Immunocryosurgery for facial, non-superficial basal cell carcinoma triggers consistent but transient cell counts alterations in the circulating innate immune cell lines

Immunocryosurgery, a fixed-time combination of imiquimod and cryosurgery,¹ is a highly effective modality for basal cell carcinomas (BCCs),² including non-superficial tumors amenable to excision.³ Previous studies, both in tissue and peripheral blood, have focused on the role of T-regulatory lymphocytes as antitumor effectors during immunocryosurgery for BCC.^{4, 5} In the present study we addressed the influence of an immunocryosurgery treatment cycle for BCC on the peripheral blood cell counts.

Institutional Review Committee approval was granted (Θ59 45/17-11-2022). Immunocryosurgery entails the application of 5% imiquimod cream once daily for 5 weeks and a single cryosurgery session on Day 14 of the treatment cycle (liquid N₂, open spray, two freeze-thaw cycles of 20 s freezing time each).¹ As previously,⁵ peripheral blood cell counts were determined at four time points: within 4 days before the initiation of treatment (“Week 0,” ‘baseline’), on the day of cryosurgery (“Week 2”), on the last day (plus a maximum of 2 days) of treatment (“Week 5”), and at 4 weeks follow up (±2 days: “Week 9”). The distributions of continuous variables are represented as medians and means±standard deviations (SD). Friedman tests (four measurements/patient) with post-hoc Dunn's tests were calculated using SPSS, with p < 0.05 indicating statistical significance.

Thirteen immunocompetent patients were included (age: 67–81 years; 11 men), each with a facial, biopsy confirmed non-superficial BCC (median BCC diameter: 15 mm, range: 7–19 mm). All tumors responded with vivid local inflammatory alterations, cleared completely after one treatment cycle and remained recurrence-free for at least 12 months. A discrete pattern of temporal variation was encountered in the counts of all leukocyte cell lines, except for monocytes, during imiquimod treatment (Table 1). The whole leukocytes and the three granulocyte subpopulations (neutrophils, eosinophils, and basophils) cell counts reduced significantly compared to baseline during imiquimod application (p < 0.0001, Friedman test) and recovered within 1 month thereafter. Also, for lymphocytes, nadir counts were observed at the end of imiquimod treatment; however, the perturbations of the lymphocyte numbers did not present a similar deviation degree as the cell lines of the innate immunity (p = 0.0153, Friedman test). Nonetheless, in no case did the cell numbers at nadir drop below the corresponding normal cell counts limits (Figure 1) or did we observe any clinical signs of immunosuppression.

Our results document a systemic, to date underrecognized immunocryosurgery effect on the peripheral blood cell counts. Thereby the role of imiquimod is central as the counts drop was evident before cryosurgery. Our findings add to the sparse data regarding the hematological impact of the topical imiquimod. Measurable blood levels of imiquimod have already been detected with 3 times weekly topical application,⁶ yet, with overall minor, though consistent hematological imprint.⁷ In line with comparable hematological findings of oral imiquimod administration,⁸ we suggest that the oscillation of the leukocyte counts in the context of immunocryosurgery might be primarily due to increased imiquimod absorption through daily drug application to inflamed and eroded skin. Taken together, our present findings advise a complete blood count before immunocryosurgery, and test repetition for patients with pathologic baseline findings 1 month after treatment. Possible drawbacks of this exploratory study are the small number of patients and the lack of control groups (healthy individuals and patients treated only with cryosurgery or imiquimod).

Recently, granulocyte cell lines, especially neutrophils and eosinophils, have emerged as potential key targets of tumor immunotherapy due to their complex, dual, pro- and contra-neoplastic effects.^{9, 10} Therefore, the currently documented hematological alterations could be further investigated as a potential component of the antineoplastic mechanism underlying the efficacy of immunocryosurgery against BCC, by including data from treatment failures, as well as from immunocryosurgery use for indications other than BCC.

G. Gaitanis study conception and design, acquisition and interpretation of data. A. Ganiatsa acquisition and analysis of data. G. Vartholomatos study design, acquisition and analysis of data. K. Seretis data analysis and interpretation. I. D. Bassukas conception and design of the study, analysis and interpretation of the data, wrote the first drafting of the article. All authors revised the manuscript for important intellectual content and approved the final version to be published.

The authors declare no conflict of interest.

All patients in this manuscript have given written informed consent for participation in the study and the use of their deidentified, anonymized, aggregated data and their case details for publication. Institutional Review Committee approval was granted (Θ59 45/17-11-2022).