DOI:http://dx.doi.org/10.7314/APJCP.2012.13.5.

2171
Improved Diagnosis of Pancreatic Diseases with Various Novel Serum Biomarkers in Nepal

RESEARCH COMMUNICATION

Improved Diagnostic Accuracy of Pancreatic Diseases with

a Combination of Various Novel Serum Biomarkers - Case
Control Study from Manipal Teaching Hospital, Pokhara,
Nepal
Mohammad Shamim Farooqui1*, Ankush Mittal2, Bibek Poudel2, Suhas Kumar
Mall1, Brijesh Sathian3, Mohammad Tarique4, Mohammad Hibban Farooqui5

Abstract
Background: Pancreatic cancer is a distressing disease with a miserable prospects and early recognition
remains a challenge due to ubiquitous symptomatic presentation, deep anatomical location, and aggressive
etiology. False positives and problems in distinguishing pancreatitis from adenocarcinoma limit the use of CA 19-9
as both disorders can present with similar symptoms and share radiographic physiognomies. This study aimed
to assess the relative increase in accuracy of diagnosing the patients with chronic pancreatitis, benign neoplasm
of pancreas and adenocarcinomas with CA 19-9, haptoglobin, and serum amyloid A in comparison to CA 19-9
alone. Materials and Methods: This hospital based case control study was carried out in the Departments of
Medicine and Biochemistry of Manipal Teaching Hospital, Pokhara, Nepal, between 1st January 2010 and 31st
December 2011. The variables assessed were age, gender, serum CA19-9, serum haptoglobulin, serum Amyloid A.
The data were analyzed using Excel 2003, R 2.8.0 Statistical Package for the Social Sciences (SPSS) for Windows
Version 16.0 (SPSS Inc; Chicago, IL, USA) and the EPI Info 3.5.1 Windows Version. Results: Out of 197 cases
of pancreatic disease, maximum number of assumed cases were of adenocarcinoma of pancreas (95). Number
of males (59) were more than females (36) in assumed cases of adenocarcinoma of pancreas. The mean values of
CA19-9 raised considerably in cases of chronic pancreatitis, benign neoplasm and adenocarcinoma of pancreas
when compared to controls. The highest augmention in CA19-9 values were in cases of adenocarcinoma of
pancreas. The p-value indicates that in cases of chronic pancreatitis, there was not significant increase in precision
of diagnosis. Conclusions: These statistics established that haptoglobin and SAA are useful in discriminating
cancer from benign conditions as well as healthy controls.
Keywords: Pancreatic diseases - serum biomarkers - Nepal

Asian Pacific J Cancer Prev, 13, 2171-2174

Introduction is CA19-9. The false positives results in distinguishing

pancreatitis from adenocarcinoma of pancreas limit the
Pancreatic cancer is a distressing disease with a use of CA 19-9 as both disorders can present with similar
miserable prospects and early recognition remains a symptoms and share radiographic physiognomies. No
challenge due to ubiquitous symptomatic presentation, single test could effectively discriminate PA from benign
deep anatomical location, and aggressive etiology. conditions. The current absence of reliable biomarker
Pancreatic cancer is currently fourth leading cause of testing for pancreatic cancer mandates the development
cancer-related deaths in the United States and on the of novel strategies for identifying and characterizing
whole 5-year survival has been accounted to be less additional biomarkers. One of novel biomarker is serum
than 5% (Greenlee et al., 2000). In 2010, an estimated amyloid A (SAA). Serum amyloid A (SAA) which shares
newly diagnosed cases of pancreatic cancer were antigenicity with amyloid AA, has been recognized as
43,140 in USA and amazingly 36,800 cases perishes acute-phase reactant (Kushner et al., 1994). The infection,
from the disease(Randall et al., 2011). The process of trauma, or presence of malignant disease, may results
pancreatic carcinogenesis is a multifactorial phenomenon in the elevation of plasma SAA (Biran et al., 1999).
(Shrikhande et al., 2009). The most widely used and best Another biomarker is haptoglobulin, which is glycoprotein
validated tumor marker for adenocarcinoma of pancreas produced in liver. The appearance of fucosylated
Manipal College of Medical Sciences, 2Department of Biochemistry, 3Department of Community Medicine, Manipal College of
1

