IV [100]; usefully, this ‘5-
biomarker panel’ has been reported to exceed the performance of CEA both in specificity and
sensitivity. Immunoproteomics - involving techniques such as immunoblotting, ELISA and

immunocapture-mass spectrometry - has also yielded candidate biomarkers in CRC, including
antibodies against inosine monophosphate dehydrogenase II [101], MUC-5A, MUC-1,
MAPKAPK3, AVCR2B, HlpA, RpL7/L12, and nucleobindin- 1. [102-106]
5.2. Lipidomics
Lipidomics is a novel ‘omics field which deals with complex large scale analysis of the full
complement of various classes of lipids and lipid networks expressed by a cell, tissue or
organism (the ‘lipidome’) and involves the high throughput systems-level identification and
quantification of lipid metabolic pathways that may be involved in disease using chromato‐
graphic methods coupled to mass spectrometry. Generally, lipids are hydrophobic molecules
that are involved in energy storage, structural components of a cell, cell signaling, endocrine
actions, signal transduction, membrane trafficking, and morphogenesis. Structurally, lipids
can be classified as: fatty acids; glycerolipids; glycerophospholipids; sphingolipids; sterol
lipids; prenol lipids; saccharolipids; and polyketides. These 8 classes of lipids can also be
further subdivided into several subclasses.
Whilst there is currently a paucity of clinically validated lipidomics biomarkers for colorectal
cancer, the prospects of this field to complement proteomics and genomics in a combined
‘systems biology’ approach for disease detection, monitoring and treatment is worth consid‐
eration since over the past two decades, numerous publications have described the perturba‐
tion of lipid metabolism and signaling in colorectal carcinogenesis. [107-110] Lipidomics
analysis of primary and metastatic colorectal cancer cell lines (SW480 and SW620) identified
600 and 694 lipids respectively in ‘shotgun’ study [111]; increased level of triglyceride lipid
and plasmacholine were observed, while a decrease in the level of C-16 containing sphingo‐
myelin, ceramide lipid and plasmenylethanolamine were observed in the metastatic CRC cell
line compared to the primary isogenic CRC cell line, implying that lipidomic biomarkers of
metastatic CRC disease might be plausible.
Empirically, polyunsaturated fatty acids have been known to be more beneficial in the
prevention of colorectal diseases compared to saturated long-chain types, but the pathways
related to this were largely unclear. The emergence of lipidomics techniques however has
revealed that metabolic control of long chain fatty acids is an important factor in development
of CRC, with short chain fatty acids from the gut microflora/ microbiome being described as
onco-preventive. [112]
Elevated level of lysophosphatidic acid - a phosphoglyceride - has been described as a
prospective cancer biomarker of ovarian tumours [113-115] but, paradoxically, a marked
decrease in the serum level of lysophosphatidylcholine has been reported in CRC. [116]
Similarly, elevated profiles of phosphatidylcholine and choline kinase activity have been
demonstrated it colon cancers [117] and a high ratio of phosphatidylcholine to phosphatidy‐
lethanolamine has been used to differential metastatic colon cancers from localized ones. [118]
Elevated levels of sphingomyelin have also been reported to characterize human colon cancer,
based on nuclear magnetic resonance (NMR) studies [119], whilst cancer cell motility was
shown to be down-regulated by the interaction between CD9 and sialoglycosphingolipid GM3
using CRC cell lines [120] and ceramides have been found to induce apoptosis in CRC cell lines
383

