MUDr. Martin Lužbeták, M.Sc.

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Could the analysis of tumor cells from the blood predict the therapeutic outcome and challenge previous treatment methods?

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Significance of tumor cells circulating in the blood

The development of targeted cancer therapies is being pursued in many studies, and research on tyrosine kinase inhibitors as well as immune checkpoint inhibitors is becoming increasingly important. However, cytotoxic chemotherapeutic agents remain the gold standard in neo-adjuvant or adjuvant treatment of tumors at various stages. The choice for mono- or combination therapy is often guideline-based without any molecular or functional indication, reflecting the limitations of such non-personalized therapies. For the treatment of metastatic breast cancer, first-line therapy is taxol, which only 30% of patients respond to. Another 30% achieve disease stagnation, and 40% do not respond at all but still go through severe side effects and toxicity of this therapy. The failure of chemotherapy is due to tumor resistance, which is random, unpredictable, and only apparent during clinical evaluation for treatment response. If the resistance and sensitivity profile of a tumor could be detected before therapy application, significantly higher response rates and subsequently higher cure rates would be the result. In previous trials of a chemosensitivity assay from isolated tumor tissue, no clinical correlation could be found with the results. Other attempts at chemo- and biologic-sensitivity testing from circulating tumor cells in the laboratory failed due to low levels of extracted tumor cells.

The authors of the study “Clinical utility of circulating tumor-associated cells to predict and monitor chemo-response in solid tumors” (*) described a method that allows the detection, extraction, and recovery of useful circulating tumor cells in the lab from peripheral blood of patients with various solid organ tumors. Using this method, tumor cells were obtained from the blood of 5090 patients with a previous diagnosis of 17 different solid organ tumors. From this collective, tumor cells were isolated from a previous tumor tissue biopsy in 230 patients. Chemosensitivity of circulating cancer cells was tested against various cytostatic agents, and the results were tested for concordance with those of chemosensitivity of cells from tumor tissue. Both primary and secondary resistance from untreated, as well as pretreated patient cases, were identified.

Study Design: Patient Description

The study results were obtained from one observational study and three intervention studies, all of which compared personalized therapy using molecular chemosensitivity analysis from tumor cells in the blood or directly from tumor tissue and conventional therapeutic approaches using cytotoxic chemotherapeutic agents. The 5090 patients were retrospectively divided into three study populations. Population 1 consists of 230 patients who first received a blood draw and then a tumor biopsy. Some of these patients were already pretreated. The second population consists of 2201 patients who received a blood draw and were already treated with a cytotoxic chemotherapeutic agent. Population 3 consists of 2734 patients who received a blood draw and whose tumor was still untreated. There is a partial overlap between the populations. From the blood draws, all mononuclear cells were epigenetically treated, whereupon cell death was induced in healthy cells with intact apoptosis signaling cascade. The tumor-associated cells thus remained. The recovered tumor cells could now be immunohistochemically characterized and assigned to the different organs and carcinomas.

The chemosensitivity assay was performed on microtitration plates containing a specific number of cells and one chemotherapeutic agent per well. Sensitivity is tested by light transmission, which correlates with apoptosis rate. Thus, the highest and lowest sensitivity could be tested for each cell line of tumor cells. In population 1, the chemosensitivity of different chemotherapeutic agents was first tested on cells from tumor biopsy and then compared with the results of circulating cancer cells. In populations 2 and 3, only those chemotherapeutic agents were tested which are used according to the guidelines for the respective cancer types. A chemotherapeutic agent was considered sensitive if cell death was detected in more than 50% of cells 12 hours after initial exposure.

Results of the different patient groups

In population 1, the results of resistance and sensitivity from tumor cells circulating in the blood and cells biopsied directly from tumor tissue were compared. The results were in 93.7% concordance.

In population 3, resistance to cytotoxic chemotherapeutic agents was detected in 58.9%, indicating innate, intrinsic resistance of tumor cells. Of 2734 patients, radiologic follow-up at six months was undertaken in 77 subjects. In 33 patients with high sensitivity to the chemotherapeutic agents tested, PET-CT detected a complete or partial response to therapy in 32 (97%) patients. Thus, the in vitro/in vivo concordance is 97%. In those 44 patients in whom no sensitivity was detected chemically, the radiological examination also showed no improvement in 18 cases (41%).

In population 2, resistance to cytotoxic chemotherapeutic agents was found in 77.8 % of patients. It follows that many tumor cells develop acquired resistance to chemotherapeutic agents already in use. In this population, 143 patients received radiological follow-up. In all of them, the examination showed worsening cancer and disease state. Laboratory analysis showed chemical matches of resistance to the drugs used in 124 patients. The in vitro/in vivo agreement here is 86.7%.

Conclusion: Promising therapeutic approach for tumor patients
Although detection of resistance is of great importance for optimized cancer treatment, no technology has yet been established for the identification of innate and prospective resistance. Current methods have long turnaround times, show low clinical agreement, and are highly invasive, requiring a large amount of biopsied tissue. The high invasiveness and rapid tumor progression make these methods obsolete.

The present study demonstrates a method to rapidly and non-invasively establish a chemosensitivity profile. In addition, not only tumor cells are detected, but also tumor-associated cells such as macrophages and fibroblasts. These contribute significantly to tumor progression, for example, by suppressing anti-tumor immunity. The epigenetic modification allows tumor cells to be rapidly harvested and, subsequently chemotherapeutic agents to be tested directly.

In addition, the study confirms that tumor cells circulating in the blood adequately reflect the tumor present. Significant correlations between radiology therapy response rates and chemosensitivity profiles were also found. Therefore, the chemosensitivity tests can accurately predict the success or failure of chemotherapy and serve as a long-term monitoring tool for tumor patients.

In addition, the study authors found high variability between individual patients with different cancer types. Therefore, chemosensitivity profiling prior to therapy initiation is advisable to avoid treatment failures, the rapid spread of cancer, and the unnecessary administration of toxic agents with significant side effects. Knowledge about the in vitro sensitivity profile enables rapid identification of resistance and could provide significantly better and personalized therapeutic approaches. The introduced method is non-invasive and cost-saving. Moreover, it conveys synchronous real-time information at diagnosis as well as continuously during therapy. The study offers a therapeutic approach that can be used immediately and may expand the range of approved chemotherapeutic agents outside their current field of use.