Relationship and Clinical Significance of ALDH1A1 and CD133 Cancer Stem Cell Markers and Mutant p53 in Lung Adenocarcinoma

doi:10.21203/rs.3.rs-948192/v1

Background

Cancer stem cells (CSCs) are major contributors to the malignant transformation of cells because of their capacity for self-renewal. Although several CSC markers have been identified in various tumor types, they are primarily used as cancer-prediction markers and for the isolation of CSC populations. Aldehyde dehydrogenase1A1 (ALDH1A1) and CD133 are promising candidate of CSC markers in non-small cell lung cancer (NSCLC). Furthermore, TP53 is frequently mutated in lung cancer, and the loss of its function is associated with malignant characteristics. However, the relationship between CSCs and mutant p53 in lung adenocarcinoma is not well-established. We examined the expression of ALDH1A1, CD133, and mutant p53 in lung adenocarcinoma patients and conducted a clinicopathological study.

Methods

The subjects of the study were 194 patients who underwent surgery for lung adenocarcinoma. ALDH1A1, CD133, and mutant p53 were immunohistochemically stained in resected specimens from these patients.

Results

Triple-negative cases without ALDH1A1, CD133, and mutant p53 expression in lung adenocarcinoma were shown to have a much better prognosis than others.

Conclusions

Our present results suggest that detection of CSC markers and mutant p53 by immunohistochemical staining may be effective in therapeutic strategies for lung adenocarcinoma.

Cancer Biology

Oncology

Stem Cell & Developmental Cell Biology

Cancer stem cells

Aldehyde dehydrogenase1A1

TP53

clinicopathological study

The recurrence rate of lung cancer is as high as 80% even in early stages that are treated with chemotherapy [1], and almost one-quarter of all cancer-related deaths are due to lung cancer [2]. Several molecular-targeted drugs and immune checkpoint inhibitors have been approved for lung cancer, but no significant improvements in survival have been observed.

Cancer stem cells (CSCs) can self-renew and differentiate [3], and may be highly resistant to chemotherapy and radiation, resulting in tumor initiation, progression, and metastasis [4]. Accumulating evidence supports the existence of a CSC phenotype in human lung cancer, and several CSC markers, such as ALDH1A1, CD133, and CD44, have been characterized in lung cancer [5].

ALDH1 proteins (mainly ALDH1A1, ALDH1A2, and ALDH1A3) are primarily localized in the cytosol of cells in various tissues and let aliphatic aldehyde oxidize at retina. Member of the ALDH family has a main biological role in cell protection through the detoxification of aldehydes, but they also regulate cell proliferation, differentiation, and survival [6]. And especially, ALDH1A1 is a putative hematopoietic stem cell marker associated with drug resistance that increased of many cancer types [7].

CD133 antigen, also known as prominin-1, is a member of the pentaspan transmembrane glycoprotein family that is specifically located to cellular protrusions. In fact, it is widely utilized as a CSC biomarker in various types of cancer, including liver, stomach, kidney, and lung cancer [8–11].

We previously reported that lung adenocarcinoma that is negative for both CD133 and ALDH1A1 had a significantly better prognosis regarding both overall survival (OS) and disease-free interval (DFI) [12]. Furthermore, we reported that patients with stage I+II lung adenocarcinoma who were negative for CD133 had a significantly better DFI [13]. Thus, there is certainly a close correlation between CSCs and lung adenocarcinoma.

TP53, one of tumor suppressor genes, has some functions in gene transcription, cell division, and DNA repair [14–16]. Conversely, TP53 gene mutations are the most common genetic abnormalities in human malignancies and are also frequently detected in primary lung cancer; mutant p53 is observed in 70% of small cell lung cancers and 47% of non-small cell lung cancers [17]. In fact, cases of surgically resectable non-small cell lung cancer (NSCLC) with mutant p53 have a significantly poorer prognosis, and it has been reported that mutant p53 can be an independent prognostic factor [18].

To date, there have been many reports about CSCs and p53, but few have investigated the association between CSCs and p53 in lung cancer. Here, we scrutinized the expression and prognosis of CSC marker-positive cells locally in resected samples from patients with lung adenocarcinoma and then explored the association between CSC markers and mutant p53 expression.

