**3. ALDH as a stem cell marker**

In theory, ALDH isozymes including ALDH1A, ALDH1A2, ALDH1A3, and ALDH3A1, which are involved in drug resistance and RA formation, are vital in protecting SCs against toxic endogenous and exogenous aldehydes and for SCs' ability to differentiate, respectively. It is unknown what ALDH isozymes are responsible for the ALDH activity that are used to identify stem cell progenitors. In the overlap gene profile of different stem cell populations, ALDH7A1, known as antiquitin, and ALDH2 were identified, consequently, and are worthy of further investigation. There is more about ALDH to be explored as a cause of its full physiological function has remained elusive. ALDH7A1 is a green pea 26g protein, which has function in regulation of turgor pressure, and has ≥50% amino acid identity with the 3 pseudogenes in the ALDH family. It also has 69% equity with ALDH2, but nevertheless has considerably lower affinity for acetaldehyde than ALDH2. However, ALDH2, which is a mitochondrial enzyme, has been widely studied mostly for its affilitation with ethanol metabolism. Yet, there might be an extent of confusion as to how ALDH2 is associated in gene profiling studies. According to the nomenclature, this enzyme indeed is ALDH1A, related to a series of events linked to the development of dopaminergic neurons through its ability to produce RA. It was reported that ALDH2 or AHD2 expression changes during differentiation of NIH-3T3 cells into adipocytes. These studies continue to focus on ALDH1A1's role in SCs and stem cell differentiation. For hematopoietic stem cell progenitors, ALDH1A1 has been a thoroughly established marker for many years. Reseach on the role of RA in granulocyte differentiation of hematopoietic SCs discovered that ALDH1A1 and ALDH1B1 catalyze cellular RA synthesis and are expressed in CD34+ hematopoietic progenitors (Russo*, et al.*, 2002, Luo*, et al.*, 2007).They also showed that ALDH1A2 or 1A3 do not show those characteristics. For the differentiation to mature granulocytes, these 2 enzymes' expressions are necessary, however their expressions are lost once the differentiation is complete. The *in vitro* disulfiram treatment in which disulfiram acts as an ALDH inhibitor may inhibit granulocytic differentiation. ALDH1A1 is found in erythrocytes and has been pointed to contribute to the aldophosphamide detoxification. A study that inhibited ALDH and retinoid signaling with diethylamino-benzaldehyde (DEAB) that reported the expansion of human HSCs probably by blocking differentiation and assisting self-renewal and HSC expansion (Marchitti*, et al.*, 2008).

8 Dehydrogenases

enzymes participate in cellular detoxification, differentiation and drug resistance through

The functional activity of ALDH has been widely used to identify and isolate CSCs found in the bone marrow (Ran*, et al.*, 2009), breast (Ginestier*, et al.*, 2007), lung (Ucar*, et al.*, 2009), ovary (Deng*, et al.*, 2010), colon (Huang*, et al.*, 2009), prostate (van den Hoogen*, et al.*, 2010),

The derivation of SCs from adult tissues, their relative ease of isolation and enormous expansion potential in culture make them attractive therapeutic candidates (Prockop*, et al.*, 2010). These cells are identified by their expression of a particular panel of surface molecules, with the presence of CD73, CD90, CD105, and the absence of CD14, CD34, CD45, and HLA-DR. They show no proliferative response from alloreactive lymphocytes because of the negligible levels of extracellular MHC class I and II determinants. SCs also have important immunomodulatory functions in all the cells involved in both the innate and

