Treatment of patients with chronic-phase CML with IM results in hematologic remissions in over 90% of chronic phase patients and complete cytogenetic remissions expected in over 80% of patients in upfront treatment.20
Despite these excellent clinical responses, the overwhelming majority of patients treated with IM remain with molecularly measurable disease. Data from the Imperial College recently reported a 5-year cumulative incidence of CMR of only 8.3% for patients treated up-front with IM.21
Perhaps more reflective of the subjects reported here, Palandri et al., found that only one of 277 late chronic phase patients resistant or intolerant to IFN-α had a durable CMR with a median of 72 months of follow-up.22
Data continue to accumulate that the quiescent, primitive Ph chromosome positive CML stem cells are insensitive to imatinib in vitro
and therefore render single agent IM as incapable of completely eradicating the disease23–25
and result in relapse in most patients who stop IM therapy.26–30
Previous reports have shown the ability to induce T cell responses in patients undergoing peptide-based vaccines that attempted to target the fusion sequence in the P210 protein unique to the CML. The majority of the early reports suggested the detection of peptide specific T cell proliferative responses and delayed-type hypersensitivity responses and not clinical responses.31
Efforts to modify the strategies through vaccine alteration or adjuvants resulted in small, but measureable changes in disease burden including several cases who achieved molecular remissions.32,33
As with all single antigen vaccine approaches targeting defined epitopes, these strategies are limited by the requirement for patients to express specific HLA class I or II molecules for peptide presentation. In contrast, the broad spectrum of tumor-associated antigens delivered by genetically modified tumor cell-based vaccines are not HLA class restricted. In addition, each patient selects the hierarchy of antigens that are recognized, increasing the potential for a response while the polyvalency of this formulation reduces the likelihood of tumor escape by antigen loss. The K562 cell line, originally derived from a patient with CML, expresses many potential antigens that may serve as immunologically important therapeutic targets. Candidate antigen targets shared by both the K562 cell line and primary CML include the BCR-ABL
fusion protein, proteinase-3,8
Wilm's tumor-1 (WT-1),9–14
In spite of these advantages, neither antibody nor T cell responses were detected against these reported candidate antigens even though they are abundantly expressed by the K562 cells (data not shown). These results mirror those reported by Lee and colleagues who identified T cell responses to autologous leukemia cells, but not to any known leukemia-associated antigens (including those examined in our study) after IM induced remission.34
We have since embarked on an unbiased screen of a cDNA expression library from K562 cells using sera from the subjects reported here. While still preliminary, we have now identified over 25 antigens expressed by CML cells that were recognized in post-, but not pre-vaccine samples. Interestingly, while some of the antigens identified are known immune targets in CML, including hyaluronan-mediated motility receptor (RHAMM)35
and enolase alpha36
, most of the targets that have been discovered in this screen are novel. Studies are ongoing to determine the frequency of immune responses to each of these antigens in larger numbers of CML patients and to examine the correlation of clinical responses with the pattern of antigen recognition.
Although our study was not designed to test the impact of IM on the immune effects seen, it is possible that concurrent use of IM may have influenced the effectiveness of the K562/GM-CSF immunotherapy. Abelson tyrsoine kinases are one of the three families of tyrosine kinases that play important roles in the development, activation, and expansion of T cells. These non-receptor kinases are activated in response to engagement of both the pre-T cell receptors and T-cell receptors and transduce signals that affect T-cell development and mature T-cell function.37,38
Limiting the function of these signaling pathways with IM and other TKIs is one mechanism that could result in immunosuppressive effects.39,40
It is just as plausible however, that IM enhanced the biologic activity of the K562/GM-CSF vaccines by its interactions at one or more critical junctures of the proposed T-cell activation pathways. IM has been shown to enhance the stimulatory capacity of antigen presenting cells to activate T cells, 41
to promote the development and activity of interferon-producing killer dendritic cells,42
and to limit the suppressive feedback loop of regulatory T-cells.43
The balance between IM's influence on the opposing arms of the immune response remains unclear, however its effects should not be discounted. These issues not withstanding, the ability of IM to hold the disease in a state of MRD affords a unique opportunity to intercede with immunotherapies that have mechanisms of action that are distinct from leukemia-intrinsic targeted therapies.
In general, the clinical importance of small changes in CML tumor burden, as measured by PCR, for remains unclear. Most PCR assays suggest that changes of up to 1-log may fall within the varibility of the assay and trends in the PCR values over time may better reflect a true clinical effect. Our study attempted to minimize the confusion in presenting the clinical responses by not simply duplicating the measures recommended by Branford, et. al.44
and Hughes, et. al45
to help analyze the IRIS study, but by presenting all of the data points to best show the trends in measures over time. The changes in tumor burden observed following immunotherapy in our study were intriguing as 13 of our patients had decreasing PCR trends following our immunotherapy, more than half of the responding patients reached the level of molecularly undetectable at some point during follow up, and 5 of these 7 responses appear durable. Despite the fact that PCR is in its relative infancy as a clinical tool, downward trends and achieving molecularly undetectable levels of disease support the biologic activity of the immunotherapy. The current study did not randomize patients between immunotherapy and observation leaving the question of ongoing molecular improvement based solely on IM therapy unanswered. Our patient population was composed of late chronic phase patients, 12 of whom had never achieved a major molecular response despite a median of over 3 years on IM, who would be unlikely to show significant changes in their tumor burdens on stable IM alone.
This pilot study demonstrated that the use of K562/GM-CSF immunotherapy is well tolerated with no serious adverse events for any of the 76 vaccine cycles in our 19 subjects. This intervention was associated with the reduction of tumor burden and the achievement of both major and complete molecular responses in CML patients with previous incomplete responses to single agent IM. Ultimately, the impact of immunotherapy will best be determined in a trial in which the persistence of undetectable disease is assessed following the planned withdrawal of IM. This design has been incorporated into our ongoing follow-up trial to this pilot study.
STATEMENT OF TRANSLATIONAL RELEVANCE
The tremendous strides by imatinib towards improving outcomes for patients with CML appear only limited by its inability to directly eliminate early progenitors that remain in most patients despite years of therapy. Targeting such minimal residual disease may lead to improved outcomes and possibly eliminate the need for lifelong therapy. Immunotherapy has the potential to target such MRD based on the clinical observations that allogenenic stem cell transplant and donor lymphocyte infusions are both capable of curing patients with CML. Our manuscript describes the clinical impact of a K562/GM-CSF whole cell vaccine at decreasing MRD in a group of late, chronic phase patients. Elimination of MRD may result in medical cures for CML.