Medical Sciences, Pokhara, Nepal 4Department of Surgery, Katihar Medical College,Katihar, India 5Department of ENT, Turaf
General Hospital, Turaf, Kingdom of Saudi Arabia *For correspondence: drfrqms@aol.com
Asian Pacific Journal of Cancer Prevention, Vol 13, 2012 2171
Mohammad S Farooqui et al
haptoglobin has been reported in various diseases such Table 1. Distribution of Cases and Controls According
as pancreatic cancer, hepatocellular carcinoma, liver to their Gender, Types and Stages
cirrhosis, gastric cancer and colon cancer (Okuyamaet Genderwise variation No. Age (Range)
al., 2006). The enhanced diagnostic precision over CA
19-9 alone and the ability to discriminate between PA Controls
and pancreatitis have become the minimal standard for Total 170 65 (34-86)
Female 68 63 (34-81)
validation of novel serum biomarkers. This study aims
Male 102 67 (37-86)
to assess the relative increase in accuracy of diagnosing Suspected Cases
the patients with chronic pancreatitis, benign neoplasm Chronic inflammation of Pancreas
of pancreas and adenocarcinomas with CA 19-9, Total 45 47 (28-70)
haptoglobin, and serum amyloid A in comparison to CA Female 21 48 (28-63)100.0
19-9 alone. Male 24 46 (29-70) 6.3
Benign tumor of pancreas
Total 57 73 (45-85)
Materials and Methods Male 24 75 (45-82) 75.0
Female 33 71 (48-85)
It was a hospital based case control study carried out in
Distribution of cases according to its type 56.3
the Department of Medicine and Biochemistry of Manipal Intraductal Papillary Mucinous Tumor 19
Teaching Hospital, Pokhara, Nepal between 1st January Mucinous Cystic Neoplasm 13
50.0
2010 and 31st December 2011. The variables collected Neuroendocrine Tumor 13
were age, gender, serum CA19-9, serum haptoglobulin, Serous Cystadenoma 6
Serum Amyloid A. Approval for the study was obtained Cystic Neuroendocrine Neoplasm 6 25.0
from the institutional research ethical committee. Solid-Cystic Papillary Neoplasm 2
Quantitative Analysis of Human Serum Amyloid A and Maligmant carcinoma of pancreas (Adenocarcinomas) 31.3

CA19-9 was performed by ELISA reader for all cases. The Total 95 72 (54-90)
Male 59 69 (57-90) 0
standard procedure was followed as per manufacturer’s
Female 36 75 (54-87)
instructions for ELISA with minor modifications (Sell.,