In theory, ALDH isozymes including ALDH1A, ALDH1A2, ALDH1A3, and ALDH3A1, which are involved in drug resistance and RA formation, are vital in protecting SCs against toxic endogenous and exogenous aldehydes and for SCs' ability to differentiate, respectively. It is unknown what ALDH isozymes are responsible for the ALDH activity that are used to identify stem cell progenitors. In the overlap gene profile of different stem cell populations, ALDH7A1, known as antiquitin, and ALDH2 were identified, consequently, and are worthy of further investigation. There is more about ALDH to be explored as a cause of its full physiological function has remained elusive. ALDH7A1 is a green pea 26g protein, which has function in regulation of turgor pressure, and has ≥50% amino acid identity with the 3 pseudogenes in the ALDH family. It also has 69% equity with ALDH2, but nevertheless has considerably lower affinity for acetaldehyde than ALDH2. However, ALDH2, which is a mitochondrial enzyme, has been widely studied mostly for its affilitation with ethanol metabolism. Yet, there might be an extent of confusion as to how ALDH2 is associated in gene profiling studies. According to the nomenclature, this enzyme indeed is ALDH1A, related to a series of events linked to the development of dopaminergic neurons through its ability to produce RA. It was reported that ALDH2 or AHD2 expression changes during differentiation of NIH-3T3 cells into adipocytes. These studies continue to focus on ALDH1A1's role in SCs and stem cell differentiation. For hematopoietic stem cell progenitors, ALDH1A1 has been a thoroughly established marker for many years. Reseach on the role of RA in granulocyte differentiation of hematopoietic SCs discovered that ALDH1A1 and ALDH1B1 catalyze cellular RA synthesis and are expressed in CD34+ hematopoietic progenitors (Russo*, et al.*, 2002, Luo*, et al.*, 2007).They also showed that ALDH1A2 or 1A3 do not show those characteristics. For the differentiation to mature

the oxidation of cellular aldehydes (Moreb*, et al.*, 1996).

and pancreas (Dembinski & Krauss, 2009).

adaptive immune responses (Nauta & Fibbe, 2007).

**3. ALDH as a stem cell marker** 

**2.2. Stem cell markers** 

The cancer stem cell theory is supported by current evidence in tumor biology, which may also provide a biological reason for the age-related survival difference. The theory demonstrates that CSCs, a small subset of tumor cells with stem cell-like properties such as epithelial-to-mesenchymal progression, are capable of differentiation and self-renewal, after which leads to formation of a heterogeneous tumor cell population. Including aldehyde dehydrogenase-1 (ALDH1) activity, CD44+/CD24-, CD133, and ITGA6, a wide range of putative breast cancer stem cell markers have been proposed. ALDH1 expression has especially demonstrated an assurance of a clinically relevant prognostic marker. In addition, the subset of CSCs is shown to be relatively insusceptible to chemo and radiotherapy by various studies. For this reason, the subpopulation of CSCs can present a statement and a therapeutic target for poor-prognostic, treatment-resistant and recurrent breast cancer. Through its role in oxidizing retinol to RA, which is a modulator of cell proliferation, ALDH1 might have a role in early differentiation of SCs and stem cell proliferation (Mieog*, et al.*, 2012).

It is possible to isolate leukemia SCs depending on the elevated ALDH activity by using the aldefluor assay. In patient samples, the researchers encountered a population of ALDH+ acute myeloid leukemia (AML) cells (Rollins-Raval*, et al.*, 2012). In most cases, the ALDH+ AML cells coexpressed CD34+ (formerly determined leukemia stem cell marker), and were introduced considerably better than the ALDH- AML cells in immunocompromised mice. In the same year, ALDH+ cells from breast cancers, which had the tumorigenic and selfrenewal features of CSCs, were shown to be possibly isolated. This innovative study displayed the potential applicability of quantifying ALDH activity in solid tumors. ALDH activity would be used successfully as a CSC marker for abundant cancers including liver, colon, lung, bone, prostate, pancreatic, head and neck, thyroid, bladder, brain, cervical and melanoma in the proceeding years. With one exception of a current study for melanoma, 35 demonstrate growing evidence recommending ALDH's activity to be a universal CSC marker. Nonetheless, as amounted by the aldefluor assay in various tissues and cancers, the cause of ALDH activity may differ. Essentially, determination of specific ALDH isoforms carried out commonly in certain cancers might have prognostic suitability. Besides their valuable function in detoxification of aldehydes, ALDHs carry out other functions such as serving as binding proteins for various molecules (e.g., androgens and cholestorol), potentially act as antioxidants by NAD(P)H production, ultraviolet light absorption and/or hydroxyl radical scavenging and ester hydrolysis.

