Background Cancer vaccines aim to generate antigen-specific CD8⁺ T cell responses but efficacy has been hampered by inefficient targeting of antigens to antigen presenting cells (APCs) for presentation on MHC-I. Cell Squeeze^® technology delivers target antigens directly into red blood cells (RBCs) using temporary cell membrane disruption via a microfluidic chip. During the Cell Squeeze^® process, phosphatidylserine is flipped to the extracellular membrane, making the RBCs appear aged. TLR3 agonist, poly I:C, is also delivered into RBCs, which serves to mature target APCs and ensure that presentation of tumor antigens occurs in an immunostimulatory context. The resultant SQZ^® activating antigen carriers (AACs) thus leverage the natural clearance mechanism of aged RBCs, via phagocytosis by professional APCs, to enhance presentation of antigens on MHC-I of endogenous APCs. SQZ-AAC-HPV is an autologous RBC cell cancer vaccine targeting HPV16 viral oncoproteins E6 and E7 using the Cell Squeeze^® technology.

Methods SQZ-AAC-HPV-101 (NCT04892043) enrolled HLA-A*02⁺patients with advanced HPV16+ cancers who had progressed after standard therapy, have an ECOG of 0–1, proper organ function, and measurable lesion(s). After whole blood collection, the cell product is manufactured in less than 24 hours with a collection-to-release time of ~1 week. SQZ-AAC-HPV was given IV q3w without a conditioning regimen. The response was assessed via RECIST 1.1 and biopsies were required at baseline and day 28 for pharmacodynamic biomarker analysis.

Results Five patients [head and neck (4), anorectal (1)] were dosed in 2 monotherapy cohorts (from 0.5 to 5.0 x10e8/kg [DP]). There were no DLTs, no Grade (G) >2 related SAEs and no related G >3 AEs. One patient in the high dose cohort died due to disease progression in cycle one. Four of the five patients remain on SQZ-AAC-HPV treatment. Three patients in the low dose cohort have undergone at least one on-treatment scan; two patients showed a best overall response of stable disease with the third patient demonstrating a confirmed complete response of their anorectal cancer. Notably all three of these patients were immune checkpoint inhibitor naïve. Paired biopsy analysis for the patient with a confirmed complete response revealed an increase in CD8 density by IHC and a post-treatment tumor microenvironment with a high density of CD8 T cells and a low density of regulatory T cells per MIBI analysis.

Conclusions SQZ-AAC-HPV treatment was generally well-tolerated and initial efficacy data are encouraging. This presentation may include safety, additional efficacy, and biomarker data from a cut-off proximal to the presentation.

Acknowledgements We thank the patients, their families, and their caregivers for their time and their willingness to participate in this study. We also thank the investigators and the investigational site staff for their contributions to the study.

Ethics Approval The study was performed in accordance with ethical principles that originated in the Declaration of Helsinki consistent with the ICH/GCP and applicable regulatory requirements. The protocol was approved by IRBs/IECs at each center. Patients provided written informed consent to participate.