PYRAZOLOPYRIMIDINES AS ANTICANCER AGENTS; SYNTHESES AND MODE OF ACTION (REVIEW ARTICLE)

Pyrazolopyrimidines are composed of a pyrazole ring fused with pyrimidine moiety, unlike the imidazole moiety in purines; they are initially reported as adenosine receptor antagonists. Different methods of synthesis of substituted pyrazolo[3,4-d ]pyrimidines and pyrazolo[1,5-a ]pyrimidines have been reported with a survey of their biological activities especially anti-cancer activity. During the last two decades, pyrazolopyrimidines have gained great attention as biologically active compounds; they are reported to have anti-cancer activity, anti-microbial, anti-inflammatory, anti-hyperuricemia, anti-viral and anti-hypertensive activities. Researchers paid great attention to pyrazolopyrimidines due to their high anti-cancer activity, so they tried to prepare new derivatives and examine their anti-cancer activity. In this review we are going to focus on the different methods used for the synthesis of pyrazolopyrimidines, their anticancer activities as well as their reported mode of action


Introduction
Cancer is a disease in which a group of cells displays uncontrolled growth, sometimes invasion which destroys adjacent tissues and sometimes metastasizes to lymph nodes or other body sites via the lymphatic system or through the bloodstream.These properties differentiate malignant tumors from benign cancer (Mandour et al., 2022;Upadhyay, 2021).

Tyrosine kinase inhibition
The tyrosine kinase enzymes (TK) regulate several physiological mechanisms, including cell proliferation, differentiation, migration and metabolism by transferring the ATP terminal phosphate to the tyrosine residues of protein substrates (Carmi et al., 2012;Ebrahimi et al., 2023;Roskoski, 2023).

Src tyrosine kinase inhibition
The Src family of tyrosine kinases (SFKs) is constituted of nine members: Src, Lck, Fyn, Yes, Hck, Blk, Fgr, Lyn and Yrk.SFKs play an important role in signalling pathways that control a diverse spectrum of biological activities, such as cell division, growth factor signalling, differentiation, survival, adhesion, migration, invasion and so on

Inhibition of urokinase plasminogen activator (uPA)
Urokinase plasminogen activator (uPA) is a serine protease enzyme that functions in the conversion of the circulating plasminogen to the active, broad-spectrum serine protease, plasmin It has been reported that the 6-methyl-1-p-tolyl-1H-pyrazolo [3,4-d]pyrimidine-4(5H)one 65 was synthesized and evaluated in-vitro for its anticancer activity against MCF-7 breast cancer and Hep-G2 liver cancer cell lines (Fig. 11).It showed 77% inhibition for MCF-7 breast cancer cell line and 63% inhibition for Hep-G2 liver cancer cell line relative to 91% ,88% produced by Doxorubicin, respectively, this antiproliferative activity was considered due to uPA inhibition (Shamroukh et al., 2014).

Cyclin-dependent kinase (CDK) inhibitor
During the last two decades, there are many promising drugs available for the treatment of cancer diseases.For example, Dinaciclib 124 was synthesized by Paruch et al. (Fig. 30) as a potent and selective cyclin-dependent kinase (CDK) inhibitor that currently undergoing clinical evaluation (Paruch et al., 2007).

Lymphocyte-specific kinase (Lck) inhibitors
Gommermann et al. have synthesized different pyrazolopyrimidine derivatives 129 from a series of pyrazolo[1,5-a]pyrimidines and were optimized to target lymphocyte-specific kinase (Lck) (Fig. 33).These derivatives exhibited significant potency against LCK cell line with IC 50 =7 nM.Compound 129a showed very potent inhibition of Lck and displayed very good selectivity against cSrc, Hck and KDR, while its acylation with glycine led to the highly active compound 129b that demonstrated a low clearance and long half-life (Gommermann et al., 2010).

Allosteric agonists for the high affinity nicotinic acid receptor GPR109A
Shen et al. have synthesized pyrazolopyrimidine derivatives and they were discovered as the first class of allosteric agonists for the high affinity nicotinic acid receptor GPR109A.In addition to its intrinsic activity, compound 132 significantly enhances nicotinic acid binding to the receptor, thereby potentiating the functional efficacy of nicotinic acid (Fig. 36) (Shen et al., 2008).In 2017, Wang et al. discovered pyrazolo[3,pyrimidine 140 as epidermal growth factor receptor (EGFR) inhibitor which displayed strong anti-proliferative effect against EGFR mutant-driven non-small cell lung cancer (NSCLC) cell lines such as H1975, PC9, HCC827, and H3255.Its molecular docking study revealed that the 4amino-pyrazolo [3,4-d]pyrimidine moiety formed two hydrogen bonds with Met793 and Gln791 in the hinge area, respectively.Additionally, the N2 and the N7 of the pyrazolo [3,4-d]pyrimidine moiety were involved in another two hydrogen bonds with Lys 745 and Gly 796, respectively through water bridges.Moreover, the pyrazole part of the pyrazolo [3,4-d]pyrimidine moiety was involved in a hydrophobic interaction with

Conclusion
This review article seeks to provide an up-to-date overview of the latest advances in the development of the synthetic approaches to construct pyrazolo[1,5a]pyrimidine and pyrazolo [3,4-d]pyrimidine skeletons which have always been of a paramount chemical significance in pharmaceutical and synthetic chemistry field.This review presented a broad range of simple and efficient synthetic methods to access a wide range of functionalized pyrazolo[1,5-a]pyrimidine and pyrazolo [3,4-d]pyrimidine scaffolds through several strategies.Also, this article clearly showed that these ring systems play an important role in medicinal chemistry being evaluated against numerous biological targets.Many studies revealed that different pyrazolopyrimidine derivatives possess extensive potential applications as antitumor agents and kinase inhibitors and have been successfully developed, marketed, and extensively used in the clinic in treating various types of cancer diseases with low toxicity, high bioavailability, and good curative effects.Much effort contributed to structural modification of clinical pyrazolopyrimidine drugs are needed to retain the advantages of these drugs and overcome their shortcomings.One significant strategy is to employ some functionality that are helpful for improving the physicochemical properties and affinity with the target sites to modify clinical drugs.This intention is to increase their biological activities and overcome drug resistances.

(Chu et al., 2024; Kumar et al., 2022). uPA
is secreted as an inactive single-chain proenzyme by many different cell types and exists in a soluble or cell associated forms by binding to a specific membrane uPA receptor (uPAR) (

Lin et al., 2013).
Elevated expression levels of urokinase is implicated in a large number of malignancies e.g.Lung cancer (

Chu et al., 2014), prostate (Vyas and Singh, 2014) and thyroid carcinoma (Horvatic Herceg et al., 2013).
Different mechanisms account for the cytotoxic effect of this class of compounds; they have been reported to act as vascular endothelial growth factor receptor inhibitors (