KRAS and H1F1a expression in colorectal cancer and its association with the tumor clinicopathological features

doi:10.21203/rs.3.rs-4993609/v1

Download PDF

Research Article

KRAS and H1F1a expression in colorectal cancer and its association with the tumor clinicopathological features

https://doi.org/10.21203/rs.3.rs-4993609/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

Background: Colorectal cancer represents one of the commonest cancers worldwide. It is ranked as the fourth commonest one accounting for nearly 10 % of all cancers. Some tumor markers may help identify the prognosis of the Colorectal cancer . One of those markers is the Kirsten Rat Sarcoma Protein (KRAS). KRAS is one of the proteins important for the transduction cascade of the epidermal growth factor (EGF) and Hypoxia inducible factor HIF1a which is essential mediators of cellular response to hypoxia, regulate gene expression for tumor angiogenesis, glucose metabolism, and resistance to oxidative stress.

Aim: To study the expression of KRAS and H1F1a and its relationship with other clinical and histopathological prognostic factors in patients with Colorectal cancer .

Methods: This is a retrospective immunohistochemical study on 55 resection specimens from 55 Colorectal cancer cases. The pathology specimens were collected from July 2019 to February 2020. Tumor tissues were prepared as formalin-fixed, paraffin-embedded specimens. The paraffin blocks were sectioned at the 5 microns thickness. Then the collected sections were stained with hematoxylin & eosin (H&E) for histopathological revision and immune-histochemical staining for KRAS and H1F1a proteins.

Results: In our sample, only 54% of cases were positive for KRAS expression, and 50.9% were positive for H1F1a. KRAS and H1F1a expression showed no statistically significant relationship with the different clinical, and histopathological parameters including age groups, sex, histological variant, and tumor stages.

Conclusion: immune-histochemical staining staining with KRAS and H1F1a could be a promising modality for screening of mutations of Colorectal cancer with less cost and comparable results to molecular studies. However, the interpretation of our results is limited by the small sample size of our population.

Colo-rectal cancer

KRAS

H1F1a

Colorectal cancer (CRC) represents one of the commonest cancers worldwide. It is ranked as the fourth commonest one accounting for nearly 10% of all cancers (1, 2). CRC affects old males mainly. However, its rates have been increasing among the younger population in different countries (1, 2). CRC is more common in developed countries compared to developing countries (3). In Egypt, CRC represents the ninth most commonest malignancy, although it is a low incidence compared to other countries (4). This signifies the importance of early diagnosis, evaluation of the prognosis, and management of those cases.

The prognosis of CRC depends on many factors including tumor stage according to the TNM staging system (5), histological type, grade, and other microscopic features (6). In addition, the margin status of the tumor (7), desmoplasia (8), host immune response (9), and other variables according to the patient’s clinical characteristics affect the tumor prognosis and treatment response (10).

Moreover, some tumor markers may help identify the prognosis of the CRC, however, there is no accurate prognostic marker and the role of the available markers is still questionable (11).

One of those markers is the Kirsten Rat Sarcoma Protein (KRAS). KRAS is one of the proteins important for the transduction cascade of the epidermal growth factor (EGF). Mutant KRAS leads to uncontrolled stimulation of this cascade that eventually leads to tumorigenesis (12, 13). Mutant KRAS is found in most CRC cases. It is estimated to occur in up to 40% of cases (14).

The clinical role of KRAS is the prediction of tumor response to certain therapeutic agents. One of those agents is the anti-EGF receptors monoclonal antibodies (anti-EGFR mAb). In cases of KRAS-positive CRC, monoclonal antibodies are ineffective in such cases. Mutant KRAS leads to unrestricted activation of the signaling cascade of the EGF. So, tumor cells with the mutant KRAS show no response to anti-EGFR mAbs (15, 16).

However, KRAS protein is still a target of anti-cancer agents as it is found in nearly 20% of all human cancers. Certain drugs target the GTP or GDP binding sites. But the role of those agents is still a query (17–19).

Hypoxia inducible factor 1 alpha (HIF1a), a factor commonly induced in hypoxic conditions, plays a crucial role in neoangiogenesis and metastasis in cancer (20).

HIF1a influences angiogenesis and the metastasis of colorectal cancer cells in experimental study (21).