Of 286 patients with lung adenocarcinoma, 194 (67.8%) were eligible for this study. The clinicopathological parameters are shown in Table 1. OS and DFI were poor in older patients (≥75 years) (p<0.05). In terms of sex, OS was much better in women (n=88, 45.4%) than in men (p<0.05), but there was no significant difference in DFI. Current smokers (n=79, 34.5%) had significantly worse OS and DFI than never and former smokers (p<0.05). In terms of tumor markers, patients with high serum CEA levels (>5 ng/ml) had significantly worse OS and DFI compared with those with normal CEA levels. Partial resection (n=25, 13.3%) conferred a worse prognosis than complete resection, and patients with advanced-stage lung adenocarcinoma had a worse prognosis than patients with early-stage disease (p<0.05). Regarding CSCs, Figure 1 shows the immunohistochemical staining of ALDH1A1, CD133, and mutant p53 in sections of resected samples. Table 2 shows the negative and positive rates of CSCs and mutant p53, as well as the significant differences between OS and DFI. Approximately 76.3% (n=148) of patients were ALDH1A1-positive. OS of the ALDH1A1-positive group was significantly worse than that of the ALDH1A1-negative group, but there was no difference in DFI (Figure 2A, B). Approximately 26.3% (n=51) of patients were CD133-positive, and the DFI of the CD133-positive group was significantly worse than that of the CD133-negative group, but there was no difference in OS (Figure 2C, D). Although approximately 51.5% (n=100) of the patients were in the mutant p53-positive group, there was no significant difference in OS or DFI compared with the mutant p53-negative group (Figure 2E, F). The group negative for both ALDH1 and CD133 (double-negative group) had a significantly better prognosis than the other groups regarding DFI, but no significant difference in OS was observed (Figure 3A, B). In patients negative for CSC markers and mutant p53 (triple-negative group), the prognosis was significantly better than that of the other groups in terms of both OS and DFI (Figure 3C, D).

Lung cancer is classified into NSCLC and SCLC according to its pathological features [19]. Among them, NSCLC is divided into three forms histologically: adenocarcinoma, squamous cell carcinoma, and large cell carcinoma [20, 21]. Also, the smoking has high death rate in one of the most important causes by lung cancer [1]. A major clinical problem associated with lung cancer is the acquired resistance of tumors to chemotherapy [22], with NSCLC patients having a 5-year survival rate of less than 20% [23]. In clinical practice, approximately 20% of NSCLCs are operable, but the 5-year survival rate remains low despite advances in chemoradiotherapy, targeted therapy, and immunotherapy for inoperable cases (7%–20%), and the recurrence rate remains high in 30%–50% [24]. However, the 5-year survival rate for localized disease is 59% [25]. From these knowledges, novel therapeutic strategies are required to overcome cancer recurrence, metastasis, and resistance to chemo- and radiotherapy.

The concept of CSCs was introduced in 1977 and was an extremely interesting topic in cancer research [26]. CSCs are an exceedingly rare population among the entire cancer cell population (less than 1% for most solid tumors) that exhibits high tumorigenicity [27]. At first, CSCs were recognized as cancer-initiating cells because they are thought to be the basic cause or species of cancer [4, 28]. However, in reality, the characteristics of CSCs include self-renewal, differentiation capability, high infiltration and migration properties, high tumorigenicity, and resistance to chemotherapy [29]. In short, the main CSC properties are as follows: (a) unique self-renewal ability to produce the daughter cells with the same stem cell characteristics (similar to normal stem cells); (b) the ability to differentiate into a variety of cancer cell lines and promote cell proliferation and overall tumor survival; and (c) high tumorigenic potential to expand and create non-CSC strains and form new tumors. Until now, CSCs of various cancer types including lung cancer were discovered through specific cell-surface proteins by large number of research groups [30].