Newly diagnosed without treatment

Distribution of cases according to stages
1990). Estimation of serum haptoglobins was done by Stage IA 4
colorimetric method that was based on the peroxidase Stage IB 8
activity of haptoglobin-methaemoglobin complexes Stage IIA 9
(Owen et al., 1960). All these laboratory parameters Stage IIB 27
were analyzed using Human reagent kits and with the Stage III 21
help of ELISA and semi autoanalyser (Humalyser 3500, Stage IV 26
Germany). Analysis was done using descriptive statistics
and Confidence Interval (CI). The data was analyzed were more than females(36) in assumed cases of
using Excel 2003, R 2.8.0 Statistical Package for the adenocarcinoma of pancreas. Number of patients
Social Sciences (SPSS) for Windows Version 16.0 (SPSS suspected of Stage IIB (27) and Stage IV(26) were
Inc; Chicago, IL, USA) and the EPI Info 3.5.1 Windows almost equal. The total number of suspected cases of
Version. benign tumor of pancreas were 57 and females were more
Inclusion criteria: Suspected cases of chronic prone to benign tumor of pancreas. Maximum number of
pancreatitis were enrolled as evidenced by 2 out of suspected cases were of Intraductal Papillary Mucinous
the following 4 criteria: (a) calculi on abdominal X-ray, Tumor(19). Furthermore ,the total number of chronic
(b) dilated pancreatic duct on ultrasound, (c) changes pancreatitis were 45 and median age was 47 years. For
of pancreatitis on ERCP and/or MRCP and (d) EUS comparison with cases, healthy controls (170) were also
appearances of pancreatitis. Mass lesions on ultrasound taken with average age of 65 yrs.
or CT scan for suspected cases of benign and malignant Table 2 illustrates that mean values of CA19-9
neoplasm of pancreas. Confirmation of pancreatic cancer raised considerably in cases of chronic pancreatitis,
was done by histological confirmation of malignancy benign neoplasm and adenocarcinoma of pancreas when
by biopsy (radiology/endoscopy or surgery) and compared to controls. The highest augmention in CA19-
establishment of benign nature of disease by follow-up 9 values were in cases of adenocarcinoma of pancreas.
of more than 3 years without any evidence of recurrence There was inconsequential increase in cases of chronic
of mass or metastatic disease. pancreatitis in comparison to controls for haptoglobulin.
Exclusion criteria: The patients suffering from any There was noteworthy enhancement in haptoglobulin
other disease except chronic pancreatitis, benign neoplasm levels in cases of benign neoplasm and adenocarcinoma
and adenocarcionma of pancreas were excluded from our of pancreas. Similarly, there was significant increase in
study. mean values of serum Amyloid A in cases of chronic
pancreatitis, benign neoplasm and adenocarcinoma of
Results pancreas when compared to controls.
Table 3 depicts that there was significant difference in
Table 1 illustrates that out of 197 cases of pancreatic cases of pancreatic adenocarcinomas and benign neoplasm
disease, maximum number of assumed cases were of of pancreas when percentage of accuracy was calculated
adenocarcinoma of pancreas(95). Number of males(59) with the help of other serum markers i.e. haptoglobulin
2172 Asian Pacific Journal of Cancer Prevention, Vol 13, 2012
 DOI:http://dx.doi.org/10.7314/APJCP.2012.13.5.2171
Improved Diagnosis of Pancreatic Diseases with Various Novel Serum Biomarkers in Nepal
Table 2. Mean Values of CA19-9, Haptoglobulin, Serum Amyloid A in Controls and Cases
Variables Controls Chronic pancreatitis Benign neoplasm Adenocarcinomas p Value
of pancreas of pancreas
CA19-9 24.74±06.38 114.11±87.76† 204.53±097.73† 561.21±315.63† 0.0001**
(<40 U/ml) (23.77,25.70) (87.74,140.48) (178.59,230.46) (496.91,625.51)
Haptoglobulin 118.53±059.78 134.22±69.83 142.58±078.56† 178.48±069.28† 0.0001**
(33 to 213 mg/dl) (109.48,127.58) (113.24,155.20) (121.73,163.43) (164.37,192.60)
Serum Amyloid A 4.41±1.35 6.86±1.50† 9.49±03.50† 15.35±03.59† 0.0001**
(<8ug/ml) (4.21,4.62) (6.41,7.31) (8.56,10.42) (14.61,16.08)
p-Value <0.001, statistically significant between controls vs cases groups, **p-Value <0.05, statistically significant
†
100.0
Table 3. Diagnostic Precision of the Haptoglobin/SAA/ combination
6.3 other 10.1
biomarkers(88.8%). In contrast to that 1
CA19-9 in Comparison to CA19-9 Alone the recognition of true positive cases in cases of benign
20.3
neoplasm(77.19%) and pancreatic adenocaercinoma
Cases CA19-9* Haptoglobulin/SAA/CA19-9* p-Value 75.0 25.0 30.0
(83.2%) had been increased in comparison of suspected
Chronic inflammation of Pancreas cases of benign neoplasm
46.8
(73.7%) and adenocarcinoma
91.1% (41/45) 88.8% (40/45) 0.153 56.3
of pancreas(80.0%)(Okuyamaet al.,2006). The synthesis 5
Benign tumor of pancreas 50.0of acute-phase protein is mainly takes place
54.2 in cancer cell
73.7% (42/57) 77.19% (44/57) 0.0001** 31.3
lines that reflect the growth of malignancy and largely 30.0
Pancreatic Adenocarcinomas
regulated by inflammation- associated cytokines such as
80.0 %(76/95) 83.2% (79/95) 0.0001**
IL-1, IL-6, and tumor necrosis factor. Serum amyloid A
* Percentage (Confirmed/suspected) 25.0
synthesis is induced in various tissues by the stimulation
38.0 100.0 3
of inflammation-related
31.3 cytokines.
23.7 Haptoglobin
31.3 is a 30.0
6
and serum amyloid along with CA19-9. The p-value glycoprotein produced in the liver. Other studies also
indicates that in cases of chronic pancreatitis, there was 0illustrates that interleukin-6 (IL-6) increases the
not significant increase in precision of diagnosis. secretion of fucosylated haptoglobin from pancreatic 75.0

