Predicting DNA-Binding Specificities of Eukaryotic Transcription Factors

Today, annotated amino acid sequences of more and more transcription factors (TFs) are readily available. Quantitative information about their DNA-binding specificities, however, are hard to obtain. Position frequency matrices (PFMs), the most widely used models to represent binding specificities, are experimentally characterized only for a small fraction of all TFs. Even for some of the most intensively studied eukaryotic organisms (i.e., human, rat and mouse), roughly one-sixth of all proteins with annotated DNA-binding domain have been characterized experimentally. Here, we present a new method based on support vector regression for predicting quantitative DNA-binding specificities of TFs in different eukaryotic species. This approach estimates a quantitative measure for the PFM similarity of two proteins, based on various features derived from their protein sequences. The method is trained and tested on a dataset containing 1 239 TFs with known DNA-binding specificity, and used to predict specific DNA target motifs for 645 TFs with high accuracy

Targeting claudin‐overexpressing thyroid and lung cancer by modified Clostridium perfringens enterotoxin

Clostridium perfringens enterotoxin (CPE) can be used to eliminate carcinoma cells that overexpress on their cell surface CPE receptors - a subset of claudins (e.g., Cldn3 and Cldn4). However, CPE cannot target tumors expressing solely CPE-insensitive claudins (such as Cldn1 and Cldn5). To overcome this limitation, structure-guided modifications were used to generate CPE variants that can strongly bind to Cldn1, Cldn2 and/or Cldn5, while maintaining the ability to bind Cldn3 and Cldn4. This enabled (a) targeting of the most frequent endocrine malignancy, namely, Cldn1-overexpressing thyroid cancer, and (b) improved targeting of the most common cancer type worldwide, non-small-cell lung cancer (NSCLC), which is characterized by high expression of several claudins, including Cldn1 and Cldn5. Different CPE variants, including the novel mutant CPE-Mut3 (S231R/S313H), were applied on thyroid cancer (K1 cells) and NSCLC (PC-9 cells) models. In vitro, CPE-Mut3, but not CPEwt, showed Cldn1-dependent binding and cytotoxicity toward K1 cells. For PC-9 cells, CPE-Mut3 improved claudin-dependent cytotoxic targeting, when compared to CPEwt. In vivo, intratumoral injection of CPE-Mut3 in xenograft models bearing K1 or PC-9 tumors induced necrosis and reduced the growth of both tumor types. Thus, directed modification of CPE enables eradication of tumor entities that cannot be targeted by CPEwt, for instance, Cldn1-overexpressing thyroid cancer by using the novel CPE-Mut3