BIOMARKERS OF GO’S CLINICAL EFFICACY

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The biomarker that has thus far been most recognized as predictor of GO’s clinical efficacy with regard to improvement of survival is, as discussed in the previous section in detail, the cytogenetic/molecular profile of the leukemia. In addition, emerging data suggest that CD33 SNPs may prove useful as biomarkers for long-term benefit of GO as well. A number of other AML cell-associated factors – including drug efflux activity, CD33 expression levels, CD33 saturation, circulating CD33 antigenic load, and perhaps methylation status of SOCS3 – have been described as predictors of short-term response, i.e. reduction in tumor cell loads and/or achievement of CR, but it is unclear whether they could also serve as predictors of long-term benefit. Moreover, it has not been studied to what degree these factors underlie the observed relationship between disease risk and GO efficacy.

Consistent with preclinical studies indicating the central role of drug efflux for GO-induced cytotoxicity, correlative studies on biospecimens from patients enrolled in GO monotherapy trials showed an association between P-glycoprotein as well as MRP1 activity with persistence of marrow blasts, failure to achieve CR, or reduced in vitro drug-induced apoptosis (143, 156). This susceptibility to drug efflux may significantly limit the efficacy of GO in clinical practice, especially for the treatment of relapsed or refractory non-APL AML. It is well established that ABC transporter activity, in particular mediated by P-glycoprotein, predicts for therapeutic failure of standard induction therapy (157); thus, the same factor that predicts for failure of conventional chemotherapy and need for salvage therapy also predicts for failure of GO. Additionally, increasing evidence links ABC drug transporters to protection of cancer stem cells – the intended targets for GO – from chemotherapeutic agents (158). In contrast to drug efflux, the effect of CD33 expression on GO efficacy was initially uncertain, and several smaller studies did not find a correlation between CD33 expression levels and response to GO (103, 122). However, later correlative studies conducted on specimens from patients enrolled on the phase 2 trials with GO monotherapy found higher CD33 expression levels on AML blasts to be associated with favorable outcome after GO monotherapy, although multivariate analyses suggested that drug efflux was the more relevant factor for clinical GO resistance than CD33 expression levels (159). A conceivable explanation for this difficulty to demonstrate a quantitative relationship between CD33 expression and clinical efficacy of GO could include the recent observation that CD33 expression levels are inversely associated with favorable cytogenetic/molecular disease features (160) or the possibility that CD33 abundance on disease-relevant stem/progenitor subsets of AML cells may not be adequately reflected by the average CD33 expression of bulk blasts. In line with the former notion, older data, again derived from pediatric AML, indicated that AML patients with high CD33 expression on AML blasts have worse outcomes than those with low CD33 expression when treated with conventional chemotherapy (161). Undoubtedly, this relationship between CD33 expression and inherent AML disease biology could modify the association between CD33 expression and GO efficacy and render such correlative studies challenging. To make matters more complex, recent data raised the possibility that some CD33 SNPs may not only be associated with CD33 expression levels but also response to GO-containing chemotherapy (162). Specifically, homozygosity for the variant allele (TT) at the coding SNP, rs12459419 (C>T; A14V), homozygosity of the reference allele (AA) at the coding SNP, rs2455069 (A>G; R69G), and the 3’ UTR SNP, rs1803254 (G>C) have been associated with significantly lower CD33 expression on AML blasts as compared to other genotypes. Moreover,
among Caucasians, homozygosity (GG) at the coding SNP, rs35112940 (G>A; R304G), was independently associated with improved relapse-free survival relative to the other genotypes in a cohort of pediatric patients treated with GO-containing multi-agent chemotherapy (AAML03P1) but not in a cohort of pediatric patients treated with multi-agent chemotherapy that did not contain GO (St. Jude AML02 trial) (163). Whether these SNPs directly influence the function of CD33 and response to GO is currently under active investigation.

Considering the relationship between GO uptake and efficacy, it is not surprising that in vitro studies have linked reduced CD33 saturation to reduced GO-induced cytotoxicity (103). While the clinically used doses of GO are typically saturating, a high CD33-antigenic load in the peripheral blood probably can act as antibody sink and lead to reduced CD33 saturation in the marrow, thereby adversely affecting GO efficacy (103). An important yet neglected aspect of CD33 uptake is the timing of GO administration. Early studies indicated that surface CD33 levels return to pretreatment levels within 72 hours after anti-CD33 antibody administration despite internalization and modulation (55, 70). This observation suggests that repeated administrations of lower, (near-)saturating doses of GO every 3 days, as clinically pioneered by the ALFA group (143-145), may enhance intracellular accumulation of the calicheamicin-g₁^I derivative over the initial biweekly administration schedule.

Finally, preliminary data suggest that the methylation status of SOCS3 may serve as biomarker of responsiveness to GO. Specifically, in an uncontrolled, retrospective study on 24 patients treated with GO alone or in combination with chemotherapy at a single institution, methylation of the SOCS3 CpG island was found in 8; there was a statistically insignificant increase in response rate (86% vs. 56%; P=0.17) and longer overall survival (25.1 vs. 10.3 months; P=0.09) in patients with SOCS3 hypermethylation (164).