Novel Targeted Combination Therapies for Peripheral T-Cell Lymphoma

Introduction: New and promising evidence indicates that Aurora A and B kinases are important drivers of peripheral T-cell non-Hodgkin lymphoma (PTCL), a rare and heterogeneous disease accounting for ~15% of non-Hodgkin lymphomas (NHLs), without a clear standard of care. Auroras are a highly conserved family of oncogenic serine/threonine protein kinases that play key regulatory roles during mitosis. They are over-expressed and dysregulated in malignancy which make them very attractive therapeutic targets. We investigated the molecular, cellular, and cytokine drivers of PTCL in patients (S1108) treated with an aurora A inhibitor, alisertib (MLN8237). It is hypothesized that patient and pre-clinical models will guide the design of better strategies to target auroras in PTCL.

Methods: Immunohistochemistry (IHC) on 22 PTCL patient samples (S1108) for level of expression of aurora A, aurora B, Myc, Notch-1, Bcl-2 and PI3Kd was performed. Nuclear positivity was determined for aurora A, B and Myc using Aperio’s Nuclear v9 Quantification algorithm. Expression of Bcl-2, Notch-1 and PI3Kd was quantified with Aperio’s Positive Pixel Count algorithm. The Raybiotech G2000 human cytokine array of 174 unique cytokines was analyzed using pre-/post-alisertib (day 8) therapy from 12 PTCL patient samples. MTS cellular cytotoxicity assays were conducted with single agent alisertib, PI3K inhibitor (MLN1117), mTor inhibitor (MLN0128) and vincristine (VCR) in TIB-48, CRL-2396 and Jurkat T-NHL cell lines. Combination studies were conducted with alisertib + MLN1117 or MLN0128 or VCR or triple combinations to ascertain IC₅₀s and synergism (Chou-Talalay). A mouse PTCL xenograft model is underway to evaluate the anti-tumor activity of alisertib, MLN1117, and alistertib + MLN1117. Mechanistic studies were conducted for target and pathway inhibition.

Results: As previously shown in patients from S0350, S1108 confirms aurora B is over-expressed in most patients (nuclear) versus aurora A (cytoplasmic). PI3Kd is over-expressed on the surface and/or the cytoplasm mainly within the tumor microenvironment. Myc expression is nuclear within a host-response type infiltrate in the tumor. Bcl-2 expression is confined to small T-cells, a host-response marker. Notch-1 expression was only observed in 2 cases within large tumor cells. Although level of expression of the 6 IHC markers did not correlate with alisertib activity, aurora B and PI3Kd are highly over-expressed in PTCL. Cytokine profiling revealed that pre-/post-alisertib is unique to each patient and are correlated to therapeutic response. Of note 4 of 12 patients had an elevated soluble TNF-RII, a poor prognostic marker in PTCL. IC₅₀s for alisertib in CRL-2396, TIB-48 and Jurkat cells: 44-64 nM, MLN1117 9-14 μM, and MLN0128 36-130 nM. Combination indices (CI) indicate strong synergism for alisertib +MLN1117, MLN0128+MLN1117 and alisertib+MLN1117+VCR. These combinations lead to target inhibition and apoptosis. The PTCL xenograft mouse safety and response data will be available at the meeting.

Conclusions: Aurora and PI3Kd inhibition + VCR holds promise as a combination targeted approach for patients with PTCL. IHC to stratify and cytokines to follow therapy are biomarkers to be considered in prospective trials in the treatment of PTCL patients.