Management of high-grade gliomas remains a complex challenge. Standard of care consists of microsurgical resection, chemotherapy and radiation, but despite these aggressive multimodality therapies the overall prognosis remains poor. A major focus of ongoing translational research studies is to develop novel therapeutic strategies that can maximize tumor cell eradication while minimizing collateral side effects. Particularly, biological intratumoral therapies have been the focus of new translational research efforts due to their inherent potential to be both dynamically adaptive and target specific. This two-part review will provide an overview of biological intratumoral therapies and summarize key advances and remaining challenges in intratumoral biological therapies for high-grade glioma. Part I focuses on discussion of the concepts of intratumoral delivery and immunotoxin therapies.