Temozolomide-fatty acid conjugates for glioblastoma multiforme: In vitro and in vivo evaluation

Glioblastoma multiforme (GBM) is the deadliest brain tumor with a poor prognosis and limited therapeutic options. Temozolomide (TMZ) is the first-line chemotherapeutic agent used for the treatment of GBM; however, it suffers from several limitations, including short half-life, rapid metabolism, <1% brain bioavailability, methyl guanine methyl transferase (MGMT) based chemoresistance, and hematological toxicities. Several approaches have been adopted to overcome these limitations, particularly by using nanotechnology-based systems, but its physicochemical properties make TMZ challenging to load into these nanocarriers. In the current research, we conjugated TMZ with different fatty acids, i.e., linoleic acid (LA), oleic acid (OA), and palmitic acid (PA), to obtain TMZ-fatty acid conjugates, which are comparatively hydrophobic, less prone to degradation and potent. These conjugates were thoroughly characterized using ¹H NMR spectroscopy, high-resolution mass spectrometry (HR-MS), and reverse phase-high performance liquid chromatography (RP-HPLC). The synthesized conjugates, namely Temozolomide-oleic acid (TOA,6R₁), Temozolomide-linoleic acid (TLA, 6R₂), and Temozolomide-palmitic acid (TPA, 6R₃), showed an IC₅₀ of 101.4, 67.97, and 672.04 μM, respectively in C₆ cells and 428.257, 366.43 and 413.69 μM, respectively in U87-MG cells. On the other hand, the free TMZ showed an IC₅₀ of >1000 μM and 564.23 μM in C₆ and U87-MG, respectively. Further, the in vivo efficacy of the TMZ-fatty acid conjugates was evaluated in the C₆-induced orthotropic rat glioblastoma model, wherein the TMZ-fatty acid conjugate showed improved survival rate (1.6 folds) and overall health of the animals. Collectively, the conjugation of fatty acids with TMZ improves its anticancer potential against glioblastoma multiforme (GBM).