In this study, we revealed that cases expressing ALDH1A1 had much worse OS than ALDH1A1-negative cases when ALDH1A1 alone was examined, but there was no significant difference in DFI. ALDH (a detoxifying enzyme that oxidizes intracellular aldehydes) is a member of a group of enzymes that protect stem cells from oxidative damage by causing the oxidation of aldehydes to carboxylic acids [31]. So far, ALDH1 expression has been assessed for the identification of CSCs in several cancer types including leukemia and breast, neural, head and neck, colon, liver, and lung cancer [32]. In addition, ALDH1 converts retinol to retinoic acid, which causes stem cell differentiation and proliferation in the early stages [33]. Furthermore, ALDH1A1 expression has been shown to be related with resistance to chemotherapy [34, 35]. Finally, a recent meta-analysis revealed that increased ALDH1A1 expression is associated with poor OS and disease-free survival in lung cancer patients [36].

In the current study, we revealed that CD133-expressing cases had a much worse DFI than CD133-negative cases when CD133 alone was investigated, but there was no significant difference in OS. CD133 is an 865-amino acid penta-span transmembrane protein that has been accepted as a principal marker of stemness in several solid tumors [37]. Some previous studies have shown that human lung cancer contains CD133-positive CSCs that can self-renew and have high tumorigenicity [38]. In addition, the expression of CD133 in NSCLC is associated with the degree of cell differentiation, lymph node metastasis, and prognosis [39]. Furthermore, CD133 expression is negatively correlated with the prognosis of patients with lung cancer because lung tumors containing CD133-positive cells are resistant to cisplatin [40]. Therefore, CD133 expression is a marker for lung CSCs. Our previous study revealed that immunohistological CD133 expression was correlated with the pathological stage of human adenocarcinoma, especially stage I+II disease. Apart from these, various studies suggested that CD133 may have an important role in regulating the expression of CSC genes by interacting with several signaling pathways. However, further investigations are necessary to understand possibility of CD133 in CSC regulation [41].

In this study, there was no clear significant difference in mutant p53 expression between OS and DFI, but mutant p53-expressing cases tended to have a poorer prognosis than negative cases. TP53 encodes the protein consisting of 393 amino acids (p53) in short arm of chromosome 17, which acts as a transcription factor that regulates the expression of other genes. And wild-type p53 has the role of suppressing the accumulation of mutant cells through the induction of apoptosis, giving anticancer drug sensitivity and radiosensitivity [42, 43]. Furthermore, the DNA binding site of p53 is a hotspot for point mutations that result in changes to its three-dimensional structure that inhibit the binding of p53 to its target DNA, and loss of transcriptional activity indicates loss of its tumor suppressor function [44, 45]. To date, various studies have revealed that abnormal p53 expression may promote the initiation and progression of CSCs [46, 47], and activation of mutant p53 was found to increase tumorigenicity by promoting symmetric self-renewal division and inhibiting macrophage accumulation [48]. Although these studies suggested that p53 could be a barrier to CSC formation, the precise mechanism by which p53 regulates CSC survival and tumorigenesis remained unclear. Other than the above, somatic mutations of TP53 occur frequently during the development of human neoplasia, and because mutant p53 proteins are often much more stable than wild-type p53 protein, mutant p53 accumulates to a high level [49].

Various studies have shown the association between ALDH and CD133, including the following: NSCLC cells with relatively high ALDH1 activity are characterized by their increased ability to proliferate, self-renew, differentiate, and express CD133 CSC marker [50, 51]. Furthermore, Jiang et al. found that ALDH expression was associated with the decreased survival of patients with stage I NSCLC and reported a high association between CD133 and ALDH1 expression. This suggests that these proteins are markers for the same tumor cell population [52].

Regarding the association between CSCs and p53, a study by Hilla et al. showed that overexpression of ALDH1A1 in colorectal cancer was associated with reduced apoptosis, which indicated the involvement of ALDH1A1 in the mediation of mutant p53-dependent chemotherapy resistance [53]. An in vitro study, wild-type p53 was also described to suppress CD133 expression transcriptionally in colon cancer. In addition, the tumor-suppressive effect of wild-type p53 in some cancer cell lines needed p53-mediated CD133 inhibition [54]. These results guessed that CD133 may be a potential target for tumor inhibition in highly tumorigenic cancers with impaired p53 function [55].

In this study, double-negative (ALDH1A1−/CD133−) cases had a much better DFI than others, but there was no significant difference in OS. Furthermore, triple-negative (ALDH1A1−/CD133−/p53−) cases had a notably better prognosis than other cases.

