Table 2.

Compilation of animal studies investigating normal tissue effects and/or tumour control after proton minibeam irradiation.

Study Aim [Ref]	Treatment Protocol	Observations
Normal tissue toxicity evaluation after mouse ear irradiation
Comparative study of mouse ear irradiation [94]	(1) Homogenous 20 MeV proton field of overall 60 Gy (2) Minibeam (4 × 4 minibeams, 0.18 × 0.18 mm2, 1.8 mm distance) of 6000 Gy peak dose	Up to 4-times greater extent of ear swelling, erythema, and desquamation in the homogenous group vs. minibeam. Hair loss and damage to the sebaceous glands observed only in the homogenously exposed group.
Minibeam size dependence of normal tissue effects in mouse ear [95]	20 MeV minibeam (4 × 4 minibeams, 1.8 mm centre-to-centre beam distance) of a 6 kGy peak dose, with various beam sizes	The ideal minibeam size for minimal side effects should be < 0.1 mm. Still, any spatial fractionation with a beam size < 1 mm leads to superior normal tissue protection as compared to homogenous irradiation.
Normal tissue toxicity and tumour control evaluation after high-grade glioma treatment in rats
The effect of proton minibeam vs. standard proton therapy on rat high-grade gliomas [96]	(1) Standard proton therapy of 100 MeV with 2 Gy/min at a 1 cm depth of 25 Gy in one fraction (2) Proton minibeam of 400 μm-wide slits and 3200 μm centre-to-centre distance, 57 Gy peak dose at a 1 cm depth	Main aim: to evaluate the reduction in neurotoxicity with proton minibeam. Rats under standard proton therapy developed significant skin damage and long-term brain damage (after 6 mo of evaluation) as compared to minimal toxicity in the minibeam group.
Tumour control of high-grade gliomas in rats with proton minibeam [97]	100 MeV proton minibeam of 400 μm-wide slits and a 3200 μm centre-to-centre distance, 70 Gy peak dose at a 1 cm depth; one dose fraction	22% disease-free long-term survival (6 mo follow-up) in the treated group. Mean survival time of the control group (untreated): 18 d; mean survival time of minibeam group: 32.5 d.
Evaluation of cerebral functions in rats with high-grade gliomas after proton minibeam [98]	100 MeV proton minibeam with a 57 Gy peak dose at a 1 cm depth. Minibeam width at 1 cm was 1100 ± 50 μm.	No locomotory, behavioural, or cognitive differences observed between proton minibeam and control (unirradiated) groups. Small growth perturbation was observed in the treated group (12.5% lower body weight and size).