CASE.1

Manufacture of gRNA used in the CRISPR-Cas9 system with 86.5% purity

Sumitomo has established its standard manufacturing method (SMM) for gRNAs, which is applicable to gRNAs ranging in length from 95 mer to 131 mer. As the SMM enables direct scale-up from a single laboratory-scale feasibility run to GMP production without trial batches, customers can benefit from time and cost savings.

Through this case presentation, we aim to highlight our achievements in the first GMP production at the brand-new oligonucleotides plant in Oita.

Background

Typically, scaling up from laboratory to GMP manufacturing is a time-consuming process. Specifically, trial batches before GMP production have a significant impact on the cost and timeline, and this may lead to a delay in development.

To address this critical issue, Sumitomo developed the SMM, which enables us to dispense with trial batches when scaling up from laboratory scale to GMP production.

In the development of genome editing for therapeutic applications, customers prefer a tight schedule from gRNA sequence selection to clinical trial. Thus, the timeline from sequence selection to GMP production is of utmost importance.

Achievements

We achieved the first GMP manufacturing of a gRNA for the CRISPR-Cas9 system.

Purity of the product： 86.5%

It takes only 6 months from sequence disclosure to complete GMP manufacturing.

We are confident in our ability to quickly deliver GMP-grade gRNAs. Collaborating with us is the quickest way for customers to bring their products to the clinic.

Please feel free to contact us and share your schedule needs. We will make every effort to meet your required timeline.

CASE.2

130 mer with over 80% purity

Sumitomo excels in delivering unmatched purity for gRNAs, an achievement that sets us apart from others in the industry. While typical gRNAs used for the CRISPR-Cas9 system are around 100 mer in length, recent technological advancements require even longer gRNAs. Our gRNA chemical synthesis technology delivers gRNAs with exceptionally high purity that meets the most rigorous regulatory standards even for gRNAs with lengths exceeding 100 mer. Through this case presentation, we aim to highlight our achievements in GMP manufacturing of a 130 mer RNA with high purity.

Background

gRNA synthesis involves several processes such as:

Detritylation

Coupling

Oxidation

Capping

The synthesized RNA is then cleaved from the support, deprotected, purified, desalted, and isolated. For example, the synthesis of RNAs with 130 mer length involves a total of 520 (130 x 4) reactions, illustrating the significant challenge in synthesizing longer RNAs.

Achievements

We successfully completed GMP manufacturing without a single day’s delay.

The GMP production was a direct scale-up from a laboratory-scale feasibility run, without any trial batches.

Product purity: 82.5%

We are confident that we are the world’s first supplier to achieve such high purity for gRNAs of this length. Purity is a critical factor in the manufacture of gRNAs for therapeutic applications. With our cutting-edge technologies, we stand as the premier supplier for customers pioneering advanced genome-editing therapy.

CASE.3

155 mer laboratory sample with 81% purity

Sumitomo is dedicated to providing gene-editing biotechnology companies with unmatched high-purity gRNA. We understand the importance of meticulous planning for future GMP production, and we are here to support your endeavors at every step.

If you are struggling to find a CMO that can provide you with very long gRNAs （>110 mer）with high purity, try Sumitomo. We are ready and have the ability to meet your needs with precision and care. You can try our laboratory samples, evaluate the actual quality and purity of our products, and decide with confidence. The quality of our small-scale samples will be scalable to GMP production.

Background

In this case presentation, one of our clients had difficulty in obtaining high-purity gRNAs of length greater than 150 mer, and they finally approached Sumitomo. To enable the client to evaluate Sumitomo’s quality, we offered a free small-scale sample for evaluation. We also offered a free analysis of samples retained in the client so that they can grasp the whole picture towards reasonable vendor selection.

Achievements

Delivery: A sample was delivered within a 2-month timeframe from sequence disclosure

Product purity: Achieved 81% purity

Weight: About 10 mg of oligonucleotides

Our unique technologies enable us to supply gRNAs that far exceed 100 mer length with unparalleled purity. Our delivery timeline for small samples like the one described above is usually about 2 months after sequence disclosure.

We invite you to experience our gRNA chemical synthesis technology firsthand.

Sumitomo is ready to facilitate a seamless transition to large-scale GMP production when the time is right for your company.

For more information or to place an order, please do not hesitate to contact us through our website. We are confident that Sumitomo's technology will add significant value to your endeavors.

CASE.4

Laboratory production of 200 mer gRNA

Sumitomo has been supplying gRNAs with the world's highest purity. To further contribute to rapidly advancing genome-editing technology, Sumitomo is taking on the challenge of synthesizing gRNAs with lengths far exceeding 100 mer.

In the synthesis of gRNA with 100 mer length, even a 0.1% difference in the cycle yield has a significant impact on the total yield. With 200 mer, the impact greatly increases. In this case presentation, we would like to showcase Sumitomo's achievement in the synthesis of high-quality 200 mer RNA, for the first time in the world.

Background

Three key elements are required for the synthesis of 200 mer RNAs. To improve the one-cycle yield, we have achieved them all.

High-quality amidites

High-quality solid supports

An appropriate analysis method

Achievements

HPLC purity result of 74%

Differentiation of a 1 mer difference around the 200 mer range by our LC analysis method

This successful result indicates the exceptional performance of our PMM amidite and solid supports. In this instance, we developed a brand-new solid support for the efficient synthesis of 200 mer RNA. While CPG is well-known solid phase support for long RNAs, Sumitomo newly developed the ZEO support, which overcomes the drawbacks of the CPG support.

Without appropriate analytical methods, confirming the success of the synthesis is impossible. In fact, by analyzing the mixture of 5’-truncated impurities and 200 mer RNA, we confirmed that the conventional Sumitomo LC-method cannot separate 199 mer and 200 mer.

Therefore, we started to optimize the LC-method for 200 mers and achieved the separation of N-1 from 200 mers. To prove our successful synthesis, we synthesized RNAs with lengths ranging from 196 mer to 200 mer, with a difference of 1 mer at the 5' end, and simultaneously loaded them into the LC column to confirm the suitability of our analysis method. The five distinct peaks corresponding to the 196 mer to 200 mer RNAs indicate that our LC analysis method can differentiate a 1 mer difference around the 200 mer range.

In the future, next-generation RNA-based technologies may require even longer RNAs. Moving forward, we will verify that the laboratory results are well-reproduced by large-scale GMP equipment and strive to further contribute to the potential of genome editing.