HIF1a play a vital role in cellular adaptation to hypoxia and regulation of the expression of the glucose metabolism genes, and cell survival, more over HIF1a and related pathways are potential therapeutic targets (22).

.Hence, in this observational immunohistochemical study, we aim to investigate the expression of KRAS and H1F1a protein in CRC patients and correlate its expression with the pathological prognostic aspects of CRC.

Study design & settings:

This is an observational retrospective immunohistochemical study on 55 resection specimens from 55 CRC cases.

Inclusion & Exclusion Criteria:

Patients with CRC at the previously mentioned pathology department during our study period were included. Patients who received preoperative chemotherapy and/or radiotherapy with complete or near-complete response were excluded. This is due to the loss of a major component of the tumor, which would negatively affect the immunohistochemical analysis. A total of five cases that met our exclusion criteria were excluded. There were only 55 cases were eligible for our study.

Clinical data of included patients:

Clinical data of the included cases were collected from the pathology reports. This included sex, age, anatomical site of the tumor, and preoperative therapy.

Histopathological Data of the specimens

The pathology reports were reviewed for histological data and these data were confirmed by H&E slides. Data included histological variant and grade, the presence of mucinous activity perineural invasion, lymphovascular invasion, T & N staging, and the respective stage group (tumor regression score (TRS) according to the 8th edition of AJCC staging system for those who received preoperative therapy.

Specimens’ preparation

Tumor tissues were prepared as formalin-fixed, paraffin-embedded specimens. The paraffin blocks were sectioned at the 5 microns thickness. Then the collected sections were stained with hematoxylin & eosin (H&E) for histopathological revision and immune-histochemical staining for KRAS and H1F1a proteins.

Immunohistochemical Staining technique and analysis:

The sections were baked at 60 degrees, deparaffinized in xylene, then rehydrated through

a series of decreasingly graded concentrations of alcohol (95–70%) ended with distilled

water. After 20 minutes of cooling, the endogenous peroxidase activity was inhibited by

incubation in 3% hydrogen peroxide (H2O2) for 5 minutes. The sections were then boiled

for 5 minutes in citrate solution, with a pH of 6, for antigen retrieval. The sections were

incubated with the respective primary antibody for 1 hour at room temperature and a

dilution of 1:50. The primary antibody used for KRAS detection was a rabbit polyclonal anti-KRAS antibody manufactured by Chongqing Biopsies, China. The primary antibody used for H1F1a detection kit was a rabbit poly coloynal antibod manufactured by abbexa USA, cat No (abx032306), Subsequently, the sections were stained for protein detection in 3, 3-diaminobenzidine for 2 minutes. Afterward, the sections were rinsed well in tap water and then counterstained with hematoxylin to stain the nuclei. Finally, the slides were cleared in xylene and the coverslips were applied. The positive control for KRAS was human pancreatic tissue in which KRAS normally shows cytoplasmic staining in the acinar cells.

KRAS expression was assessed following the methodology of Piton et al (12). KRAS expression was considered positive if at least 10% of the cells were stained by the marker.

Cytoplasmic HIF1a expression was recorded as no, weak, moderate, or strong expression with the percentage of positive tumor cells, HIF1a positivity was defined as the presence of moderate cytoplasmic staining in ≥ 50% of tumor cells or strong expression in any fraction of tumor cells(23).

Statistical Analysis:

The Microsoft Excel 2010 software and SPSS software (Statistical Package for Social Science) version 25 were used for the data entry and statistical analysis respectively. Simple descriptive statistics were used to describe the distribution of various demographic, clinical, and histologic variables as well as the immunohistochemical findings. Inferential statistics were used to determine the existence of a statistically significant relationship between immunohistochemical expression of KRAS, H1F1a and other variables in the study. The chi-square test was used for the analysis of KRAS and H1F1a expression. The level of statistical significance was set at a probability (P) value less than 0.05.

General characteristics and Histological findings of the included cases:

In this study, we included 55 specimens of colorectal adenocarcinoma from 55 CRC patients. The age ranged between 29–83 years, with median 60 years, the mean age of our study population was 56 ± 12. Most of them were females (54%) with a male to female ratio of 6:5. Most of our specimens (63.6%) were obtained from the rectosigmoid anatomical region. Among the 55 included patients only 14 patients (25.5%) received preoperative therapy.