Lastly, several isoforms (ALDH1A1, ALDH1A2, ALDH1A3 and ALDH8A1), take place via RA formation by oxidation of all-trans-retinal and 9-cis-retinal in RA cell signaling, which

has been related to the "stemness" characteristics of CSCs. Consequently, its supported by widening evidence that ALDH may be more than just a CSC marker and have an accomplishable role in CSC biology (Marcato*, et al.*, 2011).

Aldehyde Dehydrogenase: Cancer and Stem Cells 11

disassociated populations of dopaminergic neurons. Decreased levels of ALDH1A1 takes place in dopaminergic neurons of the substantia nigra of patients with Parkinson's disease (PD), as well as the ventral tegmental area in schizophrenic patients. In the central nervous system (CNS), monoamine oxidase (MAO) metabolizes dopamine to aldehyde, as it's metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL), which growing evidence suggests might be neurotoxic, and it may lead to cell death in relation to neurological pathologies when accumulated. In maintaining low intraneuronal levels of DOPAL, ALDH1A1 may undertake a critical role by catalyzing its metabolism to 3,4-dihydroxyphenylacetic acid (DOPAC). Being one of 139 genes that are differentially expressed in primary human HSCs, and through the production of RA, ALDH1A1 has been shown to promote their

These data suggest that for the therapeutic amplification of HSCs, ALDH1A1 inhibition

ALDH1A2 is a cytosolic homotetramer expressed in several embryonic and adult tissues such as brain, kidney, intestine, testis, liver, retina, lung. As ALDH1A1, ALDH1A2 also catalyzes the reaction in which both all-*trans*-retinal and 9-*cis*-retinal oxidize to RA. However, when compared with other ALDH isozymes, ALDH1A2 appears to acquire the highest specificity (*V*max/*K*m = 49 nmol·min−1·mg−1·μM−1) for all-*trans*-retinal. This characteristic may be because of an uncommon discreate loop in its active site that binds all-

Taking action in several developmental processes, ALDH1A2 might be a key regulator of RA synthesis in developing tissues. Due to defects in early heart morphogenesis, *Aldh1*α*2*−/<sup>−</sup> mice die in early embryonic stages in which they seem to lack axial rotation, incomplete neural tube closure, reduction of the trunk region and many of the properties of human DiGeorge/velocardiofacial syndrome, a disorder characterized by cleft palate, heart abnormalities and learning disabilities. During early vascular development, aberrations in endothelial cell cycle progression have also been determined in *Aldh1*α*2*−/<sup>−</sup> embryos. *Aldh1a2* has been determined as a key regulator in the development of many tissues including kidney, retina, lung, forebrain, pancreas, and spinal cord by

ALDH1A3 is a cytosolic homodimer that participates in RA synthesis, oxidizes both all*trans*-retinal and 9-*cis*-retinal (*K*m 0.2 μM for all-*trans*-retinal) to RA, and has an important role in embryonic development; including brain, retina, skeletal muscle, tooth buds, intestine, kidney, prostate, lung, liver and pancreas, it is expressed in various late-stage embryonic and adult rodent tissues. In humans, ALDH1A3 expression has been noted in stomach, salivary gland, breast, kidney and fetal nasal mucosa. *Aldh1a3*−/− mouse embryos

could potentially be used (Marchitti*, et al.*, 2008, Moore*, et al.*, 2009).

miscellanous animal models (Marchitti*, et al.*, 2008, El Kares*, et al.*, 2010).

die as a result of defects in nasal development.

differentiation.

*3.1.2. ALDH1A2* 

*3.1.3. ALDH1A3* 

*trans*-retinal in a unique manner.