In conclusion, lung adenocarcinoma negative for mutant p53 was presumed to have a good prognosis, but when the expression of mutant p53 was investigated in combination with CSC markers, the prognosis of these patients was better with statistical significance. These results suggest that mutant p53 expression may promote the expression of CSC markers and CSC activity in lung adenocarcinoma. Thus, CSC markers and mutant p53 may be effective targets in therapeutic strategies for lung adenocarcinoma. However, these functions have not yet been clarified completely and require further research. In future studies, we will elucidate the detailed mechanism involved and verify other CSC markers.

A total of 286 patients with lung adenocarcinoma who underwent complete resection or other surgery were investigated; 100 cases at Fukuoka-Wajiro Hospital from 2006 to 2012 and 186 cases at Shin-Komonji Hospital from 2010 to 2018. Clinicopathological parameters were assessed in pathological specimens. The mean age at the time of surgery was 70 years (range, 40–92 years). None of the patients received chemotherapy or radiation before surgery. Tumor staging was performed in accordance with the 8th edition of TNM classification by the International Association for the Study of Lung Cancer (IASLC). The clinicopathological parameters were examined age, sex, smoking history, tumor marker level (carcinoembryonic antigen, CEA), surgical procedure, and pathological stage. A paraffin-embedded section was prepared from the resected lung sample, and histological diagnosis was obtained by hematoxylin–eosin (HE) staining. Sectioning of lung adenocarcinoma samples was donated for immunohistochemical staining of ALDH1A1, CD133, and mutant p53 using a standard immunoperoxidase technique, as described previously [12].

ALDH1A1 staining was performed on 4-μm-thick paraffin sections using a mouse monoclonal antibody (anti-ALDH1A1; ab52492; Abcam, Cambridge, MA, USA) at a 1:100 dilution. As previously reported, the results were semi-quantitatively graded on the basis of the percentage of stained cells and the staining intensity [12].

CD133 staining was performed on 4-μm-thick paraffin sections using a mouse monoclonal antibody (anti-CD133; MAB4399-I; Millipore, Temecula, CA, USA) at a 1:200 dilution. As previously reported, the CD133 expression score was defined as the proportion of cells with strong membranous staining in tumor sections [12, 13].

Mutant p53 staining was performed on 4-μm-thick paraffin sections using a mouse monoclonal antibody (anti-p53; M700129; Agilent., Santa Clara, CA, USA) at a 1:200 dilution. The mutant p53 expression score was defined as the proportion of cells with strong membranous staining in tumor sections. The percentage of positive cells was graded as 0%–100%. Mutant p53 positivity was defined as staining in more than 20% of the tumor cells (negative, ≤20%; positive, >20%).

Stained sections were evaluated using virtual slides (Nano Zoomer-XR, Hamamatsu Photonics Co., Ltd.). One physician with experience in pathological research and two physicians with experience in clinical research evaluated the specimens and derived the mean value. All data were analyzed using the statistical software StatView (SAS Institute Inc. Cary, NC, USA). The survival curves were evaluated using the Kaplan–Meier method. P-values ≤0.05 were considered statistically significant.

This study received ethical approval for human subjects from the Shin-Komonji hospital’s research ethics committee. Informed consent was obtained from each patient. Also, all methods were carried out in accordance with relevant guidelines and regulations.

Acknowledgments

The authors would like to thank the Department of Pathology at Shinkomonji Hospital and the Laboratory of Cell and Gene Therapy, Hyogo College of Medicine, for providing specimens and for technical support. We also thank H. Nikki March, PhD, from Edanz Group (https://jp.edanz.com/ac) for editing a draft of this manuscript.

Author contributions statements

NY: conception and design, acquisition of data, analysis and interpretation of data

TS: conception and design, analysis and interpretation of data

TM: Been involved in drafting the manuscript or revising it critically for important intellectual content

TY: Been involved in drafting the manuscript or revising it critically for important intellectual content

RN: acquisition of data

TO: conception and design, analysis and interpretation of data

WM: Been involved in drafting the manuscript or revising it critically for important intellectual content

AG: conception and design, Been involved in drafting the manuscript or revising it critically for important intellectual content

Competing interests

The Author declare no competing interests.