Most of our cases (89.1%) showed the morphology of conventional colorectal adenocarcinoma. In addition, most of our cases (85.5%) were classified as grade II adenocarcinoma according to the two-tier grading system accepted by the WHO (24). Regarding tumor invasion, perineural and vascular invasion were seen in 30.9% and 4.19% of cases. As for the staging of the tumors, 83.6% of cases were classified as T3 and T4 stages. Moreover, 58.2% of cases showed positive lymph node metastasis. Table 1.

Immunohistochemical Expression of KRAS

In the current study, only 54.5% of cases were positive for KRAS expression; Fig. 1. KRAS expression was negative in the rest of the cases; Fig. 2. KRAS expression showed no statistically significant relationship with the different clinical, and histopathological parameters including age groups, sex, histological variant, and tumor stages, There was a significant statistical relationship between KRAS expression cases with H1F1a expression as 63.3% of positive H1F1a cases showed positive KRAS expression Table 2.

Immunohistochemical Expression of H1F1a

28 (50.9%) of the stydied cases were positive for H1F1a expression, staining intensity of H1F1a was weak in 21.8%, moderate in 27.3% and strong in 23.6% and absent in27.3% of the studied cases Fig. 3.

There is no statistical significant relationship between H1F1a expression and the different clinical, and histopathological parameters including age groups, sex, histological variant, and tumor stages Table 3.

Table (1): Clincopathological data of the studied cases

		Parameter	N
Sex	Male	25	45.5%
	Female	30	54.5%
	Total	55	100.0%
Site	Ascending	7	12.7%
	Cecum	6	10.9%
	Descending	4	7.3%
	Rectum	16	29.1%
	Sigmoid	19	34.5%
	Transverse	3	5.5%
	Total	55	100.0%
Variant	Classic	49	89.1%
	Mucinous	5	9.1%
	Signt Ring	1	1.8%
	Total	55	100.0%
Grade	G1	1	1.8%
	G2	47	85.5%
	G3	7	12.7%
	Total	55	100.0%
T stage	T2	9	16.4%
	T3	45	81.8%
	T4	1	1.8%
	Total	55	100.0%
N stage	N0	23	41.8%
	N1	20	36.4%
	N2	12	21.8%
	Total	55	100.0%
V. invasion	Negative	28	50.9%
	Positive	27	49.1%
	Total	55	100.0%
N. invasion	Negative	38	69.1%
	Positive	17	30.9%
	Total	55	100.0%
Therapy	Negative	41	74.5%
	Positive	14	25.5%
	Total	55	100.0%
KRAS	Negative	25	45.5%
	Positive	30	54.5%
	Total	55	100.0%
H1F1a STAINIG	0	15	27.3%
	1	12	21.8%
	2	15	27.3%
	3	13	23.6%
	Total	55	100.0%
H1F1a EXPRESION	Negative	27	49.1%
	Positive	28	50.9%
	Total	55	100.0%

Table (2): KRAS expression and relation with different clinicopathological data

		KRAS				P value
		Negative		Positive
		N	%	N	%
Sex	Male	10	40.0%	15	50.0%	0.458
	Female	15	60.0%	15	50.0%
	Total	25	100.0%	30	100.0%
Site	Ascending	2	8.0%	5	16.7%	0.561
	Cecum	3	12.0%	3	10.0%
	Descending	2	8.0%	2	6.7%
	Rectum	5	20.0%	11	36.7%
	Sigmoid	11	44.0%	8	26.7%
	Transverse	2	8.0%	1	3.3%
	Total	25	100.0%	30	100.0%
Variant	Classic	23	92.0%	26	86.7%	0.626
	Mucinous	2	8.0%	3	10.0%
	Signt Ring	0	0.0%	1	3.3%
	Total	25	100.0%	30	100.0%
Grade	G1	0	0.0%	1	3.3%	0.541
	G2	21	84.0%	26	86.7%
	G3	4	16.0%	3	10.0%
	Total	25	100.0%	30	100.0%
T stage	T2	4	16.0%	5	16.7%	0.649
	T3	21	84.0%	24	80.0%
	T4	0	0.0%	1	3.3%
	Total	25	100.0%	30	100.0%
N stage	N0	10	40.0%	13	43.3%	0.933
	N1	9	36.0%	11	36.7%
	N2	6	24.0%	6	20.0%
	Total	25	100.0%	30	100.0%
V. invasion	Negative	11	44.0%	17	56.7%	0.349
	Positive	14	56.0%	13	43.3%
	Total	25	100.0%	30	100.0%
N. invasion	Negative	19	76.0%	19	63.3%	0.311
	Positive	6	24.0%	11	36.7%
	Total	25	100.0%	30	100.0%
Therapy	Negative	18	72.0%	23	76.7%	0.692
	Positive	7	28.0%	7	23.3%
	Total	25	100.0%	30	100.0%
H1F1a	Negative	16	64.0%	11	36.7%	0.043
	Positive	9	36.0%	19	63.3%
	Total	25	100.0%	30	100.0%

