Circulating Adenoid Cystic Carcinoma associated MYB transcripts enable rapid and sensitive detection of metastatic disease in blood liquid biopsies

Abstract

Adenoid cystic carcinoma (ACC) is a rare and lethal malignancy that originates in secretory glands of the head and neck. A prominent molecular feature of ACC is the overexpression of the proto-oncogene MYB. ACC has a poor long-term survival due to its high propensity for recurrence and protracted metastasis. Currently, clinical technologies lack the efficiency to distinguish patient prognosis prior to its redevelopment. We hypothesize that metastatic ACC can be detected by monitoring tumor-specific MYB expression in patients' blood. We developed a quantitative polymerase chain reaction (qPCR) assay for MYB transcripts and screened blood samples from four patient cohorts: no history or evidence of ACC (n=23), past history of ACC and no evidence of disease (NED) for greater than three years (n=15), local ACC (n=6), and metastatic ACC (n=5). Our assay detected significantly elevated levels of MYB transcripts in the metastatic ACC cohort (p < 0.01). Receiver operating characteristic (ROC) curves comparing metastatic to NED and metastatic to local disease were significant, with p values < 0.0001 and 0.0008, respectively. Single-cell RNA sequencing (scRNA-seq) of blood from metastatic ACC identified a cluster of circulating tumor cells (CTCs) expressing MYB. Here, we report a sensitive, cost-effective, and minimally invasive diagnostic test that leverages tumor-specific signatures to screen for metastatic ACC disease, potentially enhancing detection earlier than the current clinical standard.

Keywords: Adenoid cystic carcinoma; Diagnostic biomarkers; Liquid Biopsy; MYB; rare cancer.

Figure 1:. Detection of MYB in whole blood for patients with metastatic ACC.

(A) Immunofluorescent imaging of MYB expression in primary and metastatic tumor samples. (B) Schematic of MYB translocation events that occur with NFIB, RAD51B, and TGFBR3. (C) Violin plot of relative expression of MYB primer pair spanning exons 2–3 in control, NED, local, and metastatic patients. **** = p-value < 0.0001. (D) ROC curve of NED > 3 years verse metastatic results of whole blood MYB relative expression test. (E) ROC curve of local disease verse metastatic results of whole blood MYB relative expression test.

Figure 2:. Circulating tumor cell identification in PBMCs of metastatic ACC patient.

(A) Tracking patient T6 MYB relative expression over time in whole blood (WB), PMBC, and plasma samples. (B) UMAP of 15 clusters from sample T6 PBMC single-cell sequencing experiment with cell type identifications. (C) Hematoxylin and eosin (H&E) image of primary ACC tumor used for spatial transcriptomics with circled tumor foci indicated. (D) Probability prediction of single-cell cluster of interest cell type overlaid on spatial transcriptomic primary ACC tumor sample. (E) Violin plot of MYB expression in PBMC single-cell sequencing.