Company: GAINI
Filing Date: 2025-11-06
Form Type: 424B2
Source: 0001193125-25-269767
Chunk: 2

Company: GLADSTONE INVESTMENT CORPORATION\DE
Filing Date: 2025-11-06
Form: 424B2
Chunk 2
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703) 287-5893or on the Investors section of our corporate website located at www.gladstoneinvestment.com, which, except for the documents incorporated by reference into this prospectus supplement and the accompanying prospectus, is not part of this prospectus supplement nor the accompanying prospectus. You may also call us collect at this number to request other information or to make an investor inquiry. See “ Where You Can Find More Information” on page S-39of this prospectus supplement. The SEC has not approved or disapproved of these securities or determined if this prospectus supplement or the accompanying prospectus is truthful or complete. Any representation to the contrary is a criminal offense.

|                                                     |     | Per Note |         |   |     | Total |            |
|:----------------------------------------------------|:----|:---------|--------:|:--|:----|:------|-----------:|
| Public offering price                               |     |          | 100.000 | % |     | $     | 60,000,000 |
| Sales load (underwriting discounts and commissions) |     |          |   1.500 | % |     | $     |    900,000 |
| Proceeds, before expenses, to us(1)                 |     |          |  98.500 | % |     | $     | 59,100,000 |

| (1) | Total expenses of the offering payable by us, excluding underwriting discounts and commissions, are estimated to be $0.4 million. See “Underwriting” on pageS-31of this prospectus supplement. |

Delivery of the Notes in book-entry form only through The Depository Trust Company (“DTC”) will be made on or about November 10, 2025. Sole Book-Running Manager B. Riley Securities The date of this prospectus supplement is November 6, 2025

TABLE OF CONTENTS

|                                                                                       |     | Page |      |
| Prospectus Supplement                                                                 |     |      |      |
| About this Prospectus Supplement                                                      |     |      | S-ii |
| Prospectus Supplement Summary                                                         |     |      |  S-1 |
| The Offering                                                                          |     |      |  S-4 |
| Supplementary Risk Factors                                                            |     |      |  S-9 |
| Special Note Regarding Forward-Looking Statements                                     |     |      | S-13 |
| Use of Proceeds                                                                       |     |      | S-14 |
| Capital