The main purpose of the present investigations was to provide proof of principle of feasibility, safety, and efficacy of a simple and inexpensive protocol for more effective cell-mediated immunotherapy of patients fully resistant to all available anti-cancer modalities. Based on successful cure with no severe side effects of the first 12 years old girl treated with IMAK currently nearly 30 years out with no further treatment, a physician with two children in excellent clinical condition (Slavin 2005), a total of 32 additional patients with different MDR incurable hematologic malignancies not eligible for allogeneic SCT consented to be treated with IMAK. Considering the anticipated short circulation time of non-engrafting mismatched donor-derived killer cells, it seemed reasonable to anticipate that cure could be anticipated only when IMAK will be applied against MRD or against a relatively low tumor burden. Alternatively, the procedure would have to be repeated using different donor cells, but in this pilot study only one treatment cycle with IMAK was applied. Indeed, as shown in table 3 and figure 1, it was really surprising that 22 out of 33 patients treated with IMAK using either haploidentical related donor cells or fully mismatched lymphocytes obtained from consenting unrelated donors showed evidence of response, while at least 6 of 22 patients that were observed for more than 5 years may even be considered cured.
The anticancer effects of IL-2-activated lymphocytes is not new but originally considered clinically relevant for activating patient's own anti-cancer effector cells in patients with cancer (Mule JJ, Shu S, Schwarz SL et al (1984), Rosenberg SA, Mulé JJ, Spiess PJ et al (1985), Rosenberg SA et al (1987)). Focusing on the particular role of NK cells as anti-cancer effector cells was also confirmed by many previous investigators (Kärre K (2002), Myers JA and Miller JS (2021), Chu J et al (2022)) and the use of NK cells derived from different possible sources was also suggested by Miller and colleagues (Johnson and Miller JS (2018), Tanaka and Miller (2020)). The important contribution of the present work using IMAK for treatment of multi-drug resistant cancer initially pioneered by Slavin in 1993 for successful treatment of patients with hematologic malignancies (Slavin 2015) and subsequently also for treatment of multi-drug resistant metastatic solid tumors (Slavin et al 2010) is documentation of the potential safe use of the most effective killer cells for successful treatment and even potential cure of patients considered otherwise incurable. The other special feature of IMAK in comparison with other suggested immunotherapy procedures focusing on the use of isolated NK cells is the easier and less expensive immunotherapy procedure that can be easily applied on a larger scale based on the use of the activation of unseparated lymphocytes collected by apheresis, including a mixture of both fully mismatched T and NK cells pre-activated with IL-2 for a very short period of 4 days. Infusion of IMAK was followed by continuous in vivo activation of donor lymphocytes with low-dose subcutaneous IL-2 injections for no longer than 5 days. As such, both donor’s and patient's own NK cells and T lymphocytes can play a role in induction of anti-cancer cytotoxicity and activation of immune-mediated rejection of MHC-incompatible cancer cells, respectively. Induction of forceful immune reaction against MHC-incompatible target cells represents a most effective and consistent biologic principle that was extrapolated by IMAK for maximizing direct anti-cancer cytotoxicity of the one hand, while ensuring consistent rejection of killer cells after induction of anti-cancer cytotoxicity for prevention of GVHD, on the other. The concept of combined use of both T cells and NK cells for cancer immunotherapy was recently upgraded (Slavin, in preparation) toward developing an even more sophisticated immunotherapy procedure based on the combination of elimination of existing cancer cells followed by induction of long-lasting anti-cancer immunity (Morecki et al. 2006, 2008), aiming for induction of resistance against recurrent disease (Slavin, in preparation).
Although our cumulative experience in a small number of patients in need confirmed the feasibility, safety and efficacy of immunotherapy induced by IMAK, many questions remain to be answered. First, the role of IMAK needs to be confirmed in prospective randomized clinical trials for treatment of different disease categories, both for treatment of patients with overt relapse and later on, provided that safety of IMAK treatment will be confirmed, also for patients with high-risk disease with MRD following successful response to conventional treatment. Fortunately, induction of complete remission can be accomplished by conventional 1st line chemotherapy even in patients with high-risk leukemia or lymphoma at an early stage of the disease, or if indicated following high-dose chemotherapy and supportive autologous SCT, as per the routine procedure in patients with multiple myeloma. Therefore, once the safety and efficacy of IMAK will be confirmed, eradication of MRD potentially resulting in cure could be easily accomplished at an early stage of high-risk disease before relapse occurs, when another opportunity to accomplish MRD against mutated and more resistant malignant cells may be difficult or impossible to be accomplished.
Other important parameters that need to be investigated include whether treatment outcome could be improved by targeting the mismatched killer cells with relevant monoclonal antibodies against cancer-associated cell surface antigens such as CD20, CD19, CD38 to mention just a few. Also, as shown in table 3, using NK-enriched mismatched killer cells for treatment of 3 patients with AML was also successful, suggesting that IL-2-activated NK cells may be the dominant anti-cancer effector cells. Using CD3-depletion or NK-enriched CD56 positive killer cells will certainly minimize any potential risks of GVHD in patients that may remain immunosuppressed by prior anticancer modalities or SCT, since an extended circulation time of IMAK is likely to increase the risk of GVHD. Accordingly, we need to investigate if treatment with T cell-depleted or NK-enriched killer cells is worth the extra-complicated procedure and higher cost as compared with using an unmanipulated mixture of IL-2 activated killer cells.
Another potentially important question is whether the anti-cancer effects inducible by IMAK could be improved by control of patient's negative regulators such as regulatory T cells, checkpoint inhibitors, and myeloid-derived suppressor cells. Also, in the special case of diseases caused by malignant B cells or multiple myeloma, the anti-cancer effects induced by IMAK will have to be compared to the anti-cancer effects induced by CAR-T cells. More important, it would be interesting to investigate if recurrent disease following treatment with CAR-T cells can be controlled by IMAK.
Taken together, the concept of fast and effective killing of malignant cells despite resistance to maximally tolerated doses of chemo-radiotherapy and other available anti-cancer modalities by cell therapy alone, while avoiding much more complicated allogeneic SCT and the risks of GVHD, could possibly represent a new approach for more effective yet much simpler approach for immunotherapy of otherwise incurable hematologic malignancies. Our available proof of principle should provide the basis and justification for future investigations and prospective clinical trials to confirm the safety and challenge the efficacy of cell-mediated immunotherapy based on the use of intentionally mismatched donor lymphocytes.
Remembering that no available immune reaction can compete with the capacity of mismatched lymphocytes to reject any MHC mismatched target cells, including even kilograms of organ allografts in sub-optimally immunosuppressed recipients, our prediction based on multiple studies in pre-clinical animal models and our limited clinical experience suggests that IMAK application against a true stage of MRD could result in complete eradication of all residual resistant malignant cells. Future studies are indicated to prove or disprove our working hypothesis.