Table (3): H1F1a expression and relation with different clinicopathological data

		H1F1a				P value
		Negative		Positive
		N	%	N	%
Sex	Male	13	48.1%	12	42.9%	0.694
	Female	14	51.9%	16	57.1%
	Total	27	100.0%	28	100.0%
Site	Ascending	2	7.4%	5	17.9%	0.766
	Cecum	4	14.8%	2	7.1%
	Descending	2	7.4%	2	7.1%
	Rectum	7	25.9%	9	32.1%
	Sigmoid	10	37.0%	9	32.1%
	Transverse	2	7.4%	1	3.6%
	Total	27	100.0%	28	100.0%
Variant	Classic	26	96.3%	23	82.1%	0.227
	Mucinous	1	3.7%	4	14.3%
	Signt Ring	0	0.0%	1	3.6%
	Total	27	100.0%	28	100.0%
Grade	G1	1	3.7%	0	0.0%	0.247
	G2	21	77.8%	26	92.9%
	G3	5	18.5%	2	7.1%
	Total	27	100.0%	28	100.0%
T stage	T2	4	14.8%	5	17.9%	0.572
	T3	23	85.2%	22	78.6%
	T4	0	0.0%	1	3.6%
	Total	27	100.0%	28	100.0%
N stage	N0	12	44.4%	11	39.3%	0.459
	N1	11	40.7%	9	32.1%
	N2	4	14.8%	8	28.6%
	Total	27	100.0%	28	100.0%
V. invasion	Negative	17	63.0%	11	39.3%	0.079
	Positive	10	37.0%	17	60.7%
	Total	27	100.0%	28	100.0%
N. invasion	Negative	21	77.8%	17	60.7%	0.171
	Positive	6	22.2%	11	39.3%
	Total	27	100.0%	28	100.0%
Therapy	Negative	19	70.4%	22	78.6%	0.485
	Positive	8	29.6%	6	21.4%
	Total	27	100.0%	28	100.0%

This is an immunohistochemical (IHC) study on 55 resection specimens from 55 CRC cases. KRAS protein was expressed in only 54.5% of cases and H1F1a protein was expressed in 50.9% of the studied cases. However, no significant statistical relationship was founded between KRAS and H1F1a expression and the different clinical, and histopathological parameters including age groups, sex, histological variant, and tumor stages.

However, there was a significant statistical relationship between KRAS expression cases with H1F1a expression as 63.3% of positive H1F1a cases showed positive KRAS expression.

It’s reported that KRAS mutant CRC shows difficulty in management (25). Thus, detection of KRAS mutations is of great importance when treating patients with metastatic CRC. The presence of such mutations means that the patient will not benefit from treatment with anti-EGFR monoclonal antibodies; thus, it is considered a contraindication of giving anti-EGFR monoclonal antibodies (15).

In the current study, we considered assessing the expression of KRAS protein using IHC. The results showed that 54.5% of cases showed positive expression. These are comparable to the findings of Piton et al (12). and Elsabah et al. (26). They found that KRAS was positive in 33% and 42% of their cases, respectively.

In their study, Elsabah et al (26) reported that 42.3% of their patients showed positive KRAS staining of their cytoplasm. With strong positivity in nearly 27% of them. In addition, they observed a variation in the expression of KRAS according to the histological type of colorectal carcinoma. The mucinous and signet ring types showed negative results on staining. On the other hand, 50% of the adenocarcinoma samples were positive for K-RAS. This was similar to current findings, as KRAS staining was positive in 47.3% of the conventional variants, while 60% in the mucinous type and one out of two cases of signet ring variant.