(HT-29, LOVO, and HCT-116). [121-123] Urinary phospholipids analysis using nanoflow LC-
ESI MS/MS has been previously used for the analysis of breast [124, 125]) and prostate cancer
[124], but there is a dearth of literature on the application of this method to colorectal cancer.
Interestingly though, urinary levels of metabolites of prostaglandin E2 have been used as a
biomarker for colorectal cancer risk evaluation. [126, 127] Overall, these observations of
disease-associated variation in colorectal cancer lipid profiles provide a sound precedent for
the future development of reliable lipidomics biomarkers.
5.3. Metabolomics
The relatively new field of ‘omics techniques that investigates the presence and abundance of
low molecular weight metabolites in cells and body fluids is known as ‘metabolomics’. This
new branch in the ‘omics world has emerged to address molecular biologic problems that have
hitherto not been amenable to genomics or proteomics approaches. Common specimens
compatible with metabolomics experiments include urine, serum and tissue. As is the case for
genomics, proteomics and lipidomics, the ‘metabolome’ changes depending on physiologic
and pathologic states of an individual and identification of unique metabolites provides
potentially useful insight into pathogenetic mechanism of disease.
A number of analytical techniques have been used for metabolomics research, including: gas
chromatography mass spectrometry (GC-MS); liquid chromatography mass spectrometry
(LC-MS); capillary electrophoresis mass spectrometry (CE-MS); matrix assisted laser desorp‐
tion-ionization mass spectrometry (MALDI-MS); and nuclear magnetic resonance (NMR).
[128] By way of example, metabolomics studies on CRC patient serum samples, using a
combination of proton-NMR and GC-MS techniques, was used to differentiate locoregional
CRC from metastatic types as well as to identify CRC that metastasized to the liver. [129]
Separately, in a review of eight metabolomics studies on CRC for diagnostic accuracy and
distinguishing metabolites, twelve metabolites were found to be elevated. [130] In a further
study using GC-MS, 34 endogenous metabolites were found significantly elevated in CRC
compared with health individual, whilst the serum 3-hydroxybutyric acid level was noted to
be reduced. [131] A predictive model developed in yet another study comprised of 2-hydrox‐
ybutyrate, kynurenine, cystamine and aspartic acid and was found to have specificity,
sensitivity and accuracy as high as 85%, 85%, and 85% respectively. [128] Finally, Cheng et al
[132] found evidence of dysregulation of several metabolic pathways through urine analysis
of colorectal cancer patients, whilst Qui et al [133] also observed evidence of similar pertur‐
bations in tricarboxylic acid (TCA) and tryptophan metabolic pathways. With a solid founda‐
tion, a panel of metabolite markers may ultimately be developed for metabolomic profiling of
colorectal patients as a means to improve diagnosis.
5.4. Molecular Imaging
Visualisation of precursor lesions and of malignant tissue is a major aspect of diagnosis and
monitoring of therapeutic interventions in oncology. Classically this has been carried out using
anatomical and functional technologies, such as Ultrasound (US), Computerised Tomography
(CT), and Magnetic Resonance Imaging (MRI). While these approaches have been the mainstay

of medical imaging, they are limited to information at the gross anatomical and physiological
levels respectively. Recent years have however seen the emergence of molecular imaging and
its addition to the repertoire of techniques to visualise disease.
In previous sections we have discussed how a range of different ‘omics technologies have
facilitated an in-depth view of the pathophysiology and molecular pathology of cancer. These
insights provide candidate biomarkers based on the identification of differentially expressed
genes and proteins between tumours and normal tissue. These biomarkers have either been
associated with prognosis, stage of disease, molecular subtypes, predictors of treatment
outcome, and markers of discriminating carcinoma from benign lesions at the molecular level.
In this light, molecular imaging is then the science of utilising the ever-increasing knowledge
of cancer molecular pathology to: identify the appropriate molecular targets; design molecular
constructs to selectively and specifically interact with them; and produce a visual signal that
is measurable through an imaging technology. In order for this methodology to succeed at the
molecular level, the target should be available for interaction and should be unique enough to
ensure selective and specific interactions. Furthermore, once a signal has been generated from
the appropriate interaction, it is important that a suitably sensitive imagining technology exists
in order to visualise, and quantify, signal emission.
In the context of colorectal cancer, plasma-membrane associated proteins are generally
considered good candidate targets since they are reasonably accessible and a subset of them
play vital roles in signal transduction. In a similar fashion, aberrantly expressed or deregulated
intracellular and secreted enzymes represent potentially useful targets for molecular imaging,
based on their ability to catalyse formation of spectroscopically-measurable products. Both of
these types of targets are accessible through the extensive vascularisation of the colorectum,
but more importantly these targets can be accessed via the luminal surface through topical
application of an appropriately designed molecular construct.
Molecular imaging provides tremendous opportunities to enhance the early detection of CRC,
as well as potentially aiding the demarcation of clear margins during surgical resection, as
reviewed recently by Abdullah [134], Seaman et al [135] and Akin et al [136].
By way of illustration of the potential of molecular imaging methods, colorectal cancers have
been found to naturally emit a red fluorescent signal due to the accumulation of protopor‐
phyrin IX (PpIX) in primary colorectal tumours and associated metastases located in lymph
nodes. [137] These authors postulated that endogenous PpIX accumulates as a result of
aberrant metabolic changes in the CRC cells; Kemmner et al [138] subsequently provided
supporting evidence for this hypothesis by showing that there is a significant down-regulation
of ferrochelatase (FECH) mRNA expression in gastric, colon, and rectal carcinomas, leading
to accumulation of PpIX. In an effort to utilise this information Moesta et al [137] found that
metastatically involved lymph nodes could be identified compared to all other palpable nodes;
in the context of previously untreated patients (n=24), this observation had a sensitivity of 62%
and a specificity of 78% (p < 0.0001). However, in a neoadjuvant setting there was a reduction
in PpIX fluorescence in primary tumours, and a drastic reduction of fluorescent signal in
metastases that resulted in not being able to discriminate between lymph nodes containing
385

