Synthesis and cytotoxic activity of 3-O-acyl/3-hydrazine/2-bromo/ 20,29-dibromo betulinic acid derivatives Rama Mukherjee, a,b,c Manu Jaggi, a, * Mohammad J. A. Siddiqui, b Sanjay K. Srivastava, c, * Praveen Rajendran, a Anand Vardhan b and Anand C. Burman a,b,c a Division of Experimental Oncology, Dabur Research Foundation, 22, Site IV, Sahibabad, Ghaziabad 201 010, UP, India b Division of Chemical Research, Dabur Research Foundation, 22, Site IV, Sahibabad, Ghaziabad 201 010, UP, India c Division of Medicinal Chemistry, Dabur Research Foundation, 22, Site IV, Sahibabad, Ghaziabad 201 010, UP, India Received 7 April 2004; accepted 10 May 2004 Available online Abstract—A series of 3-O-acyl, 3-hydrazine, 2-bromo, and 20,29-dibromo betulinic acid derivatives (1–27) have been synthesized and screened for in vitro cytotoxic activity on human cancer cell lines MOLT-4, JurkatE6.1, CEM.CM3, BRISTOL8, U937, DU145, PA-1, A549, and L132. A number of compounds have shown ED 50 <1 lg/mL against the cancer cell lines tested and have shown better cytotoxicity than betulinic acid. Compounds 13, 19, 20, 23,and 27 were the best derivatives and were selected as lead molecules for further development. The structure–activity relationship has been described. Ó 2004 Elsevier Ltd. All rights reserved. 1. Introduction Betulinic acid (3-b-hydroxy-lup-20(29)-en-28-oic acid) is apentacycliclupane-typetriterpeneandhasbeenshown to possess several medicinal properties including anti- cancer 1;2 and anti-HIV 3 activities. Previous reports indicated that betulinic acid was a melanoma-specific cytotoxic compound, 1;2 however, more recent evidence indicates that betulinic acid possesses a broader spec- trum of activity against other cancer cell types. 4–7 Bet- ulinic acid was shown to act through induction of apoptosis 1;2 independent of the cell’s p53 status. 7–9 A previous investigation had shown that betulinic acid inhibited the in vitro activity of aminopeptidase N, 10 an endogenous angiogenic factor and inhibited the mito- chondrial function in endothelial cells. 11 Recently, the anti-angiogenic activity of some betulinic acid deriva- tives has been reported by us. 12;13 These findings have made betulinic acid a very attractive candidate for the clinicaltreatmentofvariousformsofcancer.Asaresult, further studies have been performed to derive synthetic betulinic acid analogs in an effort to establish mean- ingful structure–activity relationships. Herewereportthesynthesisof3-O-acyl,3-hydrazine,2- bromo, and 20,29-dibromo betulinic acid derivatives (1– 27) and their in vitro cytotoxicity against a number of human cancer cell lines. For the first time, the anti-leu- kemia and anti-lymphoma activity of betulinic acid and its derivatives (1–27) is being reported here. 2. Chemistry Synthesis of betulinic acid derivatives 1–20 2;12 has been described in Scheme 1. Betulinic acid was elaborated in fourways.Infirst,betulinicacidwastreated,separately, with acetyl chloride, (S)-())-2-acetoxypropionyl chlo- ride, trimethylacetyl chloride, and propionyl chloride in methylenechloride(DCM)toaffordcorresponding3-O- acyl derivatives 1, 2, 5, and 8, respectively. Compounds 1 and 2 were reacted, separately, with methyl bro- moacetate and sodium hydride in DMF to give 17- carboxymethyl carboxylate derivatives 3 and 7, respec- tively. While in the second, betulinic acid was reacted with methyl bromoacetate, as for 3 and 7, to yield 17- carboxymethyl carboxylate derivative 4 and which, upon hydrolysis with 10% sodium hydroxide afforded the respective acid derivative 6. In the third, betulinic *Corresponding authors. Tel.: +91-120-2777901 to 25; fax: +91-120- 2777303; e-mail addresses: jaggim@dabur.com; srivastavasa@ dabur.com 0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2004.05.034 Bioorganic & Medicinal Chemistry Letters 14 (2004) 4087–4091