As for clinical grading of the tumor, 55% and 42.8% of grade 2 and grade 3 showed positive results respectively. This was relatively similar to what was reported by El Sabah et al (26), who found that 50% and 39% of grade 2 and grade 3 showed positive KRAS staining respectively. However, there was no significant difference between both types; p = 0.54 which is similar to Elsabah results; p = 0.683. Moreover, many studies found no significant relationship between KRAS mutation expression and the histological grade (30–33).

It is also of note that immunohistochemical expression of KRAS did not statistically correlate with any of the other parameters in our study. This was consistent with the results of previous studies on KRAS overexpression (29, 30). However, Sammoud et al reported a positive correlation between age and KRAS expression (31).

KRAS expression can be compared to the figures found using molecular techniques, which show that 35–46% of colorectal cancers carry KRAS mutations (32). However, Akkiprik et al. (27) failed to identify a significant relationship between KRAS overexpression and KRAS mutation. They explained this finding by the fact that during IHC stating, the used KRAS antibodies interact and detect all expressed proteins whether mutant or wild types (27). Nevertheless, in previous studies by Akkiprik et al (27) and Okulczyk et al. (29) on metastatic CRC; analysis of Dukes’ staging against KRAS mutation using PCR techniques revealed no significant difference between them.

On the other hand, other studies did not establish a relationship between K-RAS mutations and distant metastasis (28, 30, 33, 34). Moreover, Piton et al (12) inquire about the efficacy of immunohistochemistry compared to the molecular biology tools in the detection of KRAS and BRAF mutations in CRC. They found that IHC staining is of poor sensitivity (27%) and specificity (64%) for KRAS protein detection. However, IHC staining for BRAF detection showed excellent sensitivity and specificity with 100% each. This makes IHC staining a questionable modality for the detection of the K-RAS protein compared to other mutations. However, IHC seems to be a promising tool with less cost compared to molecular modalities. It is reported that molecular testing for KRAS cost up to 13 million dollars annually with 452 dollars per patient. On the other hand, IHC staining costs nearly 50 dollars per antibody test (12, 35).

H1F1a expression was found in 50.9 of cases, with no statistical significant relation with clinical and histopathological parameter. Lee et al (36), and Saka et al (37), studies found HIF1a expression was not related to clinicopathological characteristics or prognosis

Compared to study done by Zhi-Yin et al (38), who found H1F1a expression in 68.4% of cases, they reported that high HIF1a expression is related to advanced TNM stages (P = 0.025). Moreover, HIF1a expression is significantly associated with the possibility of distant metastasis, vascular invasion (P = 0.048), and might also be related to the possibility of lymphatic metastasis (P = 0.041). Depending on this finding they stated that new insights into the role of HIF1a in colon carcinoma and it could potentially work as a biomarker in the prediction of metastasis and poor prognosis.

Another study done by Baba et al (39), demonstrated that HIF1a expression was found in 19% of cases, it was significantly associated with higher colorectal cancer-specific mortality, but they did not find any significant relationship with TNM stage. They suggested that HIF1a is a biomarker with potentially important therapeutic implications.

This discrepancy might be due to different method of scoring HIF1a expression, clone used for staining and different sample sizes.

Alot of experiments is Currently concerning cancer therapy are targeting HIF1a, which is closely associated with tumor angiogenesis, metabolism, and proliferation (40).

A novel HIF1a inhibitor, IDF-11774, is proved to decrease the energy production of colorectal cancer (41). Combination treatment with a specific HIF1a inhibitor was also recommended for drug resistant colorectal cancer (42).

There was a significant statistical relationship between KRAS expression cases with H1F1a expression as 63.3% of positive H1A1a cases showed positive KRAS expression.

Baba et al (39), studied the relation between H1F1a immunohistochemical expression and Sequencing of KRAS by PCR, 41% of positive H1F1a cases showed positive KRAS mutation.

Study done by Kikuchi et al (43), has shown that HIF1a translation is regulated by KRAS and BRAF in colon cancer.

It is important to note that the IHC study is a preliminary test for KRAS and H1F1a proteins detection. Confirmation of KRAS and H1F1a expression needs PCR techniques for gene mutation detection. Yet, the immunohistochemical results of othis study need to be challenged against molecular techniques to verify whether those cases that expressed the protein do carry the mutations or not.

