AZT (3'-azido-2’, 3’-dideoxythymidine) has been shown to prolong the survival of patients infected with human immunodeficiency virus (HIV) and decrease the severity of opportunistic infections. Such studies have prompted the use of AZT to treat symptomless individuals infected with HIV in the hope of delaying or even preventing the progression to acquired immunodeficiency syndrome (AIDS). However, before chronic use of AZT in symptomless individuals is initiated, it is important to establish whether this anti-viral drug would directly alter the phenotype and functions of the cells involved in the immune system. In the current study, we observed that AZT when administered orally for 7 -14 days into DBA/2 mice at 500 - 1000 mg/kg body weight induced a dose-dependent decrease in cellularity of the thymus. AZT caused significant alterations in the thymus resulting from a significant decrease in the number of double-positive (CD4⁺CD8​​⁺) cells and an increase in the number of double-negative (CD4⁻CD8⁻) cells. Interestingly, after the i.p. administration of interleukin-2 (IL-2) simultaneously with AZT, the total cellularity of the thymus was completely reconstituted. We also observed that AZT effectively suppressed the in vivo T cell response to conaibumin and gp120 of HIV. Furthermore, the addition of AZT to in vitro cultures caused a dose-dependent decrease in T and B cell proliferative responses to mitogens at 50μM or greater concentrations. Also, AZT inhibited the generation of cytotoxic T lymphocytes when added to the culture and this inhibition was reconstituted by the addition of exogenous IL-2. Together, our studies demonstrate that AZT modulates the phenotype and function of cells of the immune system which, in turn, could have marked repercussions on immune responses of the host toward infections and cancers. Also, our data demonstrating that AZT can suppress T cell responsiveness against HIV antigens caution against chronic use of AZT in asymptomatic HIV-infected individuals.