Minna, JD. et al. Focus on lung cancer. Cancer Cell. 1, 49-52 (2002).
Islami, F. et al. Proportion and number of cancer cases and deaths attributable to potentially modifiable factors in the United States in 2014. CA Cancer J Clin. 68, 31-54 (2018).
Reya, T., Morrison, S. J., Clarke, M. F. & Weissman, I. L. Stem cells, cancer, and cancer stem cells. Nature. 414, 105-111 (2001).
Adorno-Cruz, V. et al. Cancer stem cells: Targeting the roots of cancer, seeds of metastasis, and sources of therapy resistance. Cancer Res. 75, 924-929 (2015).
Kornakiewicz, A., Czarnecka, A. M., Khan, M. I., Krasowski, P., Kotrys, A. V. & Szczylik, C. Effect of Everolimus on Heterogenous Renal Cancer Cells Populations Including Renal Cancer Stem Cells. Stem Cell Rev Rep. 14, 385–397 (2018).
Vassalli G. Aldehyde Dehydrogenases: Not Just Markers, but Functional Regulators of Stem Cells. Stem Cells Int. 2019, 3904645 (2019).
Raha, D. et al. The cancer stem cell marker aldehyde dehydrogenase is required to maintain a drug-tolerant tumor cell subpopulation. Cancer Res. 74, 3579-3590 (2014).
Zhao, Q. et al. Prognostic value of the expression of cancer stem cell-related markers CD133 and CD44 in hepatocellular carcinoma: From patients to patient-derived tumor xenograft models. Oncotarget. 7, 47431-47443 (2016).
Wen, L. et al. Prognostic value of cancer stem cell marker CD133 expression in gastric cancer: A systematic review. PLoS One. 8, e59154 (2013).
Kim, K., Ro, J. Y., Kim, S. & Cho, Y. M. Expression of stem-cell markers OCT-4 and CD133: Important prognostic factors in papillary renal cell carcinoma. Human Pathol. 43, 2109-2116 (2012).
Le, H., Zeng, F., Xu, L., Liu, X. & Huang, Y. The role of CD133 expression in the carcinogenesis and prognosis of patients with lung cancer. Mol Med Rep. 8, 1511-1518 (2013).
Miyata, T. et al. The Clinical Significance of Cancer Stem Cell Markers ALDH1A1 and CD133 in Lung Adenocarcinoma. Anticancer Res. 37, 2541-2547 (2017).
Yamashita, N. et al. Association Between CD133 Expression and Prognosis in Human Lung Adenocarcinoma. Anticancer Res. 41, 905-910 (2021).
Olovnikov, I. A., Kravchenko, J. E. & Chumakov, P. M. Homeostatic functions of the p53 tumor suppressor: regulation of energy metabolism and antioxidant defense. Semin Cancer Biol. 19, 32-41 (2009).
Ryan, K. M., Phillips, A. C. & Vousden, K. H. Regulation and function of the p53 tumor suppressor protein. Curr Opin Cell Biol. 13, 332–337 (2001).
Rodier, F., Campisi, J. & Bhaumik, D. Two faces of p53: aging and tumor suppression. Nucleic Acids Res. 35, 7475-7484 (2007).
Greenblatt, M. S., Bennett, W. P., Hollstein, M. & Harris, C. C. Mutations in the p53 tumor suppressor gene: clues to cancer etiology and molecular pathogenesis. Cancer Res. 54, 4855-4878 (1994).
Marchetti, A. et al. p53 alterations in non-small cell lung cancers correlate with metastatic involvement of hilar and mediastinal lymph nodes. Cancer Res. 53, 2846-2851 (1993).
Travis, W. D. et al. Pathology and genetics of tumours of the lung, pleura, thymus and heart (ed. Travis, W. D.) 26-67 (Lyon, 2003).
Sanders, H.R. & Albitar, M. Somatic mutations of signaling genes in non-small-cell lung cancer. Cancer Genet Cytogenet. 203, 7-15 (2010).
Liu, Q., Bai, W., Huang, F., Tang, J. & Lin, X. Downregulation of microRNA-196a inhibits stem cell self-renewal ability and stemness in non-small-cell lung cancer through upregulating GPX3 expression. Int J Biochem Cell Biol. 115, 105571 (2019).
Zakaria, N. et al. Human non-small cell lung cancer expresses putative cancer stem cell markers and exhibits the transcriptomic profile of multipotent cells. BMC Cancer. 15, 84 (2015).
Bade, B. C. & Dela Cruz, C. S. Lung Cancer 2020: Epidemiology, Etiology, and Prevention. Clin Chest Med. 41, 1–24 (2020).
Miyata, T. et al. Cancer stem cell markers in lung cancer. Personal Med Univ. 4, 40-45 (2015).
Siegel, R. L., Kimberly, K. D., Fuchs, H. E. & Jemal, A. Cancer Statistics, 2021. CA Cancer J Clin. 71, 7-33 (2021).
Bonnet, D. & Dick, J. E. Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell. Nat Med. 3, 730-737 (1997).
Pardal, R., Clarke, M. F. & Morrison, S. J. Applying the principles of stem-cell biology to cancer. Nat Rev Cancer. 3, 895-902 (2003).
Koren, E. & Fuchs, Y. The bad seed: Cancer stem cells in tumor development and resistance. Drug Resist Updat. 28, 1-12 (2016).