metastatic cells. In a follow-up study by Wan et al [139], conducted in xenografted nude mice,
a novel siRNA-mediated knockdown of FECH was used to enhance the accumulation of PpIX,
thereby increasing the endogenous fluorescence in tumour cells. While these results still need
to be developed further, and tested in a clinical trial, they do hold promise for increasing the
accuracy of early detection of primary and metastatic lesions and monitoring therapeutic
response based on the size of the visualised tumour.
In different work, differential gene expression profiling, confirmed by immunohistochemistry,
demonstrated that matrix-metalloproteases (MMPs) are differentially expressed in the context
of colorectal adenocarcinoma by macrophage subpopulations and, at times, by the tumours
themselves. MMPs are a family of zinc-dependent endopeptidases with multiple human
peptidase members. For example, MMP-9 is able to degrade specific components of the
extracellular matrix, including type IV collagen, after activation of the secreted zymogen [140]
and as a direct consequence, malignant cells are thereby able to become mobile and achieve
metastatic potential.
Of particular interest to surgical resection was the finding that colorectal adenocarcinomas
express MMP-9 via a distinct macrophage subpopulation found at the edge of primary tumours
and local lymph node metastases. [140] Fudala et al [141] have subsequently designed a dual
fluorophore beacon molecule that is specifically cleaved by MMP-9, resulting in the emission
of a specific fluorescent signal, suggesting that if a suitably non-invasive and anti-immuno‐
genic method is developed to administer the beacon molecule to an anatomical structure under
investigation, then accurate measurement of the tumour edge may be possible during surgical
resection procedures.
As can be seen from the above examples, molecular imaging holds considerable promise for
application in CRC. As this field becomes more developed and is validated through clinical
trials, it should provide improved visualisation ability coupled with the ability to quantify
specific molecules, enabling novel insights into diagnosis, prognosis, treatment response
monitoring, and underlying tumour physiology and molecular pathology in CRC.
5.5. Cancer nanotechnology
Nanotechnology as a division of engineering concerned with the manipulation of atomic and
subatomic molecules has recently found its place in the detection, staging, imaging, and
management of human cancers. Various physical, chemical and biologic principles have been
applied to improve the diagnosis and treatment using elements and molecules in the Nano-
range (~10-9
). [142]
For diagnosis of CRC, nanoparticles have been used to enhance the precision and reliability
of colonoscopy and other conventional diagnostic methods, largely resulting in earlier
detection and obviating variables such as operator skills and speed of examination. Quantum
dots (QD) and surface-enhanced Raman scattering (SERS) are two important nano- method‐
ologies used to improve tissue based diagnosis of cancer, avoiding the cumbersome protocols
and lower reliability of multiplexed tissue staining. Both methods have the ability to detect
multiple biomolecular signals in a single cancer cell. Gold and silver particles have been