Limitations

The sample size of the study represents the major limitation. It was relatively small; which makes it difficult to give a representative result. Therefore, this should be considered in the upcoming trials.

IHC staining could be a promising modality for screening of mutations of CRC with less cost and comparable results to molecular studies. However, the interpretation of our results is limited by the small sample size of our population.

CRC	Colorectal cancer
KRAS	Kirsten Rat Sarcoma Protein
EGF	epidermal growth factor
HIF1a	Hypoxia inducible factor

Ethics approval and consent to participate: This study was reviewed and approved by the ethical committee, Faculty of Medicine, Fayoum University, Fayoum, Egypt (Approval No. R588).
Consent for publication: All the authors have approved the manuscript for submission
Funding: Not applicable

Author Contribution

in this manuscript all the three authors collected the samples and patients data from the pathology labs. in their institutions. Both Hosain and Byoumi supervised and revised the preparation of the H&E and immunostained slides. the 3 authors revised and interpret the slides. Elmahdi wrote the manuscript and sent it as the corresponding author. All authors reviewed the manuscript and approved its submission.

Rawla P, Sunkara T, Barsouk A. Epidemiology of colorectal cancer: incidence, mortality, survival, and risk factors. Przegla̜d Gastroenterol. 2019;14(2):89.
Haggar FA, Boushey RP. Colorectal Cancer Epidemiology: Incidence, Mortality, Survival, and Risk Factors. Clin Colon Rectal Surg. 2009;22(4):191.
Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin [Internet]. 2021; Available from: https://pubmed.ncbi.nlm.nih.gov/33538338/
World Health Organization. Egypt Source: Globocan 2020 Number. Int agency Res cancer. 2020;895:1–2.
Bellizzi AM, Montgomery EA, Hornick JL. American Registry of Pathology Expert Opinions: Evaluation of poorly differentiated malignant neoplasms on limited samples - Gastrointestinal mucosal biopsies. Ann Diagn Pathol. 2020;44:151419.
Sato Y, Kudo S-E, Ichimasa K, Matsudaira S, Kouyama Y, Kato K, et al. Clinicopathological features of T1 colorectal carcinomas with skip lymphovascular invasion. Oncol Lett. 2018;16(6):7264–70.
Lee JM, Chung T, Kim KM, Simon NSM, Han YD, Cho MS, et al. Significance of Radial Margin in Patients Undergoing Complete Mesocolic Excision for Colon Cancer. Dis Colon Rectum. 2020;63(4):488–96.
Ueno H, Kanemitsu Y, Sekine S, Ishiguro M, Ito E, Hashiguchi Y, et al. A Multicenter Study of the Prognostic Value of Desmoplastic Reaction Categorization in Stage II Colorectal Cancer. Am J Surg Pathol. 2019;43(8):1015–22.
Haruki K, Kosumi K, Li P, Arima K, Väyrynen JP, Lau MC, et al. An integrated analysis of lymphocytic reaction, tumour molecular characteristics and patient survival in colorectal cancer. Br J Cancer. 2020;122(9):1367–1377.
Balhareth A, Aldossary MY, McNamara D. Impact of physical activity and diet on colorectal cancer survivors’ quality of life: a systematic review. World J Surg Oncol. 2019;17(1):153.
Dinu D, Dobre M, Panaitescu E, Bîrlă R, Iosif C, Hoara P, et al. Prognostic significance of KRAS gene mutations in colorectal cancer–preliminary study. J Med Life. 2014;7(4): 581–7.
Piton N, Borrini F, Bolognese A, Lamy A, Sabourin JC. KRAS and BRAF Mutation Detection: Is Immunohistochemistry a Possible Alternative to Molecular Biology in Colorectal Cancer? Gastroenterol Res Pract. 2015;2015.
Huang L, Guo Z, Wang F, Fu L. KRAS mutation: from undruggable to druggable in cancer. Signal Transduct Target Ther. 2021;6(1):386.