Nassar, D. & Blanpain, C. Cancer stem cells: Basic concepts and therapeutic implications. Annu Rev Pathol. 11, 47-76 (2016).
Tirino, V. et al. Cancer stem cells in solid tumors: an overview and new approaches for their isolation and characterization. FASEB J. 27, 13-24 (2013).
Sophos, N. A. & Vasilis, V. Aldehyde dehydrogenase gene superfamily: the 2002 update. Chem Biol Interact. 143-144, 5-22 (2003).
Ma, I. & Allan, A. L. The role of human aldehyde dehydrogenase in normal and cancer stem cells. Stem Cell Rev Rep. 7, 292-306 (2011).
Duester, G., Mic, F. A. & Molotkov, A. Cytosolic retinoid dehydrogenases govern ubiquitous metabolism of retinol to retinaldehyde followed by tissue-specific metabolism to retinoic acid. Chem Biol Interact. 143-144, 201-210 (2003).
Lee, H. E. et al. An increase in cancer stem cell population after primary systemic therapy is a poor prognostic factor in breast cancer. Br J Cancer. 104, 1730-1738 (2011).
Tanei, T. et al. Association of breast cancer stem cells identified by aldehyde dehydrogenase 1 expression with resistance to sequential Paclitaxel and epirubicin-based chemotherapy for breast cancers. Clin Cancer Res. 15, 4234-4241 (2009).
Wei, D., Peng, J. J., Gao, H., Zhang, T., Tan, Y. & Hu, Y. H. ALDH1 Expression and the Prognosis of Lung Cancer: A Systematic Review and Meta-Analysis. Heart Lung Circ. 24, 780-788 (2015).
Li, Z. CD133: a stem cell biomarker and beyond. Exp Hematol Oncol. 2, 17 (2013).
Eramo, A. et al. Identification and expansion of the tumorigenic lung cancer stem cell population. Cell Death Differ. 15, 504-514 (2008).
Qu, H., Li, R., Liu, Z., Zhang, J. & Luo, R. Prognostic value of cancer stem cell marker CD133 expression in non-small cell lung cancer: A systematic review. Int J Clin Exp Pathol. 6, 2644-2650 (2013).
Bertolini, G. et al. Highly tumorigenic lung cancer CD133+ cells display stem-like features and are spared by cisplatin treatment. Proc Natl Acad Sci USA. 106, 16281-16286 (2009).
Wei, Y. et al. Activation of PI3K/Akt pathway by CD133-p85 interaction promotes tumorigenic capacity of glioma stem cells. Proc Natl Acad Sci USA. 110, 6829-6834 (2013).
Gebitekin, C., Bayram, A. S., Tunca, B. & Balaban, S. A. Clinical significance of p53 gene mutation in T1-2N0 non-small cell lung cancer. Asian Cardiovasc Thorac Ann. 15, 35-38 (2007).
Oyama, T. et al. Molecular genetic tumor markers in non-small cell lung cancer. Anticancer Res. 25, 1193-1196 (2005).
Olszewski, M. B., Pruszko, M., Snaar-Jagalska, E., Zylicz, A. & Zylicz, M. Diverse and cancer type‑specific roles of the p53 R248Q gain‑of‑function mutation in cancer migration and invasiveness. Int J Oncol. 54, 1168-1182 (2019).
Oyama, T. et al. p53 mutations of lung cancer are not significantly affected by CYP1A1 or GSTM1 polymorphisms. Int J Oncol. 11, 305-309 (1997).
Godar, S. et al. Growth-inhibitory and tumor- suppressive functions of p53 depend on its repression of CD44 expression. Cell. 134, 62-73 (2008).
Zhang, M. et al. Identification of tumor-initiating cells in a p53-null mouse model of breast cancer. Cancer Res. 68, 4674-4682 (2008).
Xu, Y. et al. Mutated p53 Promotes the Symmetric Self-Renewal of Cisplatin-Resistant Lung Cancer Stem-Like Cells and Inhibits the Recruitment of Macrophages. J Immunol Res. 2019, 7478538 (2019).
Ramael, M. et al. Immunoreactivity for p53 protein in malignant mesothelioma and non-neoplastic mesothelium. J Pathol. 168, 371-375 (1992).
Liu, J. et al. Lung cancer tumorigenicity and drug resistance are maintained through ALDH(hi)CD44(hi) tumor initiating cells. Oncotarget. 4, 1698-1711 (2013).
Kahlert, C. et al. Expression analysis of aldehyde dehydrogenase 1A1 (ALDH1A1) in colon and rectal cancer in association with prognosis and response to chemotherapy. Ann Surg Oncol. 19, 4193-4201 (2012).
Jiang, F. et al. Aldehyde dehydrogenase 1 is a tumor stem cell-associated marker in lung cancer. Mol Cancer Res. 7, 330-338 (2009).
Solomon, H. et al. Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers. Oncogene. 37, 1669-1684 (2018).
Park, E. K. et al. Transcriptional repression of cancer stem cell marker CD133 by tumor suppressor p53. Cell Death Dis. 6, e1964 (2015).
Chen, X. et al. p53 positively regulates the expression of cancer stem cell marker CD133 in HCT116 colon cancer cells. Oncol Lett. 16, 431-438 (2018).

