Early Detection of Minimal Residual Disease in Biological Prognostic Risk Groups in Chronic Lymphocytic Leukaemia (CLL) Utilizing the Novel CD160FCA Assay

CLL is a heterogenous disease. At diagnosis, prognostication is possible using a number of clinical and biological factors, which can inform the choice of treatment. In terms of response to treatment, minimal residual disease (MRD) detection is becoming increasingly important in CLL. Utilizing the CD160FCA assay, we have evaluated the time to MRD detection for a number of established prognostic markers.

Utilizing the anti-CD160 monoclonal antibody (BY55, Coulter Immunotech, Marseille, France), we have developed a highly sensitive CD160 Flow Cytometric Assay (CD160FCA) incorporating a sequential gating strategy. Between July 2010 and July 2011, 63 patients were investigated for MRD by the routine clinical diagnostic service. 47 (73%) had completed treatment and were evaluable for long-term outcome. Response rates and event-free survival (EFS) were determined. The time to MRD detection post therapy was determined using CD160FCA for the prognostic indicators, ZAP-70, CD38, beta-2 microglobulin (B₂M) and cytogenetic abnormalities.

As expected, establishing clonality by light-chain restriction could not be reliably measured in patients with very low B-cell numbers, and hence could not reliably be used to determine MRD. The CD160FCA could reliably monitor patients for MRD, including immunotherapy with Rituximab and Campath. 28 Patients (44%) were in complete remission (CR) following therapy by current NCI guidelines. Of these patients, 9/28 (32%) were CR-MRD positive and had significantly shortened EFS compared with CR-MRD negative patients (61 days vs 957 days P<0.0001). Those patients with adverse cytogenetic markers had shorter time to disease detection (TTD): p53 mutation (30.5 days), 17pdel (61 days) and 11qdel (122 days) compared to +12 (427d), 13qdel (957d) and 13qdel with additional abnormalities (760 d). Of interest, patients with bi-allelic 13qdel had significantly shorter TTD compared to those with monoallelic 13qdel (46d vs 957d).

Likewise CD38 status (CD38+: 92d vs CD38-: 669d; P=0.03) and B₂M prior to treatment (B₂M high: 91d vs B₂M normal: 822d; P=0.004) were predictors of TTD based on residual disease assessment by the CD160FCA. 25 patients (40%) were assessed for ZAP-70 prior to commencing therapy. Those patients who were ZAP-70 positive had shorter TTD compared to those who were negative (ZAP+: 61d vs ZAP- 822d; P=0.01).

Many clinical studies base their entry criteria on clinical and biological prognostication, as this provides insights into the biology of CLL and its response to therapy. The CD160FCA is a single tube tumor specific assay for MRD detection, which is highly sensitive, independent of the therapy and can be employed throughout treatment. Patients in CR had significantly different EFS based on their MRD status following treatment using the CD160FCA. For those patients with adverse prognostic markers (including CD38, ZAP-70 and B₂M), the detection of MRD or relapsing disease using CD160FCA, is significantly shorter than those with a normal or good prognosis.

No relevant conflicts of interest to declare.