Diffuse optical tomography (DOT) has attracted attentions in the last two decades due to its intrinsic sensitivity in imaging chromophores of tissues such as hemoglobin, water, and lipid. However, DOT has not been clinically accepted yet due to its low spatial resolution caused by strong optical scattering in tissues. Structural guidance provided by an anatomical imaging modality enhances the DOT imaging substantially. Here, we propose a computed tomography (CT) guided multispectral DOT imaging system for breast cancer imaging. To validate its feasibility, we have built a prototype DOT imaging system which consists of a laser at the wavelength of 650 nm and an electron multiplying charge coupled device (EMCCD) camera. We have validated the CT guided DOT reconstruction algorithms with numerical simulations and phantom experiments, in which different imaging setup parameters, such as projection number of measurements and width of measurement patch, have been investigated. Our results indicate that an air-cooling EMCCD camera is good enough for the transmission mode DOT imaging. We have also found that measurements at six angular projections are sufficient for DOT to reconstruct the optical targets with 2 and 4 times absorption contrast when the CT guidance is applied. Finally, we have described our future research plan on integration of a multispectral DOT imaging system into a breast CT scanner.