Design, Synthesis and Anticancer Evaluation of Nitroimidazole Radiosensitisers

Abstract

The role of hypoxic tumour cells in resistance to radiotherapy, and in suppression of immune response, continues to endorse tumour hypoxia as a bona fide, yet largely untapped, drug target. Radiotherapy innovations such as stereotactic body radiotherapy herald new opportunities for classical oxygen-mimetic radiosensitisers. Only nimorazole is used clinically as a radiosensitiser, and there is a dearth of new radiosensitisers in development. In this report, we augment previous work to present new nitroimidazole alkylsulfonamides and we document their cytotoxicity and ability to radiosensitise anoxic tumour cells in vitro. We compare radiosensitisation with etanidazole and earlier nitroimidazole sulfonamide analogues and we identify 2-nitroimidazole and 5-nitroimidazole analogues with marked tumour radiosensitisation in ex vivo assays of surviving clonogens and with in vivo tumour growth inhibition.

Keywords: DNA damage; chemoradiotherapy; electron affinity; hypoxia; nitroimidazole; prodrugs; radiosensitisers; radiotherapy; sulfonamide; tumour microenvironment.

Figure 1

Nitroimidazole radiosensitisers.

Scheme 1

Synthesis of new nitroimidazole methyl sulfonamides. Reagents: (i) ClCH₂SO₂Cl, Et₃N, DCM, 81%; (ii) 2-nitroimidazole, NaI, Cs₂CO₃, DMF, 42%; (iii) HOAc/H₂O/THF, 92%; (iv) TBDMSCl, imidazole, DMF, 97%; (v) MCPBA, DCM, 96%; (vi) NaN₃, NH₄Cl, DMF, 25%; (vii) H₂, Pd/C, 100%; (viii) BrCH₂SO₂Cl, Et₃N, DCM, 54%; (ix) 2-nitroimidazole, NaI, Cs₂CO₃, DMF, 54%; (x) TBAF, THF, 61%.

Scheme 2

Synthesis of new nitroimidazole ethyl sulfonamides. Reagents: (i) NCS, HCl, MeCN, then (2,2-dimethyl-1,3-dioxolan-4-yl)methanamine, iPr₂NEt, DCM, 24%; (ii) HOAc/H₂O/THF, 85%; (iii) NCS, HCl, MeCN, then 4-(2-aminoethyl)morpholine, iPr₂NEt, DCM, 24%; (iv) tert-butyl (2-bromoethyl)carbamate, NaI, Cs₂CO₃, DMF, 67%; (v) TFA, 98%; (vi) mercaptoethanol, NCS, nBu₄NCl, water, then 32, iPr₂NEt, MeCN, 22%.

Figure 2

In vivo radiosensitisation of hypoxic cells in (A) HCT116/54C tumours and (B) UT-SCC-74B tumours by ex vivo clonogenic assay in NIH-III mice. Tumours were excised 18 h after mice were treated by i.v. injection with 2.2 mmol/kg drug alone, 10 Gy radiation (RAD) alone or 2.2 mmol/kg drug combined with 10 Gy radiation (n = 3–5), and plated to determine the number of surviving cells relative to untreated controls. ****, p < 0.0001; *, p < 0.05 for comparison of RAD vs. drug + RAD by one-way ANOVA with Sidak’s multiple comparison analysis.

Figure 3

Ex vivo clonogenic assay in HCT116/54C tumours from NIH-III mice treated with 34 or 38 at 2.2 mmol/kg by i.v. injection either 5 min before 10 Gy radiation (RAD) or 20 min after (n = 5). Tumours were collected 18 h after treatment and plated to determine the number of surviving cells relative to untreated controls. ***, p < 0.001 vs. RAD alone by one-way ANOVA with Sidak’s multiple comparison analysis.

Figure 4

Tumour growth inhibition in HCT116/54C and UT-SCC-74B tumour xenografts. NIH-III mice were treated with radiosensitisers at 2.2 mmol/kg by i.v. injection, 5 min prior to radiation, given either as a single dose at 10 Gy or fractionated over 3 successive weeks (day 0, 7 and 14) at 5 Gy. (A) Average tumour volume over time (±SEM); (B) change in bodyweight (mean ± SEM); (C) RTV⁴ survival time. N = 8–10 for treatments and 3–4 for controls.