Abstract PR005: Intraductal administration of a recombinant transferrin receptor-directed immunotoxin clears ductal carcinoma in situ in preclinical mammary in-duct (MIND) models of breast cancer
Authors: Sukumar Saraswati 
Published: December 2022

Abstract
Background: For DCIS, a preneoplastic lesion with a 15-50% chance of recurrence or progression, there is a reappraisal of the necessity of aggressive treatment with its attendant toxicities. HB21(Fv)-PE40, a conjugate of an Fv fragment of the human transferrin-receptor (TFR) mAb and a 40 kDa fragment of bacterial Pseudomonas exotoxin (PE), has potent tumor cell killing activity. HB21(Fv)-PE40 binds to abundantly expressed TFR on cancer cells; the molecule is internalized, processed, and trafficked to the cytosol. Inside the cell, PE-toxin catalyzes the transfer of ADP ribose from nicotinamide adenine dinucleotide (NAD) to elongation factor 2 (EF2), inactivates EF2, and leads to an arrest in protein translation and cell death by apoptosis. However, due to its abundant expression in the liver, the recombinant immunotoxin (RIT) is potentially toxic. We hypothesized that RIT administration via the mammary ductal opening at the teat will destroy preneoplasias, both visible and occult, in the entire ductal tree, while systemic effects will be minimal. The intraductal approach could provide a direct, safe, and highly effective treatment for DCIS. Methods: We tested this concept in two models of DCIS using an ER/PR-, HER2+ cell line derived from a DCIS, SUM225-Luc, and in the ER/PR+, HER2- breast cancer cell line, MCF-7-Luc. In both cell lines, the IC50 of the toxin was in the nM range. DCIS developed within one week in the MIND models following i.duc injection of tumor cells in the 4th pair of mammary glands of female NSG mice. On day 7, the tumor-bearing mice received vehicle, TFR mAb alone, or the HB21(Fv)-PE40 conjugate by the i.duc or i.v route three times at weekly intervals and were observed for 1-6 months. Within two weeks of the treatment, IVIS imaging signals were undetectable in the mammary gland treated with RIT i.duc in both the models (P&lt;0.0001). Mammary gland whole mount analysis, histopathology, and immunohistochemistry for Ki67, Ku80, TFR and CD31 showed that i.duc toxin rendered more than 90% of the mice tumor-free. Pharmacokinetics studies were conducted in mice with the RIT administered i.v. or i.duc. RIT was undetectable by TFR-ELISA in the blood of i.duc-treated mice. Conclusions. Overall, we have shown that HB21(Fv)-PE40 has remarkable antitumor activity by i.duc administration in two MIND models of DCIS. RIT was undetectable in blood after i.duc injection by ELISA, suggesting that the toxin remains inside the ductal system during its 30 min half-life. This preclinical study provides a solid foundation for a clinical study. Dose escalation studies of HB21(Fv)-PE40 by the i.duc route could be performed in patients with DCIS and early breast cancer who will undergo mastectomy as part of their treatment regimen. This study design will allow a detailed examination of the local (breast) and systemic effects of i.duc HB21(Fv)-PE40. It will introduce a potent agent for DCIS treatment and cancer interception in women at high risk of developing breast cancer. The contribution of the immune system remains to be explored.
Citation Format: Saraswati Sukumar. Intraductal administration of a recombinant transferrin receptor-directed immunotoxin clears ductal carcinoma in situ in preclinical mammary in-duct (MIND) models of breast cancer [abstract]. In: Proceedings of the AACR Special Conference on Rethinking DCIS: An Opportunity for Prevention?; 2022 Sep 8-11; Philadelphia, PA. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_1): Abstract nr PR005.