<h3>Purpose</h3> Chemoradiation (CRT) may modulate the intratumoral and peripheral immune milieu by serving as an <i>in situ</i> vaccine. The impact of the rapid dose delivery of brachytherapy (BT) on the T-cell repertoire has not been examined. HPV-related cancers express viral oncoproteins E6/E7, which could function as tumor antigens (Ag) and are thus ideal for investigating the impact of CRT in generating tumor-specific immunity. <h3>Methods</h3> 11 pts with cervical cancer were enrolled on a multi-institutional prospective tissue banking protocol from 1/2020—2022. Pts underwent standard external beam RT (EBRT) spanning 5-7 wks with concurrent weekly 40mg/m<sup>2</sup> cisplatin, then BT boost with either high dose rate (HDR) in 5 fx or pulse dose rate (PDR) in 2 fx. Cervical tumor swabs were taken at 5 timepoints: <b>baseline</b> (<b>T1</b>), <b>wk 1</b> (<b>T2</b>, after 5 fx), <b>wk 3</b> (<b>T3</b>, after 10-15 fx), <b>wk 5</b> (<b>T4</b>, at conclusion of EBRT and prior to 1st BT), prior to 2nd BT (<b>T4B</b>), and at <b>1st follow-up</b> (<b>T5</b>, 12 wks post-treatment). Samples underwent functional expansion of HPV Ag-specific T cells before DNA extraction and multiplex deep sequencing of the CDR3 region of TCR-β using immunoSEQ PCR (Adaptive Biotech). TCR productive frequency (freq), templates, clonality, and rearrangements were compared across timepoints. Separately, HPV-responsive T-cell clones underwent <i>ex vivo</i> expansion by incubation with HPV long peptide sequences of E6/E7 overnight (ON). TCR sequences were compared between HPV-stimulated T cells and evaluated against incubations with CEF (positive control peptides) or media (negative control). Statistical analysis was via Mann-Whitney U. <h3>Results</h3> Median age was 47 yrs (range 27-57). 6 pts (54.5%) had SCC; 5 (45.5%) had adenocarcinoma. 4 pts (36.4%) had FIGO 2018 ≤Stage IIB disease, 45.5% were Stage III, and 18.2% were Stage IVA/B. 7 pts (63.6%) received PDR BT and 4 pts (36.4%) underwent HDR BT. At median follow-up of 7.5 mo, all pts are alive and with NED. In serially sampled peritumoral microenvironment (TME) isolates, TCR productive clonality increased after first BT, culminating in a significant increase at T5 during the first post-treatment follow-up (<b>Fig 1A</b>). There was a trend (ns) toward increased productive templates throughout CRT and BT (<b>Fig 1B</b>), but this decreased by T5. Productive freq was significantly higher at T5 compared to prior to BT (T1—T4), <b>Fig 1C</b>. Productive rearrangements trended toward abundance throughout CRT and BT (ns), but decreased at T5 (<b>Fig 1D</b>). Functional HPV Ag stimulation assays of peritumoral T-cells in the full cohort did not show significant changes in the TCR repertoire after ON incubation with HPV E6/7 peptides. However, 4 pts had notable E6/E7-responsive increases in these metrics at either T4 or T4B—namely, abundance of productive templates (ranges: 1.9—30.3 fold-increase from baseline), freq (1.25—5.5), rearrangements (1.5—13.8), and clonality (1.31—3.8). <h3>Conclusions</h3> We characterized TCR remodeling during BT in this first translational analysis, demonstrating (i) increased peritumoral TCR productive freq and clonality post-BT at T5, and (ii) transient rise in productive templates and rearrangements through CRT and BT, with a numeric return to baseline post-BT at T5. Some pts had functional Ag-specific activation upon HPV stimulation during and after BT. These findings suggest that despite the cytotoxic effects of radiation, BT alters the TME to facilitate expansion of specific T-cell populations, which may contribute to treatment efficacy.
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Oct 25, 2024
Marjolein Louise Elisabeth Heidotting + 3
Open Access