Tan C, Du X. KRAS mutation testing in metastatic colorectal cancer. World J Gastroenterol. 2012;18(37):5171–80.
Saridaki Z, Georgoulias V, Souglakos J. Mechanisms of resistance to anti-EGFR monoclonal antibody treatment in metastatic colorectal cancer. World J Gastroenterol. 2010;16(10):1177–87.
Soulières D, Greer W, Magliocco AM, Huntsman D, Young S, Tsao M-S, et al. KRAS mutation testing in the treatment of metastatic colorectal cancer with anti-EGFR therapies. Curr Oncol. 2010;17 Suppl 1(Suppl 1):S31-40.
Jancík S, Drábek J, Radzioch D, Hajdúch M. Clinical relevance of KRAS in human cancers. J Biomed Biotechnol. 2010;2010:150960.
Pantsar T. The current understanding of KRAS protein structure and dynamics. Comput Struct Biotechnol J. 2020;18:189–98.
Han CW, Jeong MS, Jang SB. Understand KRAS and the Quest for Anti-Cancer Drugs. Cells. 2021;10(4).
Li H, Rokavec M, Jiang L, Horst D, Hermeking H. Antagonistic effects of p53 and HIF1A on microrna-34a regulation of ppp1r11 and stat3 and hypoxia-induced epithelial to mesenchymal transition in colorectal cancer cells. Gastroenterology 2017; 153: 505–520.
Xiang J, Sun H, Su L, Liu L, Shan J, Shen J, Yang Z, Chen J, Zhong X, Ávila MA, Yan X, Liu C, Qian C. Myocyte enhancer factor 2D promotes colorectal cancer angiogenesis downstream of hypoxia-inducible factor 1alpha. Cancer Lett 2017; 400: 117–126.
Narita T, Yin S, Gelin CF, Moreno CS, Yepes M, Nicolaou KC, Van Meir EG: Identification of a novel small molecule HIF-1_ translation inhibitor. Clin Cancer Res 2009, 15:6128–6136.
Yoshifumi Baba,* Katsuhiko Nosho,* Kaori Shima,* Natsumi Irahara,* Andrew T. Chan,† Jeffrey A. Meyerhardt,* Daniel C. Chung,†Edward L. Giovannucci,‡§ Charles S. Fuchs,*‡ and Shuji Ogino*¶. HIF1A Overexpression Is Associated with Poor Prognosis in a Cohort of 731 Colorectal Cancers. The American Journal of Pathology, Vol. 176, No. 5, May 2010.
Nagtegaal ID, Odze RD, Klimstra D, Paradis V, Rugge M, Schirmacher P, et al. The 2019 WHO classification of tumours of the digestive system. Histopathology. 2020;76(2):182–8.
Meng M, Zhong K, Jiang T, Liu Z, Kwan HY, Su T. The current understanding on the impact of KRAS on colorectal cancer. Biomed Pharmacother. 2021;140(February):111717.
Elsabah MT, Adel I. Immunohistochemical assay for detection of K-ras protein expression in metastatic colorectal cancer. J Egypt Natl Canc Inst. 2013;25(1):51–6.
Akkiprik M, Celikel CA, Düşünceli F, Sönmez O, Güllüoğlu BM, Sav A, et al. Relationship between overexpression of ras p21 oncoprotein and K-ras codon 12 and 13 mutations in Turkish colorectal cancer patients. Turkish J Gastroenterol Off J Turkish Soc Gastroenterol. 2008;19(1):22–7.
Schimanski CC, Linnemann U, Berger MR. Sensitive detection of K-ras mutations augments diagnosis of colorectal cancer metastases in the liver. Cancer Res. 1999;59(20):5169–75.
Okulczyk B, Kovalchuk O, Piotrowski Z, Myśliwiec P, Chyczewski L. Clinical usefulness of K-RAS mutation detection in colorectal cancer and in surgical margins of the colon. Rocz Akad Med Bialymst. 2004;49 Suppl 1:52–4.
Liu X, Jakubowski M, Hunt JL. KRAS gene mutation in colorectal cancer is correlated with increased proliferation and spontaneous apoptosis. Am J Clin Pathol. 2011;135(2):245–52.
Sammoud S, Khiari M, Semeh A, Amine L, Ines C, Amira A, et al. Relationship between expression of ras p21 oncoprotein and mutation status of the K-ras gene in sporadic colorectal cancer patients in Tunisia. Appl Immunohistochem Mol Morphol AIMM. 2012;20(2):146–52.
