Six drugs facing key patent expirations and potential generic entry from May to June 2022.

A challenge in anticipating generic entry is elucidating which patents and regulatory protections constrain generic entry. Presented here is a set of estimated loss of exclusivity dates for six drugs, from May through June 2022. These estimated drug patent expiration dates and generic entry opportunity dates are calculated from analysis of known patents and US regulatory protections covering drugs [1]. This methodology can be extended to ex-US jurisdictions by leveraging these estimates and tracking patent family members in other patent offices.

Two drugs facing key patent expirations and potential generic entry from July to August 2020.

A challenge in anticipating generic entry is elucidating which patents and regulatory protections constrain generic entry.

Five drugs facing key patent expirations and potential generic entry from April to May 2020.

A challenge in anticipating generic entry is elucidating which patents and regulatory protections constrain generic entry.

Where Are Drugs Invented, and Why Does It Matter?

Globalization has disrupted many industries, initially shifting unskilled labor employment from high wage-cost industrialized nations to lower wage-cost emerging economies. There has been a trend toward moving increasingly skilled work away from industrialized nations, which raises the question-has innovative research and development been affected by globalization, and why is relocation of innovation much more than a simple economic concern?

The ferric uptake regulator (Fur) protein from Bradyrhizobium japonicum is an iron-responsive transcriptional repressor in vitro.

The Fur protein represses transcription of iron-responsive genes in bacteria. The discovery that Fur is a zinc metalloprotein and the use of surrogate metals for Fe(2+) for in vitro studies question whether Fur is a direct iron sensor. In the present study, we show that the affinity of Fur from Bradyrhizobium japonicum (BjFur) for its target DNA increases 30-fold in the presence of metal, with a K(d) value of about 2 nM. DNase I footprinting experiments showed that BjFur protected its binding site within the irr gene promoter in the presence of Fe(2+) but not in the absence of metal, showing that DNA binding is Fe(2+)-dependent. BjFur did not inhibit in vitro transcription from the irr promoter using purified components in the absence of metal, but BjFur repressed transcription in the presence of Fe(2+). Thus, BjFur is an iron-responsive transcriptional repressor in vitro. A regulatory Fe(2+)-binding site (site 1) and a structural Zn(2+)-binding site (site 2) inferred from the recent crystal structure of Fur from Pseudomonas aeruginosa are composed of amino acids highly conserved in many Fur proteins, including BjFur. BjFur mutants containing substitutions in site 1 (BjFurS1) or site 2 (BjFurS2) bound DNA with high affinity and repressed transcription in vitro in an Fe(2+)-dependent manner. Interestingly, only a single dimer of BjFurS2 occupied the irr promoter, whereas the wild type and BjFurS1 displayed one- or two-dimer occupancy. We suggest that the putative functions for metal-binding sites deduced from the structure of P. aeruginosa Fur cannot be extrapolated to bacterial Fur proteins as a whole.

A novel DNA-binding site for the ferric uptake regulator (Fur) protein from Bradyrhizobium japonicum.

The Fur protein is a global regulator of iron metabolism and other processes in many bacterial species. A key feature of the model of Fur function is the recognition of a DNA element within target promoters with similarity to a 19-bp AT-rich palindromic sequence called a Fur box. The irr gene from Bradyrhizobium japonicum is under the control of Fur. Here, we provide evidence that B. japonicum Fur (BjFur) binds to the irr gene promoter with high affinity despite the absence of DNA sequence similarity to the Fur box consensus. Both Escherichia coli Fur and BjFur bound a synthetic Fur box consensus DNA element in electrophoretic gel mobility shift assays, but only BjFur bound the irr promoter. BjFur maximally protected a 30-bp region in DNase I footprinting analysis that includes three imperfect direct repeat hexamers. BjFur formed a high mobility complex and a low mobility complex with DNA in electrophoretic gel mobility shift assays corresponding to occupancy by a single dimer and two dimers or a tetramer, respectively. A mutation in the downstream direct repeat DNA sequence allowed high mobility complex formation only. In vitro transcription from the wild type irr promoter or from a mutated promoter that allowed only dimer occupancy was repressed by Fur, indicating that the dimer can be a functional repressor unit. Our findings identify a novel DNA-binding element for Fur and suggest that the Fur box consensus may not completely represent the target sequences for bacterial Fur proteins as a whole. In addition, Fur binding to a target promoter is sufficient to repress transcription in vitro.