Current functional MRI techniques have shown promising results for prediction of response to chemoradiotherapy (CRT) in rectal cancer, but lack sufficient accuracy for clinical use. There is a wide variation in performance of functional MRI in response prediction reported. Most studies describe single parameter values from either diffusion or perfusion MRI. Single parameter measurements, such as mean ADC or K^trans, do not reflect tumour heterogeneity. The purpose of this study was to prospectively evaluate the role of quantitative diffusion weighted imaging (DWI) and dynamic contrast enhanced (DCE) imaging used in combination for multi-parametric voxel-wise prediction of treatment response in rectal cancer.This study used a voxel-by-voxel multi-parametric histogram analysis strategy to assess tumour heterogeneity and its changes in response to combined chemotherapy and radiotherapy (CRT). Twenty patients with locally advanced rectal cancer undergoing preoperative CRT prospectively underwent MRI on a 3T wide bore Siemens Skyra at 3 time-points: Pre-CRT, week 3 CRT, and post-CRT. The study protocol consisted of: (i) T2-weighted images (ii) DWI using RESOLVE, which has been previously shown to be robust with respect to geometrical distortions¹. Images were acquired with b-values 50 and 800s/mm² and 1 & 3 averages. ADC maps and calculated b=1400s/mm² images were produced as part of protocol. (iii) DCE consisted of pre-contrast VIBE scans with flip angles 2º and 15º in order to calculate native T1, followed by gadoversetamide (0.1mM/kg) injection and 60 phases using TWIST with a 5s temporal resolution. ADC and K^trans parameter maps were registered to T2-weighted images. Semi-automated segmentation was used to define the volume of interest from hyperintense tumour on the b-value=1400 images. A voxel-by-voxel technique was used to produce colour coded histograms of ADC and K^trans, as well as combined scatterplots for each time-point. CRT response was defined according to histopathology tumour regression grade (TRG) (AJCC 7th Edition)².Of 20 patients, 1 had clinical stage T2N2M0, 5 had T3N0M0, 4 had T3N1M0, 7 had T3N2M0, and 3 had T4N2M0. Eight patients had a good response (TRG0-1) and 11 patients had a poor response (TRG2-3) to CRT. Pathology for 1 patient is pending. A complete protocol and analysis strategy was successfully developed which has utilized commercial, in-house developed and works-in-progress (OncoTreat) software. In good responders, the week 3 histograms and maps showed both a shift in distribution of ADC of pixels to higher values and K^trans of pixels to lower values compared to the pre-CRT histogram. The ADC histograms for a good responder shown in Figure 1 demonstrated an increase in the absolute ADC values of voxels over the time-points. For the same patient, the majority of K^trans voxel values were high (red) pre-CRT, and by week 3-CRT the K^trans histogram demonstrated a marked reduction in the absolute K^trans values of voxels (Figure 2). Figure 3 shows the scatterplots demonstrating changes in combined ADC and K^trans of voxels of segmented region over the time-points for a good responder and a poor responder. We found the calculated b-value=1400 images useful for visualization of tumour. For DCE analysis, pre-registration of flip angle to dynamic images was a crucial step in producing pixel-by-pixel T1 map, to ensure accurate voxel-by-voxel calculation of K^trans and had to be performed outside of commercial software owing to its limitations. Patient 1 (good responder) had majority of voxels with high K^trans pre-CRT. A possible explanation for this is that the high K^trans is due to a well perfused oxic tumour, which is predictive of good radiotherapy response. In contrast, Patient 2, a poor responder, had low K^trans values pre-CRT, without much change the in values of voxels over the time-points (Figure 3). The low K^trans values in this patient may be due to poor perfusion representing a hypoxic tumour, which is predictive of a radio-resistant tumour and poor response. Multi-parametric histogram analysis of ADC and K^trans appears to be a promising and feasible method of assessing tumour heterogeneity and its changes in response to CRT in rectal cancer.