Gene expression profiling upon ²¹²Pb‐TCMC‐trastuzumab treatment in the LS‐174T i.p. xenograft model

Abstract

Recent studies have demonstrated that therapy with 212Pb-TCMC-trastuzumab resulted in (1) induction of apoptosis, (2) G2/M arrest, and (3) blockage of double-strand DNA damage repair in LS-174T i.p. (intraperitoneal) xenografts. To further understand the molecular basis of the cell killing efficacy of 212Pb-TCMC-trastuzumab, gene expression profiling was performed with LS-174T xenografts 24 h after exposure to 212Pb-TCMC-trastuzumab. DNA damage response genes (84) were screened using a quantitative real-time polymerase chain reaction array (qRT-PCR array). Differentially regulated genes were identified following exposure to 212Pb-TCMC-trastuzumab. These included genes involved in apoptosis (ABL, GADD45α, GADD45γ, PCBP4, and p73), cell cycle (ATM, DDIT3, GADD45α, GTSE1, MKK6, PCBP4, and SESN1), and damaged DNA binding (DDB) and repair (ATM and BTG2). The stressful growth arrest conditions provoked by 212Pb-TCMC-trastuzumab were found to induce genes involved in apoptosis and cell cycle arrest in the G2/M phase. The expression of genes involved in DDB and single-strand DNA breaks was also enhanced by 212Pb-TCMC-trastuzumab while no modulation of genes involved in double-strand break repair was apparent. Furthermore, the p73/GADD45 signaling pathway mediated by p38 kinase signaling may be involved in the cellular response, as evidenced by the enhanced expression of genes and proteins of this pathway. These results further support the previously described cell killing mechanism by 212Pb-TCMC-trastuzumab in the same LS-174T i.p. xenograft. Insight into these mechanisms could lead to improved strategies for rational application of radioimmunotherapy using α-particle emitters.The apoptotic response and associated gene modulations have not been clearly defined following exposure of cells to α-particle radioimmunotherapy (RIT). Gene expression profiling was performed with LS-174T i.p. (intraperitoneal) xenografts after exposure to 212Pb-TCMC-trastuzumab. Differentially regulated 22 genes were identified following the stressful growth arrest conditions provoked by 212Pb-TCMC-trastuzumab, providing an informative approach toward understanding the molecular basis of tumor biology in response to α-particle radiation and leading to improved strategies for RIT using α-particle emitters. This study provides data which is among the first to describe in detail, the cellular response to α-particle irradiation in vivo.

Evaluation of cetuximab as a candidate for targeted α-particle radiation therapy of HER1-positive disseminated intraperitoneal disease

“…Targeting LS-174T tumors with 212 Pb-trastuzumab increases DNA double-stranded breaks, impairs DNA damage repair, persistent G2/M arrest and chromatin remodeling. 9,10 HER2 is expressed by an array of epithelial cancers such as pancreas (35-45%), breast (25-30%), ovarian (25-100%) and colorectal (up to 90%). [11][12][13][14] Conversely, one might also state that HER2, for example, is not well expressed in 55-65% of pancreatic cancers, and therefore identification of additional molecular targets is warranted to expand the repertoire of a-targeted RIT.…”

Application of ²¹²Pb for Targeted α-particle Therapy (TAT): Pre-clinical and Mechanistic Understanding through to Clinical Translation

“…Gene expression qRT-PCR array profiling after exposure to 212 Pb-TCMC-trastuzumab was recently reported from Yong et al In this study, 212 Pb-TCMC-trastuzumab treatment differentially regulated genes including genes involved in apoptosis ( ABL, GADD45a, GADD45r, PCBP4, and p73 ), Cell cycle ( ATM, DDIT3, GADD45a, GTSE1, MKK6, PCBP4, and SESN1 ), and damaged DNA binding (DDB) and repair (ATM, and BTG2). Furthermore, the p73/GADD45 signaling pathway mediated by p38 kinase signaling may be involved in the cellular response of tumors exposed to 212 Pb-TCMC-trastuzumab [12]. …”

“…In pre-clinical studies, TAT using 212 Pb has shown significant therapeutic efficacy in both in vitro and in vivo model systems [10–12]. A mechanistic study of 212 Pb-labeled TAT demonstrated significant impact of catastrophic dsDNA destruction as a result of high-LET 212 Pb traversal of the nucleus, with interference with DNA damage repair and perturbation of the cell cycle, leading to significant tumor cell killing [11].…”

“…Therefore, a single targeted therapeutic strategy that delivers α-particles from a source such as 212 Pb would be an innovative approach to the elusive goal of creating a magic-bullet therapy to target cancer. Recent progress in pre-clinical and mechanistic studies in in vivo tumor models has provided critical information (such as induction of apoptosis, G2/M arrest, blockage of double-strand DNA damage repair, modulation of genes) for clinical transition of 212 Pb-labeled TAT [10–12]. With the advent of the first clinical trial that employed 212 Pb as the therapeutic agent (targeted by trastuzumab), this radionuclide has finally reached a landmark position in the family of α-emitters validated as suitable for such applications [13].…”

Comparative studies on the therapeutic benefit of targeted α-particle radiation therapy for the treatment of disseminated intraperitoneal disease

“…9 Targeted α-radiation therapy has been shown to effect cell death through production of double-stranded DNA breaks, DNA crosslinking, chromosomal rearrangements, the induction of apoptosis, perturbation of the cell cycle, and blockage or downregulation of the DNA damage repair mechanism specific to double-stranded DNA breaks. 10–14 …”