Abstract: Methods of treating patients suffering from cancer using a combination of 1) an immunomodulatory agent; 2) a check point inhibitor; and 3) a cancer antigen releasing therapy are provided. In some aspects, the immunomodulatory agent is an anti-CD3 immunotoxin, the check point inhibitor inhibits the interaction of PD-1 and PD-L1 and the cancer antigen releasing therapy is radiation.

Abstract: The present invention relates to a method of expressing an immunotoxin in Pichia pastoris strain mutated to toxin resistance comprising a) growing the Pichia pastoris in a growth medium comprising an enzymatic digest of protein and yeast extract and maintaining a dissolved oxygen concentration at 40% and above; and b) performing methanol induction with a limited methanol feed of 0.5-0.75 ml/min/IO L of initial volume during induction along with a continuous infusion of yeast extract at a temperature below 17.5° C., antifoaming agent supplied up to 0.07%, agitation reduced to 400 RPM, and the induction phase extended out to 163 h.

Abstract: The present invention relates to a method of expressing an immunotoxin in Pichia pastoris strain mutated to toxin resistance comprising a) growing the Pichia pastoris in a growth medium comprising an enzymatic digest of protein and yeast extract and maintaining a dissolved oxygen concentration at 40% and above; and b) performing methanol induction with a limited methanol feed of 0.5-0.75 ml/min/IO L of initial volume during induction along with a continuous infusion of yeast extract at a temperature below 17.5° C., antifoaming agent supplied up to 0.07%, agitation reduced to 400 RPM, and the induction phase extended out to 163 h.

Abstract: The present invention relates to a method of expressing an immunotoxin in Pichia pastoris strain mutated to toxin resistance comprising a) growing the Pichia pastoris in a growth medium comprising an enzymatic digest of protein and yeast extract and maintaining a dissolved oxygen concentration at 40% and above; and b) performing methanol induction with a limited methanol feed of 0.5-0.75 ml/min/IO L of initial volume during induction along with a continuous infusion of yeast extract at a temperature below 17.5° C., antifoaming agent supplied up to 0.07%, agitation reduced to 400 RPM, and the induction phase extended out to 163 h.

Abstract: In one aspect the present invention relates to a method of expressing an immunotoxin in Pichia pastoris strain mutated to toxin resistance comprising a) growing the Pichia pastoris in a growth medium comprising an enzymatic digest of protein and yeast extract and maintaing a dissolved oxygen concentration at 40% and above; and b) performing methanol induction with a limited methanol feed of 0.5-0.75 ml/min/10 L of initial volume during induction along with a continuous inusion of yeast extract at a temperature below 17.5° C., antifoaming agent supplied up to 0.07%, agitation reduced to 400 RPM, and the induction phase extended out to 163 h.