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Polypeptides contemplated herein specifically encompass the CARs of the present disclosure, or sequences that have deletions from, additions to, and/or substitutions of one or more amino acid of a polypeptide as contemplated herein. The present invention contemplates, in part, engineered TCR and CAR polypeptides and fragments thereof, cells and compositions comprising the same, and vectors that express polypeptides. While this thing could be a body part, a type of clothing, or some inanimate object; it could also be a type of attitude or mood (Dominant, Cheerful, Submissive, etc.). Illustrative hinge domains suitable for use in the CARs described herein include the hinge region derived from the extracellular regions of type 1 membrane proteins such as CD8.alpha., CD4, CD28 and CD7, which may be wild-type hinge regions from these molecules or may be altered. For example, in particular embodiments, it may be desirable to improve the binding affinity and/or other biological properties of the engineered TCRs or CARs by introducing one or more substitutions, deletions, additions and/or insertions. Polypeptides include "polypeptide variants." Polypeptide variants may differ from a naturally occurring polypeptide in one or more substitutions, deletions, additions and/or insertions. As noted above, polypeptides of the invention may be altered in various ways including amino acid substitutions, deletions, truncations, and insertions.

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In particular embodiments, the inhibitor of the PI3K/AKT/mTOR pathway inhibits PDK-1 activation of AKT. In various embodiments, a method for maintaining the proliferation and decreasing the differentiation of restimulated T cells expressing an engineered TCR or CAR comprising: (a) contacting all or a portion of a population of proliferated T cells comprising an engineered TCR or CAR with an anti-CD3 antibody or CD3-binding fragment thereof, and an anti-CD28 antibody or CD28-binding fragment thereof, which stimulates a CD28 accessory molecule on the surface of the T cells, thereby restimulating the activated T cells to proliferate; wherein the restimulated T cells have maintained proliferation and decreased differentiation compared to the proliferation of T cells that were stimulated or restimulated in the absence of the inhibitor of the PI3K/AKT/mTOR pathway is provided. In additional embodiments, the population of activated T cells restimulated in the presence of an inhibitor of PI3K/AKT/mTOR pathway do not express CD57 or KLRG1 or express less CD57 or KLRG1 compared to a population of T cells activated and stimulated in the absence of the inhibitor of PI3K/AKT/mTOR pathway.

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