derivatized and stabilized with Raman active particles and silica respectively for used in
generating composite organic and inorganic nanoparticles (COINs) for potential improvement
of biopsy diagnosis. [143] Simultaneous Multiple Aptamers and RGD Targeting (SMART)
cancer probes have also been used to detect multiple cancer biomarker signals using currently
available imaging techniques. [144] Superconducting quantum interference device (SQUID)
sensors and magnetic relaxometry - which are both nanotechnology based techniques - have
been reported to be more accurate in the diagnosis of breast cancer than mammography and
MRI respectively. KRAS mutant alleles have been detected in gastrointestinal malignancies,
including colorectal carcinoma, using nanofluidic digital PCR which showed a better per‐
formance in detection of mutation KRAS in colorectal adenomas compared to conventional
PCR. [145] Nanoparticles have been coupled with short cancer specific oligonucleotides
(aptamers) for targeted binding to prostate specific membrane antigen positive cells in prostate
cancer cell lines. [146] Finally, serum detection of colorectal cancer has been achieved with
gold nanoparticles using SERS spectroscopy in a study which exemplified the use of this
approach as a viable, minimally invasive screening method for colorectal cancer. [147]
Nanotechnology also has significant potential in the development of targeted therapeutics for
cancer, with many studies currently at experimental- and a few at clinical validation stages.
Different types of nanostructures, including mesoporous silica nanoshells, dendrimers,
supramagnetic iron cores, nanosuspensions, gold nanoparticles, nanolipogels, nanoemul‐
sions, carbon nanotubes, titanium oxide nanoparticles, liposomes, polymeric miscelles, and
other lipid based nanoparticles have been used as drug delivery vehicles and facilitators of
targeted cancer therapies. [148-155] Although most of the cancer nanodiagnostic and nano‐
therapy studies are still in their infancy, it seems clear that nanotechnology will play an
important role in colorectal cancer diagnostics and therapeutics in the future.
6. Prospects of ‘Omics based molecular approaches in colorectal cancer
diagnosis and treatment in a developing country: A case study in Cape
Town
Groote Schuur Hospital, situated in Cape Town, South Africa, is a quaternary hospital where
the many colorectal cancer patients from the Western Province region are treated by combi‐
nations ofradiotherapy,colonicresectionandstandardchemotherapeuticregimens.However,
there are two complicating factors involved in treating these patients with the greatest efficacy:
(1) the relatively high cost of utilising platinum- or modern monoclonal antibody-based
regimens; and (2) the fact that the majority of patients present with advanced stages of disease,
typically between TNM stage II and III. As mentioned earlier, these stages of disease have
relatively high recurrence rates and as such timely diagnosis and efficacious treatment
schedules are needed to reduce disease recurrence and improve patient prognoses.
To illustrate this point, consider the cost of the standard chemotherapeutic regimen of 5-
fluorouracil (5-FU) alone versus Oxaliplatin: One cycle of Oxaliplatin costs approximately ZAR
5,000.00 (~USD 560), while one cycle of 5-FU is drastically less at a cost of ZAR 200 (~USD 22).
387

The response rate obtained with 5-FU alone can be improved by utilising Leucovorin (LV) in
a combination therapy approach. However, it has been observed that the further addition of
Oxaliplatin or Irinotecan can improve the stage II/III CRC response rates drastically to around
40 – 50%. [156, 157] The relatively recent appearance of monoclonal antibody based therapies
has also offered significant gains in treatment response rates.
South Africa is a developing nation with a limited budget for treating non-communicable
diseases such as cancer, not least because a large proportion of the country’s healthcare budget
is understandably spent on addressing the concurrent HIV/AIDS and TB epidemics (see for
example the Lancet series on “Health in South Africa” published in 2009, with particular
emphasis on the following publications: Abdool et al [158]; Chopra et al [159]; Coovadia et al
[160]; and Mayosi et al [161]). Considering these financial constraints, two possible situations
can be envisaged in which a patient treated in the public sector (i.e. subject to government
healthcare budgets) could access platinum-based regimens and/or modern biological thera‐
peutic agents:
The first opportunity for a patient to access medication with greater efficacy would be through
participation in clinical trials conducted by pharmaceutical companies either wanting to assess
their treatment in ethnically diverse cohorts (which could be required by local regulatory
authorities) or to explore a new disease indication for an existing therapeutic agent. The second
opportunity might be one afforded by the use of ‘omics approaches, leveraging biomarker
panels to provide predictive indications of whether or not a patient might benefit from a
particular chemo- or biological therapeutic agent. Furthermore, given the heterogeneity of
tumours and the unique molecular subtypes of colorectal cancer, it would be instructive to
assess the relative likelihood of recurrent disease. Such a measurement could be used to
motivate the aggressiveness of the treatment regimen prescribed.
In terms of possible cost effective benefits from the addition of platinum-based drugs to the
standard regimen of 5-FU and LV, a trial conducted in the United Kingdom found that the
relative levels of topoisomerase-1 (Topo1), assessed by routine immunohistochemistry, could
be used to identify patient subpopulations who could potentially benefit from the addition of
Oxaliplatin to their 5-FU/LV regimen [162]. Furthermore, Paré and colleagues [163] reported
that a particular polymorphism in the excision repair cross-complementing 1 ERCC1 gene
(codon 118) can predict response and overall survival in patients treated with an Oxaliplatin/
5-FU/LV regimen. Provided that these markers are further validated in a clinical setting, it then
stands to reason that a simple immunohistochemical or real-time polymerase chain reaction
assay could therefore be routinely requested to determine likely response to Oxaliplatin and
therefore to motivate additional expenditure on an Oxaliplatin-supplemented regimen. This
type of personalised approach has the obvious advantage of improving treatment efficacy, and
reducing the risk of disease recurrence with a concomitant cost saving for the hospital authority
from not having to conduct lengthy additional treatments, after possible first round treatment
failure.
Similarly, assessment of the common SNPs that are predictive of benefit from the limited array
of biological agents (e.g. the mutational status of the KRAS and BRAF genes) could provide