None
Remission
Persistence or recurrence
Newly diagnosed without treatment

Newly diagnosed with treatment

cancer tissue Both haptoglobin and serum amyloid
Discussion A are acute-phase proteins play role in angiogenesis, 56
antioxidant, cell migration, lipid metabolism, induction 50.0
Chronic pancreatitis, benign neoplasm of pancreas and of extracellular matrix degrading enzymes , conscription
pancreatic cancer are accountable for most of the burden of inflammatory cells to sites of inflammation, tumor
of exocrine pancreatic disease (Raimondi et al., 2010). growth, metastasis, and neovascularization (Narisada M et
Even though improvements in imaging techniques, it may al., 2008). Although synthesized mainly in the liver, local 25.0
be problematic to segregate inflammatory head masses, differential expression of haptoglobin and serum amyloid 31
benign grazes from malignant masses. Discrepancy A has been demonstrated in cancer tissues. Expression in
between benign (inflammatory) and malignant masses has cancer cells as well as potential roles in angiogenesis, cell 0
important therapeutic repercussions – evade unnecessary migration and extracellular matrix remodeling suggests
resection in inflammatory masses (Boll et al., 2003). that haptoglobin and SAA may directly contribute to
various complement of proteins and antigens synthesized tumorigenesis. Additionally, it is likely that levels of
by tumor cells indcate that no single common marker proteins secreted or released by the tumor will correlate
would be effective in diagnosing of disease. Numerous together and therefore mitigate the advantage of their
prospective serum and tissue markers for pancreatic cancer use in combination. Accurate diagnostic and prognostic
are presently enduring evaluation, none are sufficiently biomarkers for PA could improve outcomes through early
validated for routine clinical use (Benson et al., 2007). detection, selection of appropriate treatment strategies,
Thus, CA 19-9 which remains the serum pancreatic cancer monitoring intervention efficacy, and surveillance
marker, new markers in combination for this malignancy of groups at high-risk for developing PA. Thus, the
should be judged (Katz et al., 2008). In our present study, evaluation of serum biomarker levels not only capable
mean values of CA 19-9 in cases of chronic pancreatitis of discriminating and diagnosing pancreatic cancer from
114.11 ± 87.76 (87.74,140.48) and benign neoplasm benign conditions and chronic pancreatitis with high
204.53±97.73(178.59,230.46) were below 300U/ml. sensitivity and specificity, but also offer improved insight
The mean levels of CA19-9 in cases of adenocarcinoma into the complex network of factors involved in pancreatic
of pancreas was 561.21±315.63 (496.91, 625.51). Our tumorigenesis.
findings concurred with the findings of Rocha et al which In conclusions, these statistics established that
revealed that an elevated CA 19-9 greater than 300 U/ haptoglobin and SAA are useful in discriminating PA from
mL in the setting of head mass with chronic pancreatitis benign conditions as well as healthy controls. This study
stalwartly advocates malignancy (Rocha et al., 2007). In supports the use of combined biomarkers for improved
our present study, the mean levels of both haptoglobulin accuracy in the diagnosis of pancreatic adenocarcinomas.
and serum amyloid had found to be increased linearly with
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