Background: Glioblastoma multiforme (GBM) is the most common brain cancer. Front-line systemic therapy with temozolomide is often ineffective due to O⁶-methylguanine-DNA-methyltransferase (MGMT)-mediated resistance. Dianhydrogalactitol (VAL-083) is a bi-functional DNA alkylating agent that crosses the blood-brain barrier and overcomes resistance to MGMT in vitro. The goal of this clinical trial is to determine an appropriate dose for phase III trials in refractory GBM. Methods: Open-label, single-arm phase I/II dose-escalation study in patients with histologically-confirmed GBM, previously treated with radiation and must have failed both bevacizumab and temozolomide, unless contraindicated. The study utilizes a 3+3 dose-escalation design. Patients receive VAL-083 IV on days 1, 2, and 3 of a 21-day cycle. Tumor response is assessed according to RANO criteria prior to every other 21-day treatment cycle. Results: 25 patients have been enrolled across 8 dose cohorts ranging from 1.5 to 50mg/m²/d. A dose limiting toxicity consisting of grade 4 thrombocytopenia was observed at dose level 8 (50mg/m²/d). The DLT-related symptoms resolved rapidly and spontaneously without concomitant treatment. Prior to this, other treatment related toxicities have been mild to moderate and included two grade 1 lymphopenias and one grade 1 thrombocytopenia. Maximum tolerated dose (MTD) will be determined based on 3+3 design. Three patients had a response (stable disease or partial response) reporting improved clinical signs (maximum response of 84 wks). Pharmacokinetic analyses show dose-dependent linear systemic exposure with a short plasma 1-2h terminal half-life; Cmax ranged from 1130–739 ng/mL (7.7–5.1µM) at 40mg/m²/d. Compared to historical trials, the present regimen delivers substantively more drug by Cmax and dose intensity. A dose intensity of 25 mg/m²/wk in combination with radiation was previously shown superior to radiation alone against GBM; a dose intensity of 50 mg/m²/wk is achieved in the current trial. Conclusions: VAL-083 dosing may be limited by myelosuppression. The MTD will be studied in a Phase II expansion and possible phase III registration trial. Clinical trial information: NCT01478178