Downregulation of Neuronal and Dendritic Connexin36-Made Electrical Synapses Without Glutamatergic Axon Terminals in Spinal Anterior Horn Cells From the Early Stage of Amyotrophic Lateral Sclerosis

Connexin36 (Cx36) forms gap junctions between neurons, which are called electrical synapses, enabling adjacent neurons to communicate directly. The participation of chemical synapses in neurodegeneration in amyotrophic lateral sclerosis (ALS) has long been indicated, but it remains unclear whether electrical synapses are involved in the pathogenesis of ALS. We performed extensive immunopathological analyses using mutant superoxide dismutase 1 (SOD1G93A) transgenic mice and their littermates to investigate whether Cx36-made electrical synapses are affected in motor neuron diseases. We found that in the lamina IX of the lumbar spinal cord from wild type mice, about half of the Cx36 puncta existed independently of chemical synapse markers, while the rest coexisted with chemical synapse markers, such as vesicular glutamate transporter 1 (VGLUT1), which is a glutamatergic axon terminal marker, and/or glutamate decarboxylase 65 (GAD65), which is a GABAergic axon terminal marker. Cx36 single or Cx36/GAD65 double positive puncta, but not VGLUT1-containing puncta, were preferentially decreased on neuronal and dendritic surfaces of the anterior horn cells in the early stage of SOD1G93A ALS mice. Moreover, in five human autopsied sporadic ALS cases with bulbar or upper limb onset, Cx36 immunoreactivity was diminished in the proximal dendrites and neuropils of well-preserved large motor neurons in the lumbar anterior horns. These findings suggest that downregulation of neuronal and dendritic Cx36 in the spinal anterior horns commonly occurs from the early stage of hereditary and sporadic ALS. Cx36-made electrical synapses without glutamatergic signaling appear to be more vulnerable than other chemical synapses and electrical synapses with glutamatergic signaling in the early stage of motor neuron degeneration, suggesting involvement of Cx36-made electrical synapses in the pathogenesis of human ALS

Remote Stereoinduction in the Organocuprate-Mediated Allylic Alkylation of Allylic <i>gem</i>-Dichlorides: Highly Diastereoselective Synthesis of (<i>Z</i>)‑Chloroalkene Dipeptide Isosteres

Highly diastereoselective synthesis of (<i>Z</i>)-chloroalkene dipeptide isosteres has been achieved by 1,4-asymmetric induction in the organocuprate-mediated allylic alkylation adjacent to the chiral center of allylic <i>gem</i>-dichlorides. The reaction proceeds with a variety of heterocuprates prepared from CuCN and various organometallic reagents. It allows rapid construction of valuable architectures of l,d-type and l,l-type (<i>Z</i>)-chloroalkene dipeptide isosteres from the corresponding (<i>E</i>)- and (<i>Z</i>)-allylic <i>gem</i>-dichlorides in high yields, with excellent (<i>Z</i>)-selectivity and diastereoselectivity

Delivery of a Proapoptotic Peptide to EGFR-Positive Cancer Cells by a Cyclic Peptide Mimicking the Dimerization Arm Structure of EGFR

A cyclic decapeptide, CQTPYYMNTC (<b>1</b>), which mimics the dimerization arm of the EGF receptor (EGFR), was previously found to be captured into cells. We have sought to investigate the promising potential of this peptide as an intracellular delivery vehicle directed to EGFR-positive cells. The selectivity of peptide <b>1</b> to the EGFR was confirmed by a positive correlation between the expression level of the receptor and the cellular uptake of peptide <b>1</b> as shown by siRNA knockdown of the EGFR. The proapoptotic domain (PAD) peptide ([KLAKLAK]₂) has limited use due to a deficiency of cell membrane permeability resulting from cationic sequences and lack of specificity for cancer cells. As a proof-of-concept study, the cellular delivery of the PAD peptide was challenged by conjugation with peptide <b>1</b>. The cellular uptake of a conjugated peptide <b>2</b>, which was composed of peptide <b>1</b>, the PAD peptide, and a linker cleavable with a protease, was evaluated by treatment of an EGFR-positive lung carcinoma cell line, A549. Significant suppression of proliferation by peptide <b>2</b> was shown in the results of a cell viability assay. The PAD peptide alone had no effect on the cells. The results suggest that peptide <b>1</b> is a promising lead compound as a new intracellular delivery vehicle for therapeutically effective peptides

Stereoselective Formation of Trisubstituted (<i>Z</i>)‑Chloroalkenes Adjacent to a Tertiary Carbon Stereogenic Center by Organocuprate-Mediated Reduction/Alkylation

A robust and efficient method for the synthesis of trisubstituted (<i>Z</i>)-chloroalkenes is described. A one-pot reaction of γ,γ-dichloro-α,β-enoyl sultams involving organocuprate-mediated reduction/asymmetric alkylation affords α-chiral (<i>Z</i>)-chloroalkene derivatives in moderate to high yields with excellent diastereoselectivity, and allylic alkylation of internal allylic <i>gem</i>-dichlorides is also demonstrated. This study provides the first examples of the use of allylic <i>gem</i>-dichlorides adjacent to the chiral center for novel 1,4-asymmetric induction

Synthesis of a Chloroalkene Dipeptide Isostere-Containing Peptidomimetic and Its Biological Application

The first rapid and efficient chemical synthesis of a cyclic Arg-Gly-Asp (RGD) peptide containing a chloroalkene dipeptide isostere (CADI) is reported. By a developed synthetic method, an <i>N</i>-<i>tert</i>-butylsulfonyl protected CADI was obtained utilizing diastereoselective allylic alkylation as a key reaction. This CADI was also transformed into an <i>N</i>-Fmoc protected CADI in a few steps. The CADI was used in Fmoc-based solid-phase peptide synthesis. The first synthesis of a CADI-containing cyclic RGD peptide was successful, and the synthesized CADI-containing peptidomimetic was found to be a more potent inhibitor against integrin-mediated cell attachment than the parent cyclic peptide