SHIP1 Is Present but Strongly Downregulated in T-ALL, and after Restoration Suppresses Leukemia Growth in a T-ALL Xenotransplantation Mouse Model.

Acute lymphoblastic leukemia (ALL) is the most common cause of cancer-related death in children. Despite significantly increased chances of cure, especially for high-risk ALL patients, it still represents a poor prognosis for a substantial fraction of patients. Misregulated proteins in central switching points of the cellular signaling pathways represent potentially important therapeutic targets. Recently, the inositol phosphatase SHIP1 (SH2-containing inositol 5-phosphatase) has been considered as a tumor suppressor in leukemia. SHIP1 serves as an important negative regulator of the PI3K/AKT signaling pathway, which is frequently constitutively activated in primary T-ALL. In contrast to other reports, we show for the first time that SHIP1 has not been lost in T-ALL cells, but is strongly downregulated. Reduced expression of SHIP1 leads to an increased activation of the PI3K/AKT signaling pathway. SHIP1-mRNA expression is frequently reduced in primary T-ALL samples, which is recapitulated by the decrease in SHIP1 expression at the protein level in seven out of eight available T-ALL patient samples. In addition, we investigated the change in the activity profile of tyrosine and serine/threonine kinases after the restoration of SHIP1 expression in Jurkat T-ALL cells. The tyrosine kinase receptor subfamilies of NTRK and PDGFR, which are upregulated in T-ALL subgroups with low SHIP1 expression, are significantly disabled after SHIP1 reconstitution. Lentiviral-mediated reconstitution of SHIP1 expression in Jurkat cells points to a decreased cellular proliferation upon transplantation into NSG mice in comparison to the control cohort. Together, our findings will help to elucidate the complex network of cell signaling proteins, further support a functional role for SHIP1 as tumor suppressor in T-ALL and, much more importantly, show that full-length SHIP1 is expressed in T-ALL samples.

A Lack of Effectiveness in the ATM-Orchestrated DNA Damage Response Contributes to the DNA Repair Defect of HPV-Positive Head and Neck Cancer Cells.

Patients with human papillomavirus-positive squamous cell carcinoma of the head and neck (HPV+ HNSCC) have a favorable prognosis compared to those with HPV-negative (HPV-) ones. We have shown previously that HPV+ HNSCC cell lines are characterized by enhanced radiation sensitivity and impaired DNA double-strand break (DSB) repair. Since then, various publications have suggested a defect in homologous recombination (HR) and dysregulated expression of DSB repair proteins as underlying mechanisms, but conclusions were often based on very few cell lines. When comparing the expression levels of suggested proteins and other key repair factors in 6 HPV+ vs. 5 HPV- HNSCC strains, we could not confirm most of the published differences. Furthermore, HPV+ HNSCC strains did not demonstrate enhanced sensitivity towards PARP inhibition, questioning a general HR defect. Interestingly, our expression screen revealed minimal levels of the central DNA damage response kinase ATM in the two most radiosensitive HPV+ strains. We therefore tested whether insufficient ATM activity may contribute to the enhanced cellular radiosensitivity. Irrespective of their ATM expression level, radiosensitive HPV+ HNSCC cells displayed DSB repair kinetics similar to ATM-deficient cells. Upon ATM inhibition, HPV+ cell lines showed only a marginal increase in residual radiation-induced γH2AX foci and induction of G2 cell cycle arrest as compared to HPV- ones. In line with these observations, ATM inhibition sensitized HPV+ HNSCC strains less towards radiation than HPV- strains, resulting in similar levels of sensitivity. Unexpectedly, assessment of the phosphorylation kinetics of the ATM targets KAP-1 and Chk2 as well as ATM autophosphorylation after radiation did not indicate directly compromised ATM activity in HPV-positive cells. Furthermore, ATM inhibition delayed radiation induced DNA end resection in both HPV+ and HPV- cells to a similar extent, further suggesting comparable functionality. In conclusion, DNA repair kinetics and a reduced effectiveness of ATM inhibition clearly point to an impaired ATM-orchestrated DNA damage response in HPV+ HNSCC cells, but since ATM itself is apparently functional, the molecular mechanisms need to be further explored.