Testicular germ cell tumors (TGCTs) are exceptionally sensitive to genotoxic chemotherapy, resulting in a high cure rate for the young men presenting with these malignancies. However, this treatment is associated with significant toxicity, and a subset of malignant TGCTs demonstrate chemoresistance. Mixed nonseminomas often contain pluripotent embryonal carcinoma (EC) cells, the cancer stem cells (CSCs) of these tumors. We hypothesized that differentiation therapy, a treatment strategy which aims to induce differentiation of tumor-propagating CSCs to slow tumor growth, could effectively treat mixed nonseminomas without significant toxicity. The FDA-approved antipsychotic thioridazine and the agricultural antibiotic salinomycin are two drugs previously found to selectively target CSCs, and here we report that these agents differentiate EC cells in vitro and greatly reduce their tumorigenic potential in vivo. Using a novel transformed induced pluripotent stem cell allograft model and a human xenograft model, we show that thioridazine extends the survival of tumor-bearing mice and can reduce the number of pluripotent EC cells within tumors. These results suggest that thioridazine could be repurposed as an alternative TGCT treatment that avoids the toxicity of conventional chemotherapeutics.

As global temperatures rise due to climate change, crops are becoming increasingly vulnerable to failure. We tested the accessibility of thermodynamic phenotypic diversity within limited positional changes using 2 ⁹ =512 mutants of a terpene synthase (TPS), Tobacco 5-epi-Aristolchene synthase. First, we measured the thermal unfolding curves of each mutant and found that mutations shifted the T _m s both higher and lower, including a cohort of mutants that failed to fold. The low correlation coefficient between the T _m s of these mutants and the proportions of each terpene product output by the enzymes revealed that thermostability and product output are independent traits. Maximum Noise Entropy analyses were used to analyze the impact of the 9 mutational positions on thermostability, revealing that three of these positions were mainly responsible for the trait of increased T _m . These positions form a functional network as measured by the nonlinearity of their combined effects on thermostability. Unexpectedly, the strongly destabilizing positions combine nonlinearly to ameliorate each other’s deleterious effects resulting in a synergistic dampening. Taken together, our study shows the high potential for specialized metabolic enzyme engineering but also reveals a complex interconnected system of amino acids that will continue to evade perfect predictability.