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Purpose: Combination of radiation with radiosensitizing chemotherapeutic agents improves outcomes for locally advanced rectal cancer. Current treatment includes 5-fluorouracil–based chemoradiation prior to surgical resection; however pathologic complete response varies from 15% to 20%, prompting the need to identify new radiosensitizers. Exportin 1 (XPO1, also known as chromosome region 1, CRM1) mediates the nuclear export of critical proteins required for rectal cancer proliferation and treatment resistance. We hypothesize that inhibition of XPO1 may radiosensitize cancer cells by altering the function of these critical proteins resulting in decreased radiation resistance and enhanced antitumoral effects.Experimental Design: To test our hypothesis, we used the selective XPO1 inhibitor, selinexor, to inhibit nuclear export in combination with radiation fractions similar to that given in clinical practice for rectal cancer: hypofractionated short-course radiation dosage of 5 Gy per fraction or the conventional long-course radiation dosage of 1 Gy fractions. Single and combination treatments were tested in colorectal cancer cell lines and xenograft tumor models.Results: Combination treatment of radiotherapy and selinexor resulted in an increase of apoptosis and decrease of proliferation compared with single treatment, which correlated with reduced tumor size. We found that the combination promoted nuclear survivin accumulation and subsequent depletion, resulting in increased apoptosis and enhanced radiation antitumoral effects.Conclusions: Our findings suggest a novel therapeutic option for improving radiation sensitivity in the setting of rectal cancer and provide the scientific rationale to evaluate this combination strategy for clinical trials. Clin Cancer Res; 22(7); 1663–73. ©2015 AACR.

Supplementary Figure 1. Selinexor induces HT29 and LoVo cell apoptosis; Supplementary Figure 2. Combination of radiation and Selinexor reduces tumor cell radiation survival; Supplementary Figure 3. Body weight of female nude mice; Supplementary Figure 4. mRNA Survivin levels in in vivo models of CRC.

Delivery of small interfering RNA (siRNA) as a therapeutic tool is limited due to critical obstacles such as the cellular barrier, the negative charges of the siRNA molecule, and its instability in serum. Several siRNA delivery systems have been constructed using cell-penetrating peptides (CPPs) since the CPPs have shown a high potential for oligonucleotide delivery into the cells, especially by forming nanoparticles. In this study, we have developed a new family of short (15mer or 16mer) tryptophan-(W) and arginine-(R) rich Amphipathic Peptides (WRAP) able to form stable nanoparticles and to enroll siRNA molecules into cells. The lead peptides, WRAP1 and WRAP5, form defined nanoparticles smaller than 100 nm as characterized by biophysical methods. Furthermore, they have several benefits as oligonucleotide delivery tools such as the rapid encapsulation of the siRNA, the efficient siRNA delivery in several cell types, and the high gene silencing activity, even in the presence of serum. In conclusion, we have designed a new family of CPPs specifically dedicated for siRNA delivery through nanoparticle formation. Our results indicate that the WRAP family has significant potential for the safe, efficient, and rapid delivery of siRNA for diverse applications.

Purpose: Combination of radiation with radiosensitizing chemotherapeutic agents improves outcomes for locally advanced rectal cancer. Current treatment includes 5-fluorouracil–based chemoradiation prior to surgical resection; however pathologic complete response varies from 15% to 20%, prompting the need to identify new radiosensitizers. Exportin 1 (XPO1, also known as chromosome region 1, CRM1) mediates the nuclear export of critical proteins required for rectal cancer proliferation and treatment resistance. We hypothesize that inhibition of XPO1 may radiosensitize cancer cells by altering the function of these critical proteins resulting in decreased radiation resistance and enhanced antitumoral effects. Experimental Design: To test our hypothesis, we used the selective XPO1 inhibitor, selinexor, to inhibit nuclear export in combination with radiation fractions similar to that given in clinical practice for rectal cancer: hypofractionated short-course radiation dosage of 5 Gy per fraction or the conventional long-course radiation dosage of 1 Gy fractions. Single and combination treatments were tested in colorectal cancer cell lines and xenograft tumor models. Results: Combination treatment of radiotherapy and selinexor resulted in an increase of apoptosis and decrease of proliferation compared with single treatment, which correlated with reduced tumor size. We found that the combination promoted nuclear survivin accumulation and subsequent depletion, resulting in increased apoptosis and enhanced radiation antitumoral effects. Conclusions: Our findings suggest a novel therapeutic option for improving radiation sensitivity in the setting of rectal cancer and provide the scientific rationale to evaluate this combination strategy for clinical trials. Clin Cancer Res; 22(7); 1663–73. ©2015 AACR.

A study investigating the mechanisms underlying lateral inhibition and lineage plasticity in the mouse small intestine crypts in vivo finds that crypt cells maintain a permissive chromatin state upon which a transcription factor acts to determine lineage specification, and this is the basis of lateral inhibition. Intestinal crypts are the focus of intensive study, stimulated in part by the recent discovery of distinct stem-cell populations and markers. Ramesh Shivdasani and colleagues have studied mechanisms underlying lateral inhibition and lineage plasticity in mouse small intestine crypts in vivo. They find that crypt cells maintain a permissive chromatin state upon which a transcription factor acts to determine lineage specification, and that this is the basis of lateral inhibition. Cells differentiate when transcription factors bind accessible cis-regulatory elements to establish specific gene expression programs. In differentiating embryonic stem cells, chromatin at lineage-restricted genes becomes sequentially accessible1,2,3,4, probably by means of ‘pioneer’ transcription factor activity5, but tissues may use other strategies in vivo. Lateral inhibition is a pervasive process in which one cell forces a different identity on its neighbours6, and it is unclear how chromatin in equipotent progenitors undergoing lateral inhibition quickly enables distinct, transiently reversible cell fates. Here we report the chromatin and transcriptional underpinnings of differentiation in mouse small intestine crypts, where notch signalling mediates lateral inhibition to assign progenitor cells into absorptive or secretory lineages7,8,9. Transcript profiles in isolated LGR5+ intestinal stem cells10 and secretory and absorptive progenitors indicated that each cell population was distinct and the progenitors specified. Nevertheless, secretory and absorptive progenitors showed comparable levels of H3K4me2 and H3K27ac histone marks and DNase I hypersensitivity—signifying accessible, permissive chromatin—at most of the same cis-elements. Enhancers acting uniquely in progenitors were well demarcated in LGR5+ intestinal stem cells, revealing early priming of chromatin for divergent transcriptional programs, and retained active marks well after lineages were specified. On this chromatin background, ATOH1, a secretory-specific transcription factor, controls lateral inhibition through delta-like notch ligand genes and also drives the expression of numerous secretory lineage genes. Depletion of ATOH1 from specified secretory cells converted them into functional enterocytes, indicating prolonged responsiveness of marked enhancers to the presence or absence of a key transcription factor. Thus, lateral inhibition and intestinal crypt lineage plasticity involve interaction of a lineage-restricted transcription factor with broadly permissive chromatin established in multipotent stem cells.