Bylsma LC, Gillezeau C, Garawin TA, Kelsh MA, Fryzek JP, Sangaré L, et al. Prevalence of RAS and BRAF mutations in metastatic colorectal cancer patients by tumor sidedness: A systematic review and meta-analysis. Cancer Med. 2020;9(3):1044–57.
Dahabreh IJ, Terasawa T, Castaldi PJ, Trikalinos TA. Systematic review: Anti-epidermal growth factor receptor treatment effect modification by KRAS mutations in advanced colorectal cancer. Ann Intern Med. 2011;154(1):37–49.
Milano G, Etienne-Grimaldi M-C, Dahan L, Francoual M, Spano J-P, Benchimol D, et al. Epidermal growth factor receptor (EGFR) status and K-Ras mutations in colorectal cancer. Ann Oncol Off J Eur Soc Med Oncol. 2008;19(12):2033–8.
Muirhead D, Aoun P, Powell M, Juncker F, Mollerup J. Pathology economic model tool: a novel approach to workflow and budget cost analysis in an anatomic pathology laboratory. Arch Pathol Lab Med. 2010;134(8):1164–9.
Lee SJ, Kim JG, Sohn SK, Chae YS, Moon JH, Kang BW, Park JS, Park JY, Choi GS. No association of the hypoxia-inducible factor-1alpha gene polymorphisms with survival in patients with colorectal cancer. Med Oncol 2011; 28: 1032–1037.
Saka B, Ekinci O, Dursun A, Akyurek N. Clinicopathologic and prognostic significance of immunohistochemical expression of HIF-1α, CXCR4 and CA9 in colorectal carcinoma. Pathol Res Pract 2017; 213: 783–792.
Zhi-Yin Huang1,2, Lin-Hao Zhang2, Chong Zhao1,3, Rui Liu1,2,3, Huan Tong2, Can Gan2, Tian Lan2, Cheng-Wei Tang1,2,3, Jin-Hang Gao1,2,3. High HIF-1α expression predicts poor prognosis of patients with colon adenocarcinoma. Int J Clin Exp Pathol 2018;11(12):5635–5646.
Baba Y, Nosho K, Shima K, Irahara N, Chan AT, Meyerhardt JA, Chung DC, Giovannucci EL, Fuchs CS, Ogino S. HIF1A overexpression is associated with poor prognosis in a cohort of 731 colorectal cancers. Am J Pathol 2010; 176: 2292–2301.
Kheshtchin N, Arab S, Ajami M, Mirzaei R, Ashourpour M, Mousavi N, Khosravianfar N, Jadidi-Niaragh F, Namdar A, Noorbakhsh F, Hadjati J. Inhibition of HIF-1α enhances anti-tumor effects of dendritic cell-based vaccination in a mouse model of breast cancer. Cancer Immunol Immunother 2016; 65: 1159–1167
Ban HS, Kim BK, Lee H, Kim HM, Harmalkar D, Nam M, Park SK, Lee K, Park JT, Kim I, Lee K, Hwang GS, Won M. The novel hypoxia-inducible factor-1alpha inhibitor IDF-11774 regulates cancer metabolism, thereby suppressing tumor growth. Cell Death Dis 2017; 8: e2843.
Wang Y, Lei F, Rong W, Zeng Q, Sun W. Positive feedback between oncogenic KRAS and HIF-1αlpha confers drug resistance in colorectal cancer. Onco Targets Ther 2015; 8: 1229–1237.
Kikuchi H, Pino MS, Zeng M, Shirasawa S, Chung DC: Oncogenic KRAS and BRAF ifferentially regulate hypoxia-inducible factor-1_ and – 2_ in colon cancer. Cancer Res 2009, 69:8499–8506.

No competing interests reported.

Download PDF

Version 1

posted

You are reading this latest preprint version

KRAS and H1F1a expression in colorectal cancer and its association with the tumor clinicopathological features

Status:

Version 1

Abstract

Figures

BACKGROUND

METHODS

Study design & settings:

Inclusion & Exclusion Criteria:

Clinical data of included patients:

Histopathological Data of the specimens

Specimens’ preparation

Immunohistochemical Staining technique and analysis:

Statistical Analysis:

RESULTS

General characteristics and Histological findings of the included cases:

Immunohistochemical Expression of KRAS

Immunohistochemical Expression of H1F1a

DISCUSSION

Limitations

CONCLUSION

Abbreviations

Declarations

Author Contribution

References

Additional Declarations

Status:

Version 1