Relative Biological Effectiveness (RBE) of [64Cu]Cu and [177Lu]Lu-NOTA-panitumumab F (ab')2 radioimmunotherapeutic agents vs. γ-radiation for decreasing the clonogenic survival in vitro of human pancreatic ductal adenocarcinoma (PDAC) cells.

INTRODUCTION: Our objective was to compare [64Cu]Cu-NOTA-panitumumab F(ab')2 and [177Lu]Lu-NOTA-panitumumab F(ab')2 radioimmunotherapy (RIT) agents for decreasing the clonogenic survival fraction (SF) in vitro of EGFR-positive human pancreatic ductal adenocarcinoma (PDAC) cell lines and estimate the relative biological effectiveness (RBE) vs. γ-radiation (XRT). METHODS: EGFR-positive PDAC cell lines (AsPC-1, PANC-1, MIAPaCa-2, Capan-1) and EGFR-knockout PANC-1 EGFR KO cells were treated in vitro for 18 h with (0-19.65 MBq; 72 nmols/L) of [64Cu]Cu-NOTA-panitumumab F(ab')2 or [177Lu]Lu-NOTA-panitumumab F(ab')2 or XRT (0-8 Gy) followed by clonogenic assay. The SF was determined after culturing single treated cells for 14 d. Cell fractionation studies were performed for cells incubated with 1 MBq (72 nmols/L) of [64Cu]Cu-NOTA-panitumumab F(ab')2 or [177Lu]Lu-NOTA-panitumumab F(ab')2 for 1, 4, or 24 h to estimate the time-integrated activity (Ã) on the cell surface, cytoplasm, nucleus and medium. Radiation absorbed doses in the nucleus were calculated by multiplying à by S-factors calculated by Monte Carlo N Particle (MCNP) modeling using monolayer cell culture geometry. The SF of PDAC cells was plotted vs. dose and fitted to a linear quadratic model to estimate the dose required to decrease the SF to 0.1 (D10). The D10 for RIT agents were compared to XRT to estimate the RBE. DNA double-strand breaks (DSBs) caused by [64Cu]Cu-NOTA-panitumumab F(ab')2 or [177Lu]Lu-NOTA-panitumumab F(ab')2 continuous exposure for 5 h or 20 h were probed by immunofluorescence for γ-H2AX. Relative EGFR expression of PDAC cells was assessed by flow cytometry (scored + to +++) and cell doubling times for untreated cells were determined. RESULTS: The D10 for [64Cu]Cu-NOTA-panitumumab F(ab')2 ranged from 9.1 Gy (PANC-1) to 39.9 Gy (Capan-1). The D10 for [177Lu]Lu-NOTA-panitumumab F(ab')2 ranged from 11.7 Gy (AsPC-1) to 170.8 Gy (Capan-1). The D10 for XRT ranged from 2.5 Gy (Capan-1) to 6.7 Gy (PANC-1 EGFR KO). D10 values were not correlated with EGFR expression over a relatively narrow range (++ to +++) or with cell doubling times. Based on D10 values, PANC-1 EGFR KO cells were 1.6-fold less sensitive than PANC-1 cells to [64Cu]Cu-NOTA-panitumumab F(ab')2 and 1.9-fold less sensitive to [177Lu]Lu-NOTA-panitumumab F(ab')2. The RBE for [64Cu]Cu-NOTA-panitumumab F(ab')2 ranged from 0.06 for Capan-1 cells to 0.45 for PANC-1 cells. The RBE for [177Lu]Lu-NOTA-panitumumab F(ab')2 ranged from 0.015 for Capan-1 cells to 0.28 for AsPC-1 cells. DNA DSBs were detected in PDAC cells exposed to [64Cu]Cu-NOTA-panitumumab F(ab')2 or [177Lu]Lu-NOTA-panitumumab F(ab')2 but were not correlated with the SF of the cells. CONCLUSIONS: We conclude that at the same dose delivered to the cell nucleus [64Cu]Cu-NOTA-panitumumab F(ab')2 and [177Lu]Lu-NOTA-panitumumab F(ab')2 were less radiobiologically effective than XRT for decreasing the SF of human PDAC cells, but [64Cu]Cu-NOTA-panitumumab F(ab')2 was more cytotoxic than [177Lu]Lu-NOTA-panitumumab F(ab')2 except for AsPC-1 cells which were more sensitive to [177Lu]Lu-NOTA-panitumumab F(ab')2. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: This study demonstrates that higher radiation doses may be required for RIT than XRT to achieve radiobiologically equivalent effects when used to treat PDAC.

Floral organ abscission in Arabidopsis requires the combined activities of three TALE homeodomain transcription factors.

Floral organ abscission is a separation process in which sepals, petals, and stamens detach from the plant at abscission zones. Here, we investigated the collective role of three amino-acid-loop-extension (TALE) homeobox genes ARABIDOPSIS THALIANA HOMEOBOX GENE1 (ATH1), KNAT6 (for KNOTTED LIKE from Arabidopsis thaliana) and KNAT2, which form a module that patterns boundaries under the regulation of BLADE-ON-PETIOLE 1 and 2 (BOP1/2) co-activators. These TALE homeodomain transcription factors were shown to maintain boundaries in the flower, functioning as a unit to coordinate the growth, patterning, and activity of abscission zones. Together with BOP1 and BOP2, ATH1 and its partners KNAT6 and KNAT2 collectively contribute to the differentiation of lignified and separation layers of the abscission zone. The genetic interactions of BOP1/2 and ATH1 with INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) were also explored. We showed that BOP1/2 co-activators and ATH1 converge with the IDA signalling pathway to promote KNAT6 and KNAT2 expression in the abscission zone and cell separation. ATH1 acts as a central regulator in floral organ abscission as it controls the expression of other TALE genes in abscission zone cells.