Table 1. Clinicopathological parameters and prognosis

Parameters		Number (%)	p-value (OS)	p-value (DFI)
Total		194
Age	<75 years	124 (64.0)	<0.0001	0.0034
Age	≥75 years	70 (36.0)	<0.0001	0.0034
Sex	Female	88 (45.4)	0.0006	0.2810
Sex	Male	106 (54.6)	0.0006	0.2810
Smoking history	Never and former	115 (59.3)	<0.0001	0.0084
Smoking history	Current	79 (34.5)	<0.0001	0.0084
Serum CEA	Normal	127 (65.5)	<0.0001	<0.0001
Serum CEA	High	67 (34.5)	<0.0001	<0.0001
Surgical procedure	Complete resection	169 (86.7)	<0.0001	0.5470
Surgical procedure	Partial resection	25 (13.3)	<0.0001	0.5470
Pathological stage	I	133 (68.6)	<0.0001	<0.0001
	II	22 (11.3)
	III	26 (13.4)
	IV	13 (6.7)

Table 2. Results and prognosis of immunohistochemical staining of CSCs and mutant p53

		Number (%)	p-value (OS)	p-value (DFI)
ALDH1A1	Negative	46 (23.7)	0.0137	0.0714
	Positive	148 (76.3)
CD133	Negative	143 (73.7)	0.4428	0.0285
	Positive	51 (26.3)
Mutant p53	Negative	94 (48.5)	0.1094	0.1262
	Positive	100 (51.5)

No competing interests reported.

Relationship and Clinical Significance of ALDH1A1 and CD133 Cancer Stem Cell Markers and Mutant p53 in Lung Adenocarcinoma

Status:

Version 1

Abstract

Background

Methods

Results

Conclusions

Figures

Introduction

Results

Discussion

Methods and Materials

Declarations

References

Tables

Additional Declarations

Status:

Version 1