an indication of whether or not the comparatively expensive anti-epidermal growth factor
receptor mAb therapy should be prescribed.
As discussed above, a number of gene expression panels could also be used to predict patient
prognosis and the associated likelihood of disease recurrence, with results being used to design
a personalised treatment regimen in which the aggressiveness of the treatment schedule
correlates with the severity of disease, the most likely prognosis and the likelihood of recur‐
rence. In this context, Genomic Health’s Oncotype DX® Colon Cancer Assay costs ~USD 3,200
[45], which translates to approximately ZAR 28,560 in South Africa; this assay could provide
a very useful clinical metric of the likelihood of recurrence, particularly in complicated TNM
stage II cases and stage III cases where recurrence rates are still unfavorably high, but in reality
such a test is beyond the financial means available for treatment of CRC disease at a public
facility in South Africa today, not least since its cost dwarfs even that of Oxaliplatin-based
treatments. The prospects for wide uptake of the Oncotype DX® Colon Cancer Assay in South
Africa therefore seem remote.
As the complexities of biopsy heterogeneity are addressed by the biomedical community, it is
thus clear that local validation studies of simple and cost-effective, assays will be necessary to
ensure that the prognostic and treatment outcome biomarkers reported in the literature apply
to the diverse ethnicities in South Africa. Given the financial constraints imposed by the
government funded healthcare system, it appears that relatively inexpensive techniques such
as PCR based SNP assays would be well-suited to the public sector since, despite being a
simple, low cost assay, the results could have a profound impact on treatment outcomes for
the patients afflicted with this disease. Importantly, if utilised correctly such a molecular
diagnostics strategy could actually result in significant cost savings mid-term for the hospitals
administering care by avoiding the complexities of treatment failures.
7. Conclusion
‘Omics based techniques represent novel, scientifically sound approaches to the diagnostic
and therapeutic aspects of managing colorectal cancer patients, but there remain very signif‐
icant challenges regarding their uptake and wide utilisation in developing world healthcare
settings, primarily due to financial considerations. None-the-less, surgeons, clinicians, basic
medical researchers and all other healthcare workers at the cutting edge of colorectal cancer
management need to remain abreast of the prospects and potential effectiveness of integrating
molecular approaches in to colorectal cancer management. The old paradigm where patients
had no active choice or participation in their disease management, with treatment choices
being exclusively the decision of the clinician, is under threat today since many patients now
have access to information about emerging therapeutic options via the internet. It is therefore
important that surgeons and clinicians, in spite of their invariably tight schedules, consider
some form of participation in basic medical research in order to contribute clinical perspectives
as well as to improve their understanding of molecular approaches to diagnosis and treatment.
389

Acknowledgements
HA thanks the ICGEB for a PhD scholarship; RWG thanks the National Research Foundation
(NRF), South Africa, for a PhD scholarship; JB thanks the NRF for a Research Chair.
Author details
Henry Adeola1,2
, Ryan William Goosen1
, Paul Goldberg1,3
and Jonathan Blackburn1
1 Institute of Infectious Disease & Molecular Medicine, Department of Clinical Laboratory
Science, Faculty of Health Sciences, University of Cape Town, South Africa
2 International Centre for Genetic Engineering and Biotechnology, Cape Town, South Africa
3 Surgical Gastroenterology Unit, Department of Surgery, Groote Schuur Hospital, Cape
Town, South Africa
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Dr Adeola Henry_Colorectal cancer book chapter 2014

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